Antibody-dependent enhancement and neutralization against CVB4 investigated in vitro and in silico through an agent-based model.

The infection with coxsackievirus B4 (CVB4) can be enhanced in vitro by antibodies directed against the viral capsid protein VP4. In peripheral blood mononuclear cells, antibody-dependent enhancement (ADE) of CVB4 infection leads to the production of interferon alpha (IFN-α). To investigate ADE of CVB4-induced production of IFN-α, an agent-based model was constructed with enhancing and neutralizing antibodies. The model recapitulates viral neutralization and ADE in silico. The enhancing and neutralizing activities of serum samples were evaluated in vitro to confront the model predictions with experimental results. Increasing the incubation time of CVB4 with serum samples improves virus neutralization in silico as well as in vitro. It also results in ADE at lower antibody numbers in silico, which is confirmed in vitro with IFN-α production at lower serum concentrations. Furthermore, incubation of CVB4 with serum at a low temperature does not induce IFN-α production in vitro. Thus, taken together our results suggest that enhancing antibodies bind cryptic epitopes, more accessible with longer incubation time and at higher temperature due to changes in capsid conformation, consistent with previous results indicating that enhancing antibodies are anti-VP4 antibodies.

Enterocin DD14 can inhibit the infection of eukaryotic cells with enveloped viruses.

Bacteriocins are ribosomally synthesized bacterial peptides endowed with antibacterial, antiprotozoal, anticancer and antiviral activities. In the present study, we evaluated the antiviral activities of two bacteriocins, enterocin DD14 (EntDD14) and lacticaseicin 30, against herpes simplex virus type 1 (HSV-1), human coronavirus 229E (HCoV-229E) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Vero, Huh7 and Vero E6 cells, respectively. In addition, the interactions of these bacteriocins with the envelope glycoprotein D of HSV-1 and the receptor binding domains of HCoV-229E and SARS-CoV-2 have been computationally evaluated using protein-protein docking and molecular dynamics simulations. HSV-1 replication in Vero cells was inhibited by EntDD14 and, to a lesser extent, by lacticaseicin 30 added to cells after virus inoculation. EntDD14 and lacticaseicin 30 had no apparent antiviral activity against HCoV-229E; however, EntDD14 was able to inhibit SARS-CoV-2 in Vero E6 cells. Further studies are needed to elucidate the antiviral mechanism of these bacteriocins.

Combating coxsackievirus B infections.

Coxsackieviruses B (CVB) are small, non-enveloped, single-stranded RNA viruses belonging to the Enterovirus genus of the Picornaviridae family. They are common worldwide and cause a wide variety of human diseases ranging from those having relatively mild symptoms to severe acute and chronic pathologies such as cardiomyopathy and type 1 diabetes. The development of safe and effective strategies to combat these viruses remains a challenge. The present review outlines current approaches to control CVB infections and associated diseases. Various drugs targeting viral or host proteins involved in viral replication as well as vaccines have been developed and shown potential to prevent or combat CVB infections in vitro and in vivo in animal models. Repurposed drugs and alternative strategies targeting miRNAs or based on plant extracts and probiotics and their derivatives have also shown antiviral effects against CVB. In addition, clinical trials with vaccines and drugs are underway and offer hope for the prevention or treatment of CVB-induced diseases.

[Coxsackievirus B infection and pathogenesis of type 1 diabetes]. / Infection à coxsackievirus B et pathogenèse du diabète de type 1.

Epidemiological and experimental studies suggest that enteroviruses (EV) and particularly coxsackieviruses B (CVB) are likely to trigger or accelerate the onset of islet autoimmunity and the development of type 1 diabetes (T1D) in genetically susceptible individuals. Several mutually non-exclusive mechanisms have been proposed to explain the involvement of CVB in the pathogenesis of T1D. CVB can infect and persist in the intestine, thymic cells, monocytes/macrophages, ductal cells and pancreatic ß-cells, which leads to structural or functional alterations of these cells. A chronic inflammatory response and disruption of tolerance towards ß-cells due to CVB infections are able to promote the recruitment and activation of pre-existing autoreactive T-cells and the destruction of ß-cells. Vaccine or therapeutic strategies to control EV infections have been developed and open perspectives for the prevention or treatment of T1D.

Pancreatic beta cells persistently infected with coxsackievirus B4 are targets of NK cell-mediated cytolytic activity.

It has been suggested that the persistence of coxsackieviruses-B (CV-B) in pancreatic beta cells plays a role in the pathogenesis of type 1 diabetes (T1D). Yet, immunological effectors, especially natural killer (NK) cells, are supposed to clear virus-infected cells. Therefore, an evaluation of the response of NK cells to pancreatic beta cells persistently infected with CV-B4 was conducted. A persistent CV-B4 infection was established in 1.1B4 pancreatic beta cells. Infectious particles were found in supernatants throughout the culture period. The proportion of cells containing viral protein VP1 was low (< 5%), although a large proportion of cells harbored viral RNA (around 50%), whilst cell viability was preserved. HLA class I cell surface expression was downregulated in persistently infected cultures, but HLA class I mRNA levels were unchanged in comparison with mock-infected cells. The cytolytic activities of IL-2-activated non-adherent peripheral blood mononuclear cells (PBMCs) and of NK cells were higher towards persistently infected cells than towards mock-infected cells, as assessed by an LDH release assay. Impaired cytolytic activity of IL-2-activated non-adherent PBMCs from patients with T1D towards infected beta cells was observed. In conclusion, pancreatic beta cells persistently infected with CV-B4 can be lysed by NK cells, implying that impaired cytolytic activity of these effector cells may play a role in the persistence of CV-B in the host and thus in the viral pathogenesis of T1D.

The relationship between SARS-CoV-2 infection and type 1 diabetes mellitus.

Environmental factors, in particular viral infections, are thought to have an important role in the pathogenesis of type 1 diabetes mellitus (T1DM). The COVID-19 pandemic reinforced this hypothesis as many observational studies and meta-analyses reported a notable increase in the incidence of T1DM following infection with SARS-CoV-2 as well as an association between SARS-CoV-2 infection and the risk of new-onset T1DM. Experimental evidence suggests that human ß-cells express SARS-CoV-2 receptors and that SARS-CoV-2 can infect and replicate in ß-cells, resulting in structural or functional alterations of these cells. These alterations include reduced numbers of insulin-secreting granules, impaired pro-insulin (or insulin) secretion, and ß-cell transdifferentiation or dedifferentiation. The inflammatory environment induced by local or systemic SARS-CoV-2 infection might result in a set of signals (such as pro-inflammatory cytokines) that lead to ß-cell alteration or apoptosis or to a bystander activation of T cells and disruption of peripheral tolerance that triggers autoimmunity. Other mechanisms, such as viral persistence, molecular mimicry and activation of endogenous human retroviruses, are also likely to be involved in the pathogenesis of T1DM following SARS-CoV-2 infection. This Review addresses the issue of the involvement of SARS-CoV-2 infection in the development of T1DM using evidence from epidemiological, clinical and experimental studies.

Markers of Epstein-Barr Virus Infection in Patients with Multiple Sclerosis.

Viral infections have been suspected of being involved in the pathogenesis of certain autoimmune diseases for many years. Epstein-Barr virus (EBV), a DNA virus belonging to the Herpesviridae family, is thought to be associated with the onset and/or the progression of multiple sclerosis (MS), systemic lupus erythematosus, rheumatoid arthritis, Sjögren's syndrome and type 1 diabetes. The lifecycle of EBV consists of lytic cycles and latency programmes (0, I, II and III) occurring in infected B-cells. During this lifecycle, viral proteins and miRNAs are produced. This review provides an overview of the detection of EBV infection, focusing on markers of latency and lytic phases in MS. In MS patients, the presence of latency proteins and antibodies has been associated with lesions and dysfunctions of the central nervous system (CNS). In addition, miRNAs, expressed during lytic and latency phases, may be detected in the CNS of MS patients. Lytic reactivations of EBV can occur in the CNS of patients as well, with the presence of lytic proteins and T-cells reacting to this protein in the CNS of MS patients. In conclusion, markers of EBV infection can be found in MS patients, which argues in favour of a relationship between EBV and MS.

The Fight against Poliovirus Is Not Over.

Poliovirus (PV), the virus that causes both acute poliomyelitis and post-polio syndrome, is classified within the Enterovirus C species, and there are three wild PV serotypes: WPV1, WPV2 and WPV3. The launch of the Global Polio Eradication Initiative (GPEI) in 1988 eradicated two of the three serotypes of WPV (WPV2 and WPV3). However, the endemic transmission of WPV1 persists in Afghanistan and Pakistan in 2022. There are cases of paralytic polio due to the loss of viral attenuation in the oral poliovirus vaccine (OPV), known as vaccine-derived poliovirus (VDPV). Between January 2021 and May 2023, a total of 2141 circulating VDPV (cVDPV) cases were reported in 36 countries worldwide. Because of this risk, inactivated poliovirus (IPV) is being used more widely, and attenuated PV2 has been removed from OPV formulations to obtain bivalent OPV (containing only types 1 and 3). In order to avoid the reversion of attenuated OPV strains, the new OPV, which is more stable due to genome-wide modifications, as well as sabin IPV and virus-like particle (VLP) vaccines, is being developed and offers promising solutions for eradicating WP1 and VDPV.

Viruses and Endocrine Diseases.

Viral infections have been frequently associated with physiological and pathological changes in the endocrine system for many years. The numerous early and late endocrine complications reported during the current pandemic of coronavirus disease 2019 (COVID-19) reinforce the relevance of improving our understanding of the impact of viral infections on the endocrine system. Several viruses have been shown to infect endocrine cells and induce endocrine system disturbances through the direct damage of these cells or through indirect mechanisms, especially the activation of the host antiviral immune response, which may lead to the development of local or systemic inflammation or organ-specific autoimmunity. In addition, endocrine disorders may also affect susceptibility to viral infections since endocrine hormones have immunoregulatory functions. This review provides a brief overview of the impact of viral infections on the human endocrine system in order to provide new avenues for the control of endocrine diseases.

Vemurafenib Inhibits Acute and Chronic Enterovirus Infection by Affecting Cellular Kinase Phosphatidylinositol 4-Kinase Type IIIß.

Enteroviruses are one of the most abundant viruses causing mild to serious acute infections in humans and also contributing to chronic diseases like type 1 diabetes. Presently, there are no approved antiviral drugs against enteroviruses. Here, we studied the potency of vemurafenib, an FDA-approved RAF kinase inhibitor for treating BRAFV600E mutant-related melanoma, as an antiviral against enteroviruses. We showed that vemurafenib prevented enterovirus translation and replication at low micromolar dosage in an RAF/MEK/ERK-independent manner. Vemurafenib was effective against group A, B, and C enteroviruses, as well as rhinovirus, but not parechovirus or more remote viruses such as Semliki Forest virus, adenovirus, and respiratory syncytial virus. The inhibitory effect was related to a cellular phosphatidylinositol 4-kinase type IIIß (PI4KB), which has been shown to be important in the formation of enteroviral replication organelles. Vemurafenib prevented infection efficiently in acute cell models, eradicated infection in a chronic cell model, and lowered virus amounts in pancreas and heart in an acute mouse model. Altogether, instead of acting through the RAF/MEK/ERK pathway, vemurafenib affects the cellular PI4KB and, hence, enterovirus replication, opening new possibilities to evaluate further the potential of vemurafenib as a repurposed drug in clinical care. IMPORTANCE Despite the prevalence and medical threat of enteroviruses, presently, there are no antivirals against them. Here, we show that vemurafenib, an FDA-approved RAF kinase inhibitor for treating BRAFV600E mutant-related melanoma, prevents enterovirus translation and replication. Vemurafenib shows efficacy against group A, B, and C enteroviruses, as well as rhinovirus, but not parechovirus or more remote viruses such as Semliki Forest virus, adenovirus, and respiratory syncytial virus. The inhibitory effect acts through cellular phosphatidylinositol 4-kinase type IIIß (PI4KB), which has been shown to be important in the formation of enteroviral replication organelles. Vemurafenib prevents infection efficiently in acute cell models, eradicates infection in a chronic cell model, and lowers virus amounts in pancreas and heart in an acute mouse model. Our findings open new possibilities to develop drugs against enteroviruses and give hope for repurposing vemurafenib as an antiviral drug against enteroviruses.

Coxsackievirus Infection and Associated Diseases.

Coxsackieviruses (CV) are ubiquitous and widespread single-stranded RNA viruses belonging to the Picornaviridae family and the genus Enterovirus, which also includes poliovirus (PV), the best known of the enteroviruses (EV) [...].

Persistent coxsackievirus B infection and pathogenesis of type 1 diabetes mellitus.

Enteroviruses are believed to trigger or accelerate islet autoimmunity in genetically susceptible individuals, thereby resulting in loss of functional insulin-producing ß-cells and type 1 diabetes mellitus (T1DM). Although enteroviruses are primarily involved in acute and lytic infections in vitro and in vivo, they can also establish a persistent infection. Prospective epidemiological studies have strongly associated the persistence of enteroviruses, especially coxsackievirus B (CVB), with the appearance of islet autoantibodies and an increased risk of T1DM. CVB can persist in pancreatic ductal and ß-cells, which leads to structural or functional alterations of these cells, and to a chronic inflammatory response that promotes recruitment and activation of pre-existing autoreactive T cells and ß-cell autoimmune destruction. CVB persistence in other sites, such as the intestine, blood cells and thymus, has been described; these sites could serve as a reservoir for infection or reinfection of the pancreas, and this persistence could have a role in the disturbance of tolerance to ß-cells. This Review addresses the involvement of persistent enterovirus infection in triggering islet autoimmunity and T1DM, as well as current strategies to control enterovirus infections for preventing or reducing the risk of T1DM onset.

Fighting Enteroviral Infections to Prevent Type 1 Diabetes.

Enteroviruses (EVs), especially coxsackieviruses B (CVB), are believed to trigger or accelerate islet autoimmunity in genetically susceptible individuals that results in type 1 diabetes (T1D). Therefore, strategies are needed to fight against EV infections. There are no approved antiviral drugs currently available, but various antiviral drugs targeting viral or host cell proteins and vaccines have recently shown potential to combat CVB infections and may be used as new therapeutic strategies to prevent or reduce the risk of T1D and/or preserve ß-cell function among patients with islet autoantibodies or T1D.

Predictive Value of Immune Cells in the Risk of Gestational Diabetes Mellitus: A Pilot Study.

Aims: Immunological and biochemical parameters are gaining more and more importance in the prognosis of diabetes and its complications. Here, we assessed the predictive power of immune cells correlated with biochemical parameters in gestational diabetes mellitus (GDM). Materials and Methods: Immune cells and serum biochemical parameters were determined in women with GDM and pregnant controls. Receiver operating characteristics (ROC) curve analyses were conducted to assess the optimal cutoff and value of ratios of immune cells to biochemical parameters for predicting GDM. Results: Blood glucose, total cholesterol, LDL-cholesterol and triglycerides were significantly increased whereas HDL-cholesterol decreased in women with GDM compared to pregnant controls. Glycated hemoglobin, creatinine, transaminase activities did not significantly differ between both groups. Total leukocyte, lymphocyte and platelet numbers were significantly high in women with GDM. Correlation tests showed that ratios of lymphocyte/HDL-C, monocyte/HDL-C and granulocyte/HDL-C were significantly higher in women with GDM than in pregnant controls (p = 0.001; p = 0.009 and p = 0.004 respectively). Women with a lymphocyte/HDL-C ratio greater than 3.66 had a 4-fold increased risk of developing GDM than those with lower ratios (odds ratio 4.00; 95% CI: 1.094 - 14.630; p=0.041). Conclusion: Our study showed that ratios of lymphocyte, monocyte and granulocyte to HDL-C might represent valuable biomarkers for GDM and in particular, lymphocyte/HDL-C ratio exhibited a strong predictive power for GDM risk.

Low Serum Levels of Interferon Alpha in COVID-19 Patients Are Associated with Older Age.

Innate immune response, especially type 1 interferon (IFN) response is considered to play a substantial role in the outcome of SARS-CoV-2 infection. A reduced and delayed IFN response has been associated with progression to severe COVID-19. In this study, we investigated levels of circulating IFNα and serum neutralizing activity in COVID-19 patients admitted to the intensive care unit. We found a significant association of levels of IFNα with age (p = 0.007). This association has also been observed in a cohort of COVID-19 outpatients with mild infection (p = 0.02). The impact of senescence on IFN response can explain the higher susceptibility of the elderly to severe COVID-19.

Persistence of Coxsackievirus B4 in Pancreatic ß Cells Disturbs Insulin Maturation, Pattern of Cellular Proteins, and DNA Methylation.

Coxsackievirus-B4 (CV-B4) can persist in pancreatic cell lines and impair the phenoytpe and/or gene expressions in these cells; however, the models used to study this phenomenon did not produce insulin. Therefore, we investigated CV-B4 persistence and its consequences in insulin-producing pancreatic ß cells. The insulin-secreting rat ß cell line, INS-1, was infected with CV-B4. After lysis of a large part of the cell layer, the culture was still maintained and no additional cytopathic effect was observed. The amount of insulin in supernatants of cell cultures persistently infected with CV-B4 was not affected by the infection; in fact, a larger quantity of proinsulin was found. The mRNA expression of pro-hormone convertase 2, an enzyme involved in the maturation of proinsulin into insulin and studied using real-time reverse transcription-polymerase chain reaction, was inhibited in infected cultures. Further, the pattern of 47 cell proteins analyzed using Shotgun mass spectrometry was significantly modified. The DNA of persistently infected cell cultures was hypermethylated unlike that of controls. The persistent infection of INS-1 cells with CV-B4 had a deep impact on these cells, especially on insulin metabolism. Cellular changes caused by persistent CV-B4 infection of ß cells can play a role in type 1 diabetes pathogenesis.

Fluoxetine Can Inhibit SARS-CoV-2 In Vitro.

An outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) resulted in the coronavirus disease pandemic, drastically affecting global health and economy. Though the understanding of the disease has improved, fighting the virus remains challenging. One of the strategies is repurposing existing drugs as inhibitors of SARS-CoV-2. Fluoxetine (FLX), a selective serotonin reuptake inhibitor, reportedly inhibits the replication of RNA viruses, especially Coxsackieviruses B (CVB), such as CV-B4 in vitro and in vivo. Therefore, in this study, we investigated the in vitro antiviral activity of FLX against SARS-CoV-2 in a model of acute infection. When 10 µM of FLX was added to SARS-CoV-2-infected Vero E6 cells, the virus-induced cytopathic effect was not observed. In this model, the level of infectious particles in the supernatant was lower than that in controls. The level was below the limit of detection of the assay up to day 3 post-infection when FLX was administered before viral inoculation or simultaneously followed by daily inoculation. In conclusion, FLX can inhibit SARS-CoV-2 in vitro. Further studies are needed to investigate the potential value of FLX to combat SARS-CoV-2 infections, treat SARS-CoV-2-induced diseases, and explain the antiviral mechanism of this molecule to pave way for novel treatment strategies.

Effect of Coxsackievirus B4 Infection on the Thymus: Elucidating Its Role in the Pathogenesis of Type 1 Diabetes.

The thymus gland is a primary lymphoid organ for T-cell development. Various viral infections can result in disturbance of thymic functions. Medullary thymic epithelial cells (mTECs) are important for the negative selection of self-reactive T-cells to ensure central tolerance. Insulin-like growth factor 2 (IGF2) is the dominant self-peptide of the insulin family expressed in mTECs and plays a crucial role in the intra-thymic programing of central tolerance to insulin-secreting islet ß-cells. Coxsackievirus B4 (CVB4) can infect and persist in the thymus of humans and mice, thus hampering the T-cell maturation and differentiation process. The modulation of IGF2 expression and protein synthesis during a CVB4 infection has been observed in vitro and in vivo in mouse models. The effect of CVB4 infections on human and mouse fetal thymus has been studied in vitro. Moreover, following the inoculation of CVB4 in pregnant mice, the thymic function in the fetus and offspring was disturbed. A defect in the intra-thymic expression of self-peptides by mTECs may be triggered by CVB4. The effects of viral infections, especially CVB4 infection, on thymic cells and functions and their possible role in the pathogenesis of type 1 diabetes (T1D) are presented.

Th2-Immune Polarizing and Anti-Inflammatory Properties of Insulin Are Not Effective in Type 2 Diabetic Pregnancy.

BACKGROUND: The implication of the immune system in the physiopathology of pregnancy complicated by diabetes has been reported. Here, we investigated the effects of insulin treatment on the frequencies of immune cell subpopulations as well as T cell-derived cytokines in type 2 diabetic (T2D) pregnancy compared to gestational diabetes mellitus (GDM). METHODS: Fifteen (15) women with GDM, twenty (20) insulin-treated T2D pregnant women, and twenty-five (25) pregnant controls were selected. Immune cell subpopulation frequencies were determined in blood using flow cytometry. The proliferative capacity of T cells was performed, and serum and cell culture supernatant cytokine levels were also quantified. RESULTS: The frequencies of total CD3+ and CD4+ T cells and nonclassical monocytes significantly increased in insulin-treated T2D pregnant women compared to pregnant controls. The proportions of CD4+ T cells as well as B cells were significantly higher in women with GDM than in pregnant controls. GDM was associated with high frequencies of total CD3+ and CD4+ T cells and B cell expansion, suggesting a concomitant activation of cellular and humoral immunity. Concomitantly, Th1/Th2 ratio, determined as IFN-γ/IL-4, was shifted towards Th1 phenotype in women with GDM and insulin-treated T2D pregnant women. Besides, isolated T cells elicited similar proliferative capacity in the three groups of women. Insulin-treated T2D pregnant women and women with GDM exhibited a low serum IL-10 level, without any change in the number of Treg cells. CONCLUSION: Our study showed that, despite insulin treatment, pregnant women with T2D displayed a proinflammatory status consistent with high proportions of CD3+ and CD4+ T cells, upregulation of Th1 cytokines, and low IL-10 production, suggesting a reduced immune-suppressive activity of regulatory T cells. However, GDM, although associated with proinflammatory status, has shown increased humoral immunity consistent with high proportion of CD19+ B cells. Thus, the lack of response to insulin in diabetes during pregnancy and clinical implications of these immunological parameters deserves further investigations.

Enteroviral Pathogenesis of Type 1 Diabetes: The Role of Natural Killer Cells.

Enteroviruses, especially group B coxsackieviruses (CV-B), have been associated with the development of chronic diseases such as type 1 diabetes (T1D). The pathological mechanisms that trigger virus-induced autoimmunity against islet antigens in T1D are not fully elucidated. Animal and human studies suggest that NK cells response to CV-B infection play a crucial role in the enteroviral pathogenesis of T1D. Indeed, CV-B-infected cells can escape from cytotoxic T cells recognition and destruction by inhibition of cell surface expression of HLA class I antigen through non-structural viral proteins, but they can nevertheless be killed by NK cells. Cytolytic activity of NK cells towards pancreatic beta cells persistently-infected with CV-B has been reported and defective viral clearance by NK cells of patients with T1D has been suggested as a mechanism leading to persistence of CV-B and triggering autoimmunity reported in these patients. The knowledge about host antiviral defense against CV-B infection is not only crucial to understand the susceptibility to virus-induced T1D but could also contribute to the design of new preventive or therapeutic approaches for individuals at risk for T1D or newly diagnosed patients.