Pluripotent stem cells (PSCs) can differentiate into three germ layers and diverse autolo-gous cell lines. Being among the most commonly used large domesticated hoofed animals and serving as a vital source of food and bioreactors, cattle are among the many species in which PSCs have been developed. Bovine PSCs (bPSCs) are the "seed cells" of regenerative science and have immense potential to advance species propagation, genetic engineering, and disease treatment. Moreover, they can aid in screening new drugs for efficacy and toxicity, model human diseases, and establish a sustainable agricultural system that can cater to the needs of the growing global population. Establishing stable bPSCs in vitro has been a critical scientific challenge, and researchers have made numerous attempts to address this issue. This review primarily delves into the current research progress of bPSCs, the challenges faced, and their potential applications.

The cellular cytoskeleton provides the cell with a mechanical rigidity which allows mechanical interaction between cells and the extracellular environment. The actin structure plays a key role in mechanical events like motility, or establishment of cell polarity. From the earliest stages of development, as represented by ex vivo expansion of naïve embryonic stem cells (ESCs), the critical mechanical role of the actin structure is becoming recognized as a vital cue for correct segregation and lineage control of cells and as a regulatory structure that controls several transcription factors. Naïve ESCs have a characteristic morphology and the ultrastructure that underlies this condition remains to be further investigated. Here, we investigate the 3D actin cytoskeleton of naïve mouse ESCs using super resolution optical reconstruction microscopy (STORM). We investigate the morphological, cytoskeletal and mechanical changes in cells cultured in 2i or Serum/LIF media reflecting a homogenous preimplantation cell state and a state that is closer to embarking on differentiation. STORM imaging showed that the peripheral actin structure undergoes a dramatic change between the two media conditions. We also detected micro-rheological differences in the cell periphery between the cells cultured in these two media correlating well with the observed nano-architecture of the ESCs in the two different culture conditions. These results pave the way for linking physical properties and cytoskeletal architecture to cell morphology during early development.

The differentiation of ESCs into cardiomyocytes in vitro is an excellent and reliable model system for studying normal cardiomyocyte development in mammals, modeling cardiac diseases, and for use in drug screening. Mouse ESC differentiation still provides relevant biological information about cardiac development. However, the current methods for efficiently differentiating ESCs into cardiomyocytes are limiting. Here, we describe a “WNT Switch” method to efficiently commit mouse ESCs into cardiomyocytes using the small molecule WNT signaling modulators CHIR99021 and XAV939 in vitro. This method significantly improves the yield of beating cardiomyocytes, reduces number of treatments, and is less laborious.

Mouse embryonic stem cells (mESCs) possess remarkable characteristics of unlimited self-renewal and pluripotency, which render them highly valuable for both fundamental research and clinical applications. A comprehensive understanding of the molecular mechanisms underlying mESC function is of utmost importance. The Human Silence Hub (HUSH) complex, comprising FAM208A, MPP8, and periphilin, constitutes an epigenetic silencing complex involved in suppressing retroviruses and transposons during early embryonic development. However, its precise role in regulating mESC pluripotency and differentiation remains elusive. In this study, we generated homogenous miniIAA7 tagged Mpp8 mouse ES cell lines. Upon induction of MPP8 protein degradation, we observed impaired proliferation and reduced colony formation ability of mESCs. Furthermore, this study unveils the involvement of MPP8 in regulating the activity of the LIF/STAT3 signaling pathway and Nanog expression in mESCs. Finally, we provide compelling evidence that degradation of the MPP8 protein impairs the differentiation of mESC.

Far-red fluorescent reporter genes can be used for tracking cells non-invasively in vivo using fluorescence imaging. Here, we investigate the effectiveness of the far-red fluorescent protein, E2-Crimson (E2C), for tracking mouse embryonic cells (mESCs) in vivo following subcutaneous administration into mice. Using a knock-in strategy, we introduced E2C into the Rosa26 locus of an E14-Bra-GFP mESC line, and after confirming that the E2C had no obvious effect on the phenotype of the mESCs, we injected them into mice and imaged them over 9 days. The results showed that fluorescence intensity was weak, and cells could only be detected when injected at high densities. Furthermore, intensity peaked on day 4 and then started to decrease, despite the fact that tumour volume continued to increase beyond day 4. Histopathological analysis showed that although E2C fluorescence could barely be detected in vivo at day 9, analysis of frozen sections indicated that all mESCs within the tumours continued to express E2C. We hypothesise that the decrease in fluorescence intensity in vivo was probably due to the fact that the mESC tumours became more vascular with time, thus leading to increased absorbance of E2C fluorescence by haemoglobin. We conclude that the E2C reporter has limited use for tracking cells in vivo, at least when introduced as a single copy into the Rosa26 locus.

Mixed lineage leukemia 1 (MLL1) introduces 1-, 2- and 3-methylation into histone H3K4 through the evolutionarily conserved set domain. In this study, bovine embryonic stem cells (bESCs, named bESCs-F7) were established from the in vitro fertilized (IVF) embryos by Wnt signaling inhibition, while its contribution to endoderm in vivo is limited. To improve the quality of bESCs, MM-102, an inhibitor of MLL1, was applied to the culture. The results showed that MLL1 inhibition along with GSK3 and MAP2K inhibition (3i) at the embryonic stage did not affect bESCs establishment and pluripotency. MLL1 inhibition improves the pluripotency and differentiation potentials of bESCs via up-regulation of stem cell signaling pathways such as PI3K-Akt and WNT. MLL1 inhibition decreases H3K4me1 modification at promoters in bESCs and altered the distribution of DNA methylation in bESCs. In summary, MLL1 inhibition enables bESCs to acquire better pluripotency, and its application may provide high quality pluripotent stem cells for domestic animals.

Malignant melanoma (MM) is the most fatal form of skin cancer, accounting for 90% of all skin cancer deaths. Multiple dysregulated cellular pathways are involved in MM, including the MAPK/ERK and PI3K/PTEN/AKT pathways, which can be targeted with novel targeted inhibitors. However, these blockades can be overcome through the activation of alternative pathways, leading to treatment resistance. The high immunogenicity of MM has been exploited using checkpoint inhibitors resulting in improved survival outcomes of patients with advanced MM. However, their side effect profile and prohibitive cost remain a challenge and the survival outcomes for patients with metastatic MM remain poor. Treatment resistance has been attributed to the presence of cancer stem cells (CSCs), a small subpopulation of pluripotent, highly tumorigenic cells proposed to drive cancer progression, metastasis, and treatment resistance. CSCs reside within the MM tumor microenvironment (TME) regulated by the immune system, and the paracrine renin-angiotensin system (RAS), expressed in many cancer types, including MM. This article reviews the current treatment of advanced MM with targeted therapy and immune checkpoint blockers, highlights the emergence of mRNA vaccine. It also discusses the MM TME and its relationship with CSCs, and the local immune system and the paracrine RAS. It highlights the regulation of the MAPK/ERK and PI3K/AKT/mTOR pathways by the RAS via prorenin receptor and angiotensin II receptor 1, and how this relates to the CSCs and treatment resistance. This underscores the potential multi-modal therapeutic strategy by concurrent targeting of multiple components of the TME.

The stromal vascular cell fraction (SVF) of visceral and subcutaneous adipose tissue (VAT and SAT) has increasingly come into focus in stem cell research, since these compartments represent a rich source of multipotent adipose-derived stem cells (ASCs). ASCs exhibit a self- renewal potential and differentiation capacity. Our aim was to study the different expression of embryonic stem cell markers NANOG, SOX2 and OCT3/4 and to evaluate if there exists a hierarchal role in this network in ASCs derived from both SAT and VAT. ASCs were isolated from SAT and VAT biopsies of 72 consenting patients (23 men, 47 women; age 45 ± 10; BMI between 25 and 30 range) undergoing elective open-abdominal surgery. Sphere-forming capability was evaluated by plating cells in low adhesion plastic. Stem cell markers CD90 and CD105 were analyzed by flow cytometry and stem cell transcription factors NANOG, SOX2 and OCT3/4 were detected by immunoblotting and Real-Time PCR. NANOG, SOX2 and OCT3/4 interplay was explored by gene silencing. ASCs from VAT and SAT confirmed their mesenchymal stem cell (MSC) phenotype expressing the specific MSC markers CD90, CD105, NANOG, SOX2 and OCT3/4. NANOG silencing induced a significant OCT 3/4 (70% ± 0.05) and SOX2 (75% ± 0.03) down-regulation whereas SOX2 silencing did not affect NANOG gene expression. Adipose tissue is an important source of MSC, and siRNA experiments endorse a hierarchical role of NANOG in the complex transcription network that regulates pluripotency and plasticity.

Infertility” is a condition defined by the failure to establish a clinical pregnancy after 12 months of regular, unprotected sexual intercourse or due to an impairment of a person’s capacity to reproduce either as an individual or with his/her partner’. The authors have set out to succinctly investigate, explore and assess infertility treatments harnessing the potential of stem cells to effectively and safely treat infertility, in addition to the legal and regulatory complexities at the heart of stem cell research, with an overview of the legislative state of affairs in six major European countries. In couples who cannot benefit from assisted reproductive technologies (ART) to treat their infertility, stem cells-based approaches have been shown to be a highly promising approach. Nonetheless, lingering ethical and immunological uncertainties require more conclusive findings and data before such treatment avenues can become mainstream and applied large scale. The isolation of human embryonic stem cells (ESCs) is ethically controversial, since their collection involves the destruction of human embryonic tissue. Overall, stem cell research has resulted in important new breakthroughs in the treatment of infertility. The effort to untangle the complex web of ethical and legal issues associated with such therapeutic approaches will have to rely on evidence-based, broadly shared standards, guidelines and best practices to make sure that the procreative rights of patients can be effectively reconciled with the core values at the heart of medical ethics.

Simple Summary: This review explores the origin of Leukemic Stem-Like Cells (LSLCs) from (Very) Small Leukemic Stem-Like Cells ((V)SLSLCs) and Cancer Stem Cells (CSCs) from trans-formed Very Small Embryonic-Like Stem Cells (VSELSCs). Understanding this relation is a step toward uncovering the mechanisms behind cancer and leukemia initiation and development. We explore how these very small precursor cell stages might contribute to the initiation and progres-sion of cancer as well as leukemia with a focus on Acute Myeloid Leukemia (AML) as a model. Abstract: Recent experimental findings indicate that CSCs originate from transformed VSELSCs, which are early precursor stages of healthy stem cells. This discovery marks a crucial step in understanding the mechanisms that drive cancer development and initiation. Our team's follow-up studies on leukemia, lung cancer, and healthy embryonic kidney cells have, for the first time, identified stag-es resembling very small precursor stem cells in continuously growing cell lines. These findings suggest the existence of (Very) Small Cancer Stem Cells ((V)SCSCs) or (V)SLSCs within respective subpopulations of CSCs and LSCs in vivo. In this review, we explore theoretical examples of how leukemia progresses through various stages, highlighting the role of these very small precursor cells as fundament primitive stages. We also discuss the potential implications of further research into these unique cellular stages for advancing cancer and leukemia treatment and prevention strategies.

Human pluripotent stem cells (hPSCs) have widespread potential biomedical applications. There is a need for large-scale in vitro production of hPSCs, and optimal culture methods are vital in achieving this. Physiological oxygen (2% O2) improves key hPSCs attributes, including genomic integrity, viability, and clonogenicity, however, its impact on hPSC metabolism remains un-clear. Here, Selected Ion Flow Tube-Mass Spectrometry (SIFT-MS) was used to detect and quantify metabolic Volatile Organic Compounds (VOCs) in the headspace of hPSCs and their differentiated progeny. hPSCs were cultured in either 2% O2 or 21% O2. Media was collected from cell cultures and transferred into glass bottles for SIFT-MS measurement. The VOCs acetaldehyde and dimethyl sulfide (DMS)/ethanethiol were significantly increased in undifferentiated hPSCs compared to their differentiating counterparts, and these observations were more apparent in 2% O2. Pluripotent marker expression was consistent across both O2 concentrations tested. Transcript levels of ADH4, ADH5, and CYP2E1, encoding enzymes involved in converting ethanol to acetaldehyde, were upregulated in 2% O2, and chemical inhibition of ADH and CYP2E1 decreased acetaldehyde levels in hPSCs. Acetaldehyde and DMS/ethanethiol may be indicators of altered metabolism pathways in physiological oxygen culture conditions. The identification of non-destructive biomarkers for hPSC characterization has the potential to facilitate large-scale in vitro manufacture for future biomedical application.

Mammalian fertilization initiates the reprogramming of oocytes and sperm, leading to the formation of a totipotent zygote. The zygotic genome remains transcriptionally silent throughout this intricate process, undergoing the maternal-to-zygotic transition (MZT) and subsequent zygotic genome activation (ZGA). Histone modifications are pivotal in shaping cellular identity and gene expression in many mammals. Recent advances in chromatin analysis enable a detailed exploration of histone modifications during ZGA. This review delves into conserved and unique regulatory strategies, providing essential insights into the dynamic changes in histone modifications and their variants during ZGA in mammals. The objective is to explore recent advancements in leading mechanisms related to histone modifications governing this embryonic development phase in depth. These considerations will be useful for informing future therapeutic approaches that target epigenetic regulation in diverse biological contexts, ranging from regenerative medicine to cancer research. It will also contribute to the extensive areas of evolutionary and developmental biology and possibly lay the foundation for future research and discussion on this seminal topic.

The peanut (Arachis hypogaea L.) is recognized as one of the most important legume crops globally for its use in human food; it is widely distributed and cultivated in tropical and subtropical regions. The purpose of this study was to evaluate the cryopreservation of five peanut varieties conserved in the INIAP Germplasm Bank, testing cryopreservation methods, evaluating the germination percentage of whole seeds and embryonic shoots. Subsequently, two quantitative variables, shoot length and root, were evaluated. The average germination percentage of varieties and treatments was higher when embryonic axes were isolated with 99.31% than 86.06% seeds. The best germination percentage of the five varieties for seeds and embryonic shoots was obtained by the Peruvian variety with 88.13% and 92.50%. The best treatments by variety for the germination of whole seeds and embryonic axes were obtained by the treatment (desiccation and NL) for whole seeds (GS2) with 95.42% and embryonic axes with 92.83%. Ageing and cryopreservation treatments positively affected germination and seedling vigor in whole seeds and embryonic axes. The two quantitative variables, shoot and root length showed variability between the five varieties; significant differences were observed between the four treatments evaluated for whole seeds and embryonic axes. The three treatments for whole seeds (GS1, GS2 GS3) and the non-cryopreserved control treatment (GSC), as well as the treatments for embryonic axes (GEA1, GEA2 GEA3) and the non-cryopreserved control treatment (GEAC), obtained good survival. They germinate whole seeds and embryonic axes with sprout development (aerial part) and root formation. With the most effective treatments for whole seeds (GS2) and embryonic axes (GEA2), the cryopreservation of the national peanut collection of the INIAP Germplasm Bank could be started.

To evaluate whether autologous mesenchymal cells, adipose derived stem cells and platelet-rich plasma, grafted into the supracoroideal space by surgical treatment according to Limoli retinal restoration technique (LRRT), can produce growth factors in order to exert a beneficial effect in retinitis pigmentosa (RP) patients. Twenty-one eyes underwent surgery and divided based on retinal foveal thickness ≤ 190 or >190 µm into group A and group B, respectively. The specific LRRT triad was grafted in a deep scleral pocket above the choroid of each eye. At 6-month follow-up, group B showed an improvement in residual close-up visus and sensitivity at microperimetry compared to group A. After an in-depth review of molecular biology studies concerning degenerative phenomena underlying the etiopathogenesis of RP, it can be confirmed that further research is needed on tapeto-retinal degenerations both from a clinical and molecular point of view to obtain better functional results. In particular, it is necessary to increase the number of patients, extend observation times, and treat subjects in the presence of still trophic retinal tissue to allow adequate biochemical and functional catering.

Temperature is a crucial environmental factor that affects embryonic development, particularly for marine organisms with long embryonic development periods. However, the sensitive period of embryonic development and the role of autophagy/apoptosis in temperature regulation in cephalopods, enigmatic creatures, remain unclear. In this study, we cultured embryos of Sepiella japonica, a typical species in the local area of the East China Sea, at different incubation temperatures (18 ℃, 23 ℃, and 28 ℃) to investigate various developmental aspects, including morphological and histological characteristics, mortality rates, the duration of embryonic development, and expression patterns of autophagy-related genes (LC3, BECN1, Inx4) and apoptosis marker genes (Cas3, p53) at 25 developmental stages. Our findings indicate that embryos in the high-temperature (28 ℃) group had significantly higher mortality and embryonic malformation rates than those in the low-temperature (18 ℃) group. Furthermore, high temperature (28 ℃) shortened the duration of embryonic development by 7 days compared to the optimal temperature (23 ℃), while low temperature (18 ℃) caused a delay of 9 days. Therefore, embryos of S. japonica were more intolerant to high temperatures (28 ℃), emphasizing the critical importance of maintaining an appropriate incubation temperature (approximately 23 ℃). Additionally, our study observed, for the first time, that the Early blastula, Blastopore closure, and Optic vesicle to Caudal end stages were the most sensitive stages. During these periods, abnormalities in the expression of autophagy-related and apoptosis-related genes were associated with higher rates of mortality and malformations, highlighting the strong correlation and potential interaction between autophagy and apoptosis in embryonic development under varying temperature conditions.

Endometriosis is a common finding among women with infertility and women who are diag-nosed with endometriosis are almost twice as likely to experience infertility. Mechanisms by which endometriosis causes infertility remain poorly understood. In this review we evaluate the current literature on the impact of endometriosis on oocyte and embryo quality. The presence of endometriosis evidently reduces ovarian reserve, oocyte quality and embryo quality, however this does not appear to translate to a clear clinical impact. Analysis of data from large assisted reproduction technology registries has shown that women with endometriosis have a lower oo-cyte yield but no reduction in reproductive outcomes. There is a need for future studies in the form of well-designed randomised controlled trials to further evaluate the role of surgical and medi-cal treatment options in women with endometriosis undergoing assisted conception.

The gene sets of the two cells produced by each division of the fertilized egg are different, and these differences gradually accumulate. When the similarity between a certain cell and a certain type of cell is below a threshold, the differentiation ability in that direction is lost. When all cells can only have one developmental direction, the expression of zygotic genes is turned on. When a stem cell differentiates, it first divides into two cells. One of two cells, similar to the next level cell, becomes the next level cell, while the other, similar to the mother cell, continues to become the other stem cell. When a cell undergoes dedifferentiation, it expresses a group of genes. When the gene set is similar to the upper level stem cell, the cell undergoes dedifferentiation and becomes the upper level stem cell.

Copper, which can potentially be a highly toxic agent, is an essential nutrient due to its role as a co-factor for cuproenzymes and participation in signaling pathways. In mammals, the liver is a central organ that controls copper turnover throughout the body: copper absorption, distribution, and excretion. In ontogenesis, there are two types of copper metabolism: embryonic and adult, which maintain the balance of copper in each of these periods, respectively. In the liver cells, these types are characterized by specific expression patterns and activity levels of the genes encoding ceruloplasmin, which is the main extracellular ferroxidase and copper transporter and proteins mediating ceruloplasmin metalation. In newborns, the molecular-genetic mechanisms responsible for copper homeostasis and the ontogenetic switch from embryonic to adult copper metabolism are highly adapted to milk ceruloplasmin as a dietary source of copper. In the mammary gland cells, the level of ceruloplasmin gene expression and the alternative splicing of its pre-mRNA govern the amount of ceruloplasmin in milk, and thus, the amount of copper absorbed by the newborn is controlled. In the newborns, absorption, distribution, and accumulation copper are adapted to milk ceruloplasmin. In the newborns, which are not breast-fed at the early stages of postnatal development, the control for alimentary copper balance is absent. We tried to focus on the neonatal consequences of a violation of the balance of copper in the mother / newborn system. Although there is still much to be learned, the time to pay attention to this problem came because the neonatal misbalance of copper may provoke the development of copper related disorders for future life.

It has been proposed that tumourigenicity was an intrinsic feature of embryonic/germ cell developmental axis as well as embryonic/germ cell-related genes play a crucial role in tumourigenicity. Our previous studies indicated that primordial germ cell (PGC)-like potential could be reactivated in tumourigenesis. In this study, 4T1, 168FARN and 67NR cells which originated from the same mouse breast cancer were studied and the results indicated that the acquisition of embryonic/germ cell-like state is essential for tumourigenicity. We further demonstrated that somatic to PGC-like transformation (SPLT) was activated in 4T1 cells and that inhibition of PGC-like cell formation by depleting pluripotency and/or PGC specification-related genes markedly repressed SPLT and the tumourigenicity. Collectively, our findings reveal that tumourigenicity is linked to the acquisition of PGC-like state through SPLT in 4T1 cells, provide new insight into deeper understanding the biological nature of tumours and novel therapeutical strategies for cancer targeting.

The use of C-type natriuretic peptide (CNP) in the interaction with the oocyte and in the temporary postponement of spontaneous meiosis resumption has already been well described. However, its action in pre-implantation developmental stage embryos remains to be understood. Thus, our study aimed to detect the presence of the canonical CNP receptor (natriuretic peptide receptor, NPR2) in germinal vesicle (GV), metaphase II (MII), presumptive zygotes (PZ), morula (MO), and blastocyst (BL) stage embryos and, later, to observe possible modulations on the embryos when co-cultured with CNP. In the Experiment I, we detected and quantified the NPR2 on the above-mentioned embryo stages. Following, Experiment II, intended to test different concentrations (100, 200, or 400 nM of CNP) at different times of inclusion in the in vitro culture (IVC; inclusion from the beginning, i.e. day 1, or from day 5). In Experiment III, it was used 400 nM of CNP on day 1 (D1) in the IVC, which did no demonstrate to be embryotoxic, and it showed potentially promising results in blastocyst production rate, when compared to the control. Thus, we analyzed the embryonic development rates of bovine embryos (D7) and hatching kinetics (D7, D8, and D9). Subsequently, morula and blastocyst were collected and evaluated for transcript abundance to competence and quality (apoptosis, oxidative stress, proliferation and differentiation) and lipid metabolism. Differences with probabilities less than P < 0.05, and/or Foldchange (FC) ˃1.5 were considered significant. We demonstrate the presence of NPR2 until the blastocyst development stage when there was a significant decrease in membrane receptors. There was no statistical difference in production rate after co-culture with 400 nM CNP. However, when we evaluated the abundance of morula transcripts, there was an upregulated transcription in ADCY6 (P=0.057), and downregulated transcripts, in BMP15 (P= 0.013), ACAT1 (P= 0.040), CASP3 (P=0.082). In addition, a total of 12 transcriptions in morula presented variation FC ˃1.5. In blastocysts, the treatment with CNP induced upregulation in BID, CASP3, SOX2, HSPA5 transcripts, and downregulation in BDNF, NLRP5, ELOVL1, ELOVL4, IGFBP4, and FDX1 transcripts (FC >1.5). Thus, our study identified and quantified the presence of NPR2 in bovine pre-implantation embryos. Furthermore, 400 nM of CNP in IVC, a concentration not previously described in the literature, modulated some transcripts related to embryonic metabolism and it was not embryotoxic morphologically.

The objective of this study was to evaluate the effect of in ovo injection of L-arginine (L-Arg) into Ross broiler eggs at different embryonic developmental stages on their survival, hatchability, and body weight (BW). Additionally, we have analyzed the levels of serum glutamic-oxaloacetic transaminase (SGOT) and serum glutamic-pyruvic transaminase (SGPT), protein expression of heat shock proteins (HSPs), also we have the determined micronuclei (MN) and nuclear abnormality (NA). Results showed that survival and hatching rates as well as body weight were increased on the 14th day incubation compared to 8th and 18th day incubation at lower concentration of L-Arg. Moreover, the levels of SGOT and SGPT were also significantly (P < 0.05) increased at 14th day incubation at the same concentration (100μg/μl/egg) of injection. In addition, IgM levels were increased on the 14th day incubation compared to other days. The protein expressions of HSP-47, HSP-60, and HSP-70 in the liver were significantly down-regulated whereas the expression of myogenin and MyoD were significantly up-regulated on the 14th day after incubation in treated with all different doses such as 100μg, 1000μg and 2500μg/μl/egg namely 3T1, 3T2 and 3T3 respectively. However, the treatment with low dose of L-Arg down-regulated expression levels of those proteins on the 14th day incubation. Histopathology of liver by hematoxylin and eosin (H&E) straining showed that the majority of liver damage, specifically intracytoplasmic vacuoles, were observed in 3T1, 3T2, and 3T3. The minimum dose of 100 μg/ml/egg on the 14th day of incubation significantly prevented intracytoplasmic vacuole damages. These results demonstrate that in ovo administration of L-Arg at (100μg/μl/egg) may be an effective method to increase chick BW, hatch rate, increasing muscle growth related proteins and promote the immune response through increasing IgM on the 14th day of incubation period.

In bovine embryos, microRNA (miRNA) expression has been profiled at each stage of early development in-vitro. miRNAomic analysis of spent media has the potential to reveal characteristics of embryo health, however, applications are limited without categorizing miRNA profiles by embryo quality. Time-lapse imaging has shown the timing of embryo development in-vitro may be indicative of their developmental potential. The aim of the study was to profile miRNAs in the spent media of slow and fast-growing bovine embryos throughout the pre-implantation period. Bovine cumulus-oocyte-complexes were aspirated from ovaries, fertilized, and cultured to blastocyst stage of development. At 2-cell, 8-cell, and blastocyst stage, each microdrop of 30 presumptive-zygotes were classified as slow or fast-growing based on the percentage of embryos that had reached the desired morphological stage. Following hybridization on a GeneChip miRNA 4.0 array, comparative analysis was conducted between spent media of slow and fast-growing embryos. In total, 34 differentially expressed miRNAs were identified between the comparison groups, with 14 of the miRNAs detected in the 2-cell samples, 7 miRNAs detected in the 8-cell samples, and 12 miRNAs detected in the blastocyst samples. The results demonstrate distinct miRNAs populations can be identified between slow and fast-growing embryos, highlighting novel biomarkers of developmental potential at each stage of pre-implantation development.

The onset of embryonic heart beating may be regarded as the defining feature for the beginning of personal human life. Clarifying the timing of the 1^st human heartbeat, therefore, has religious, philosophical, ethical, and medicolegal implications. This article reviews the historical and contemporary sources of knowledge on the beginning of human heart activity. Special attention is given to the problem of the determination of the true age of human embryos and to the problem of visualization of the human embryonic heart activity. It is shown that historical and current textbook statements about the onset of blood circulation in man do not derive from observations on living human embryos but derive from the extrapolation of observations on animal embryos to the human species. This fact does not preclude the existence of documented observations on human embryonic heart activity: Modern diagnostic (ultrasound) and therapeutic (IVF) procedures facilitate the visualization of early embryonic heart activity in precisely dated pregnancies. Such studies showed that the human heart started its pumping action during the 4^th post-fertilization week. A small number of direct observations on the heart activity of aborted human embryos were reported since the 19^th century but did not receive much recognition by embryologists.

The chicken embryo is a widely used experimental animal-model in many studies such as developmental biology and to study the physiological responses and adaptation to altered environments as well as for cancer and neurobiology research. Embryonic heart rate is an important physiological variable useful as an index reflecting the embryo's natural activity and is considered one of the most difficult parameters to measure. An acceptable measurement technique of embryonic heart rate should provide a reliable cardiac signal quality while maintaining adequate gas exchange through the eggshell along the incubation and embryonic developmental period. In this paper, we presented a detailed design and methodology for a non-invasive PPG-based prototype (Egg-PPG) for real-time and continuous monitoring of embryonic heart rate during incubation. An automatic embryonic cardiac wave detection algorithm, based on normalised spectral entropy, is described. The developed algorithm successfully estimated the embryonic heart rate with 98.7% accuracy. We believe that the developed overall system presented in this paper is showing a promising solution for non-invasion, real-time monitoring of embryonic cardiac signal, which can be used in both experimental studies (e.g., developmental embryology and cardiovascular research) and in industrial incubation applications.

Somatic cell nuclear transfer (SCNT) has great potential in the replication of high commercial value animals, threatened wild species for conservation purposes, and transgenic animals for biomedical purposes. However, SCNT has a low success rate, due to intrinsic factors of the technique itself, which will lead to low rates of embryonic development and epigenetic alterations in the cloned embryos. The objective of the present study was to evaluate the effect of OF-UF on the intracellular concentration of ROS, GSH, and the development of Ovis aries cloned and parthenogenetic embryos. The results did not show a beneficial effect on the development of parthenogenetic and cloned embryos at concentrations of 0.5% OF-UF. Furthermore, at 1% OF-UF, an adverse effect was observed in cloned embryos at the blastocyst stage and 2% OF and UF in parthenogenetic embryos in the first divisions. A decrease in ROS and GSH levels was observed in the parthenogenetic blastocysts treated with 1% OF-UF, but not in the clones, in which a higher concentration of GSH and a similar concentration of ROS was observed. No effect was observed of OF-UF on embryonic development and redox balance on sheep clone embryos by Handmade Cloning.

The members of the IgLON superfamily of cell adhesion molecules facilitate fundamental cellular communication during brain development, maintain functional brain circuitry, and are associated with several neuropsychiatric disorders. Usage of alternative promoter-specific 1a and 1b mRNA isoforms in Lsamp, Opcml, Ntm and the single promoter of Negr1 in the mouse and human brain has been previously described. To determine the precise spatiotemporal expression dynamics of Lsamp, Opcml, Ntm isoforms and Negr1, in the developing brain, we generated isoform-specific RNA probes and carried out in situ hybridization in the developing (embryonic, E10.5, 13.5, 17; post natal, P0) and adult mouse brains. We show that promoter-specific expression of IgLONs is established early during pallial development (at E10.5), where it remains throughout its differentiation through adulthood. In the diencephalon, midbrain and hindbrain, strong expression patterns are initiated a few days later and begin fading after birth, being only faintly expressed during adulthood. Thus, the expression of specific IgLONs in the developing brain may provide the means for regionally specific functionality as well as for specific regional vulnerabilities. The current study will therefore improve the understanding of how IgLON genes are implicated in the development of neuropsychiatric disorders.

Turner syndrome (TS) is a chromosomal disorder that is caused by a missing or structurally ab-normal second sex chromosome. Subjects with TS are at an increased risk of developing intrauterine growth retardation, low birth weight, short stature, congenital heart diseases, infertility, obesity, dyslipidemia, hypertension, insulin resistance, type 2 diabetes mellitus, metabolic syndrome, and cardiovascular diseases (stroke and myocardial infarction). The underlying pathogenetic mechanism of TS is unknown. The assumption that X chromosome-linked gene haploinsufficiency is associated with the TS phenotype is questioned since such genes have not been identified. Thus, other pathogenic mechanisms have been suggested to explain this phenotype. Morphogenesis encompasses a series of events that includes cell division, the production of migratory precursors and their progeny, differentiation, programmed cell death, and integration into organs and systems. The precise control of the growth and differentiation of cells is essential for normal development. The cell cycle frequency and the number of proliferating cells are essential in cell growth. 45,X cells have a failure to proliferate at a normal rate, leading to a decreased cell number in a given tissue during organogenesis. A convergence of data indicates an association between a prolonged cell cycle and the phenotypical features in Turner syndrome. This review aims to examine old and new findings concerning the relationship between a prolonged cell cycle and TS phenotype. These studies reveal a diversity of phenotypic features in TS that could be explained by reduced cell proliferation. The implications of this hypothesis for our understanding of the TS phenotype and its pathogenesis are discussed. It is not surprising that 45,X monosomy leads to cellular growth pathway dysregulation with profound deleterious effects on both embryonic and later stages of development. The prolonged cell cycle could represent the beginning of the pathogenesis of TS, leading to a series of phenotypic consequences in embryonic/fetal, neonatal, pediatric, adolescence, and adulthood life.

Transitions from seed to seedling represent a critical developmental step in the life cycle of higher plants, dramatically affecting plant ontogenesis and stress tolerance. The release from dormancy to acquiring the germination ability is defined by a balance of phytohormones, with a substantial contribution of abscisic acid (ABA), which inhibits germination. We studied the embryonic axes of Pisum sativum L. before and after radicle protrusion. Our previous work compared RNA sequencing-based transcriptomics in the embryonic axes isolated before and after radicle protrusion. The current study aims to analyze ABA-dependent gene regulation during the transition of embryonic axes from germination to post-germination stage. First, we determined the levels of abscisates (ABA, phaseic acid, dihydrophaseic acid, and neo-phaseic acid) using the ultra-high-performance liquid chromatography-tandem mass spectrometry. Second, we made a detailed annotation of ABA-associated genes using RNA sequencing-based transcriptome profiling. Finally, we analyzed the DNA methylation pattern in the promoters of the PsABI3, PsABI4, and PsABI5 genes. We showed that changes in the abscisate profile are characterized by the accumulation of ABA catabolites, and the ABA-related gene profile is accompanied by the upregulation of genes controlling the seedling development and the downregulation of genes controlling the water deprivation. The expression of ABI3, ABI4 and ABI5, which encode the crucial transcription factors during late maturation, was downregulated by more than 20-fold, and their promoters exhibited a high level of methylation already at the late germination stage. Thus, although ABA remains important, other regulators seems to be involved in the transition from seed to seedling.

Ferroptosis is a unique variety of non-apoptotic cell death, driven by massive lipid oxidation in an iron-dependent manner. Since Ferroptosis was introduced as a concept in 2012, it was shown it's essential role in the pathogenesis in neurodegenerative diseases and an important role in therapy-resistant cancer cells. Thus, a detailed molecular understanding of both canonical and alternative ferroptosis pathways are required. There is a set of widely used chemical agents to modulate ferroptosis using different pathway targets: Erastin blocks cystine-glutamate antiporter, system xc-; ML210 directly inactivate GPX4; L-buthionine sulfoximine (BSO) inhibits γ-glutamylcysteine synthetase, an essential enzyme for glutathione synthesis de novo. Most studies were focused on lipidomic profiling of model systems undergoing death in a ferroptotic modality.

The early embryonic heart is a multi-layered tube consisting of (1) an outer myocardial tube; (2) an inner endocardial tube; and (3) an extracellular matrix layer interposed between myocardium and endocardium, called “cardiac jelly” (CJ). During the past decades, research on CJ has mainly focused on its molecular and cell biological aspects. This review focuses on the morphological and biomechanical aspects of CJ. Special attention is given to (1) the spatial distribution and fiber architecture of CJ; (2) the morphological dynamics of CJ during the cardiac cycle; and (3) the removal/remodeling of CJ during advanced heart looping stages, which leads to the formation of ventricular trabeculations and endocardial cushions. CJ acts as a hydraulic skeleton displaying striking structural and functional similarities with the mesoglea of jellyfish. CJ not only represents a filler substance, facilitating end-systolic occlusion of the embryonic heart lumen. Its elastic components antagonize the systolic deformations of the heart wall and thereby power the refilling phase of the ventricular tube. Non-uniform spatial distribution of CJ generates non-circular cross sections of the opened endocardial tube (initially elliptic, later deltoid), which seem to be advantageous for valveless pumping. Endocardial cushions arise from non-removed remnants of the original CJ.

Seed longevity is a crucial trait for the seed industry and genetic resource preservation. To develop excellent cultivars with extended seed lifespans, it is important to understand the mechanism of keeping seed germinability long-term and to find useful genetic resources as prospective breeding materials. This study was conducted to identify the best cultivars with a high and stable seed longevity trait in the germplasm of rice (Oryza sativa L.) and to analyse the correlation between seed longevity and embryonic RNA integrity. Seeds from 69 cultivars of the world rice core collection selected by the NIAS in Japan were harvested in different years and subjected to long-term storage or controlled deterioration treatment (CDT). The long-term storage (4 °C, RH under 35%, 10 years) was performed on seeds harvested in 2010 and 2013. The seeds harvested in 2016 and 2019 were used for CDT (36 °C, RH of 80%, 40 days). Seed longevity and embryonic RNA integrity were estimated by a decrease in germination rate and RNA integrity number (RIN) after long-term storage, or CDT. The RIN value was obtained by electrophoresis of total RNA extracted from seed embryos. Seeds of ‘Vandaran (Indica)’, ‘Tupa 729 (Japonica)’, and ‘Badari Dhan (Indica)’ consistently showed higher seed longevity and embryonic RNA integrity both under long-term storage and CDT conditions, regardless of the harvest year. A strong correlation (R2 = 0.93) was observed between germination rates and RIN values of the seeds after long-term storage or CDT among nine cultivars selected based on differences in their seed longevity. The study findings revealed the relationship between rice seed longevity and embryo RNA stability and suggested potential breeding materials, including both Japonica and Indica cultivars, for improving rice seed longevity.

New technologies have resulted in a better understanding of blood and lymphatic vascular heterogeneity at the cellular and molecular levels. Meanwhile, we still need to learn more about the heterogeneity of the cardiovascular and lymphatic systems between different species at the anatomical and functional levels. Even the deceptively simple question of the functions of fish lymphatic vessels has yet to be conclusively answered. The most common interpretation assumes a similar dual setup of the vasculature in zebrafish and mammals: on the one hand, there is the cardiovascular, circulatory system, and on the other hand, the lymphatic vascular system, in which the unidirectional flow is derived from surplus interstitial fluid and returned into the cardiovascular system. A competing interpretation questions the identity of the lymphatic vessels in fish as at least some of them receive their flow from arteries via specialised anastomoses, neither requiring an interstitial source for the lymphatic flow nor stipulating unidirectionality. In this alternative view, the “fish lymphatics” are a specialised subcompartment of the cardiovascular system, called the “secondary vascular system” (SVS). Many of the contradictions found in the literature appear to stem from the fact that the SVS develops in part or its entirety from an embryonic lymphatic vascular system by transdifferentiation. Future research needs to establish the extent of embryonic transdifferentiation of lymphatics into SVS blood vessels. Similarly, more insight is needed into the molecular regulation of vascular development in fish. Most fish do feature more than the five VEGF genes and three VEGF receptor genes that we know from mice or humans, and the relative tolerance of fish to whole genome and gene duplications could underlie the evolutionary diversification of the vasculature. This review discusses the key elements of the fish lymphatics versus the SVS and attempts to draw a picture coherent with the existing data, including the phylogenetic knowledge.