Cellular Processes Caused by Human Herpesviruses Infection

1. Introduction

Human herpesviruses are large, enveloped, double-stranded DNA viruses that cause a variety of diseases and establish lifelong latent infections in the majority of the global population. The Herpesviridae family comprises nine viruses that are capable of causing human infections and is divided into three subfamilies: Alphaherpesvirinae, Betaherpesvirinae, and Gammaherpesvirinae[1]. Alphaherpesvirinae consists of Herpes simplex virus I (HSV-1), HSV-2, and Varicella zoster virus (VZV). Betaherpesvirinae includes Human cytomegalovirus (HCMV), Human herpesvirus 6A (HHV-6A), HHV-6B, and HHV-7. Epstein–Barr virus (EBV) and Kaposi's sarcoma herpesvirus (KSHV) belong to Gammaherpesvirinae.

Vestibular neuritis (VN) is a clinical condition in the field of otolaryngology characterized by acute and persistent peripheral vertigo caused by unilateral vestibular afferent nerve block. The cause of VN is still unknown, but the most common hypothesis is viral infection or reactivation, particularly by HSV (Table 1). Several researchers have reported herpes virus infection in the vestibular ganglion of patients with vestibular neuritis[2,3,4,5,6,7]. Arbusow reported the presence of HSV-1 DNA in both the human vestibular ganglion and vestibular nuclei[8], suggesting the potential migration of the virus to the human vestibular labyrinth[5]. Furthermore, HSV-1 DNA or HSV latency-related transcripts have been detected in vestibular ganglia removed during surgery in patients with Meniere's disease[4,9]. Herpesviruses have a tendency to invade sensory neurons, establish a latency period, and can be reactivated to cause disease. The initial infection or reactivation of the herpesvirus can profoundly affect cellular processes in the host. Mice inoculated with HSV-1 and HSV-2 into the middle ear exhibited hearing loss and vestibular dysfunction. HSV infection was observed in columnar epithelial cells of the infected mice in the stria vascularis, leading to apoptosis in a portion of the infected cells, while many uninfected cells in the spiral organ of Corti also underwent apoptosis. While vestibular ganglion cells did not undergo apoptosis, some of the cells experienced functional loss[10]. Mice inoculated with HSV-1 after auricle also exhibited vestibular dysfunction along with the death of vestibular ganglion cells[11]. In addition to the damage caused by virus infection to cells, the killing of host cells by immune cells may also contribute to the pathogenesis of vestibular neuritis. The coexistence of CD8+ cells and HSV-1 has been observed in the vestibular ganglion cells of patients with vestibular neuritis.

This article provides a summary of the alterations in cellular processes post-infection, using HSV-1 and HCMV as exemplars, with the aspiration that this knowledge will aid in the treatment of VN and other diseases instigated by human herpesviruses.

2. Herpes Simplex virus Type 1

2.1. The process of HSV-1 entering host cells, replication and assembly

HSV-1 has a spherical shape. The complete HSV-1 virus comprises double-stranded DNA, a nucleocapsid, teguments, envelope proteins, and a lipid envelope (Figure 1A)[12]. The nucleocapsid shell exhibits a symmetrical three-dimensional icosahedral structure. HSV-1 is primarily transmitted through close contact[13]. The entry of herpesvirus into human cells for receptor binding and membrane fusion requires the involvement of multifunctional viral glycoproteins on its surface[14,15,16]. HSV-1 carries a minimum of 12 different glycoproteins. The viral fusion protein glycoprotein B (gB) and the hetero-oligomers glycoprotein H/glycoprotein L (gH/gL) constitute the core entry glycoproteins of the herpesvirus[17]. Briefly, when HSV is adsorbed on the cell membrane surface, the initial non-specific binding between glycoprotein gC and/or gB and the heparan sulfate mucin (HSPG) on the cell surface reduces the spatial distance between the viral envelope and the cell membrane. gD can specifically bind to Herpes virus entry mediator (HVEM), nectin-1, nectin-2, or 3-O-sulfated heparan sulfate (3-OS-HS). This binding further initiates gH and gL. Subsequently, gH-gL transmits signals to gB [18]. gB undergoes a conformational change, inserts into the host cell membrane, and then refolds to fuse the cellular and viral membranes together (Figure 1B). The refolding of multiple gB trimers creates pores in the membrane, initiating the fusion process between the viral envelope and cell membrane. This process may enable the viral nucleocapsid and DNA to enter the cytoplasm and translocate to the nucleus[19,20].

The transportation of viral capsids and vesicles carrying viral glycoproteins in the cytoplasm is closely linked to microtubules, and their translocation along axons depends on microtubules[21,22,23,24]. HSV-I utilizes microtubules and actin for retrograde entry into cells along axons, as well as for retrograde transport during virus assembly and exit[25]. There are two types of axonal transport: fast and slow[26,27]. Fast axonal transport occurs in both cis and retrograde directions, transporting mitochondria, neurotransmitters, channel proteins, and more. In contrast, slow axonal transport occurs in a paracrine direction, transporting cytoskeletal components such as neurofilaments, microtubule proteins, and actin[26,28]. HSV-1 is actively directed to spread from neurons through the axonal cytoskeleton and molecular motors. Studies using time-lapse microscopy have shown that HSV-1 undergoes rapid axonal flow in both directions[29,30]. After the nucleocapsid is transported to the surrounding area of the nucleus, it can interact with the nuclear pore complex, and then dsDNA is injected into the nucleus through the nuclear pore[31].

Figure 2. Schematic diagram of intracellular replication and assembly of herpesviruses. Once the HSV-1 nuclear capsid enters the cell, it binds to the motor protein associated with the microtubule. Subsequently, the nucleocapsid is transported towards the nucleus via microtubules. Upon reaching the vicinity of the nucleus, the HSV-1 capsid injects its genomic DNA into the nucleus through the nuclear pore complex. The injected viral genomic DNA is targeted to the PML body within the nucleus. The replicating viral genomic DNA assembles with a nucleocapsid that consists of early proteins synthesized in the nucleus and late proteins synthesized in the cytoplasm. Subsequently, it crosses the nuclear membrane and enters the cytoplasm. During this process, the virus is initially coated with an envelope, which may originate from the inner membrane of the nuclear envelope. Subsequently, the viruses lose the initial envelope through fusion with the outer nuclear membrane and are released into the cytoplasm without an envelope. Upon arrival in the cytoplasm, the capsid is subsequently reenveloped within an intracellular organelle, where it acquires its mature envelope and completes tegumentation. The capsid undergoes secondary envelopment before being released from the cell. During this process, the nucleocapsid, which is now associated with tegument proteins, buds into the membrane of a cytoplasmic organelle, resulting in the formation of an enveloped virion inside a vesicle. The origin of organelle membranes in the secondary envelope is still controversial, with some suggesting that they may originate from membrane tubes derived from recycled endosomes or vesicles from the trans Golgi network. Viruses that have completed the secondary envelopment are released through exocytosis.

Figure 2. Schematic diagram of intracellular replication and assembly of herpesviruses. Once the HSV-1 nuclear capsid enters the cell, it binds to the motor protein associated with the microtubule. Subsequently, the nucleocapsid is transported towards the nucleus via microtubules. Upon reaching the vicinity of the nucleus, the HSV-1 capsid injects its genomic DNA into the nucleus through the nuclear pore complex. The injected viral genomic DNA is targeted to the PML body within the nucleus. The replicating viral genomic DNA assembles with a nucleocapsid that consists of early proteins synthesized in the nucleus and late proteins synthesized in the cytoplasm. Subsequently, it crosses the nuclear membrane and enters the cytoplasm. During this process, the virus is initially coated with an envelope, which may originate from the inner membrane of the nuclear envelope. Subsequently, the viruses lose the initial envelope through fusion with the outer nuclear membrane and are released into the cytoplasm without an envelope. Upon arrival in the cytoplasm, the capsid is subsequently reenveloped within an intracellular organelle, where it acquires its mature envelope and completes tegumentation. The capsid undergoes secondary envelopment before being released from the cell. During this process, the nucleocapsid, which is now associated with tegument proteins, buds into the membrane of a cytoplasmic organelle, resulting in the formation of an enveloped virion inside a vesicle. The origin of organelle membranes in the secondary envelope is still controversial, with some suggesting that they may originate from membrane tubes derived from recycled endosomes or vesicles from the trans Golgi network. Viruses that have completed the secondary envelopment are released through exocytosis.

DNA viruses, such as herpesviruses, replicate in specific inclusions within the nucleus. These inclusions, referred to as viral replication compartments (VRCs), are the sites where viral DNA replication, viral transcription, and virion assembly take place [32,33,34]. Compartmentalization is an essential feature in living organisms. Cellular organisms typically utilize cell membranes to partition cells into compartments. Moreover, eukaryotic cells possess membrane-free compartments, such as stress granules and Pbodies[35,36]. Certain compartments exhibit liquid properties and are formed through a process known as liquid-liquid phase separation (LLPS), analogous to the formation of oil droplets in water[37]. There is a hypothesis suggesting that the nuclear viral replication compartments (VRCs) of DNA viruses, such as HSV-1, are also phase-separated condensates[33,34,38]. Michael Seyffert demonstrated that the HSV-1 transcription factor ICP4 has the ability to induce protein condensation, thereby imparting liquid-like properties to the VRC[39].

Primary HSV-1 infections generally occur in the epithelial cells of oral and anal mucosa[25]. Following infections of the skin or mucosa innervated by sensory nerves, HSV-1 can undergo retrograde axonal transport to neuronal cell bodies. It can then establish lifelong latency within the dorsal root ganglia（DRG）[40,41] and trigeminal ganglia (TG) and can be reactivated, resulting in tissue damage. Clinical manifestations of HSV-1 infection are changeable, depending on host immune function and mode of viral transmission[42].

2.2. Host cell processes caused by HSV-1 infection

HSV-1 belongs to the lysogenic family of viruses, and its lytic replication results in the destruction of host cells. Cell aggregation is observed almost immediately after cells are infected with HSV-1, and the severity tends to increase with the number of infections [43]. According to Roizman et al [43], herpes simplex virus infection can lead to the production of multinucleated cells, which result from the fusion of functional cells with different phenotypic characteristics. In HSV-1-infected cells, Avitabile et.al [44] found that microtubules are partially broken, especially at the cell periphery, where the connection between the microtubule network and the plasma membrane appears to be lost. Subsequently, the microtubules form bundles around the nucleus, resulting in a near-spherical shape of the cells [44]. Heeg et al observed that infection with high doses of various strains of HSV-1 for two and a half hours resulted in cell rounding, accompanied by the breakdown of actin-containing microfilaments and the appearance of knob-like protuberances containing actin at the cell periphery[45]. Hampar et.al reported that HSV-1 infection of cells causes chromosome breaks, translocations, and fusions[46]. Roizman et al reported that protein synthesis must precede viral DNA synthesis in the early stages of HSV-infected cells[47]. Both functional and structural proteins required for viral proliferation are produced by the host cell's translation system. HSV has been observed to decrease protein synthesis and mRNA levels in host cells with the expression level of viral proteins rapidly increasing, accompanied by the rapid degradation of previously existing polyribosomes and some host cell mRNA[48]. Aubert and Blaho summarize that the manifestations of HSV-1 infection include (i) the loss of matrix binding proteins on the cell surface, leading to detachment; (ii) modifications of membranes; (iii) cytoskeletal destabilizations; (iv) nucleolar alterations; and (v) chromatin margination and aggregation or damage, as well as (vi) a decrease in cellular macromolecular synthesis[49].

Cellular autophagy, apoptosis, and necrosis pathways are crucial cellular processes that are interconnected to restrict the spread of pathogens by eliminating infected cells [50]. Viral proteins can interact with these signaling molecules, disrupting downstream signal transduction and promoting viral replication and spread. Dufour et al. demonstrated that the ribonucleotide reductase R1 subunit of HSV inhibits Caspase8, thereby protecting cells from apoptosis induced by tumor necrosis factor (TNF) α and Fas ligand [51]. Furthermore, the research group demonstrated that this HSV protein disrupts the structural domain interactions of the Toll interleukin (IL)-1 receptor, thereby inhibiting poly I:C-induced apoptosis in HeLa cells [52]. Moreover, in addition to inducing the formation of filopodia in infected cells to facilitate viral transmission through cell-to-cell contact [53], Us3 proteins disable Bad by inhibiting its phosphorylation [54], thereby safeguarding the cell against DNA fragmentation, nuclear disintegration, and apoptosis [53,55].

Autophagy is a crucial cellular process that involves the self-degradation and recycling of cellular components, including the cell membrane, cytoplasm, and organelles. It plays a role in eliminating misfolded proteins, damaged organelles, and intracellular pathogens. However, certain HSV proteins, such as US11 and ICP34.5, interfere with cellular autophagy. US11 is a ribosome-associated double-stranded RNA-binding protein that directly interacts with PKR [56]. On the other hand, ICP34.5 consists of a C-terminal structural domain and an N-terminal structural domain. The C-terminal domain recruits protein phosphatase 1 (PP-1) to inhibit PKR-mediated phosphorylation of eLF2α [57], while the N-terminal domain directly interacts with Beclin-1 to block autophagy [58]. In summary, there are multiple pathways through which the host induces apoptosis and autophagy in infected cells, and HSV employs its own proteins to interfere with certain steps in these pathways to protect the survival of infected cells and facilitate its own replication and dissemination.

2.3. Immune cell process caused by HSV-1 infection

The intrinsic and innate immune responses serve as the first line of defense against viral infections, including HSV. They work together to limit the spread of viral replication until the body develops an adaptive immune response. The intrinsic immune response is particularly effective during the initial HSV-1 infection and also contributes to the subsequent adaptive immune response [59].The innate immune response is initiated through the cellular expression of pattern recognition receptors (PRRs), which detect pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns [60,61]. This recognition stimulates the secretion of interferon (IFN) α, β, or γ, along with other cytokines [62]. These cytokines can act in an autocrine and paracrine manner and play a crucial role in controlling HSV infection and coordinating innate and adaptive immune responses.Among the PRRs, Toll-like receptors (TLRs) are involved in detecting HSV nucleic acids and proteins. TLRs 2, 3, and 9 are the major TLRs responsible for HSV detection [61,63]. Interaction between PAMPs and TLRs leads to IFN secretion [62]. TLR2 recognizes viral glycoproteins, TLR3 senses double-stranded RNA (dsRNA) produced during HSV replication, and TLR9 recognizes HSV DNA. TLR2 interacts with gH and gL on the viral envelope and signals through myeloid differentiation factor 88 (MyD88) [64,65]. TLR2 activation promotes the expression of pro-inflammatory cytokines, exerting antiviral effects. However, studies on TLR2-deficient mice infected with HSV have shown that these mice exhibit fewer symptoms and longer survival than wild-type mice, suggesting that TLR2 activation may have harmful effects on the host [66,67].TLR3 recognition of dsRNA plays a protective role against herpes simplex virus encephalitis (HSE) in children [68]. Defects in the TLR3 response in the central nervous system (CNS) have been observed in approximately 5% of children with HSE [69]. Mouse experiments suggest that astrocytes rely on TLR3 to mediate resistance to HSV infection [70]. However, another study demonstrated that TLR3-deficient neurons and oligodendrocytes were more susceptible to HSV-1 infection compared to control cells, indicating the importance of TLR3 in protecting neuronal cells from HSV infection [71].TLR9 is significant for certain cell types, such as plasmacytoid dendritic cells (pDC), where the absence of TLR9 results in impaired IFN responses [72,73].

Figure 3. Pattern diagram of immune cell response process triggered by PRR signal triggered by HSV-1 infection. Inducing the secretion of inflammatory cytokines or IFN through the TLR signaling pathway (left, middle) or RLR signaling pathway (right). In the TLR signaling pathway, TLR2 recognizes signals induced by HSV-1 envelope proteins, such as gB or gD. The signal is transmitted to the cytoplasm, where MyD88 binds to the cytoplasmic domain of TLR2, leading to the activation of transcription factors like NF-κB. This activation promotes the translocation of P50/P65 into the nucleus and increases the expression of inflammatory cytokines and IFN-1. Additionally, TLR3, TLR7/8, and TLR9 signaling are activated by dsRNA, ssRNA, or CpG DNA, respectively, in endosomes. These signals activate IRF-3, IRF-7, and NF-κB, ultimately resulting in increased expression of inflammatory cytokines, IFN-1, IFN-III, and interferon-stimulated genes (ISGs). In the RLR signaling pathway, RIG-I and MDA5, which contain N-terminal caspase activation and recruitment domains, recruit and activate the mitochondrial antiviral signaling (MAVS) protein to mediate signal transduction. The activated MAVS protein further activates downstream signaling, promoting the expression of inflammatory cytokines and IFN.Both pathways contribute to the immune response against HSV-1 infection by triggering the production of inflammatory cytokines and interferons, which play crucial roles in controlling viral replication and coordinating innate and adaptive immune responses.

Figure 3. Pattern diagram of immune cell response process triggered by PRR signal triggered by HSV-1 infection. Inducing the secretion of inflammatory cytokines or IFN through the TLR signaling pathway (left, middle) or RLR signaling pathway (right). In the TLR signaling pathway, TLR2 recognizes signals induced by HSV-1 envelope proteins, such as gB or gD. The signal is transmitted to the cytoplasm, where MyD88 binds to the cytoplasmic domain of TLR2, leading to the activation of transcription factors like NF-κB. This activation promotes the translocation of P50/P65 into the nucleus and increases the expression of inflammatory cytokines and IFN-1. Additionally, TLR3, TLR7/8, and TLR9 signaling are activated by dsRNA, ssRNA, or CpG DNA, respectively, in endosomes. These signals activate IRF-3, IRF-7, and NF-κB, ultimately resulting in increased expression of inflammatory cytokines, IFN-1, IFN-III, and interferon-stimulated genes (ISGs). In the RLR signaling pathway, RIG-I and MDA5, which contain N-terminal caspase activation and recruitment domains, recruit and activate the mitochondrial antiviral signaling (MAVS) protein to mediate signal transduction. The activated MAVS protein further activates downstream signaling, promoting the expression of inflammatory cytokines and IFN.Both pathways contribute to the immune response against HSV-1 infection by triggering the production of inflammatory cytokines and interferons, which play crucial roles in controlling viral replication and coordinating innate and adaptive immune responses.

The adaptive immune response plays a crucial role in managing HSV infection and reactivation. Cell-mediated immunity, particularly involving T cells, is a key component of the adaptive immune response. After viral infection, cells present antigens to CD8+ T cells through surface major histocompatibility complex (MHC) class 1 molecules. This triggers the elimination of infected cells, limiting viral spread. T cells have been found to play a major role in the adaptive immune response to HSV. Specific T cells have been identified in sensory ganglia of infected individuals and in active and latent lesions of patients [74,75,76,77,78]. Following acute HSV infection, the percentage of blood-specific T cells is lower in infected individuals [79,80]. HSV-specific CD8+ T cells in the blood express high levels of cytolytic molecules when re-exposed to viral antigens [81]. CD4+ T cells recognize HSV-1 proteins and express cytokines associated with helper T cell type 1 (Th1)/Th0-like responses with cytolytic potential [80,82].

HSV-1 is capable of establishing a latency period in the dorsal root ganglia (DRG) of severely combined immunodeficient mice, even when CD8+ memory T cells are transplanted prior to infection. However, the presence of T cells reduces the number of infected DRG neurons, potentially limiting HSV-1 reactivation [83,84]. In mouse models, the rate of in vitro reactivation of trigeminal ganglia (TG) is directly correlated with viral ganglionic load, rather than the number of specific CD8+ T cells [85]. Specific CD8+ and CD4+ T cells are also present in the TG following human HSV-1 infection [74,75]. The infiltrating T cells in human infected TGs are characterized as memory effector T cells and surround the cell bodies and axons of neurons [74,86]. In mouse models, memory CD8+ T cells express interferon-gamma (IFN-γ), which prevents HSV replication in neurons and inhibits neuronal apoptosis, potentially promoting the survival of neurons and HSV-1 silencing and latency [87,88,89]. The mechanism of CD4+ and CD8+ T cell recognition of latently infected neurons is not fully understood. It is possible that there may be limited viral gene expression that can be recognized by T cells, allowing CD8+ T cell recognition and reactivation, along with potentially low levels of neuronal MHC class I molecule expression [90,91]. Additionally, satellite cells can act as antigen-presenting cells and express T-cell suppressor molecules to control HSV-1 latency without damaging neurons [92]. HSV also employs various strategies to inhibit antigen presentation and modulate adaptive immune responses. For example, the viral protein ICP47 blocks antigen presentation, and ICP34.5 inhibits autophagy, which is involved in antigen presentation [93]. Furthermore, HSV can inhibit antibody responses by interacting with antibodies and complement components, inhibiting antibody-dependent cell-mediated cytotoxicity [94]. These mechanisms suggest that HSV can modulate the adaptive immune response and influence the pathogenesis of the infection.

3. Human Cytomegalovirus

3.1. The process of HCMV entering host cells, replication and assembly

HCMV can be transmitted through various routes, including vertical transmission, contact transmission, and sexual transmission [95,96,97]. The virus infects different types of cells, such as leukocytes, fibroblasts, epithelial cells, and endothelial cells [98].HCMV entry into host cells involves several viral envelope glycoprotein complexes. Initially, three complexes were identified: gC-I, gC-II, and gC-III, which play a crucial role in viral entry [99]. gC-II is composed of glycoproteins M (gM) and N (gN), encoded by UL100 and UL73, respectively. It can attach to host cells through interaction with heparan sulfate proteoglycans on the cell surface [100,101]. The gH/gL complex, present in all herpesviruses, along with gB, forms the core membrane fusion machinery. gC-III is a heterotrimer consisting of the gH/gL complex linked to gO via a disulfide bond, commonly referred to as a trimer. Additionally, a pentameric complex consisting of gH/gL bound to three small glycoproteins encoded by UL128, UL130, and UL131A was discovered [103,104,105]. Binding of these complexes to the receptor transmits signals to gB, triggering membrane fusion [104,105]. gC-I is a homotrimer of gB, and different regions of gB are involved in fusion and/or interactions with proteins that trigger fusion [106,107]. After insertion into the target cell membrane, gB refolds into its post-fusion conformation, creating a fusion pore through which the viral capsid can enter the cell. Lateral interactions between gB trimers may contribute to expanding the fusion pore [17,108]. Several cell surface proteins, including platelet-derived growth factor receptor alpha (PDGFRα), epidermal growth factor receptor (EGFR), and integrins, have been reported to be associated with gB receptors [109,110,111,112].

HCMV entry into epithelial and endothelial cells occurs after endocytosis and requires binding to the pentameric complex neurofibrillary protein 2 (NRP 2) to trigger gB [113,114]. In fibroblasts, the trimers bind to PDGFRα and trigger gB at the plasma membrane [115,116,117].

After HCMV infection, the viral genome translocates to the nucleus. One of the first proteins expressed by HCMV is the Immediate Early 1 (IE 1) protein, which disrupts promyelocytic leukemia (PML) nuclear bodies and eliminates their inhibitory effect on viral gene transcription [118]. Replication and transcription of viral DNA occur in viral replication compartments (VRCs). The UL112-113 protein of HCMV plays a role in generating VRCs around the viral genome by liquid-liquid phase separation (LLPS), creating a pro-replicative environment [119]. Additionally, during the early stages of infection, most of the periplasmic proteins and glycoproteins are expressed in the cytoplasm and accumulate in specific cytoplasmic compartments called viral assembly compartments (AC) [120]. Cytoplasmic capsids are transported to the AC, where they form infectious mature particles.

3.2. Host cell processes caused by HCMV infection

HCMV infects organisms by entering through mucosal surfaces and subsequently disseminates to various tissues and organs within the body. Following invasion of the human body by HCMV, the majority of viral replication takes place locally. In 1975, Stagno initially reported a correlation between HCMV viral load and human disease, describing it as a threshold relationship. If the local viral load surpasses a critical threshold, the virus can disseminate via the bloodstream, resulting in damage to multiple target organs. The pathogenic risk associated with various target organs is intricately linked to their respective local viral loads. If the local immune response of a target organ fails to adequately control viral replication below a certain threshold, damage to the corresponding organ may ensue. Simultaneously, the local immune status also influences the threshold at which the virus becomes pathogenic. Strengthening the local immune status results in an increased threshold for viral pathogenicity, rendering the organ more resilient to the virus. Conversely, a weakened local immune status reduces the threshold for viral pathogenicity, making the local organ more susceptible to disease.

Primary infection of healthy individuals by HCMV is typically asymptomatic but can establish a lifelong latent infection in the host, which may lead to disease upon reactivation. The maintenance of latency and reactivation are regulated through the coordinated actions of both viral and cellular factors. HCMV establishes latency in hematopoietic stem cells and hematopoietic progenitor cells (HPCs) and persists in myeloid lineage cells. During latency, viral gene expression, including the major immediate early promoter (MIEP) that controls replication by regulating the IE gene, is reduced. The repression of viral genome transcription is also regulated through direct repression of IE gene expression and the expression of IE regulatory proteins. Buehler proposed a model in which pUL135 and pUL138, along with EGFR, form a molecular switch that regulates latency and replication status in HCMV infection. Upon viral entry and stimulation of the MEK/ERK signaling pathway, EGR-1 stimulates UL 138 expression, inhibiting viral replication to promote the establishment of latency. UL135 promotes EGFR recycling and counteracts the inhibitory effect of UL138, thereby promoting reactivation from latency and viral replication.

The regulation of latency also involves the modulation of host cell signaling. Recent studies have provided evidence that HCMV miRNAs play a role in modulating cellular signaling pathways and mediating latency in HPCs. For instance, Pan discovered miR-UL148, an HCMV-encoded miRNA, is expressed early during HPCs infection and can downregulate an IE activator, thereby promoting the establishment of latency. Diggins also found that miR-US25-1 can target the RhoA signaling axis to control cell proliferation. Moreover, HCMV protects HPCs from apoptosis and enhances HPCs survival through the synergistic effects of UL7 with miR-US5-1 and miR-UL112.

Virus reactivation can be triggered by various factors. For example, stimulation of stem cells moving to the periphery by granulocyte colony-stimulating factor can induce viral reactivation. Viral miRNAs, such as miR-US5-2 and miR-US22, as well as viral proteins, including UL135 and UL138, can also regulate reactivation by controlling EGFR. Additionally, the viral protein UL7 acts as a ligand for the Fms-like tyrosine kinase 3 receptor (Flt-3R), and binding to Flt-3R triggers the differentiation of HPCs and monocytes, thereby stimulating HCMV reactivation.

3.3. Immune cell process caused by HCMV infection

Innate immunity serves as the first line of defense against HCMV. Upon initial contact with host cells, viral envelope proteins stimulate an innate immune response. The TLR-1/TLR-2 heterodimer acts as a functional receptor for HCMV, recognizing glycoproteins gB and gH. Recognition of HCMV by TLR2 activates the nuclear factor-kappaB (NF-κB) pathway, leading to the production of inflammatory cytokines. However, HCMV gene products interfere with TLR activity. For example, HCMV miR-UL112-3p modulates the TLR2/NF-κB signaling pathway and downregulates TLR2 expression. HCMV-encoded proteins, such as US7 and US8, also inhibit the TLR pathway, thereby antagonizing innate immunity. NK cells and IFNs play crucial roles in natural immune defense against HCMV. TLR9 recognition of natural interferon-producing cells and dendritic cells (DCs) leads to the secretion of cytokines like IL-12, which activates the antiviral activity of NK cells. Activated NK cells can control HCMV replication through cytolytic and non-cytolytic mechanisms. They utilize perforin to lyse infected cells and secrete lymphotoxin α and TNF, which induce IFN-β expression in target cells. IFN-γ produced by NK cells confers resistance to HCMV infection in other cells. The combined action of IFN-β and IFN-γ inhibits HCMV replication without causing cell lysis after IE gene expression. HCMV employs various strategies to evade NK cell elimination. The periplasmic protein pp65 inhibits NK cell cytotoxicity by interacting with the activation receptor NKp30. HCMV actively downregulates NKG2D ligands, including UL16 binding proteins (ULBP) 1/2/6 and MHC class I-associated chain B (MICB), mediated by the glycoprotein UL16. UL148A proteins downregulate MICA, while HCMV-miR-UL112 specifically suppresses MICB expression, resulting in reduced NKG2D binding and decreased NK cell killing.

Adaptive immunity provides long-lasting protection against primary HCMV infection and latent activation in healthy individuals. The role of antibodies in protecting against HCMV infection is still a topic of debate. While individuals with HCMV-specific T cells may not be completely protected from viral infection and disease episodes, this suggests that humoral immunity, particularly neutralizing antibodies, may play a role in resistance to HCMV infection. Studies have shown that HCMV immune serum is capable of neutralizing fibroblast and epithelial/endothelial cell infections. However, it is important to note that neutralizing antibodies are found at much higher levels in epithelial/endothelial cells compared to fibroblasts. In one study, the titers of neutralizing antibodies that effectively blocked HCMV infection were 128-fold higher in epithelial/endothelial cells than in fibroblasts. Additionally, immunoreactivity induced by HCMV immunoglobulin preparations was 50-fold higher in epithelial cells compared to fibroblasts. These findings suggest that neutralizing antibodies may be more effective in protecting against HCMV infection in epithelial and endothelial cells compared to fibroblasts. However, further research is needed to fully understand the role of antibodies in HCMV immunity and their effectiveness in different cell types.

CD8+ T cells play a crucial role in the immune response to HCMV by recognizing a wide range of structural, early, and late antigens, including HCMV-encodedomodulators. The most dominant antigens targeted by HCMV-specific CD8+ T cells are UL123 (IE-1), UL122 (IE-2), and UL83 (pp65). These cytotoxic CD8+ T cells recognize HCMV antigenic peptides presented on MHC class I molecules by antigen-presenting cells (APCs). They can inhibit intracellular viral replication by secreting IFN-γ or TNF-α, and they can also directly kill infected cells by releasing granzymes and perforins. Additionally, HCMV-specific CD8+ T cells undergo oligoclonal replication, leading to their accumulation with age. In fact, they can constitute more than 40% of the CD8+ T cell pool, a phenomenon known as "memory expansion," which is also observed in HCMV-specific CD4+ T cell responses. The clonal amplification and differentiation of cytotoxic CD8+ T cells during HCMV infection support the notion that HCMV may contribute to immune senescence. CD4+ T cells are also essential in controlling HCMV infection. They recognize MHC class II viral antigens presented by APCs and secrete INF-γ and IL-2, which inhibit viral replication. Furthermore, CD4+ T cells induce the proliferation of CD8+ T cells and macrophages. There are subpopulations of HCMV-specific effector CD4+ T cells that directly counteract viral infection, as described by Costa-García. Regulatory T (Treg) cells are involved in the immune response to HCMV as well. A study by Aandahl et al. demonstrated that depletion of CD25+ Treg cells from peripheral blood mononuclear cells enhances the immune response of CD8+ T cells to HCMV antigens.

HCMV has evolved various strategies to evade host immunity, with one of the main strategies being the targeting of MHC class I presentation of viral proteins to CD8+ T cells. The HCMV IE gene US3 encodes an endoplasmic reticulum (ER)-resident glycoprotein that retains protein-loaded MHC class I in the ER, preventing their transport to the cell surface. US6 interacts with the transporters associated with antigen processing in the ER, inhibiting the intracellular transport of MHC class I molecules. Additionally, US2 and US11 downregulate the expression of MHC class I heavy chain.

HCMV also hampers antigen presentation through the MHC class II pathway. US2 leads to the degradation of HLA-DR-α and HLA-DM-α, essential proteins in the MHC class II pathway. US3 can mediate the degradation of DR-α and DM-α and inhibit the assembly of MHC class II complexes. Normally, IFN-γ activates the MHC class II transactivator gene (CIITA) to promote MHC class II expression. However, HCMV inhibits MHC class II expression by blocking IFN-γ signal transduction through the Jak/Stat pathway. Moreover, HCMV regulates CIITA transcript levels in mature Langerhans-type dendritic cells and myeloid cell lines, reducing MHC class II biosynthesis.

Another evasion mechanism employed by HCMV is the production of multiple functional homologues. For instance, UL111A encodes a homologue of human IL-10 called cmvIL-10, which interacts with the human IL-10 receptor. IL-10 is an immunosuppressive cytokine that inhibits the ability of antigen-presenting cells (APCs) to present antigens to T cells, thereby suppressing the immune response[184].

4. Conclusion

The presence of herpesviruses, including HSV-1 and HCMV, has been detected in surgically removed tissues from patients with Meniere's disease, and viral infection is considered a leading hypothesis for the development of vestibular neuritis. However, the exact role of herpesviruses in the pathogenesis of vestibular neuritis is still not fully understood. In this paper, we aim to summarize the effects of herpesvirus infections, particularly HSV-1 and HCMV, on host cells and immune system processes. This information can be valuable in predicting neuronal cell damage, as well as the infiltration and killing of immune cells following viral infection. We hope that this review will stimulate further work and efforts to advance the prevention and treatment of diseases like vestibular neuritis that are potentially caused by viral infections.

5. Patents