The quantification of human DNA extracts from forensic samples plays a key role in the forensic genetics process. Quantification guarantees maximum efficiency and avoids repeated analyses, over amplified samples, or unnecessary examinations. Here, we present our experience using an extremely sensitive real-time-based method with the Quantifiler® Trio DNA quantification kit, which we have explored in the ISO/IEC17025 accredited test validation process conducted in our laboratory. In this study, our focus was on establishing a minimum threshold beyond which DNA typing is unnecessary, by defining the correlation of negative quantification results with the absence of genetic profiles resulting from DNA typing. We investigated the results in the range in which the technique is used (50-0.005 ng/μl). In the validation process of the internally accredited method, we used data from the commercial quantification system, based on the average values of the standard curve using a wide range of samples, including artificially degraded samples. This method was extensively used continuously for nine years to quantify the DNA extract coming from a wide range of samples from criminal cases. The validation allowed us to verify the limits of the technique, especially in the use of the minimum quantities necessary to obtain genetic profiles acceptable for sending to the Italian DNA database. This optimization of the protocol for analysing forensic samples allows us to optimise the protocol for analysing forensic samples. However, the technique is used in our laboratory with a completely manual preparative phase, and we have decided to investigate the contribution of human error more accurately. Thus, in our experience, we have found that it is preferable to use quantification ranges rather than exact limits before deciding how to analyse the extracts via PCR. In addition, here we present some preliminary data related to the analysis of certain samples that initially appeared negative during quantification, applying a concentration method for maximum yield from forensic samples of judicial interest. Our aim is to ensure that no source of evidence is lost in the reconstruction of a criminal event.

Recently concluded, large-scale cancer genomics studies involving multiregion sequencing of primary tumors and paired metastases appear to indicate that many or most cancer patients have one or more “clonal" mutations in their tumors. Clonal mutations are those that are present in all of a patient’s cancer cells. Achilles Therapeutics is currently the only company specifically targeting clonal mutations. However, they are doing so with tumor-derived T cells. To address the potential limitations of immunotherapy, I have devised another approach for exploiting clonal mutations, which I call “Oncolytic Vector Efficient Replication Contingent on Omnipresent Mutation Engagement” (OVERCOME). The ideal version of OVERCOME would likely employ a bioengineered facultative intracellular bacterium. The bacterium would initially be attenuated, but (transiently) reverse its attenuation upon clonal mutation detection.

Fusarium stalk rot (FSR) caused by the Fusarium species complex is an economic threat to maize cultivation all over the world. We investigated the population structure and genetic diversity of Fusarium spp. obtained from five major maize-growing regions of India. The Tef-1α locus was used for phylogenetic analysis of geographically distinct isolates of Fusarium verticillioides, F. andiyazi, F. proliferatum, F. nygamai, and F. acutatum causing FSR. Geographical separation among four local populations contributed to 7.87% variance, whereas 92.12% of the variance was within popula-tions, suggesting a predominant influence of local adaptation or stochastic events like genetic drift within populations, with geographical separation playing a lesser but significant role in shaping genetic diversity. Genetic differentiation statistics between Fusarium species showed lower gene flow from haplotypes except in the population of F. acutatum and F. andiyazi having high GST values. In contrast, a high Kxy was reported, indicating elevated genetic differentiation between populations. The haplotype network analysis revealed the presence of eight distinct haplotypes which reflected pathogenic evolution and adaptive potential of Fusarium spp. The results offer a comprehensive framework for discussing the implications of genetic diversity in pathogen management and the evolutionary dynamics of the Fusarium spp. causing FSR of maize in the Indian subcontinent.

Abnormalities in sperm nuclei and chromatin can interfere with normal fertilization, embryonic development, implantation, and pregnancy. We aimed in this study to determine the impact of H2BFWT gene variants in sperm DNA on ICSI outcomes in couples undergoing ART treatment. One hundred and nineteen partners were divided into pregnant (G1) and non-pregnant (G2) groups. After semen analysis, complete DNA was extracted from purified sperm samples. The sequence of the H2BFWT gene was amplified by PCR and then subjected to Sanger sequencing. The results showed that there are three mutations in this gene: rs7885967, rs553509 and rs578953. rs553509 and rs578953 showed significant differences in the distribution of alternative and reference alleles between G1 and G2 (P = 0.0004 and P = 0.0020, respectively). However, there was no association between these SNPs and the studied parameters. This study is the first to shed light on the connection between H2BFWT gene variants in sperm DNA and pregnancy after ICSI therapy. This is a pilot study, so further investigations about these gene variants at the transcriptional and translational levels, in a larger cohort of patients, are needed to determine its functional consequences and to understand the mechanism of how the pregnancy is affected by sperm DNA.

We previously reported nano-pulse treatment (NPT), a pulsed power technology, resulted in 4T1-luc mammary tumor elimination and a strong in situ vaccination, thereby completely protecting tumor-free animals against a second live tumor challenge. The mechanism whereby NPT mounts effective antitumor immune responses in the 4T1 breast cancer predominantly immunosuppressive tumor microenvironment (TME) remains unanswered. In this study, orthotopic 4T1 mouse breast tumors were treated with NPT (100 ns, 50 kV/cm, 1000 pulses, 3 Hz). Blood, spleen, draining lymph nodes and tumors were harvested at 4-hour, 8-hour, 1-, 3-, 7-day. Additional studies were conducted over 3-month posttreatment intervals for the analysis of frequencies, death and functional markers of various immune cells in addition to suppressor function of regulatory T cells (Tregs). NPT was verified to elicit strong in situ vaccination (ISV) against breast cancer and promoted both acute and long-term T cell memory. NPT abolished immunosuppressive dominance systemically and in the TME by substantially reducing Tregs, myeloid-derived suppressor cells (MDSCs), and tumor-associated macrophages (TAMs). NPT induced apoptosis in Tregs and TAMs. It also functionally diminished the Treg suppression capacity, likely explained by the downregulation of activation markers, particularly 4-1BB and TGFβ, and a phenotypic shift from predominantly activated (CD44+CD62L-) to naïve (CD44-CD62L+) Tregs. Importantly, NPT selectively induced apoptosis in activated Tregs and spared effector CD4+ and CD8+ T cells. These changes were followed by a concomitant rise in CD8+CD103+ tissue-resident memory T cells and TAM M1 polarization. These findings indicate that NPT effectively switches the TME and secondary lymphatic systems from an immunosuppressive to an immunostimulatory state allowing cytotoxic T-cell function and immune memory formation to eliminate cancer cells and account for the NPT in situ vaccination.

Recently, a concept known as μTRISTAN, which involves the acceleration of μ+, has been proposed. This initiative has led to considerations of a new design for a neutrino factory. Additionally, leveraging the polarization of μ+, measurements of T violation in neutrino oscillations are also being explored. In this paper, we present analytical expressions for T violation in neutrino oscillations within the framework of standard three flavor neutrino oscillations, a scenario involving nonstandard interactions, and a case of unitarity violation. We point out that examining the energy spectrum of T violation may be useful for probing new physics effects.

A ~3-kb deletion-type DNA copy-number variation (CNV, esv3587290) located at intron 7 of the VANGL1 gene (1p13.1, MIM*610132) has been proposed as a genetic factor for developing lupus nephritis (LN) in adult systemic lupus erythematosus (SLE) patients across European-descent populations, but its replication in other ethnicities has been inconsistent and its association with LN in childhood-onset SLE (cSLE) remains unknown. Here, we performed an exploratory association study in a sample of 66 unrelated cSLE Mexican patients (11 males, 55 females; ages 7.8 to 18.6 years). Two stratified groups were compared: cSLE patients with (N=37) or without (N=29) LN, as diagnosed by renal biopsy (N=15), proteinuria (N=33), urinary protein:creatinine ratio >0.2 (N=34), and erythrocyturia and/or granular casts in urinary sediment (N=16). For esv3587290 CNV genotyping, we performed an end-point PCR assay with breakpoint confirmation by Sanger sequencing. We also determined the allelic frequencies of the esv3587290 CNV in 181 deidentified ethnically-matched individuals (reference group). The obtained genotypes were tested for Hardy-Weinberg equilibrium (HWE) by 2 test. Associations between LN and esv3587290 CNV were tested by calculating the Odds Ratio (OR) and using Pearson's 2 tests with a 95% confidence interval and p≤0.05. Contrary to the results described in European-descent populations, the esv3587290 CNV showed a significant protective effect against LN development (OR 0.14, 95% CI 0.066-0.31, p≤0.05), whereas the wild-type homozygous VANGL1 genotype showed a risk trend for LN development (OR 2.15, 95% CI 0.7711-6.18, p≤0.05). Finally, we characterized the precise breakpoint of the esv3587290 CNV as NG_016548.1(NM_138959.3):c.1314+1339_1315-897del (https://databases.lovd.nl/shared/variants/0000918418#00025811) in our population. This report supports the notion that a broad genetic heterogeneity underlies the susceptibility for developing LN.

Lymphocyte collection by apheresis for CAR-T production usually does not include mobilized blood using Granulocyte Colony Stimulating Factor (G-CSF) due to the widespread knowledge that it causes a decrease in the number and functionality of lymphocytes. However, it is used for stem cell transplant, which is a common treatment for hematological malignancies. The growing demand for CAR therapies (CAR-T and NK-CAR), both in research and clinic, makes necessary to evaluate whether mobilized PBSC products may be potential candidates for use in such therapies. This review collects recent work that experimentally verifies the role and functionality of T and NK lymphocytes, and the generation of CAR-T, from apheresis after G-CSF mobilization. As discussed, T cells do not vary significantly in their phenotype, the ratio of CD4+ and CD8+ remains constant and the different sub-populations remain stable. In addition, its expansion and proliferation rate are invariant regardless mobilization with G-CSF, as well as the secretion of proinflammatory cytokines and the cytotoxic ability. Therefore, cells mobilized before apheresis are postulated as a new alternative source of T cells for adoptive therapies that will serve to alleviate high demand, increase availability, and take advantage of the substantial number of existing cryopreserved products.

Type 2 diabetes mellitus (T2DM) is a risk factor for male infertility, but the underlying molecular mechanisms remain unclear. Advanced glycation end products (AGEs) are pathogenic molecules for diabetic vascular complications. Here, we investigated the effects of DNA-aptamer raised against AGEs (AGE-Apt) on testicular and sperm abnormalities in a T2DM mouse model. KK-Ay (DM) and wild-type (non-DM) 4- and 7-week-old male mice were sacrificed to collect the testes and spermatozoa for immunofluorescence, RT-PCR, and histological analyses. DM and non-DM 7-week-old mice were subcutaneously infused with AGE-Apt or control aptamer for 6 weeks and then sacrificed. Plasma glucose, testicular AGEs, and RAGE gene expression in 4-week-old DM mice and plasma glucose, testicular AGEs, oxidative stress, and pro-inflammatory gene expressions in 7-week-old DM mice were higher than those in age-matched non-DM mice; the latter of which was associated with seminiferous tubular dilation. AGE-Apt did not affect glycemic parameters, but it inhibited the seminiferous tubular dilation, reduced the number of testicular macrophages and apoptotic cells, and restored the decrease in sperm concentration, motility, and viability of 13-week-old DM mice. Our findings suggest that AGEs-Apt may improve sperm abnormality by suppressing the AGE–RAGE-induced oxidative stress and inflammation in the testes of DM mice.

Mitochondrial DNA copy number (mtDNAcn) is associated with mitochondrial function contributing various diseases. Understanding of the association between mtDNAcn and infant neurodevelopment, as well as the role of racial disparities, remains unknown. A cohort study was conducted with 55 preterm infants and a single blood sample was collected from each infant during their NICU stay to measure mtDNAcn. NICU Network Neurobehavioral Scale (NNNS) and Bayley Scale of Infant Developmental testing Edition III was assessed during NICU until 2 years of corrected age (CA). Linear regression models were performed to investigate the relationship between the clinical characteristics, neurobehavioral outcomes and mtDNAcn. Majority of our subjects were male, White, non-Hispanic, had C-section, and without preterm premature rupture of membrane (PPROM). Increased mtDNAcn was found associated with younger birth gestational age (GA), nonoptimal neurodevelopment. However, the opposite associations between mtDNAcn and neurodevelopmental outcomes were observed between Black and White infants up to 1 year of CA. Adverse early life experience, together with increased mtDNAcn in White infants, and decreased mtDNAcn in Black infants may be considered as significant positive predictors of poor early life neurodevelopmental outcomes in infants.

The risk of malignant transformation of induced pluripotent stem cells (iPSC)-derived cells in response to DNA damage is a potential concern as the tumor suppressor genes TP53 and CDKN2A are transiently inactivated during reprogramming. Here, we evaluated the integrity of cellular senescence pathways and DNA double-strand break (DSB) repair in iPSC-derived human fibroblasts (i-HF) compared to their parental skin fibroblasts (HF). Using a variety of functional assays, we show that the capacity of i-HF to enter senescence and repair DSB is not compromised after damage induced by ionizing radiation (IR) or overexpression of H-RASV12. Still, i-HF lines are transcriptionally different from their parental lines, showing enhanced metabolic activity and higher expression of p53-related effector genes. As a result, i-HF lines generally exhibit increased sensitivity to various stresses, have an elevated senescence-associated secretory phenotype (SASP) and cannot be immortalized unless p53 expression is knocked down. In conclusion, while our results suggest that i-HF are not at a greater risk of transformation, their overall hyperactivation of senescence pathways may impede their function as a cell therapy product.

There is significant need for development of a new environmentally friendly, extraction-free sample collection medium that can effectively preserve and protect genetic material for point-of-care, and/or self-collection, home-collection, and mail-back testing. Systematic Evolution of Ligands by Exponential Enrichment (SELEX) was used to create anti-ribonuclease (RNase) Deoxyribonucleic acid (DNA) aptamers against purified RNase A conjugated to paramagnetic carboxylated beads. Following 8 rounds of SELEX carried out under various stringency conditions, e.g., selection using Xtract-Free™ (XF) specimen collection medium, and elevated ambient temperature of 28 ºC, a panel of 5 aptamers was chosen following bioinformatic analysis using next-generation sequencing. The efficacy of aptamer inactivation of RNase was assessed by monitoring Ribonucleic acid (RNA) integrity by fluorometric and real-time RT-PCR analysis. Inclusion of aptamers in reaction incubations resulted in an 8,800 to 11,200-fold reduction of RNase activity, i.e., digestion of viral RNA compared to control. Thus, anti-RNase aptamers integrated into XF collection medium as well as other commercial reagents and kits have great potential for ensuring quality intact RNA for subsequent genomic analysis.

Retinoblastoma (Rb) is a rare cancer that nonetheless represents the most common ocular malignancy in children. Rb exists as familiar and sporadic cancer and the increasing incidence of the sporadic one and its pathogenesis is mostly unknown. To date the management of retinoblastoma requires intensive chemotherapy and sometimes invasive surgery in order to preserve vision and quality of life in Rb patients. The discovery of the epigenetic nature of non-hereditable/sporadic Rb led to the development of high-throughput precision investigation strategies for the study of its epigenetic landscape. Here we investigated the effect of the demethylating 5-Aza-2’- deoxycytidine (Decitabine; DAC) agent on preclinical models to assess its possible use as anti-cancer drug in the treatment of Rb patients. We found that DAC treatment induced cell cycle arrest and apoptosis in Weri-Rb-1 cells. We investigated the variation of differential gene expression profile in Rb cells in relation to time of exposure to DAC with respect to controls. Network map analysis evidences 15 hub/driver genes, which share pathways of regulation of TNF-, FAS-, p53-dependent apoptotic signaling and NF-kB pro-survival pathways. Furthermore, a computing expression profile analysis of publicly available datasets of patients’ primary tumors and normal retina cell-derived organoids vs WeriWeri-Rb-1 DEGs depicted the relevancy of these 15 hub genes on retinoblastoma genesis. The antiproliferative effect of DAC treatment on Rb was confirmed in vivo, both in subcutaneous xenograft and orthotopic models also showing the changes the gene methylation status in treated Weri-Rb-1, shedding a new light on the reprogramming antiproliferative dynamics induced by the epigenetic therapy and on the role of this therapeutic approach to spare children sight and life.

Early detection of neurological conditions is critical for timely diagnosis and treatment. Identifying cellular-level changes is essential for implementing therapeutic interventions prior to symptomatic disease onset. However, monitoring brain tissue directly through biopsies is invasive and high-risk. Bodily fluids such as blood or cerebrospinal fluid contain information in many forms, including proteins and nucleic acids. Particularly, cell-free DNA (cfDNA) has potential as a versatile neurological biomarker. Yet, our knowledge of cfDNA released by brain tissue and how cfDNA changes in response to deleterious events within the brain is incomplete. Mapping changes in cfDNA to specific cellular events is difficult in vivo, where many tissues contribute to circulating cfDNA. Organoids are tractable systems for examining specific changes consistently in a human background. However, few studies have investigated cfDNA released from organoids. Here, we examined cfDNA isolated from cerebral organoids. We found that cerebral organoids release quantities of cfDNA sufficient for downstream analysis with droplet-digital PCR and whole-genome sequencing. Further, gene ontology analysis of genes aligning with sequenced cfDNA fragments revealed association with terms related to neurodevelopment and autism spectrum disorder. We conclude that cerebral organoids hold promise as tools for the discovery of cfDNA biomarkers related to neurodevelopmental and neurological disorders.

KLEPTOSE® CRYSMEB is a methylated cyclodextrin derivative displaying less methylated group substitution than randomly methylated cyclodextrin. It has demonstrated an impact on atherosclerosis and neurological diseases. linked in part to cholesterol complexation and immune response, but the impact on inflammatory cascade pathways was not clear. Thus, the impact of KLEPTOSE® CRYSMEB on various pharmacological targets was assessed using human umbilical vein endothelial cells in physiological and inflammatory conditions, followed by screening against twelve human primary cell-based systems designed to model complex human tissue and disease biology of the vasculature, skin, lung, and inflammatory tissues using the BioMAP® Diversity PLUS® Panel. Finally, the anti-inflammatory mechanism was investigated on peripheral blood mononuclear cells to evaluate anti-inflammatory or pro-resolving properties. The results showed that KLEPTOSE® CRYSMEB can modulate the immune system in vitro and potentially manage vascular issues by stimulating the expression of molecules involved in the crosstalk between immune cells and other cell types. It showed anti-inflammatory effects driven by pro-inflammatory cytokine secretion inhibition and could have different impacts on different tissue types. Moreover, this cyclodextrin showed no clear impact on pro-resolving lipid mediators. Finally, it appeared that the mechanism of action of KLEPTOSE® CRYSMEB seems to not be shared by well-known anti-inflammatory molecules.

Malaria parasites increase their host erythrocyte’s permeability to obtain essential nutrients from plasma and facilitate intracellular growth. In the human P. falciparum pathogen, this increase is mediated by the plasmodial surface anion channel (PSAC) and has been linked to CLAG3, as encoded by one member of the conserved clag multigene family. Whether paralogs encoded by other clag genes also contribute to PSAC formation or serve unrelated roles remains unknown. Here, we generated transfectant lines carrying epitope-tagged versions of each CLAG. Each paralog colocalized with CLAG3, with concordant trafficking via merozoite rhoptries to the host erythrocyte membrane of newly-invaded erythrocytes. Each also exists within infected cells in at least two forms: an alkaline-extractable soluble form and a form integral to the host membrane. Like CLAG3, CLAG2 has a variant region cleaved by extracellular protease but CLAG8 and CLAG9 are protease-resistant. Paralog knockout lines, generated through CRISPR/Cas9 transfection, exhibited uncompromised growth in PGIM, a modified medium with more physiological nutrient levels; this finding is in marked contrast to a recently reported CLAG3 knockout parasite. CLAG2 and CLAG8 knockout lines exhibited compensatory increases in transcription of remaining clags and associated rhoph genes, yielding increased PSAC-mediated uptake to specific solutes. We also report distinct transport properties of these knockout lines, suggesting partially overlapping roles of CLAG paralogs in PSAC activity. We propose that the nutrient channel pore is formed by a higher order assembly of CLAG proteins at the host membrane.

Public transportation scheduling aims to optimize the allocation of transportation resources, enhance transportation efficiency, and increase passenger satisfaction, which is crucial for building a sustainable urban transportation system. As a complement to public transportation, shared bikes provide users with a solution for the last mile of travel, compensating for the lack of flexibility in public transportation and helping to improve its utilization rate. Due to the characteristics of shared bikes, such as peak usage periods in the morning and evening and significant fluctuations in demand across different areas, the optimization of shared bike dispatch can better meet user needs, thereby reducing vehicle vacancy rates and increasing operating revenue. Addressing this issue, this paper designs a comprehensive decision-making approach for spatio-temporal demand prediction and bike dispatch optimization. For demand prediction, we designed a T-GCN based bike demand prediction model. In terms of dispatch optimization, we considered factors such as dispatch capacity, distance restrictions, and dispatch costs, and designed an optimization solution based on genetic algorithms. Finally, this paper validates the approach using shared bike operating data, shows that T-GCN can effectively predict short-term demand for shared bikes. Meanwhile, the optimization model based on genetic algorithms provides a complete dispatch solution verifying the model’s effectiveness. The shared bike dispatch approach proposed in this paper, which combines demand prediction with resource scheduling. This scheme can also be extended to other transportation scheduling problems with uncertain demand, such as store replenishment delivery and intercity inventory dispatch.

Sepsis is a life-threatening syndrome triggered by infection and accompanied by high mortality, with antimicrobial resistances (AMRs) further escalating clinical challenges. Rapid and reliable detection of causative pathogens and AMRs are key factors for fast and appropriate treatment, in order to improve outcome in septic patients. However, current sepsis diagnostics based on blood culture is limited by low sensitivity and specificity while current molecular approaches fail to enter clinical routine. Therefore, we developed a Suppression PCR-based selective enrichment sequencing approach (SUPSETS), providing a molecular method combining multiplex suppression PCR with Nanopore sequencing to identify most common sepsis-causative pathogens and AMRs using plasma cell-free DNA. Applying only 1 mL of plasma, we target eight pathogens across three kingdoms and ten AMRs in a proof-of-concept study. SUPSETS was successfully tested on first ten clinical samples and revealed comparable results to clinical metagenomics while clearly outperforming blood culture. Several clinically relevant AMRs could be detected additionally. Furthermore, SUPSETS provided first pathogen and AMR-specific sequencing reads within minutes of starting sequencing, thereby potentially decreasing time to results to 11 - 13 hours and suggesting diagnostic potential in sepsis.

The enhancement of early diagnosis of lung cancer is mandatory to improve prognosis. We observed the increase cancer stem cells (CSC) and circulating tumor cells (CTC) in blood of mice on d3 after Lewis lung carcinoma (LLC) cell implantation before disease symptom. Besides we have developed an approach to reprogramming immune cells by inhibiting the MAPK/ERK pathway through MEKi and the PD-1/PD-L1 immune checkpoint signaling pathway. We hypothesized that reprogramming of spleen CD8+ T-cells could also create a population of immune cells with high antitumor activity. We reprogrammed CD8+ T-cells derived from the spleen of C57BL/6 mice (rsCD8+T-cells). The rsCD8+T-cells were able to migrate to lung and were more resistant after the exhaustion in vitro in comparison with naïve sCD8+T-cells. In the orthotopic LLC model, the rsCD8+T-cells therapy increased the amount of proliferating CD8+ and CD4+ T-cells in blood and lung from mice. The amount of CSC decreased in the blood and lung of mice treated with rsCD8+ T-cells. A morphological study revealed a decrease in the number of metastases in lung tissue. The antitumor effects of rsCD8+T-cells are based on the activation of the host immune response by increasing the populations of CD8+ and CD4+ T-cells and apoptosis of CSCs.

The sperm heads of frozen/thawed horse spermatozoa were injected into ovulated mouse oocytes where they decondensed and formed paternal pronuclei in which the DNA damage has been evaluated. The motile spermatozoa exhibit none or minor level of DNA damage and the xenogenic zygotes cleave to the two-cell embryonic stage with regular nuclei and no micronuclei (MNs). On the other hand, the immotile spermatozoa exhibit typically high levels of DNA damage and blastomeres in two-cell stage embryos contain multiple MNs. Moreover, extremely high sperm DNA damage prevents even the cleavage to the two-cell stage.Keywords: keyword 1; keyword 2; keyword 3 (List three to ten pertinent keywords specific to the article yet reasonably common within the subject discipline.)

The evolutionary cancer cell biology ECCB explains the deep homology existent between the life cycle of cancer and protists. Due to their origin and sensitivity to oxygen, all non-gametogenic germ lines and germ stem cells (GSCs), including human adult stem cells (ASCs) and cancer stem cells (CSCs), suffer severe DNADSB damage when exposed to stress oxygen content above 6.0% O2 (Urgermline hyperoxia) as occurring in tissue and bloodstream. As a result, GSCs/ASCs/ CSCs lose their stemness potential and convert from a functional asymmetric cyclic phenotype (ACD phenotype) to a dysfunctional symmetric phenotype capable of defective symmetric proliferation (DSCD) that is, however, unable to repair genomic degradation. Genome reconstruction occurs through evolutionary mechanisms evolved by the common ancestor of amoebozoans, metazoans, and fungi. They complete the cycle by removing the damage, reconstructing the functional genome, and reinstating the ASC phenotype. The stemness life cycle acts not only during carcinogenesis, tumorigenesis, and recurrence but also in the life cycle of protists.

Much research has been conducted to determine how hair regeneration is regulated, as this could provide therapeutic, cosmetic, and even psychological interventions for hair loss. The current study focused on the hair growth effect and effective utilization of fatty oil obtained from Bryde's whales through high-throughput DNA microarray approach in conjunction with immunohistochemical observations. The research also examined the mechanisms and factors involved in hair growth. In an experiment using female C57BL/6J mice, the vehicle control group (VC: propylene glycol: ethanol: water), the positive control group (MXD: 3% minoxidil), and the experimental group (WO: 20% Whale Oil) were topically applied to the back of the mouse. The results showed that 3% MXD and 20% WO were more effective than VC in promoting hair growth, especially 20% WO. Furthermore, in hematoxylin and eosin-stained skin tissue, an increase in the number of hair follicles and subcutaneous tissue thickness was observed with 20% WO. Whole-genome transcriptomic analysis also showed that 20% WO may have lower stress- and inflammation-related responses than 3% MXD. Therefore, whale oil can be expected to be used as a safe hair growth agent.

This study introduces the MuhdoAge clock, a novel saliva-based epigenetic clock, designed to predict biological age with strong correlations in a healthy, disease-free cohort. Leveraging a comprehensive dataset from Muhdo Health, this research analyses saliva samples utilising the Illumina MethylationEPIC array. Through robust statistical methods, 237 significant CpG sites were identified that correlate with age-related methylation changes, exhibiting a coefficient of determination (R²) of 0.652 after outlier removal, with tiered algorithmic design improving this further to 0.726 in a new dataset that combined pathologic and healthy individuals (n=2682). Comparatively, the MuhdoAge clock was evaluated against other known epigenetic clocks, demonstrating high precision in predicting biological age within healthy individuals (R² = 0.878, n=1844), emphasising its potential utility in non-invasive, large-scale applications. The clock shows promise for integration into public health strategies for monitoring and potentially mitigating age-related decline, facilitated by its ease of use and accessibility through saliva sampling. Furthermore, this study highlights the impact of lifestyle and environmental factors on biological ageing, highlighting the importance of personalised health interventions. Future research is to focus on longitudinal studies to validate these findings and explore the mechanisms underlying epigenetic age acceleration.

Abstract Precision medicine is essentially based on reliable and non-invasive biomarkers. Aim of this review has been to describe the newest biomarkers in the field of kidney transplantation and the kidney rejection, one of the commonest and severe complications. Standard of care tools to identify acute rejection are largely full of errors and of drawbacks. In recent years new and reliable biomarkers have been found. They are avoid of risks, non-invasive and able to detect rejection even in those frequent cases in which acute rejection are clinically asymptomatic and not otherwise identifiable. In recent years several biomarkers have been identified. Very recently have been found new relevant biomarkers able to diagnose rejection with high positive predictive value and low negative predictive value. These are the donor derived cell free DNA found in the recipient, the gene expression profile of the donor found in the recipient and the urinary cytokines that express modification in the graft tissue. The aim of this study has been to find the most recent findings in the literature on this topic and to describe the utility and possible limitation of such new biomarkers on kidney rejection.

Background: The effectiveness of the anti-COVID-19 vaccine remains a matter to be clarified. Methods: The humoral and cellular immunity after the administration of three doses of the Pfizer–BioNTech and Oxford AstraZeneca vaccines against SARS-CoV-2 over one year and the appearance of post-vaccination COVID-19 have been studied. Results: Anti-SARS-CoV-2 IgG and IgA antibodies showed a progressive increase throughout the vaccination period. This increase was greatest after the third dose. The highest levels were observed in subjects who had anti-SARS-CoV-2 antibodies prior to vaccination. There was an increase in CD4+ αβ, CD8+ γδ and TEM CD8+ γδ T cells, and a decrease in apoptosis in CD4+CD8+ and CD56+ αβ and γδ T cells. Post-vaccination SARS-CoV-2 infection was greater than 60%. The symptoms of COVID-19 were very mild and were related to γδ T cell deficit, specifically CD8+ TEMRA and CD56+ γδ TEM, as well as lower pre-vaccine apoptosis levels. Conclusions: Those results demonstrate the important role that γδ T cells play in SARS-CoV-2 vaccine immunization.

Banana bunchy top virus (BBTV) was recently detected in Mozambique and appears to be limited to the provinces of Gaza, Maputo and Zambezia, but it has great potential to spread to other provinces. Despite its importance, nothing is known about the BBTV isolates that occur in Mozambique. In this study, the sequences of the S and R genes of 40 isolates chosen as representatives of samples collected previously from eleven farms of the four administrative posts of the district of Chóckwè, Gaza province, were sequenced and analyzed. The S-DNA nucleotide sequences of the analyzed isolates were highly conserved, with identity ranging from 97% to 100%. The same was observed for the R-DNA sequences, with most identities ranging between 98% and 100% among the isolates from Chókwè and above 90% when compared to the isolates from GenBank. The phylogenetic analysis showed that the Mozambican BBTV isolates belong to the Pacific-Indian Oceans (PIO) group, showing greater proximity to the isolate JQ820453 from Malawi than to the isolates from sub-Saharan countries, which were grouped in a distinct subclade. This is the first study conducted to determine the molecular characteristics of BBTV isolates present in Mozambique.

Histone demethylases, enzymes responsible for removing methyl groups from histone proteins, have emerged as critical players in regulating gene expression and chromatin dynamics, thereby influencing various cellular processes. LSD2 and LSD1 have attracted considerable interest among these demethylases because of their associations with cancer. However, while LSD1 has received significant attention, LSD2 has not been recognized to the same extent. In this study, we conduct a comprehensive comparison between LSD2 and LSD1, with a focus on exploring LSD2's implications. While both share structural similarities, LSD2 possesses unique features. Functionally, LSD2 shows diverse roles, particularly in cancer, with tissue-dependent roles. Additionally, LSD2 extends beyond histone demethylation, impacting DNA methylation and DNA damage repair pathways. This study underscores the distinct roles of LSD2, providing insights into their contributions to cancer and other cellular processes.

Multiple sclerosis is an autoinflammatory condition that results in damage of myelinated neurons in affected patients. While disease-modifying treatments have been successful in slowing the progression of relapsing remittent disease, most patients still progress to secondary progressive disease that is largely unresponsive to disease-modifying treatments. Similarly, there is currently no effective treatment for patients with primary progressive MS. Innate and adaptive immune cells in the CNS play a critical role in initiating an autoimmune attack and in maintaining the chronic inflammation that drives disease progression. In this review, we will focus on recent insights into the role of T cells with regulatory function in suppressing the progression of MS, and, more importantly, in promoting the remyelination and repair of MS lesions in the CNS. We will discuss the exciting opportunity to genetically reprogram regulatory T cells to achieve immune suppression and enhance repair locally at sites of tissue damage, while retaining a fully competent immune system outside the CNS. In the future, reprogramed regulatory T cells with defined specificity and function may provide life medicines that can persist in patients and achieve lasting disease suppression after one cycle of treatment.

L-Theanine, a unique non-protein amino acid, is an important bioactive component of green tea. Previous studies have shown that L-theanine has many potent health benefits, such as anti-anxiety effect, regulation of immune response, relaxing neural tension, reducing oxidative damage. However, little is known concerning whether L-theanine can improve the clearance of mitochondrial DNA (mtDNA) damage in organisms. Here, we reported that L-theanine treatment increased ATP production and improved mitochondrial morphology to extend the lifespan of UVC-exposed nematodes. Mechanistic investigations showed that L-theanine treatment enhanced the removal of mtDNA damage and extended lifespan by activating autophagy, mitophagy, mitochondrial dynamics, and mitochondrial unfolded protein response (UPRmt) in UVC-exposed nematodes. In addition, L-theanine treatment also up-regulated the expression of genes related to mitochondrial energy metabolism in UVC-exposed nematodes. Our study provided a theoretical basis for the possibility that tea drinking may prevent mitochondrial related diseases.

Triple-negative breast cancer (TNBC) is an aggressive and highly metastatic type of tumor. TNBC is often enriched in tumor-infiltrating neutrophils (TINs), which support cancer growth in part by counteracting tumor-infiltrating lymphocytes (TILs). Prior studies identified the enhancer of zeste homolog 2 (EZH2) as a pro-tumor methyltransferase in primary and metastatic TNBC. We hypothesized that EZH2 inhibition in TNBC cells per se would exert anti-tumor activity by altering the tumor immune microenvironment. To test this hypothesis, we used CRISPR to generate EZH2-knockout (KO) and overexpressing lines from parent (wild-type -WT) 4T1 cells, an established murine TNBC model. In vitro, EZH2 KO and OE cells showed little change in replicative capacity and invasiveness, but marked changes in surface marker profile and cytokine/chemokine secretion compared to WT cells. On in vivo injection, EZH2 KO cells showed significantly reduced primary tumor growth and a 10-fold decrease in lung metastasis compared to WT cells, while EZH2 OE cells were unchanged. Compared to WT tumors, TIN:TIL ratios were greatly reduced in EZH2 KO tumors but unchanged in EZH2 OE tumors. Our findings show a tumor-intrinsic role of EZH2 in the 4T1 TNBC model in regulating TIN/TIL poise, primary tumor progression and lung metastasis in vivo.

As new vaccines reduce P. falciparum (Pf) cases, clinicians can focus on rescuing more children with cerebral malaria (CM). Adjunctive “therapeutically-rational exchange transfusion” (T-REX) refers to using special “Pf-resistant” donor red blood cells (RBCs) that secrete nano-sized RBC extracellular vesicles (EVs). Can some RBC EVs rapidly dislodge Pf-infected RBCs (iRBCs) sequestered in blood vessels and reduce inflammation? Does post-exchange “rebound parasitemia” reflect sequestration-reversal? We reviewed case-reports, EV and liposome studies, expert perspectives, and time-sequence data. In the AQUAMAT study, median coma-recovery time was 20 hours for artesunate. In contrast, a CM patient treated with type-O T-REX quickly began speaking (during the procedure). For other rapid-recovery CM patients, donor RBC variables had not been specified. Despite rapid Pf-killing, drugs need 20 hours to substantially reduce iRBC cytoadhesion while EVs can selectively bind distant inflamed tissues within 15 minutes. RBCs stressed by blood-bank storage and Pf infection secrete EVs. Post-exchange “rebound parasitemia” can occur within 60 minutes and may reflect release of sequestered iRBCs. Sequestration-reversal can be assessed by monitoring Pf parasitemia and total Pf biomass. New data suggest clinicians, working with transfusion- and laboratory-medicine physicians, can, and should, assess T-REX options and publish their findings to help clarify how RBC EVs may serve as therapeutic “decoy ligands.”

The mechanism of immune activation and de-activation is very important. Host should trigger effective immunity against pathogen infection. And, host cannot trigger immunity against self-antigens that will lead to detrimental autoimmune disorders. Spleen and liver have opposite roles in mediating immune responses. Spleen is the organ involving in immune activation, and liver is the organ involving in immune de-activation. Sympathetic nerve system and parasympathetic nerve system have opposite roles in mediating immune responses. Sympathetic nerve system is involving in pro-inflammatory, and parasympathetic nerve system is involving in anti-inflammatory. ACTH and glucocorticosteroid have opposite roles in mediating immune responses. ACTH is involving in activating adaptive immunity with inhibiting innate immunity, and glucocorticosteroid is involving in activating innate immunity with inhibiting adaptive immunity. The molecular chaperons, heat shock proteins, which are induced by fever, have the vital roles in immune activation. On the contrary, IgD B cells and gamma-delta T cells are causing immune de-activation via the mechanism of clonal anergy. These mechanism can help us the know the work of immunological pathways better for better control of infectious diseases and autoimmune disorders.

Histone deacetylases inhibitors (HDACi) have shown promising therapeutic outcomes in haematological malignancies such as leu-kaemia, multiple myeloma and lymphoma, with dis-appointing results in solid tumours when used as monotherapy. As a result, combination therapies either with radiation or other deoxyribonucleic acid (DNA) damaging agents have been suggested as ideal strategy to improve their efficacy in solid tumours. Numerous in vitro and in vivo studies have demonstrated that HDACi can sensitise malignant cells to both electromagnetic and particle types of radiation by inhibiting DNA damage repair. Although the radiosensitising ability of HDACi has been reported as early as 1990s, the mechanisms of radiosensitisation are yet to be fully understood. This review brings forth the various protocols used to sequence the administration of radiation and HDACi treatments in the different studies. The possible contribution of these various protocols on the ambiguity that surrounds the mechanisms of radiosensitisation is also highlighted.

Post-operative chemotherapy is still required for the treatment of glioblastoma (GBM), and nanocarrier-based drug delivery has been identified as one of the most effective methods. However, the blood-brain barrier (BBB) and non-specific delivery to non-tumor tissues can significantly limit drug accumulation in tumor tissues and cause damage to nearby normal tissues. This study describes a targeted cancer therapy approach that uses AS1411 aptamer-conjugated nanospheres, 100-300 nm in size, loaded with doxorubicin (Dox) to selectively identify tumor cells overexpressing nucleolin (NCL) proteins. The study demonstrates that the active target model, which employs aptamer-mediated drug delivery, is more effective than nonspecific enhanced permeability and maintenance (EPR)-mediated delivery and passive drug delivery in improving drug penetration and maintenance in tumor cells. Additionally, the study shows the potential for anticancer effects through 3D spheroidal and in vivo GBM xenograft models. The DNA-protein hybrid nanospheres utilized in this study offer numerous benefits, such as efficient synthesis, structural stability, high drug loading, dye labelling, biocompatibility, and biodegradability. When combined with nanospheres, the 1411 aptamer has been shown to be an effective drug delivery carrier for precisely targeting tumors. This combination has the potential to produce anti-tumor effects in active targeted therapy of GBM.

This review aims to explore the intricate relationship between epigenetic mechanisms, stress, and affective disorders, focusing on how early life experiences and coping mechanisms contribute to susceptibility to mood disorders. Epigenetic factors play a crucial role in regulating gene expression without altering the DNA (deoxyribonucleic acid) sequence, and recent research has revealed associations between epigenetic changes and maladaptive responses to stress or psychiatric disorders. A scoping review of 33 studies employing the PRISMA-S (Preferred Reporting Items for Systematic Reviews and Meta-Analyses – Statement) guidelines investigates the role of stress-induced epigenetic mechanisms and coping strategies in affective disorders' occurrence, development, and progression. The analysis encompasses various stress factors, including childhood trauma, work-related stress, and dietary deficiencies, alongside epigenetic changes such as DNA methylation and altered gene expression. Findings indicate that specific stress-related genes frequently exhibit epigenetic changes associated with affective disorders. Moreover, the review examines coping mechanisms in patients with bipolar disorder and major depressive disorder, revealing mixed associations between coping strategies and symptom severity. While active coping is correlated with better outcomes, emotion-focused coping may exacerbate depressive or manic episodes. Overall, this review underscores the complex interplay between genetic predisposition, environmental stressors, coping mechanisms, and affective disorders. Understanding these interactions is essential for developing targeted interventions and personalized treatment strategies for individuals with mood disorders. However, further research is needed to elucidate specific genomic loci involved in affective disorders and the clinical implications of coping strategies in therapeutic settings.

The rock joint fissure slope in cold regions is prone to deformation and failure problems such as relaxation, tension and collapse under freeze-thaw (F-T) cycles. Based on the ideology of clean production and promoting the structural integrity of geotechnical projects in cold areas, this study designed and prepared aeolian-sand (AS) to replace part of river sand as a filling mortar material by taking advantage of the low cost and easy availability of AS. Investigations were carried out to determine the impact of varying proportions of AS in mortar on the microstructure and shear properties of the mortar-rock interface under F-T cycles. Therefore, F-T cycle testing, nuclear magnetic resonance (NMR) testing and shear testing were carried out on grout with different proportions of AS. The results show that the right proportion of AS replacement could enhance the shear deformation resistance and F-T resistance of the mortar-rock interface, and 30% proportion is the optimal proportion. By increasing of F-T cycles, the three types of specimens showed different degrees of particle spalling, shedding and cracking. The slurry-rock interface layer in the A (0%) group exhibited the most severe F-T deterioration, while the B (15%) group showed a lesser degree, and the C (30%) group performed the best. T2 spectrum curve and total spectrum area change law from the microscopic point of view to verify the 30% AS proportion mortar-rock interface F-T resistance is the strongest, followed by 15% AS proportion mortar, the weakest is ordinary mortar. The degradation of the mortar-rock bond is primarily due to the conversion of solid to liquid phases and the movement of internal water caused by fluctuating temperatures. The extent of F-T deterioration in the slurry-rock interface layer is greatly affected by the varying proportions of AS. The shear strength and shear stiffness of specific proportion of AS decreased with the increase of F-T cycles. For same F-T cycles, the shear strength and shear stiffness of group C (30%) were the highest, followed by group B (15%) and group A (0%). The grout material not only reinforces the F-T joint fissure slope, but also reduces the cost of grout reinforcement and promotes the clean production of geotechnical structures.

Theoretical and experimental approaches have been applied to study the polymer physics under-lying compaction of DNA in the bacterial nucleoid. Knowledge of the compaction mechanism is necessary to obtain a mechanistic understanding of the process of segregation of replicating chromosome arms (replichores) during the cell cycle. In the first part of this review light microscope observations are discussed that have demonstrated that the nucleoid has a lower refractive index and thus a lower density than the cytoplasm. A polymer-physical explanation for this phenomenon was given by the theory of Theo Odijk pro-posed in 1998. By assuming a phase separation between nucleoid and cytoplasm and by imposing equality of osmotic pressure and chemical potential between the two phases, a minimal energy situation is obtained in which soluble proteins are depleted from the nucleoid, thus explaining its lower density. The theory is compared with recent views for DNA compaction that are based on mere exclusion of polyribosomes from the nucleoid or on the transcriptional activity of the cell. These new views raise the question of whether they can still explain the lower refractive index or density of the nucleoid. In the second part of this review the question is discussed how DNA segregation occurs in Esche-richia coli, in the absence of the so-called active ParABS system, present in the majority of bacteria. How is entanglement prevented of nascent chromosome arms, generated at the origin in the pa-rental network of the E. coli nucleoid? The observations of the groups of Sherratt and Hansen in 2006, that the four nascent chromosome arms synthesized in the initial replication bubble, segre-gate to opposite halves of the sister nucleoids implies that extensive intermingling of daughter strands does not occur. Based on the hypothesis that leading and lagging replichores synthesized in the replication bubble, fold into micro-domains that do not intermingle, a passive "four-excluding-arms model" for segregation is proposed. This model implies that the key for seg-regation already lies in the structure of the replication bubble at the very start of DNA replication; it explains the different patterns of chromosome arms as well as the segregation distances between replicated loci as experimentally observed.

Abstract: Congenital cytomegalovirus (cCMV) infection poses significant risks to fetal development, particularly affecting the nervous system. This study reports a fetal autopsy case, examining cCMV infection and focusing on CMV DNA measurements in various fetal organs before formalin fixation, a novel approach for comprehensive CMV DNA evaluations in fetal organs affected by cCMV. A 20-week-old male fetus was diagnosed with cCMV following the detection of CMV DNA in ascites obtained via abdominocentesis in utero. After termination of pregnancy, multiple organs of the fetus, including the cerebrum, thyroid gland, heart, lungs, liver, spleen, kidneys, and adrenal glands, were extracted and examined for CMV DNA loads using real-time polymerase chain reaction. Histopathological examination involved hematoxylin-eosin and CMV-specific immunostaining. A correlation was found between CMV DNA loads and pathology, with higher CMV-infected cell numbers observed in organs positively identified with both staining methods, exhibiting CMV DNA levels of ≥1.0 × 104 copies/mL, compared to those detected solely by CMV-specific immunostaining, where CMV DNA levels ranged from 1.0 x 103 to 1.0 x 104 copies/mL. These results highlight a quantifiable relationship between organ infection extent and CMV DNA concentration, providing insights into cCMV pathogenesis and potentially informing future diagnostic and therapeutic strategies for cCMV infection.

Graft injury affects over 50% of liver transplant (LT) recipients, but non-invasive biomarkers to diagnose and guide treatment are currently limited. We aimed to develop a biomarker of graft injury by integrating serum metabolomic profiles with clinical variables. Serum from 55 LT recipients with biopsy confirmed metabolic-dysfunction associated steatohepatitis (MASH), T-cell mediated rejection (TCMR) and biliary complications was collected and processed using a combination of LC-MS/MS assay. The metabolomic profiles were integrated with clinical information using a multi-class Machine Learning (ML) classifier. The efficacy of the model was assessed through the evaluation of the Out-of-Bag (OOB) error estimate. Our ML model yielded an overall accuracy of 79.66% with an OOB estimate of the error rate at 19.75%. The model exhibited a maximum ability to distinguish MASH, with an OOB error estimate of 7.4% compared to 22.2% for biliary and 29.6% for TCMR. The metabolites serine and serotonin emerged as the topmost predictors. When predicting binary outcomes using 3 models: Biliary (biliary vs. rest), MASH (MASH vs. rest) and TCMR (TCMR vs. rest), the AUCs were 0.882, 0.972 and 0.96 respectively. Our ML tool integrating serum metabolites with clinical variables shows promise as a non-invasive, multi-class serum biomarker of graft pathology.

Immune responses to influenza (flu) antigens reflect memory to prior infections or vaccinations, which might influence immunity to new flu antigens. Memory of past antigens has been termed “original antigenic sin” or more recently “immune imprinting” and “seniority”. We have researched a comparison between the immune response to live flu infections and inactivated flu vaccina-tions. A brief history of antibody generation theories is presented culminating in new findings about the immune-network theory suggesting that a network of clones exists between an-ti-idiotypic antibodies and T cell receptors. Findings regarding the 2009 pandemic flu strain and immune responses to it are presented, including memory B cells and conserved regions within the hemagglutinin protein. The importance of CD4+ memory T cells and cytotoxic CD8+ T cells responding to both infections and vaccinations are discussed and compared. Innate immune cells like natural killer (NK) cells and macrophages are discussed regarding their roles in adaptive immune responses. Antigen presentation via macroautophagy processes are described. New vaccines in development are mentioned along with the results of some clinical trials. The manu-script concludes with how repeated vaccinations are impacting the immune system and a sketch of what might be behind the imprinting phenomenon, including future research directions.

In chronic myeloid leukemia (CML), a clonal proliferative disorder of granulocytic lineage, the 90%‐95% of cases are characterized by t(9;22) (q34.1;q11.2) translocation, leading to the Philadelphia chromosome (1). Rarely, CML patients directly present a blast crisis, mostly of myeloid origin (2, 3). Herein, we describe one rare CML case, showing an unusual early T lymphoid blastic crisis.

DNA Topoisomerase IIa (Top2A) is a nuclear enzyme that is a cancer drug target, and there is interest in identifying novel sites on the enzyme to inhibit cancer cells more selectively and to reduce off-target toxicity. The C-terminal domain (CTD) is one potential target, but it is an intrinsically disordered domain, which prevents structural analysis. Therefore, we set out to analyze the sequence of Top2A from 105 species using bioinformatic analysis including the PSICalc algorithm, entropy analysis, and other approaches. Our results demonstrate that large (10th-order) interdependent clusters are found including non-proximal positions across the major domains of Top2A. Further, CTD-specific clusters of 3rd, 4th, and 5th-order were identified that included positions that had been previously analyzed via mutation and biochemical assays. Some of these clusters coincided with positions that when mutated either increased or decreased relaxation activity. Finally, sites of low entropy (i.e., high information content) were identified and mapped as key positions in the CTD. Included in the low entropy sites are phosphorylation sites and charged positions. Together, these results help to build a clearer picture of the critical positions in the CTD and provide potential sites/regions for further analysis.

Circulating cell-free DNA (cfDNA) refers to small fragments of DNA molecules released after programmed cell death and necrosis in several body fluids such as blood, saliva, urine, and cerebrospinal fluid. The discovery of cfDNA has revolutionized the field of non-invasive diagnostics in the oncologic field, in prenatal testing, and in organ transplantation. Despite the potential of cfDNA and the solid results published in recent literature, several challenges remain, represented by low abundance, need for highly sensitive assays and analytical issues. In this review, the main technical advances in cfDNA analysis are presented and discussed, with a comprehensive examination of the current available methodologies applied in each field. Considering the potential advantages of cfDNA, this biomarker is increasing its consensus among clinicians, as it consents to monitor patients’ conditions in an easy and non-invasive way, offering a more personalized care. Nevertheless, cfDNA analysis is still considered a research marker to be further validated, and very few centers are implementing its analysis in the real-life assistance. As technical improvements are enhancing the performances of cfDNA analysis, its application will transversally improve patients’ quality of life.

The idea of CUAD (contact unmodified antisense DNA) biotechnology and the use of oligonucleotide insecticides (briefly, olinscides, or DNA insecticides) was proposed on lepidopterans and gradually disseminated on hemipteran pests. As the next-generation platform for creation of insecticides, it has a number of unique characteristics (short unmodified antisense DNA as contact insecticide, insect rRNAs as target, DNA containment as mechanism of action) that distinguish it from existing classes of chemical insecticides. Oligonucleotide insecticides possess low carbon footprint, high safety for non-target organisms, rapid biodegradability, avoidance of target-site resistance, ensuring prosperity for agricultural countries that will implement this innovation. As the next-generation class of insecticides olinscides create opportunities that have never existed before in crop protection. Using flexible CUAD platform, any farmer, botanical garden or pest control company is capable of creating its own unique arsenal of insecticides, well-tailored for a particular population of insect pest. Most technological innovations start out very expensive and don't work well, but eventually start working well and become very cheap. CUAD biotechnology has come a long way for 16 years and is very close to the point to be implemented on a large scale against certain groups of pests. In this article we provide basic knowledge of use of oligonucleotide pesticides and list pests (mainly from suborder Sternorrhyncha, Hemiptera) which were successfully controlled by us recently.

As the global arid climate intensifies and the groundwater becomes increasingly scarce, drought stress has become a major challenge to plant growth and development worldwide, especially causing yield reduction and extinction of cultivated crops. In a large number of drought resistance studies, DNA methylation has been found dynamically associated with response to drought stress in plants by regulating gene expression and developments in plants, but the specific mechanisms are not yet clear. In this article, the current advances of research have been summarized on DNA methylation response to drought stress in various plant species, in terms of methylation detection methods, mechanisms of pattern changes (DNA de novo methylation, DNA maintenance methylation, and DNA demethylation), and its response to drought stress. With Most of the studies showed significant changes in genome or gene promoter methylation levels in plants subjected to drought stress, but the identification of the genes and pathways involved is still in the minority. The aim is to provide references for in-depth research on the response of plants to drought stress through epigenetics, and to uncover drought resistance genes regulated by DNA methylation, specific signaling pathways, and molecular mechanisms of action.

A co-curing resin system consisting of 9368 epoxy resin for prepreg and 6808 epoxy resin for resin transfer molding (RTM) was developed. A corresponding preparation method of novel polymer composite bolted T-joint with internal skeleton and external skin was proposed based on prepreg-RTM co-curing process, and novel T-joints were fabricated. A series of conventional configuration T-joints based on RTM process and T-joints made of 2A12 aluminum alloy were prepared simultaneously. Bending performances were studied on these T-joints experimentally. The results indicate that 9368 epoxy resin and 6808 epoxy resin exhibit good compatibility in rheological and thermophysical properties. The novel T-joints prepared with prepreg-RTM co-curing process show no obvious fiber local winding or resin-rich regions inside, and the interface quality between internal skeleton and external skin is excellent. The main failure modes of the novel T-joint under bending load include separation of skin and skeleton and fracture along the thickness on the base panel; the skeleton carries the main bending load, but there is still load transfer between external skin and internal skeleton through their interface. The internal damages of the novel T-joint are highly consistent with surface damages observed visually, facilitating the detection and timely discovery of damages. The initial stiffness, damage initiation load, and ultimate load of the novel T-joint are 1.65 times, 5.89 times, and 3.45 times that of conventional T-joint, respectively. When considering the influence of density, the relative initial stiffness and relative ultimate load of the novel T-joint are 1.44 times and 2.07 times that of aluminum alloy T-joint, respectively.

To elucidate the cellular immune response to coronavirus disease (COVID-19) among children, we assessed cellular immunity in 8 children post-vaccination for COVID-19 and 11 children after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, using the T-SPOT®.COVID assay for both the spike (S) and nucleocapsid (N) proteins. In the vaccinated group, the T-SPOT®.COVID assay for the S protein yielded positive results in all eight children. In contrast, in the post-infection group, the assay for the N protein was positive in 5 of 11 children, with 3 of these 5 children requiring hospital admission, including 2 who needed mechanical ventilation. Therefore, the T-SPOT®.COVID assay is valuable for assessing cellular immunity against COVID-19, and most children infected with COVID-19 may not develop such immunity unless the disease severity is significant.

DNA damage causes the mutations that are the principal source of genetic variation. DNA damage detection and repair mechanisms therefore play a determining role in generating the genetic diversity on which natural selection acts. Speciation, it is commonly assumed, occurs at a rate set by the level of standing allelic diversity in a population. The process of speciation is driven by a combination of two evolutionary forces: genetic drift and ecological selection. Genetic drift takes place under conditions of relaxed selection, and results in a balance between rates of mutation and rates of genetic substitution. These two processes, drift and selection, are necessarily mediated by a variety of mechanisms guaranteeing genome stability in any given species. One of the outstanding questions in evolutionary biology concerns the origin of the widely varying phylogenetic distribution of biodiversity across the tree of life, and how the forces of drift and selection contribute to shaping that distribution. The following examines some of the molecular mechanisms underlying genome stability and the adaptive radiations that are associated with biodiversity and the widely varying species richness and evenness in the different eukaryotic lineages.

Lumpy skin disease (LSD), caused by a virus within the Poxviridae family and Capripoxvirus genus, induces nodular skin lesions in cattle, spreading through direct contact and insect vectors, significantly affecting global cattle farming. Despite available vaccines, their effectiveness is limited by poor bioavailability and adverse effects. Our study aimed to identify potential inhibitors targeting LSD-associated DNA polymerase protein, selecting LSDV039 for further investigation through comprehensive analysis and computational methods. Virtual screening revealed rhein and taxifolin as potent binders among 380 phytocompounds, with respective affinities of -8.965 and -7.195 kcal/mol. Canagliflozin and tepotinib exhibited strong affinities (-9.858 and -8.856 kcal/mol) among 718 FDA-approved antiviral drugs. Molecular dynamics simulations of canagliflozin, tepotinib, rhein, and taxifolin highlighted taxifolin's superior stability and binding energy. Rhein displayed compactness in RMSD and RMSF but fluctuations in Rg and SASA, while canagliflozin demonstrated stability compared to tepotinib. The study highlights the promising potential of repurposed drugs and phytocompounds as LSD therapeutics. However, extensive validation through in-vitro, in-vivo, and clinical trials is crucial for their practical application.

Ajuga arabica is an essential medicinal plant found naturally in Saudi Arabia, recognized for its therapeutic characteristics. Nevertheless, research on the genetic diversity and population arrangement of this species in Saudi Arabia is deficient. In this study, we conducted a comparative examination of DNA and gene sequences to evaluate the genetic diversity and population configuration of Ajuga arabica in Saudi Arabia. Our findings demonstrate that there are high levels of genetic diversity within populations, but low levels of genetic differentiation among populations. Moreover, we discovered proof of gene flow among populations, suggesting that there are no significant genetic barriers to gene flow. Our results offer valuable insights into the genetic diversity and population structure of Ajuga arabica in Saudi Arabia, which can support conservation efforts and promote the sustainable use of this valuable medicinal plant.

Psoriasis is a multi-factorial immune-mediated disease that affects approximately 2% of the UK population and 2.5% worldwide. The condition can impact multiple organs, including the skin and joints. In moderate to severe cases, systemic and topical medications may not provide adequate relief, and as a result, biologic treatments are often administered as a monotherapy or in combination with other treatments. While biologic treatments are more effective than systemic and topical medications in treating psoriasis, patients require close supervision and monitoring while receiving this type of treatment. Liposomes are highly versatile and can be employed for therapy and research applications. They enhance drug retention at the site of administration, thereby reducing systemic side effects. Our study was the first to investigate the genotoxicity of secukinumab, an ideal biologic treatment for psoriasis (a human IgG1k antibody, anti-IL17A), in bulk and liposome nanoparticles on the lymphocytes of psoriasis patients compared to healthy individuals. We evaluated and com-pared the genotoxicity of secukinumab in bulk and liposomal form using the Comet and micronucleus assays. Both assays demonstrated that secukinumab in both forms did not ex-hibit genotoxicity and reduced DNA damage following treatment of the lymphocytes from healthy individuals and psoriatic patients with secukinumab bulk and liposomes format. Both concentrations of secukinumab used (2.1 and 2.8 µg/mL) effectively decreased H2O2-induced DNA damage in both groups to nearly the level of the negative control. Secukinumab bulk and liposome form significantly decreased H2O2-induced damage and efficiently attenuated its adverse effects both in the Comet (p<0.0001) and micronucleus assays (p<0.01). Overall, secukinumab in both forms exhibited protective and an-ti-genotoxic effects by demonstrating its potential to reduce DNA damage caused by oxida-tive stress and by not inducing any further damage in the lymphocytes of either healthy in-dividuals or patients.

The positional cloning of single nucleotide polymorphisms (SNPs) of the neutrophil cytosolic factor 1 (Ncf1) gene, advocating that a low oxidative burst drives autoimmune disease, demands an understanding of the underlying molecular causes. A cellular target could be T cells, which have been shown to be regulated by reactive oxygen species (ROS). However, the pathways by which ROS mediate T cell signaling remain unclear. The adaptor molecule linker for activation of T cells (LAT) is essential for coupling T cell receptor-mediated antigen recognition to downstream responses, and it contains several cysteine residues that have previously been suggested to be involved in redox regulation. To address the possibility that ROS regulate T cell-dependent inflammation through LAT, we established a mouse strain with cysteine-to-serine mutations at positions 120 and 172 (LATSS). We found that redox regulation of LAT through C120 and C172 mediate its localization and phosphorylation. LATSS mice had reduced numbers of double-positive thymocytes and naïve peripheral T cells. Importantly, redox insensitivity of LAT enhanced T cell-dependent autoimmune inflammation in collagen-induced arthritis (CIA), a mouse model of rheumatoid arthritis (RA). This effect was reversed on an NCF1-mutated (NCF1m1j), ROS-deficient, background. Overall, our data shows that LAT is redox-regulated, acts to repress T cell activation, and is targeted by ROS induced by NCF1 in antigen-presenting cells (APCs).

Recent advancements in molecular diagnostics have highlighted the potential of circulating cell-free DNA (ccfDNA) as a biomarker in various diseases. This study aims to explore the role of ccfDNA in the early diagnosis, prognosis, and therapeutic monitoring of diseases, with a focus on oncology. We conducted a comprehensive review and analysis of current literature, combined with experimental approaches involving advanced genomic sequencing and bioinformatics techniques. Our research systematically characterizes the molecular composition of ccfDNA, including its origins, size distribution, and methylation patterns, across different disease states. Furthermore, we investigate the dynamics of ccfDNA release and clearance in the bloodstream, elucidating mechanisms that could influence its diagnostic accuracy. In the context of cancer, we analyze the correlation between tumor-derived ccfDNA and disease progression, response to therapy, and overall survival. Our findings indicate that ccfDNA holds promise as a non-invasive biomarker for early disease detection, real-time monitoring of treatment efficacy, and potentially, predicting relapse. This study contributes to the growing field of liquid biopsies and opens new avenues for personalized medicine, emphasizing the need for standardized protocols in ccfDNA analysis to maximize its clinical utility. The implications of our findings extend beyond oncology, suggesting potential applications of ccfDNA in other pathological conditions.

DNA barcoding has emerged as a transformative tool in the realm of biodiversity assessment, facilitating rapid and precise species identification. This review article delves into the evolving landscape of DNA barcoding, with a particular focus on the emergence of mini-barcodes and alternative genes as innovative approaches. Traditional DNA barcoding, anchored in the COI gene, has significantly advanced our understanding of species diversity; however, inherent challenges persist, especially when dealing with degraded samples or taxonomic groups where COI falls short. The introduction of mini-barcodes, characterized by their reduced sequence length, has revolutionized DNA barcoding by expanding its applicability. We explore the utility of mini-barcodes in various scenarios, ranging from DNA extraction from ancient or fragmented specimens to in-field identification using portable sequencing technologies. Case studies and practical insights into their effectiveness are presented, emphasizing their role in overcoming traditional limitations. Alternative genes, a complementary facet of DNA barcoding, offer a novel avenue for enhancing taxonomic resolution and accuracy. This review showcases instances where researchers have harnessed alternative genes to overcome COI's constraints, particularly in diverse taxonomic groups, such as fungi, plants, and microorganisms. The selection of these genes is discussed, along with their potential to uncover hidden biodiversity and improve species delimitation. Examining the broader implications of mini-barcodes and alternative genes, we unveil their pivotal roles in advancing conservation efforts, forensic science, and citizen science initiatives. The adaptable nature of these approaches empowers a diverse range of stakeholders, from scientists conducting critical research to concerned citizens contributing to our collective understanding of the natural world. Looking ahead, we envision an exciting future for DNA barcoding, as these innovations continue to evolve. Integration with cutting-edge sequencing technologies, data analysis methods, and the harnessing of big data hold immense potential. Nonetheless, challenges related to standardization, scalability, and data sharing persist.

The study titled "Molecular Choreography of DNA: Insights and Innovations" explores the intricate dance of DNA, shedding light on its structure, function, and regulation. This comprehensive review delves into the historical significance of DNA research, from Watson and Crick's double helix discovery to recent innovations in genomics and molecular biology. The review highlights DNA's roles in replication, repair, gene regulation, and chromatin dynamics, emphasizing the precision and complexity of its molecular choreography. Innovations like CRISPR-Cas9 gene editing and advanced sequencing technologies have revolutionized our understanding of DNA, opening new avenues for scientific exploration. This study aims to provide readers with a deeper appreciation of DNA's central role in life's choreographed processes.

Candidate vaccines against African swine fever virus (ASFV) based on naturally attenuated or genetically modified viruses have the potential to generate protective immune responses, although there is no consensus on what defines a protective immune response against ASFV. Studies, espe-cially in sensible host species, focused on unravelling protective mechanisms will contribute to the development of safer and more effective vaccines. The present study provides a detailed analysis of phenotypic and functional data on cellular responses induced by intradermal immunization and subsequent boosting of domestic pigs with the naturally attenuated field strain Lv17/WB/Rie1, as well as the mechanisms underlying protection against intramuscular challenge with the virulent genotype II Armenia/07 strain. The transient mild to moderate increase of IL-8 and IL-10 in serum observed after immunization might be directly correlated with survival. Protection was also associated with a robust ASFV-specific polyfunctional memory T cell response, where CD4CD8 and CD8 T cells were identified as the main cellular sources of virus-specific IFNγ and TNFα. In parallel to cytokine response, these T-cell subsets also showed specific cytotoxic activity as evidenced by the increased expression of the CD107a degranulation marker. Along with virus-specific multifunctional CD4CD8 and CD8 T responses, the increased levels of antigen experienced cytotoxic CD4 T cells observed after challenge in immunized pigs might also contribute to control virulent infection by killing mechanisms targeting infected antigen-presenting cells. Future studies should elucidate whether the memory T-cell responses evidenced in the present study persist and provide long-term protection against further ASFV infections.

Abstract Background In tumors, somatic mutagenesis presumably drives DNA damage response (DDR) via altered regulatory pathways increasing genomic instability and proliferative activity. These considerations led to the standard therapeutic strategy against cancer: the disruption of mutation activated DNA repair pathways of tumors. Purpose Justifying that cancer cells are not enemies to be killed, but rather they are ill human cells having the remnants of physiologic regulatory pathways. Results 1. Genomic instability and cancer development may be originated from the flaw of estrogen signal rather than excessive estrogen signaling. 2. Healthy cells with genomic instability exhibit somatic mutations helping DNA restitution. 3. Somatic mutations in tumor cells aim the restoration of DNA damage rather than further genomic derangement. 4. In tumors, estrogen signal drives the pathways of DNA stabilization leading to apoptotic death. 5. In the peritumoral cellular infiltration, the genomic damage of tumor induces inflammatory cytokine secretion and increased estrogen synthesis. In the inflammatory cells, increased growth factor receptor (GFR) signal confers unliganded activation of estrogen receptors (ERs). 6. In tumor cells responsive to genotoxic therapy, constitutive mutations help the upregulation of estrogen signal and consequential apoptosis. In tumors non responsive to genotoxic therapy, the possibilities for ER activation via either liganded or unliganded pathway, are exhausted leading to farther genomic instability and unrestrained proliferation. Conclusion Understanding the real character and behavior of human tumors at molecular level suggests that we should learn the genome repairing methods of tumors and follow them by supportive therapy rather than provoking additional genomic damages.

This review article delves into the pivotal role of DNA in advancing criminal investigations, presenting a comprehensive analysis of its impact on modern justice systems. DNA, as the genetic blueprint of individuals, has revolutionized forensic science, augmenting the accuracy and reliability of criminal investigations. This study explores the historical evolution of DNA analysis, from its initial discovery to the development of sophisticated techniques like polymerase chain reaction (PCR), short tandem repeat (STR) profiling, and next-generation sequencing (NGS). The article elucidates the intricate process of DNA collection, preservation, and analysis, highlighting its significance in establishing identity, proving guilt or innocence, and resolving cold cases. The utilization of DNA databases and international cooperation in cross-border investigations are also examined, emphasizing their contributions to solving complex criminal cases. Moreover, the ethical, legal, and privacy considerations surrounding DNA usage in criminal justice are critically evaluated. Through a comprehensive literature review, this article synthesizes empirical evidence showcasing how DNA evidence has exonerated wrongfully convicted individuals and provided closure to victims' families. It further elucidates how technological advancements have extended DNA analysis to trace evidence, touch DNA, and forensic genealogy, thereby broadening its application in criminal investigations.

The exploration of three-dimensional (3D) DNA structures marks a pivotal shift in genomics, offering unprecedented insights into the intricacies of cellular regulation, disease genesis, and the potential for novel diagnostic and therapeutic strategies. This study delves into the dynamic nature of 3D genome organization, investigating how these structures influence cellular functions in response to environmental changes and over time. Through a combination of real-time imaging and advanced computational models, we aim to unravel the complex interplay between 3D DNA structures, non-coding RNAs, and chromatin organization, and their collective impact on gene expression and cellular differentiation. A significant focus is placed on understanding the role of 3D DNA structures in the development and progression of genetic disorders. By employing comprehensive genomic and epigenomic profiling, this research seeks to identify structural aberrations linked to diseases, paving the way for the development of targeted diagnostics and therapies. Additionally, the study explores the potential of genome editing technologies, such as CRISPR-Cas systems, in correcting these structural abnormalities, while also addressing the ethical, legal, and social implications (ELSI) of such interventions. Integrating data from various genomic analyses, we propose the development of advanced computational models to predict the effects of 3D DNA structures on gene expression and cellular function. This integrative approach not only promises to enhance our understanding of genomic regulation but also sets the stage for groundbreaking advances in genomics-based diagnostics and therapeutics. The findings of this study have the potential to revolutionize our approach to treating genetic disorders, offering a new frontier in personalized medicine.

The epigenetic mechanism of DNA methylation is essential for regulating gene expression and adaptation in plants, especially when exposed to various environmental stressors such as drought, heat, and nutrient availability. The impact of these stressors can lead to alterations in DNA methylation patterns and gene expression, which this study aims to investigate. Specifically, we are interested in understanding how DNA methylation plays a role in controlling the expression of stress-response genes, as well as genes involved in plant growth and development. To accomplish this, we are analyzing the changes in DNA methylation patterns and gene expression in response to different stressors like drought and heat. Our research findings suggest that DNA methylation is a vital regulator of plant adaptation to environmental stress, and we hope to gain further insight into these mechanisms by identifying genes that exhibit differential expression in response to stress and analyzing their DNA methylation status. This study offers valuable insights into the development of crops that can thrive under changing climate conditions.

Barcoding of DNA is a species identification technique that utilizes a small DNA fragment from a specific gene or genes known as "sequencing." The idea behind DNA barcoding is that a single sequence can be used to specifically identify an organism to species by comparison with a reference library of these DNA sections. Herbal adulteration can be monitored using DNA barcodes that utilize medicinal plants such as Atropa belladonna. Because it is challenging to distinguish between species solely based on morphology, other plants belonging to the same genus are replacing it in trade. Such replacement with plants not utilized for medical purposes could have negative side effects. Also, Atropa belladonna's positive effects can be completely lost. Taxonomists also have trouble distinguishing the genuine Atropa belladonna plant from cultivated items. As a result, DNA barcoding can now be used to overcome this challenge. The development of DNA barcodes may help with trade control, which protects and conserves endangered species from export outside of the country due to their economic and biological significance. In this review, we discuss the DNA barcoding of Atropa belladonna plant in the medicinal plants field.

Diffuse midline gliomas are among the deadliest human cancers and have had little progress in treatment in the last 50 years. Cell cultures of these tumors have been developed recently, but it is unknown the degree to which such cultures retain the characteristics of the source tumors. DNA methylation profiling offers a powerful tool to look at genome-wide epigenetic changes that are biologically meaningful and can help assess the similarity of cultured tumor cells to their in vivo progenitors. Paraffinized diagnostic tissue from three diffuse intrinsic pontine gliomas with H3 K27M mutations was compared with subsequent passages of neurosphere cell cultures from those tumors. Each cell line was passaged 3-4 times and analyzed with DNA methylation arrays and standard algorithms that provided comparison of diagnostic classification and cluster analysis. All samples tested maintained high classifier scores and clustered within the reference group of H3 K27M-mutant diffuse midline gliomas. There was a gain of 1q in all cell lines, with two cell lines initially manifesting the gain of 1q only during culture. In vitro cell cultures of H3 K27M-mutant gliomas maintain high degrees of similarity in DNA methylation profiles to their source tumor, confirming their fidelity even with some chromosomal changes.

Advancements in genomics and molecular biology have propelled our understanding of genetic information far beyond the iconic double helix structure of DNA. This comprehensive review delves into the intricate world of DNA and RNA networks, exploring the complexities that extend beyond the classical model. We investigate the dynamic interactions, regulatory mechanisms, and functional implications of the broader genetic landscape, encompassing both coding and non-coding elements. From chromatin architecture to the diverse array of non-coding RNAs, this exploration seeks to provide a holistic perspective on the multifaceted roles played by DNA and RNA in cellular processes. The three-dimensional organization of genetic material within the cell has emerged as a crucial determinant of cellular function. Chromatin architecture, comprising DNA, histones, and other associated proteins, plays a dynamic role in regulating gene expression. We delve into the intricate choreography of chromatin remodeling, deciphering the interplay between transcription factors, epigenetic modifications, and higher-order structures that modulate gene activity. The exploration of chromatin dynamics unveils a nuanced understanding of how the genome responds to environmental cues, cellular signals, and developmental processes. Moreover, the once-dismissed non-coding portion of the genome has emerged as a key regulator of cellular function. Various classes of non-coding RNAs, such as microRNAs, long non-coding RNAs, and circular RNAs, have been implicated in diverse cellular processes, ranging from fine-tuning gene expression to modulating chromatin structure. The review critically examines the functional roles of these non-coding RNAs, shedding light on their involvement in cellular homeostasis, development, and disease. As I traverse this landscape beyond the double helix, our review synthesizes current knowledge on the intricacies of DNA and RNA networks. By exploring the regulatory mechanisms, functional implications, and emerging paradigms, we aim to inspire a deeper appreciation for the molecular choreography that underlies the cellular symphony of life. Furthermore, we discuss the implications of these findings in the context of disease, highlighting potential diagnostic and therapeutic avenues that arise from a comprehensive understanding of DNA and RNA networks. In doing so, this exploration not only broadens our theoretical understanding but also opens new frontiers for translational research in genetics and molecular medicine.

DNA barcoding has emerged as a powerful tool in the field of taxonomy, transforming the way we identify and classify species. This review article aims to provide an up-to-date overview of the advancements in DNA barcoding techniques, their applications, and their impact on taxonomy and biodiversity studies. We will explore the evolution of DNA barcoding, the development of standardized barcode regions, and the challenges and limitations associated with this methodology. Additionally, we will delve into the diverse range of taxonomic groups for which DNA barcoding has been applied, highlighting key case studies and their contributions to our understanding of biodiversity. Furthermore, we will discuss the integration of DNA barcoding with other molecular and bioinformatic approaches, such as metabarcoding and machine learning, to enhance species identification and biodiversity assessment. This review aims to provide a comprehensive perspective on how DNA barcoding is reshaping the field of taxonomy and its implications for conservation, ecology, and evolutionary biology.

Lynch syndrome (LS) is an inherited cancer predisposition disorder associated with an elevated risk of various epithelial cancers. Despite sharing the same pathogenic variant (PV), Lynch syn-drome variant heterozygotes (LSVH) exhibit considerable phenotypic variability in cancer risk. The role of Human Leukocyte Antigen (HLA) in modifying cancer risk prompted our investiga-tion into whether HLA variations act as genetic modifiers influencing age at cancer diagnosis in a unique cohort of LSVH carrying a PV in the hMLH1 gene in South Africa. Within our extensive LS cohort, 426 individuals carried the same hMLH1 PV (MLH1:c.1528C>T). We selected 100 indi-viduals with the greatest diversity in age at cancer diagnosis and the oldest unaffected individu-als for high-throughput HLA genotyping of 12 HLA class I and II loci using next-generation se-quencing. Statistical analyses employed Kaplan-Meier survival analyses with Logrank tests and Cox proportional hazards. Following the robust application of statistical correction methods, six HLA alleles (3.2%) were significantly associated with a young age at cancer diagnosis. Notably, HLA-B*15:17 and HLA-DPB1*55:01 correlated significantly with very young colorectal cancer (CRC) diagnosis (Mean age: 21y [17-25]; HR = 71.59; q

Gold nanoparticles (AuNPs) exhibit improved optical and spectral properties compared to bulk materials, making them suitable for detection of DNA, RNA, antigens, and antibodies. Here, we describe a simple, selective, and rapid non-cross linking detection assay, using approx. 35 nm spherical Au nanoprobes, for a common mutation occurring in exon 19 of the epidermal growth factor receptor (EGFR), associated with non-small cell lung cancer cells. AuNPs were synthesized based on the seed-mediated growth method and functionalized with a specific 16 bp thiolated oligonucleotide using a pH-assisted method. Both AuNPs and Au nanoprobes proved to be highly stable and monodisperse by ultraviolet-visible spectrophotometry, dynamic light scattering (DLS) and electrophoretic light scattering (ELS). Our results indicate a detection limit of 1.5 µg mL−1 using a 0.15 nmol dm−3 Au nanoprobe concentration. In conclusion, this work is an effective possibility for a straight-forward, fast, and inexpensive alternative for the detection of DNA sequences related to lung cancer, leading to a potential platform for an early diagnosis of lung cancer patients.

A hybrid material, consisting of commercially available nitrocellulose (NC) membrane non-covalently modified with amino-polyethylene glycol functionalized reduced graphene oxide (NH2-PEG-rGO) nanoparticles, was successfully synthesized for oligonucleotide extraction. Fourier Transform Infrared Spectroscopy (FTIR) confirmed the modification of the NC membrane, revealing characteristic peaks of both compounds NC and NH2-PEG-rGO. Scanning Electron Microscopy (SEM) exhibited morphological changes in the NC/NH2-PEG-rGO hybrid membrane, marked by the introduction of NH2-PEG-rGO particles, resulting in a distinctly smothered surface compared to the porous surface of the NC control membrane. Wettability assays revealed hydrophobic behavior for the NC/NH2-PEG-rGO hybrid membrane, with a water contact angle exceeding 90°, contrasting with the hydrophilic behavior characterized by a 16.7° contact angle in the NC membrane. The performance of the NC/NH2-PEG-rGO hybrid membrane was evaluated for the extraction of ssDNA with fewer than 50 nucleotides from solutions containing various ionic species (MnCl2, MgCl2, and MnCl2/MgCl2). The NC/NH2-PEG-rGO hybrid membrane exhibited optimal performance when incubated in MgCl2, presenting the highest fluorescence emission at 525 relative fluorescence units (r.f.u.). This corresponds to the extraction of approximately 610 pg (≈13%) of the total oligo-DNA, underscoring the efficacy of the pristine material, which extracts 286 pg (≈6%) of oligo-DNA in complex solutions.

This review article offers an in-depth look at DNA, its discovery, structure, functions, and the significant role it plays in modern biomedical research. It begins by revisiting the landmark discovery of the DNA double helix by scientists Watson and Crick, a structure resembling a twisted ladder that forms the blueprint of life. The review explains the basic processes of DNA replication (copying DNA), transcription (converting DNA to RNA), and translation (turning RNA into proteins), which are fundamental to understanding how genetic information is passed on and used by living organisms. The article then explores gene regulation and epigenetics, which involve how genes are turned on or off and how environmental factors can affect gene expression without changing the DNA sequence itself. It examines how DNA interacts with various proteins and modifiers, which play crucial roles in these processes. A major focus is on the advancements in DNA sequencing technologies. From the initial Sanger sequencing method, which was labor-intensive and time-consuming, to the highly efficient next-generation sequencing (NGS) and the latest single-molecule sequencing techniques, the review covers how these technologies have revolutionized genomics and personalized medicine. It also highlights the importance of computational genomics and bioinformatics, fields that deal with analyzing and interpreting the vast amounts of data generated by DNA sequencing. The review discusses DNA's crucial role in understanding human diseases. It covers DNA-based research in cancer, rare genetic disorders, infectious diseases, and pharmacogenomics (how genes affect a person's response to drugs). It emphasizes studies that have led to new ways of diagnosing and treating diseases. The article looks at the future of DNA research. It talks about emerging technologies like CRISPR-Cas9, a revolutionary gene-editing tool that allows scientists to modify DNA with high precision. Synthetic biology, an interdisciplinary field that combines biology and engineering for designing and constructing new biological parts and systems, is also discussed. Furthermore, DNA nanotechnology, which involves designing DNA molecules for technological uses, is explored. Finally, the review addresses ethical issues in DNA research, such as privacy concerns and debates over genome editing.

Using multi-color flow cytometry analysis, we studied the immunophenotypical differences between leukemic blasts/LSCs from patients with AML/MDS and hematopoietic stem and progenitor cells (HSPCs) from patients in complete remission (CR) following their successful treatment. The panel of markers included CD34, CD38, CD45RA, CD123 as representatives for a hierarchical hematopoietic stem and progenitor cell (HSPC) classification as well as programmed death ligand 1 (PD-L1). Rather than restricting the evaluation on a 2- or 3-dimensional analysis, we applied a t-distributed stochastic neighbor embedding (t-SNE) approach to get a deeper insight and segregation between leukemic and normal HPSCs. For that purpose, we created a t-SNE map, which resulted in the visualization of 27 cell clusters based on their similarity concerning the composition and intensity of antigen expression. Two of these clusters were “leukemia-related” containing a great proportion of CD34⁺/CD38^- hematopoietic stem cells (HSCs) or CD34⁺ cells with a strong coexpression of CD45RA/CD123, respectively. CD34⁺ cells within the latter cluster were also highly positive for PD-L1 reflecting their immunosuppressive capacity. Beyond this proof of principle study, the inclusion of additional markers will be helpful to refine the differentiation between normal HSPCs and leukemic cells, particularly in the context of minimal disease detection and antigen-targeted therapeutic interventions. Furthermore, we suggest a protocol for the assignment of new cell ensembles in quantitative terms, via a numerical value, the Pearson coefficient, based on a similarity comparison of the t-SNE pattern with a reference.

A set of polymer composite bolted T-joints with novel configuration consisting of internal skeleton and external skin was fabricated using a prepreg-RTM co-curing molding process. Experiments were conducted to study their mechanical properties under bending load. A finite element model with a polymer resin area between the skin and skeleton was established and verified by the experimental results. Then, the damage propagation process and failure mechanism of the joint and the influence of three factors related to the layer characteristics of the skin and skeleton were investigated by the validated models. The results show that the bending stiffness and the yield limit load of the novel composite T-joint are 0.81 times and 1.65 times that of the 2A12 aluminum T-joint respectively, while at only 55.4% of its weight. The damage of the joint initiates within the resin area and leads to a degradation of the joint bending performances. The preferred stacking sequence of the skeleton is [0/+45/90/-45]ns when primarily subjected to bending loads. The decrease in bending performances is within 5% with the inclining angle of the skeleton less than 12 degrees. The more layers of 90° in the skin, the better bending performances of the joints, while the more layer of 0°, the poorer bending performances.

A Friedmann-Lemaitre-Robertson-Walker space-time model with all curvatures k=0,±1 is explored in f(R,T) gravity. The solutions are obtained via the parametrization of the scale factor that leads to a model transiting from a decelerated universe to an accelerating one. The physical features of the model are discussed and analysed.

DNA polymerases replicate cell genomes and/or participate in the maintenance of genome integrity. DNA polymerases sharing high sequence homology with E. coli DNA polymerase I (pol I), have been grouped in Family A. Pol I participates in Okazaki fragment maturation and in bacterial genome repair. Since its discovery in 1956, pol I has been extensively studied, primarily to get deeper insights into the mechanism of DNA replication. As research on DNA polymerases advances, many novel functions of this group of polymerases are uncovered. For example, human DNA polymerase θ (a Family A DNA pol) has been shown to synthesize DNA using RNA as a template, a function typically attributed to retroviral reverse transcriptase. A heightened interest in drug discovery against pol θ has emerged due to its roles in cancer. Likewise, Pol I family enzymes also appear attractive drug-development targets against microbial infections. Development of antimalarial compounds targeting apicoplast apPOL, an ortholog of Pol I, further extends the druggability of this family of enzymes. Here, we summarize reported drug-development efforts against Family A polymerases, and future perspective regarding these enzymes as antibiotic targets. Current techniques such artificial intelligence can be used to facilitate the goal of new drugs.

Coccomyxa subellipsoidea KJ (C-KJ) is a green alga with unique immunoregulatory characteristics. Here, we investigated the mechanism underlying the modification of T cell function by C-KJ components. The water-soluble extract of C-KJ was fractionated into protein (P) and sugar (S) fractions [acidic (AS), basic (BS), and neutral (NS)]. These fractions were used for the treatment of peripheral blood mononuclear cells stimulated with toxic shock syndrome toxin-1. Transcriptome analysis revealed that both P and AS enhanced the expression of the genes encoding metallothionein (MT) family proteins, inflammatory factors, and T helper (Th) 17 cytokine and suppressed that of those encoding Th2 cytokines in stimulated T cells. The kinetics of MT1 and MT2A gene expression showed a transient increase in MT1 and maintenance of MT2A mRNA after T cell stimulation in the presence of AS. The kinetics of Th17-related cytokine secretion in the early period were comparable to those of MT2A mRNA. Furthermore, our findings revealed that static, a STAT-3 inhibitor, significantly suppressed MT2A gene expression. These findings suggest that the expression of MTs is involved in the immune regulatory function of C-KJ components, which is partially regulated by Th17 responses, and may help develop innovative immunoregulatory drugs or functional foods.

Dried blood spot (DBS) testing on confetti is one of the most common methods used to collect and extract DNA from field samples for molecular epidemiology of malaria studies. Here, we investigated whether ethanol or other solutions used to clean scissors are a source of sample contamination. DBS were prepared from 3 drops of blood from two Plasmodium falciparum cultures spotted on confetti, and 3 solutions (water, ethanol and DNase) were used to clean scissors between spots during DNA extraction. For each cleaning solution, two blank confetti were used as negative controls. Samples were analyzed by PCR-based genotyping of merozoite surface proteins 1 & 2 (msp-1 and msp-2), and P. falciparum chloroquine resistance transporter (Pfcrt). A total of 15 samples were analyzed. Based on msp-1 and msp-2 amplification, P. falciparum was detected on blank confetti when scissors were washed with ethanol. Based on Pfcrt amplification, DNA of P. falciparum was detected on blank confetti when scissors were cleaned with ethanol or water. No P. falciparum genes were detected on blank confetti when scissors were cleaned with DNase. Scissors cleaned with DNase prevented cross-contamination between samples during processing of dried blood spots, whereas ethanol (which is commonly used) fails to avert cross-contamination.

Analysis of complex DNA mixtures is limited by the selected genetic markers characterized and assay technology. Small panels of short tandem repeats (STRs) are the current standards in most countries. Extension of STRs with hundreds of single nucleotide polymorphisms (SNPs) is now commercially available. The current set of extension SNPs focus on phenotypic and biogeographic ancestry informative SNPs. Larger panels with thousands of SNPs have been developed that can identify up to 10 contributors to forensic DNA samples. Herein, multiple sets of SNPs were selected with the focus of increasing the upper limit of detectable contributors. Larger SNP panel sets with lower minor allele frequency (MAF) alleles can detect large numbers of contributors to in silico mixtures of randomly selected profiles from the 1,000 Genomes project. These results provide a path to enable characterization of complex DNA mixtures that are currently impossible to analyse.

Acute and chronic graft versus host disease (a/cGVHD) are major complications of allogeneic stem cell transplantation and a frequent cause of non relapse-mortality. The treatment of a/cGVHD consists of steroid administration at full dose. In the case of aGVHD, around 60 % of patients with grade III/IV fails to respond (steroid-resistant, SR), requiring a second line therapy. Extracorporeal photopheresis (ECP) represents a good option for second line treatment, leading to multiorgan response in several patients. Here, we have evaluated the performances of a cytofluorimetric quality control of the first ECP procedure in 29 patients with severe SR aGVHD who underwent ECP. The quality control represents a check control of in vitro cell apoptosis/death induction in collected and irradiated CD3+ cells by ECP. Our analysis shows that in more than 70 % quality controls a relevant apoptotosis/death of CD3+ cells was observed after cell irradiation, as compared to non-irradiated controls. Albeit observed in a small patients’ series, the occurrence of relevant in vitro effect after ECP relates to less procedures and/or to a better survival on ECP. Age, disease, gender, aGVHD grade and time from transplant to the first ECP had not impact on ECP efficacy in this patients’ series.

Aquaculture is rapidly becoming one of the pivotal sectors in the farm economy, driven by the increasing demand for high-quality animal protein at an affordable cost, especially with the escalating human population. However, the expansion of high-density fish populations also brings forth a challenge – the rapid transmission and spread of infectious disease agents among them. To combat this, vaccination is emerging as a reliable and standardized method for providing immunity against viral and bacterial outbreaks. The ideal vaccine is expected to be safe, effective, economical, and easily administered. The fish vaccination industry continually publishes new information on fish immunology and vaccinology, contributing to the improvement of vaccine formulation and efficacy. This review aims to offer insights into the current status of bacterial, viral, and parasitic diseases, discuss existing vaccinations, and address potential industry-threatening diseases like infectious edwardsiellosis, motile aeromonas septicemia, TiLV, salmon anemia, vibriosis, and white spot disease. Technological advancements have played a crucial role in enhancing our understanding of fish immunological mechanisms, leading to improved vaccine administration and the development of recombinant live attenuated, subunit, DNA, and RNA vaccines. However, challenges such as oral tolerance, vaccine degradation, and stressful environments persist, impacting vaccine efficacy. Addressing these challenges and gaining a deeper understanding of the fish immune system and host-pathogen interactions will be pivotal for future improvements, contributing to the sustainability of aquaculture and enhancing global food security.

The initiation reactions of DNA synthesis are central processes during human chromosomal DNA replication. They are separated into two main processes: the initiation events at replication origins, the start of the leading strand synthesis for each replicon, and the numerous initiation events taking place during lagging strand DNA synthesis. In addition, a third mechanism is the re-initiation of DNA synthesis after replication fork stalling, which takes place when DNA lesions hinder the progression of DNA synthesis. The initiation of leading strand synthesis at replication origin is regulated at multiple levels, from the origin recognition to the assembly and activation of replicative helicase, the Cdc45-MCM2-7-GINS (CMG) complex. In addition, the multiple interactions of the CMG complex with the eukaryotic replicative DNA polymerases such as DNA polymerase α-primase (Pol α), DNA polymerase δ and ε at replication forks play pivotal roles in the mechanism of initiation reactions including the initiation of signalling unperturbed and stalled replication forks, ‘replication stress’ events, via ATR (ATM-Rad 3-related protein kinase). All these processes are essential for the accurate transfer of the cells’ genetic information to their daughters. Failures and dysfunctions in these processes gives rise to genome instability followed by genetic diseases including cancer. Thus, in their review ‘Hallmarks of Cancer: New Dimensions‘ Hanahan and Weinberg (2022) called genome instability a fundamental function in the development process of cancer cells. In recent years, the understanding of the initiation processes and mechanisms of human DNA replication has made substantial progress at all levels which will be discussed in the review.

Objectives: Metabolic interactions amongst mutated genes in myelodysplastic syndrome (MDS) offer promising avenues for novel anticancer treatments. Our comprehensive study delves into these mutational and transcriptomic landscapes, pinpointing hallmark gene mutations and deregulated gene expression that could influence MDS patients’ metabolomes. Methods: The study utilized a retrospective, cross-sectional approach, employing mutational data on the cBio Cancer Genomics Portal conducted and reported by the University of Tokyo in 2011 and multi-center MDS cohorts regulated by Wellcome Trust Sanger Institute, United Kingdom, all in 2020. For transcriptomic data, we selected three publicly accessible independent cohorts on the Gene Expression Omnibus (GEO) database (GSE114922 in Wellcome Trust Centre for Human Genetics, United Kingdome, GSE63569 in University of Oxford, United Kingdome, and GSE183328 in CIMA, Spain) held in 2015, 2018, and 2022, respectively. To compile clinical, mutational, and transcriptomic data on MDS patients from multiple datasets and studies. This meta-analysis included genomic data derived from cellular genomics sources to assess mutations in specific genes, alongside an examination of transcriptomic data from three separate datasets that have been previously published. Results: DNMT3A presented a 20% mutation frequency, playing a pivotal role in MDS metabolomics. The DNMT3A gene mutations displayed significant mutual exclusivity with the SRSF2, ASXL1, JAK2, and TP53 genes. The mutational analysis also showed that the gene expression landscape in MDS is associated with alterations to DNA methylation pathways. Conclusion: This analysis suggests a potential therapeutic niche. Identifying signature genes in MDS that have metabolic and methylation affiliations could illuminate the disease's intricate biology and inspire novel treatments.

Euler sums are alternating (or level two) extension of multiple zeta values (MZVs). Kaneko and Tsumura initiated the study of multiple T-values (MTVs), another level two generalization, by restricting the summation indices in the definition of MZVs to a fixed parity pattern. In this paper, we shall study finite MTVs and their alternating versions which are level two and level four variations of finite MZVs, respectively. We conjecture that all finite MZVs are in the Q-span of finite MTVs which in turn apparently lie in the span of finite Euler sums, and the inclusions are both proper. We shall first provide some structural results for Euler sums of small weights, guided by the author's previous conjecture that the finite Euler sum space of weight w is isomorphic to a quotient Euler sum space of weight w. Then, by utilizing some well-known properties of the classical alternating MTVs, we shall derive a few important Q-linear relations among the finite alternating MTVs, including the reversal, linear shuffle and sum relations. We then compute the upper bound for the dimension of the Q-span of weight w finite (alternating) MTVs for w<9, both rigorously using the newly discovered relations and numerically aided by computers.

Our goal was to determine the cellular immune response (CIR) among a random sample of the Borriana COVID-19 cohort (Spain) to identify its associated factors and their relationship with infection, reinfection and sequelae. We conducted a nested case-control study using a randomly selected sample of 225 individuals age 18 and older, including 36 individual naïve to SARS-CoV-2 infection, and 189 infected patients. We employed flow cytometry-based immunoassays for intracellular cytokine staining, utilizing Wuhan and BA.2 antigens and chemoluminescence microparticle immunoassay for detection SARS-CoV-2 antibodies. Logistic regression models were used. A total of 215 (95.6%) participants exhibited T-cell response (TCR) to at least one antigen. Positive responses of CD4+ and CD8+ T cells were 89.8% and 85.3% respectively. No difference in CIR was found for naïve and infected patients. Patients who experienced sequelae exhibited higher CIR than those without. A positive correlation was observed between TCR and Anti-Spike IgG levels. Factors positive associated with TCR included A-blood group, number of SARS-CoV-2 vaccine doses received, and Anti-N IgM; factors inversely related were the time elapsed since the last vaccine dose or infection, and B-blood group. These findings contribute valuable insights into the nuance immune landscape shaped by SARS-CoV-2 infection and vaccination.

Bioinformatics is a rapidly developing field enabling scientific experiments through computer models and simulations. Considering the vast databases of biological data available, it is ex-tremely important to develop efficient methods and algorithms for their processing. Sequence comparison is the best method for studying the evolutionary interaction between genes. It is based on alignment – the process of arranging two or more sequences to achieve the maximum level of identity and degree of similarity. The paper presents a new version of the algorithm for pairwise DNA sequences alignment, based on a new method called CAT, where a dependency with a previous match and the closes neighbor are taken in consideration to increase uniqueness of the CAT profile and to reduce possible collisions, i.e. two or more sequence having same CAT profiles. This makes proposed algorithm suitable for finding exact match of a concrete DNA se-quence in a big set of DNA data faster. The generation of CAT profiles is made once before data has been uploaded to the database, allowing the profiles to be used as metadata for the sequenc-es. It consists of an algorithm to calculate a CAT profile against the selected reference sequences and an algorithm to compare two sequences based on the calculated CAT profiles. Improve-ments in generation of the CAT profiles, are detailed described in the paper. Block scheme, pseudo code tables and figures were updated according to the proposed new version and ex-perimental results. Experiments have been carried out with the new version of the method and different datasets to align DNA sequences based on the CAT method. New experimental results in terms of collisions, speed, and efficiency of the proposed solutions are presented. Experiments related to the performance comparison with Needleman-Wunsch were re-executed with the new version of the algorithm to confirm that we have the same performance. An analysis of the per-formance of the proposed CAT based algorithm against Knuth–Morris–Pratt algorithm, which has a complexity of O(n) and is one of the most widely used for searching biological data, was performed. The impact of prior matching dependencies on uniqueness for generated CAT pro-files is investigated. The analysis of the experimental results obtained by sequence alignment shows a small deviation of the proposed algorithm based on the CAT method, which can be ig-nored if this deviation is acceptable at the expense of performance. The time efficiency of the CAT algorithm remains constant, regardless of the length of the sequences. Therefore, the ad-vantage of the proposed method is its fast processing in the alignment of large sequences, for which the execution of the exact algorithms takes a long time. The example code realization of the CAT Method, under the protection of the GNU General Public License v3.0, can be accessed on GitHub at: https://github.com/HristoS/CATSequenceAnalysis.

This paper offers a theoretical and experimental examination of the concurrent production of electricity and heat using photovoltaic thermal (PV/T) technology. The efficiency performance of the PV/T system is meticulously analyzed using MATLAB/Simulink software environments. Notably, the proposed PV/T system shows reliable performance, and its validity is confirmed through experimental validation on a test stand with a single PV/T panel positioned at a 45-degree angle to the horizontal and a 0-degree azimuth angle.

Oak woodlands are a keystone ecosystem that may not be sustainable under ongoing fire management regimes and climate change. For example, in 2014, sassafras saplings started to emerge in degraded woodlands at the 1200 ha Drury Conservation Area as part of research at the Bull Shoals Field Station in Taney County, Missouri where burning was initiated in 1999. After Spring (March) 2021 burnt, we examined trend in the potential community and ecosystem effects, we assessed sapling density, height, and specific leave weigh (SLW) of white oak and sassafras (<1.0 m) from fall 2021 through fall 2022. In fall 2021, sassafras density was 560/0.1ha with mean height of 25.42cm and SLW of 0.00436g/cm² while white Oak density was 171/0.1ha with mean height of 10.2cm and SLW of 0.00680g/cm. However, in fall 2022, Sassafras density was 582/0.1ha with mean height of 71.689cm and SLW of 0.00366 g/cm² meanwhile, white Oak density was 213/0.1ha with mean height of 50.544cm and SLW of 0.00563 g/cm². We analyzed mass loss for sassafras and white oak after buried for 198 days. The weight loss for sassafras was 0.857g while oak was 0.790g for 61 days of buried. Subsequent sassafras weight loss was 1.942g, 3.343g, 4.222g and 4.82g while that of white Oak was 1.877g, 2.862g, 3.344g and 4.28g for day 105, 153 and 198 respectively. The nutrients composition and carbon sequestration of sassafras and white oak species need to be determined to better understand the effect of their dominance in fire management regime of degraded oak woodland in Southwest Missouri Ozarks.

Beyond factors such as smoking and obesity, the risk of getting gastrointestinal cancer is passed down from parents to offspring, inherited by a gene mutation, predisposing them to develop cancer in their lifetime. Here we describe the clinical history of family members affected by gastrointestinal pathologies often leading to cancer. In the epidemiological study, the cohort of members of this family shared a high risk of developing gastrointestinal diseases. The subjects were monitored from May 2006 to December 2017 by collecting periodically clinical and endoscopic data, and determining both auto-modification of lymphocyte Poly(ADP-ribose)Polymerase as early signal of DNA damage, and erythrocyte membrane lipid composition (Fat Profile). Both biomarkers were demonstrated to be non invasive and easy tools, to monitor over time family members, first the oldest members (nine siblings), and thereafter their offspring, whose cell status evolved from physiological even to cancerous one. All subjects developed gastrointestinal pathologies of different kind and seriousness. Some diseases evolved to cancer, sometimes as a sudden and lethal event. The results of the two molecular approaches, agreed and even predicted the clinical and imaging paths. This cohort study supported the hypothesis that both noninvasive molecular analyses can predict altered cell states and support clinical and imaging tests.

The allosteric SERCA (Sarcoplasmic/Endoplasmic Reticulum Ca2+-ATPase) activator CDN1163 has been recently added to the group of pharmacological tools for probing SERCA function. We chose to investigate the effects of the compound on T lymphocyte Ca2+ stores, using the well-described Jurkat T lymphocyte as a reliable cell system for Ca2+ signaling pathways. Our study identified the lowest concentrations of the SERCA inhibitors thapsigargin (TG) and 2,5-di-(tert butyl)-1,4-benzohydroquinone (tBHQ) capable of releasing Ca2+, permitting the differentiation of the TG-sensitive SERCA 2b Ca2+ store from the tBHQ-sensitive SERCA 3 Ca2+ store. We proceeded to test the effects of CDN1163 on Ca2+ stores, examining specific actions on the SERCA 2b and SERCA 3 Ca2+ pools using our low dose SERCA blocker regimen. In contrast to previous work, we find CDN1163 exerts complex time-sensitive and SERCA isoform-specific actions on Ca2+ stores. Surprisingly, short-term exposure (0-30 minutes) to CDN1163 perturbs T cell Ca2+ stores by suppressing Ca2+ uptake with diminished Ca2+ release from the SERCA 2b-controlled store. Concomitantly, we find evidence for a SERCA-activating effect of CDN1163 on the SERCA 3 regulated store, given the observation of increased Ca2+ release inducible by low dose tBHQ. Intriguingly, longer-term (&gt;12 hours) CDN1163 exposure reversed this pattern, with increased Ca2+ release from SERCA 2b-regulated pools, yet decreased Ca2+ release responses from tBHQ-sensitive SERCA 3 pool. Our results reveal differential effects of CDN1163 on Ca2+ stores regulated by distinct SERCA isoforms, prompting the need for careful interpretation of the compound’s effects, particularly in cells expressing multiple SERCA isoforms.

The ability of recombinant, SARS-CoV-2 Spike (S) protein to modulate the production of two COVID-19 relevant, pro-inflammatory cytokines (IL-6 and IFN-γ) in PBMC cultures of healthy, pre-Covid-19 subjects was investigated. We observed that cytokine production was largely and diversely modulated by the S protein depending on antigen or mitogen stimulation, as well as on the protein source, insect (S-in) or human (S-hu) cells. While both proteins co-stimulated cytokine production by polyclonally CD3-activated T cells, PBMC activation by the mitogenic lectin Concanavalin A (Con A) was up modulated by S-hu protein and down-modulated by S-in protein. These modulatory effects were likely mediated by the S glycans as demonstrated by direct Con A-S binding experiments and use of yeast mannan as Con A binder. While being ineffective in modulating memory antigenic T cell responses, the S proteins and mannan were able to induce IL-6 production in unstimulated PBMC cultures and upregulate the expression of Mannose Receptor (CD206), a marker of anti-inflammatory M2 macrophage. Our data point to a relevant role of N-glycans, particularly N-mannosidic chains, decorating the S protein in the immunomodulatory effects here reported. These novel biological activities of the S glycan ectodomain may add to the comprehension of Covid-19 pathology and immunity to SARS-CoV-2.

Before the emergence of SARS-CoV-1, MERS-CoV, and most recently SARS-CoV-2, al-ready four other coronaviruses have been circulating in the human population. The alpha coronaviruses NL63 and 229E and the beta coronaviruses OC34 and HKU1. These circulating coronaviruses all cause mild respiratory illness during the winter seasons and most people are already infected in early life. Could antibodies and/or T cells, es-pecially against the beta coronaviruses have offered some form of protection against SARS-CoV-2? Related is the question whether survivors of SARS-CoV-1 or MERS-CoV would be relatively protected against SARS-CoV-2. More importantly, would humoral and cellular immunological memory, generated during the SARS-CoV-2 pandemic, ei-ther by infection or vaccination offer protection against future coronaviruses? Or rather than protection, could antibody dependent enhancement have taken place, a mecha-nism by which circulating corona antibodies enhance the severity of COVID-19? An-other related phenomenon, the original antigen sin, would also predict that the effec-tivity of the immune response to future coronaviruses would be impaired because of reactivation of memory against irrelevant epitopes. Current available evidence indicates that latter scenarios are highly unlikely and that especially cytotoxic memory T cells directed against conserved epitopes of human coronaviruses could at least offer partial protection against future coronaviruses.

Purpose: Pancreatic cancer (PAAD) is one of the most lethal malignancies. Immunotherapy is largely ineffective in PAAD. T-cell exhaustion contributes to the resistance for immunotherapies. We investigated the prognostic potential of T cell exhaustion-related genes (TEXGs). Methods: Single-cell RNA (scRNA) sequencing dataset from TISCH, and bulk sequencing datasets from TCGA and GTEx were applied to screen differently expressed TEXGs. Kaplan-Meier survival, LASSO regression, and Univariate/Multivariate Cox regression analysis were performed to construct TEXGs risk model. This model was used to predict the prognosis, tumor immune microenvironment, and immunotherapy response. PAAD cohorts from ICGC and GSE71729 datasets were used to validate this risk model. Pan-cancer expression of SPOCK2 was performed in TISCH database. Results: A six-gene (SPOCK2, MT1X, LIPH, RARRES3, EMP1, and MEG3) risk model was constructed. Low-risk PAAD patients had longer survival time in both training dataset (TCGA-PAAD, n=178) and validation datasets (ICGC-PACA-CA, ICGC-PAAD-US, and GSE71729, n=412). Multivariate Cox regression analysis demonstrated that the risk score was an independent prognostic variable for PAAD. High-risk patients correlated with cancer-related KEGG pathway enrichment, higher mutation frequency of KRAS and TP53 genes, and immunosuppressive status. Immunohistochemistry staining confirmed the changes of TEXGs in clinical samples. Moreover, pan-cancer scRNA sequencing datasets from TISCH analysis indicated that SPOCK2 might be a novel marker of exhausted CD8+ T cells. Conclusion: In conclusion, we established and validated a T cell exhaustion-related prognostic signature for PAAD patients. Moreover, our study suggests that SPOCK2 may be a novel marker of exhausted CD8+ T cells.

Nucleocytoplasmic large DNA viruses (NCLDVs) have massive genomes and particle sizes compared to other known viruses. NCLDVs, including poxviruses, encode ATPases of the FtsK/HerA superfamily to facilitate genome encapsidation. However, their biochemical and structural characteristics are yet to be discerned. In this study, we demonstrate that the viral ATPases are significantly shorter than their bacterial homologs, representing the minimal ATPase core of the FtsK/HerA superfamily. We analyzed the sequence and structural features of the vaccinia virus A32 protein and determined their roles in the stability of the A32 protein and its ATPase activity. We sought to purify A32 by various techniques and noted that recombinant A32 expressed in E. coli is highly insoluble and unstable in solution, possibly because of the disordered C-terminus. N-terminal fusion with the thioredoxin solubility tag could alleviate this issue to some extent, but subsequent tag cleavage results in increased susceptibility to precipitation and degradation. We also predicted a conserved coiled-coil motif (CCM) towards the C-terminus of VV A32. ATPase activity of A32 is known to increase in the presence of DNA. Our comparative analysis of wildtype A32 versus CCM mutants suggests that this DNA dependence of A32’s ATPase activity is likely regulated by the CCM. Since CCM is also known to facilitate protein oligomerization, these findings provide new opportunities for further detailed characterization of A32.

The Bunyavirales order includes at least fourteen families with diverse but related viruses, which are transmitted to vertebrate hosts by arthropod or rodent vectors. These viruses are responsible for an increasing number of outbreaks worldwide and represent a threat to public health. Infection in humans can be asymptomatic, or it may present with a range of conditions from a mild, febrile illness to severe hemorrhagic syndromes and/or neurological complications. There is a need to develop safe and effective vaccines, a process requiring better understanding of the adaptive immune responses involved during infection. This review highlights the most recent findings regarding T cell and antibody responses to the five bunyavirus families with known human pathogens (Peribunyaviridae, Phenuiviridae, Hantaviridae, Nairoviridae, and Arenaviridae). Future studies that define and characterize mechanistic correlates of protection against bunyavirus infections or disease will help inform the development of effective vaccines.

To better understand how immunity develops against SARS-CoV-2 during the pandemic period of 2020 to 2022, we analyzed the immune response of a small group of university staff and students who were either infected or vaccinated. We investigated the levels of receptor-binding domain (RBD)-specific and nucleocapsid (N)-specific IgG and IgA antibodies in serum and saliva samples taken early (around 10 days after infection or vaccination) and later (around 1 month later), as well as N-specific T-cell responses. One patient who had been infected in 2020 developed serum RBD and N-specific IgG antibodies, but declined 8 months later, then mRNA vaccination in 2021 produced a higher level of anti-RBD IgG than natural infection. In the vaccination of naïve individuals, vaccines induced anti-RBD IgG, but it declined after 6 months. A third vaccination boosted the IgG level again, albeit to a lower level than after the second. In 2022, when the Omicron variant became dominant, familial transmission occurred even among vaccinated people. In infected individuals, the levels of serum anti-RBD IgG antibodies increased later, while anti-N IgG peaked earlier. The N-specific activated T cells expressing IFN- or CD107a were detected early. In saliva, SARS-CoV-2-specific IgG but not IgA was detected. Interestingly, two individuals showed a temporary peak in RBD- and N-specific IgA antibodies in their saliva on the second day after infection. Thus, SARS-CoV-2 infection triggers typical immune responses in humans against acute viral infections, but mucosal immune responses, which are temporarily induced early during infection, may be more important for protection than current intramuscular vaccines.

Environmental DNA (eDNA) sampling has emerged as a powerful approach for monitoring biodiversity in the context of diverse terrestrial and aquatic ecosystems, and has broad potential for both environmental surveys and species management. eDNA studies focusing on plant and animal species have capitalized on analytical methodologies developed originally for metagenomic studies of microorganisms, and on technological improvements in sample collection. However, while eDNA studies have proliferated rapidly, efforts to standardize sampling and analytical methods are still nascent. Critically, fundamental data concerning the distribution and persistence of eDNA in different ecological contexts are lacking, complicating both confidence in species detection and comparability between studies. We propose that key deficits could be addressed by using well-designed synthetic DNA standards or tracers, which are widely used in fields ranging from medical diagnostics to hydrology. In-field application of a standardized mixture of barcoded synthetic DNA fragments of varying lengths and concentrations that are recovered concommitantly with eDNA sampling should provide critical yet missing data on environmental DNA distribution and turnover, as well as an internal molecular standard to enhance both species detection and comparability between eDNA studies.

With end-to-end encryption for video streaming services becoming more popular, network administrators face new challenges in preserving network performance and user experience. Video ads may cause traffic congestion and poor Quality of Experience. Because of the natural variation in user interests and network situations, traditional algorithms for increasing QoE may face limitations. To solve this problem, we suggest a novel method that uses user facial emotion recognition to deduce QoE and study the effect of ads. We use open-access Face Emotion Recognition (FER) datasets and extract facial emotion information from actual observers to train machine learning models. Participants were requested to watch ad videos and provide feedback, which will be used for comparison, training, testing, and validation of our suggested technique. Our tests show that our approach beats the ITU-T P.1203 standard in terms of accuracy by 37.1%. Our method provides a hopeful answer to the problem of increasing user engagement and experience in video streaming services.

The molecular mechanism how the infecting DNA of bacteriophage T4 is passing from the capsid through the bacterial cell wall and entering the cytoplasm is essentially unknown. After adsorption, the short tail fibers of the infecting phage extend from the baseplate and trigger the contraction of the tail sheath, leading to a puncturing of the outer membrane by the tail tip needle composed of the proteins gp5.4, gp5 and gp27. To explore the events that occur in the periplasm and at the inner membrane, we have constructed T4 phage that have a modified gp27 in their tail tip with a His-tag. Shortly after infection with these phages, the cells were chemically cross-linked, solubilised, and the cross-linked products were affinity purified on a nickel column. The co-purified proteins were identified by mass spectrometry, and we found that predominantly the inner membrane proteins DamX, SdhA and PpiD were cross-linked. The same partner proteins were identified when purified gp27 was added to Escherichia coli spheroplasts suggesting a direct protein-protein interaction.

In archaea, the interaction of HerA helicase and NurA nuclease is required for 3' end resection during DNA double-stranded break repair, and its disruption results in unfaithful DNA repair. Interactions transpire by the four conserved domains, including the N-terminal HAS domain (HerA-ATP synthase), which serves as a binding site for the exonuclease NurA. As a HerA hexamer between its two subunits, the conserved C-terminal brace extension functions as an adjacent brace. DBD serves as a location for DNA binding, which causes the DNA strands to be propelled into the HerA pocket to begin unwinding. The putative pore entrance is made up of four helix coils, which provide a suitable charged surface for guiding DNA into the pore. The crystal structure of the complex HerA-NurA allows for the identification of key residues for protein-protein interaction. This allows us to predict that most archaeal HerA orthologs will be able to interact with one another. Archaeal HerA is a homo-hexameric assembly featuring a DNA-binding domain that is conserved. It also features distinctive ATP and NurA binding domains. Following the structure and sequence conservation criteria, we find one HerA ortholog in Pyrobaculum calidifontis and multiple HerA paralogs, but the role of none is established yet.Interestingly, orthologous HerA is part of an Mre11-Rad50 gene neighborhood while paralogous HerA is not, which could be misleading. As a result, it sought to describe the criteria that may be used as a possible fingerprint for identifying orthologous genes, as well as the cloning procedures and tactics that would aid in the confirmation of the HerA helicase in P. calidifontis. Their confirmation is based on PCR cloning and characterization following HerA construct creation.

The analysis of autosomal polymorphism (STR) or Y chromosome polymorphisms (Y-STR) are usually used for the study and deconvolution of mixed DNA profiles in forensic genetics, accompanying data interpretation with biostatistical evaluations (e.g. RMP, RMNE, LR). Sometimes, however, some mixed DNA profiles are so complex that the autosomal and Y markers do not allow a correct discrimination and identification. In this work is reported a robbery case in which the analysis of the polymorphic markers of the X chromosome was applied to the mixed profiles obtained from the traces, thus falling outside the classic relationship use of the X-STRs. The aim of the authors is, in fact, to encourage the classic use of X-STRs not only in parental relationships, but also in purely forensic cases for interpreting complex mixed DNA profiles. Their application in case resolution could be more decisive than STR/Y-STR.

Human RAD51 protein (HsRad51)-promoted DNA strand exchange, a crucial step in homologous recombination, is regulated by proteins and calcium ions. The activator protein Swi5-Sfr1 and Ca2+ ions stimulate different reaction steps and induce a perpendicular orientation of DNA bases in the presynaptic complex. To investigate the importance of base orientation in the strand exchange reaction, we examined the Ca2+ concentration dependence of strand exchange activities and structural changes in the presynaptic complex. Our results show that optimal D-loop formation (strand exchange with closed circular DNA) requires Ca2+ concentrations greater than 5 mM, while 1 mM is sufficient for strand exchange between two oligonucleotides. The structural change, which is evidenced by an increase in fluorescence intensity of poly(dεA) (a poly(dA) analog), reaches a plateau at 1 mM Ca2+. Meanwhile, the linear dichroism signal intensity at 260 nm, which is indicative of rigid perpendicular DNA base orientation, requires >2 mM Ca2+ for saturation and thus correlates with the stimulation of D-loop formation. Therefore, Ca2+ exerts two different effects. Thermal stability measurements suggest that HsRad51 binds two Ca2+ ions with KD values of 0.3 mM and 2.5 mM, implying that one step is stimulated by one Ca2+ bond and the other by two Ca2+ bonds. We further discuss the parallels between Mg2+ activation of RecA and Ca2+ activation of HsRad51.

Parkinson’s disease (PD) is one of the most common neurodegenerative disorders globally, and leads to an excessive loss of dopaminergic neurons in the substantia nigra of the brain. Circulating cell-free DNA (ccf-DNA) are double-stranded DNA fragments of different sizes and origins that are released into the serum and cerebrospinal fluid (CSF) due to cell death (i.e., necrosis and apoptosis) or are actively released by viable cells via exocytosis and NETosis. Methods: Using droplet digital polymerase chain reaction (ddPCR) we comprehensively analyzed and distinguished ccfDNA and ccf mtDNA in the serum and CSF of PD and control patients. Results: The quantitative analysis of serum ccf-DNA in PD patients demonstrated significant increase of ccf mtDNA and ccfDNA compared to healthy control patients and significantly higher copy of ccf mtDNA when compared to ccfDNA. Next, the serum ccf mtDNA levels significantly increased in male PD patients compared to healthy male controls. Furthermore, CSF ccf mtDNA in PD patients increased significantly compared to ccfDNA, and ccf mtDNA decreased in PD patients compared to healthy controls. These decreases were not statistically significant, but were in agreement with previous data. In-terestingly, ccf mtDNA increased in healthy control patients in both serum and CSF as compared to ccfDNA. Limitations: the small sample size of serum and CSF were the main limitations of this study. Conclusion: To the best of our knowledge, this is the first comprehensive study on serum and CSF of PD patients using ddPCR and indicating the distribution of the copy number of ccf mtDNA, as well as ccfDNA. If validated, we suggest that ccf mtDNA has greater than ccfDNA potential to lead the development of novel treatments for PD patients.

This study investigated the impact of seven-strain probiotic supplementation on the frequency of T helper 17 (Th17) cells, regulatory T (T-reg) cells, Treg/Th17 ratio, and their gene expression in individuals with severe traumatic brain injury (STBI). Probiotics can influence the gut microbiota, modulate the immune response, and prevent or alleviate certain immune diseases. However, it is necessary to clarify whether probiotics can effectively impact Th cells in STBI patient. Despite the great attention to improving the immune system in most diseases, there is currently no accurate mechanism for the use of probiotics in diseases with damage to the head injury, which can be a basis for the use of probiotics. The primary objective of this study is to address this research gap. The present study was a randomized, double-blind, placebo-controlled trial that enrolled 48 STBI patients. They were treated with probiotics or placebo: 4 capsule/day for 14 days. The pre- and post-study evaluation included examining the frequency of Treg, Th17 cells, as well as the expression of associated immunological parameters (Foxp3 and RORγt) and Treg/Th17 ratio in both groups. No significant differences were found between the two groups in terms of subject characteristics (P>0.05). GCS scores at 14 days were significantly higher in the probiotic compared to the placebo group. The probiotic group showed a significant decrease in Th17 cell and RORγt gene expression (P = 0.022 and P< 0.001, respectively), a notable increase in T-reg and FoxP3 gene expression (P= 0.025 and P< 0.001, respectively), and a higher Treg/Th17 ratio compared to the placebo group. Our study provides promising evidence of the beneficial effects of seven-strain probiotic supplementation on Th cells in individuals with STBI.