Human intestinal organoid models for celiac disease research.

Celiac disease pathogenesis, in addition to immune cell component, encompasses pathogenic events also in the duodenal epithelium. In celiac disease patients, exposure to dietary gluten induces drastic changes in epithelial differentiation and elicit cellular response to inflammatory cytokines. The autoantigen in celiac disease, transglutaminase 2 (TG2) enzyme, has been also suggested to play its pathogenic gliadin deamidation event in the intestinal epithelium. Therefore in vitro epithelial cell-line models have been exploited in the past to study these pathogenic mechanisms, but they are hampered by their simplistic nature lacking proper cell-type composition and intestinal environ. Moreover, these cell models harbor many cancer-related mutations in tumor suppressor genes making them unsuitable for studying cell differentiation. Intestinal organoids provide a near-native epithelial cell model to study pathogenic agents and mechanisms related to celiac disease. Here we describe protocols to initiate and maintain celiac patient-derived organoid cultures and how to grow them in alternative ways allowing their exploitation in different kind of experiments.

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.

HLA-DQ-conferred risk for type 1 diabetes does not alter neutralizing antibody response to a widely used enterovirus vaccine, the poliovirus vaccine.

This study investigated whether children with HLA-DQ-conferred risk for type 1 diabetes (T1D) have an altered immune response to the widely-used enterovirus vaccine, namely poliovirus vaccine, and whether initiation of autoimmunity to pancreatic islets modulates this response. Neutralizing antibodies induced by the inactivated poliovirus vaccine against poliovirus type 1 (Salk) were analysed as a marker of protective immunity at the age of 18 months in a prospective birth cohort. No differences were observed in antibody titers between children with and without genetic risk for T1D (odds ratio [OR] = 0.90 [0.83, 1.06], p = 0.30). In the presence of the genetic risk, no difference was observed between children with and without islet autoimmunity (OR = 1.00 [0.78, 1.28], p = 1.00). This did not change when only children with the autoimmunity before 18 months of age were included in the analyses (OR = 1.00 [0.85, 1.18], p = 1.00). No effect was observed when groups were stratified based on autoantigen specificity of the first-appearing autoantibody (IAA or GADA). The children in each comparison group were matched for sex, calendar year and month of birth, and municipality. Accordingly, we found no indication that children who are at risk to develop islet autoimmunity would have a compromised humoral immune response which could have increased their susceptibility for enterovirus infections. In addition, the proper immune response supports the idea of testing novel enterovirus vaccines for the prevention of T1D among these individuals.

Serum 25-hydroxyvitamin D and fatty acids in relation to the risk of microbial infections in children: The TRIGR Divia study.

BACKGROUND & AIMS: Nutrient status may affect the risk of microbial infections and play a role in modulating the immune response against such infections. The aim of this study was to determine whether serum 25-hydroxyvitamin D [25(OH)D] and serum fatty acids in infancy are associated with microbial infections by the age of 18 months. METHODS: Altogether 576 newborn infants from Trial to Reduce IDDM in the Genetically at Risk (TRIGR) born between 2002 and 2007 were included. The concentration of 25(OH)D vitamin and proportions of 26 fatty acids (presented as % of total fatty acids) were analyzed in cord blood serum and in sera taken at 6, 12, and 18 months of age. The cord blood samples and mean of 6-18-month values were used as exposures. Infections were detected by screening IgG antibodies against 10 microbes using enzyme immunoassay and antibodies against 6 coxsackievirus B serotypes by plaque neutralization assay in serum samples taken at 18 months of age. RESULTS: A higher proportion of n-3 polyunsaturated fatty acids (PUFAs) and especially long-chain n-3 PUFAs at birth and at the age of 6-18 months was associated with decreased risk of coxsackievirus B2 infection unadjusted and adjusted for region, case-control status, and maternal type 1 diabetes. Higher proportion of docosapentaenoic acid (DPA, 22:5 n-3) at birth was associated with a decreased risk of respiratory syncytial virus infection. 25(OH)D vitamin concentration was not consistently associated with the risk of infections. When only infected children were included docosahexaenoic acid (DHA, 22:6 n-3) and arachidonic acid (20:4 n-6) proportions were positively associated with IgG antibody levels against influenza A virus. 25(OH)D vitamin concentration showed an inverse association with rotavirus IgG levels among children with rotavirus seropositivity. CONCLUSIONS: In young children with increased susceptibility to type 1 diabetes, long-chain n-3 PUFAs may influence the risk of viral infections and immune response against the infections. However, this association may depend on the type of virus suggesting virus-specific effects.

Coxsackievirus B infections are common in Cystic Fibrosis and experimental evidence supports protection by vaccination.

Viral respiratory tract infections exacerbate airway disease and facilitate life-threatening bacterial colonization in cystic fibrosis (CF). Annual influenza vaccination is recommended and vaccines against other common respiratory viruses may further reduce pulmonary morbidity risk. Enteroviruses have been found in nasopharyngeal samples from CF patients experiencing pulmonary exacerbations. Using serology tests, we found that infections by a group of enteroviruses, Coxsackievirus Bs (CVBs), are prevalent in CF. We next showed that a CVB vaccine, currently undergoing clinical development, prevents infection and CVB-instigated lung damage in a murine model of CF. Finally, we demonstrate that individuals with CF have normal vaccine responses to a similar, commonly used enterovirus vaccine (inactivated poliovirus vaccine). Our study demonstrates that CVB infections are common in CF and provides experimental evidence indicating that CVB vaccines could be efficacious in the CF population. The role of CVB infections in contributing to pulmonary exacerbations in CF should be further studied.

Detection of enterovirus RNA in peripheral blood mononuclear cells correlates with the presence of the predisposing allele of the type 1 diabetes risk gene IFIH1 and with disease stage.

AIMS/HYPOTHESIS: Enteroviral infection has been implicated consistently as a key environmental factor correlating with the appearance of autoimmunity and/or the presence of overt type 1 diabetes, in which pancreatic insulin-producing beta cells are destroyed by an autoimmune response. Genetic predisposition through variation in the type 1 diabetes risk gene IFIH1 (interferon induced with helicase C domain 1), which encodes the viral pattern-recognition receptor melanoma differentiation-associated protein 5 (MDA5), supports a potential link between enterovirus infection and type 1 diabetes. METHODS: We used molecular techniques to detect enterovirus RNA in peripheral blood samples (in separated cellular compartments or plasma) from two cohorts comprising 79 children or 72 adults that include individuals with and without type 1 diabetes who had multiple autoantibodies. We also used immunohistochemistry to detect the enteroviral protein VP1 in the pancreatic islets of post-mortem donors (n=43) with type 1 diabetes. RESULTS: We observed enhanced detection sensitivity when sampling the cellular compartment compared with the non-cellular compartment of peripheral blood (OR 21.69; 95% CI 3.64, 229.20; p<0.0001). In addition, we show that children with autoimmunity are more likely to test positive for enterovirus RNA than those without autoimmunity (OR 11.60; 95% CI 1.89, 126.90; p=0.0065). Furthermore, we found that individuals carrying the predisposing allele (946Thr) of the common variant in IFIH1 (rs1990760, Thr946Ala) are more likely to test positive for enterovirus in peripheral blood (OR 3.07; 95% CI 1.02, 8.58; p=0.045). In contrast, using immunohistochemistry, there was no correlation between the common variant in IFIH1 and detection of enteroviral VP1 protein in the pancreatic islets of donors with type 1 diabetes. CONCLUSIONS/INTERPRETATION: Our data indicate that, in peripheral blood, antigen-presenting cells are the predominant source of enterovirus infection, and that infection is correlated with disease stage and genetic predisposition, thereby supporting a role for enterovirus infection prior to disease onset.

Persistent coxsackievirus B1 infection triggers extensive changes in the transcriptome of human pancreatic ductal cells.

Enteroviruses, particularly the group B coxsackieviruses (CVBs), have been associated with the development of type 1 diabetes. Several CVB serotypes establish chronic infections in human cells in vivo and in vitro. However, the mechanisms leading to enterovirus persistency and, possibly, beta cell autoimmunity are not fully understood. We established a carrier-state-type persistent infection model in human pancreatic cell line PANC-1 using two distinct CVB1 strains and profiled the infection-induced changes in cellular transcriptome. In the current study, we observed clear changes in the gene expression of factors associated with the pancreatic microenvironment, the secretory pathway, and lysosomal biogenesis during persistent CVB1 infections. Moreover, we found that the antiviral response pathways were activated differently by the two CVB1 strains. Overall, our study reveals extensive transcriptional responses in persistently CVB1-infected pancreatic cells with strong opposite but also common changes between the two strains.

Coxsackievirus B Vaccines Prevent Infection-Accelerated Diabetes in NOD Mice and Have No Disease-Inducing Effect.

Enteroviruses, including the Coxsackievirus Bs (CVB), have been implicated as causal agents in human type 1 diabetes. Immunization of at-risk individuals with a CVB vaccine provides an attractive strategy for elucidating the role of CVBs in the disease etiology. Previously, we have shown that an inactivated whole-virus vaccine covering all CVB serotypes (CVB1-6) is safe to administer and highly immunogenic in preclinical models, including nonhuman primates. Before initiating clinical trials with this type of vaccine, it was also important to address 1) whether the vaccine itself induces adverse immune reactions, including accelerating diabetes onset in a diabetes-prone host, and 2) whether the vaccine can prevent CVB-induced diabetes in a well-established disease model. Here, we present results from studies in which female NOD mice were left untreated, mock-vaccinated, or vaccinated with CVB1-6 vaccine and monitored for insulitis occurrence or diabetes development. We demonstrate that vaccination induces virus-neutralizing antibodies without altering insulitis scores or the onset of diabetes. We also show that NOD mice vaccinated with a CVB1 vaccine are protected from CVB-induced accelerated disease onset. Taken together, these studies show that CVB vaccines do not alter islet inflammation or accelerate disease progression in an animal model that spontaneously develops autoimmune type 1 diabetes. However, they can prevent CVB-mediated disease progression in the same model.

Genetic Adaptation of Coxsackievirus B1 during Persistent Infection in Pancreatic Cells.

Coxsackie B (CVB) viruses have been associated with type 1 diabetes. We have recently observed that CVB1 was linked to the initiation of the autoimmune process leading to type 1 diabetes in Finnish children. Viral persistency in the pancreas is currently considered as one possible mechanism. In the current study persistent infection was established in pancreatic ductal and beta cell lines (PANC-1 and 1.1B4) using four different CVB1 strains, including the prototype strain and three clinical isolates. We sequenced 5' untranslated region (UTR) and regions coding for structural and non-structural proteins and the second single open reading frame (ORF) protein of all persisting CVB1 strains using next generation sequencing to identify mutations that are common for all of these strains. One mutation, K257R in VP1, was found from all persisting CVB1 strains. The mutations were mainly accumulated in viral structural proteins, especially at BC, DE, EF loops and C-terminus of viral capsid protein 1 (VP1), the puff region of VP2, the knob region of VP3 and infection-enhancing epitope of VP4. This showed that the capsid region of the viruses sustains various changes during persistency some of which could be hallmark(s) of persistency.

Bradykinin as a Probable Aspect in SARS-Cov-2 Scenarios: Is Bradykinin Sneaking out of Our Sight?

The new virus SARS-CoV-2 is savagely spreading out over the world. The biologic studies show that the target receptor for the virus might be angiotensin-converting enzyme 2 (ACE2). This peptide is responsible for converting angiotensin II (Ang II), which is a profoundly active peptide, into Ang 1-7 with quite a balancing barbell function. It is emphasized that the direct target of the virus is ACE2 underlining the obvious difference with ACE. Nevertheless, we hypothesized that a back load build up effect on Ang II may usurp the ACE capacity and subsequently leave the bradykinin system unabated. We think there are clinical clues for dry cough and the presumed aggravating role of ACE inhibitors like captopril on the disease process. Thereby, we speculated that inhibition of bradykinin synthesis and/or blockade of bradykinin B2 receptor using Aprotinin/ecallantide and Icatibant, respectively, may hold therapeutic promise in severe cases and these molecules can be advanced to clinical trials.

Eradication of persistent coxsackievirus B infection from a pancreatic cell line with clinically used antiviral drugs.

BACKGROUND: Persistent enterovirus infections create a difficult therapeutic challenge in immunocompromised patients and may also contribute to the development of chronic diseases including type 1 diabetes, cardiomyopathies, post-polio syndrome and chronic fatigue syndrome. OBJECTIVES: To study the ability of antiviral drugs to eradicate such infection in vitro to evalaute their potential in the treatments of these patients. STUDY DESIGN: We set out to evaluate several licensed or clinically tested drugs which have shown some anti-enterovirus activity in previous studies for their ability to cure persistent infection established by two different coxsackievirus B1 strains in a pancreatic cell line (PANC-1 cells). RESULTS: Among all tested drugs Enviroxime, Fluoxetine, concentrated human IgG product (Hizentra) and Pleconaril were able to eradicate persistent Coxsackievirus B1 infection. The effect Enviroxime, Hizentra and Pleconaril varied between the two virus strains. CONCLUSIONS: The identified drugs are feasible candidates for clinical trials among patients with persistent coxsackievirus B infections or chronic enterovirus-associated diseases.

Enhancing and neutralizing anti-coxsackievirus activities in serum samples from patients prior to development of type 1 diabetes.

BACKGROUND: Studies in prospective cohorts have suggested that enterovirus infections are associated with the appearance of islet autoantibodies that precede later appearance of type 1 diabetes (T1D). It was shown that in addition to an antibody-mediated anti-coxsackievirus (CV)-B neutralizing activity of serum from patients with T1D, there was also enhancing anti-CV-B activity in vitro. In this study, the patterns of enhancing and neutralizing anti-CV activities were analysed from consecutive serum samples collected from children who were followed from birth until they developed T1D in the Finnish Type 1 Diabetes Prediction and Prevention (DIPP) study and compared to those in non-diabetic control children. METHODS: The titres of serum neutralizing activity were analysed against those CVs which were detected in the stools in these children (CV-B3, CV-B5 or CV-A4) using plaque assay. The enhancing activity of these serum samples was analysed by measuring interferon-alpha (INF-α) production in cultures of peripheral blood mononuclear cells (PBMC) inoculated with a mixture of these viruses and diluted serum. RESULTS: A sustained anti-CV enhancing activity was observed in consecutive serum samples in patients with T1D. The pattern of responses differed between children who developed T1D and control children. In patients, the anti-CV enhancing activity was predominant or even exclusive over the neutralizing activity, whereas in controls the enhancing and neutralising activities were more balanced or the neutralizing activity was largely predominant. CONCLUSIONS: Evaluating the anti-enterovirus neutralizing and enhancing activity of serum samples can be useful to investigate further the relationship between enteroviruses and the development of T1D.

Efficacy and safety of chloroquine and hydroxychloroquine for COVID-19: A comprehensive evidence synthesis of clinical, animal, and in vitro studies.

Background: The world is facing a pandemic of COVID-19, a respiratory disease caused by a novel coronavirus which is now called SARS-CoV-2. Current treatment recommendations for the infection are mainly repurposed drugs based on experience with other clinically similar conditions and are not backed by direct evidence. Chloroquine (CQ) and its derivative Hydroxychloroquine (HCQ) are among the candidates. We aimed to synthesize current evidence systematically for in vitro, animal, and human studies on the efficacy and safety of chloroquine in patients with COVID-19. Methods: The Cochrane Library, Google Scholar, PubMed (via Medline), Embase, Scopus, and Web of Science, MedRxiv, clinical trial registries including clinicaltrials.gov, ChiCTR (Chinese Clinical Trial Registry), IRCT (Iranian Registry of Clinical Trials), and the EU Clinical Trials Register. We used the Cochrane tool for risk of bias assessment in randomized studies, the ROBINS tool for non-randomized studies, and the GRADE methodology to summarize the evidence and certainty in effect estimates. Results: The initial database searching retrieved 24,752 studies. Of these, 15,435 abstracts were screened and 115 were selected for full-text review. Finally, 20 human studies, 3 animal studies, and 4 in vitro studies were included in this systematic review. The risk of bias within studies was unclear to high and the overall certainty in evidence-based on GRADES- was very low. HCQ may be effective in clinical improvement in a subset of patients with COVID-19. However, the frequency of adverse events was higher in patients taking HCQ compared to standard of care alone. In contrast, animal studies, did not report any adverse effects. Furthermore, clear benefit of the drug in the survival of the animals has been reported. Most in vitro studies indicated a high selectivity index for the drug and one study that used a human coronavirus reported blockage of virus replication. Conclusion: Current evidence background is limited to six poorly conducted clinical studies with inconsistent findings which fail to show significant efficacy for HCQ. Safety data is also limited but the drug may increase adverse outcomes. Routine use of the drug is not recommended based on limited efficacy and concerns about the drug safety especially in high-risk populations.

Combination of three virus-derived nanoparticles as a vaccine against enteric pathogens; enterovirus, norovirus and rotavirus.

Enteric viruses cause diverse infections with substantial morbidity and mortality in children, rotavirus (RV) and norovirus (NoV) being the leading agents of severe pediatric gastroenteritis. Coxsackie B viruses (CVB) are common enteroviruses (EV), associated with increased incidence of severe neonatal CVB disease with potentially fatal consequences. To prevent majority of childhood gastroenteritis, we have developed a non-live NoV-RV combination vaccine consisting of NoV virus-like particles (VLPs) and RV oligomeric rVP6 protein that induced protective immune responses to NoV and RV in mice. Moreover, rVP6 acted as an adjuvant for NoV VLPs. Here, we investigated a possibility to include a third enteric virus-derived antigen in the candidate NoV-RV vaccine, by adding recombinant nanoparticles derived from EV CVB1. To examine immunogenicity of EV-NoV-RV vaccine, BALB/c mice were immunized intramuscularly twice with 10 µg CVB1 VLPs, GII.4 VLPs and rVP6 nanotubes, either separately or combined. To evaluate the adjuvant effect of rVP6 on EV responses, mice received 0.3 µg CVB1 VLPs with or without 10 µg rVP6. Comparable serum IgG antibodies were detected whether the antigens were administered separately or in combination. Each formulation generated IgG1 and IgG2a antibodies, indicating a mixed Th2/Th1-type response. CVB1 VLPs skewed the isotype distribution slightly towards IgG1 subtype, while EV-NoV-RV combination vaccine induced unbiased Th1/Th2 responses to CVB1. Each antigen also induced T cell mediated immunity measured by IFN-γ secretion to specific stimulants ex vivo. Antisera raised by single antigens and combined formulation also exhibited strong neutralizing ability against CVB1 and NoV GII.4. Further, rVP6 showed an adjuvant effect on CVB1 responses, sparing the VLP dose and homogenizing the responses. Finally, the results support inclusion of additional antigens in the candidate NoV-RV combination vaccine to combat severe childhood infections and confirm adjuvant effect of rVP6 nanostructures.

A comparative study of the effect of UV and formalin inactivation on the stability and immunogenicity of a Coxsackievirus B1 vaccine.

Type B Coxsackieviruses (CVBs) belong to the enterovirus genus, and they cause both acute and chronic diseases in humans. CVB infections usually lead to flu-like symptoms but can also result in more serious diseases such as myocarditis, aseptic meningitis and life-threatening multi-organ infections in young infants. Thus, CVBs have long been considered as important targets of future vaccines. We have previously observed CVB1 capsid disintegration and virus concentration decrease with 12-day long formalin inactivation protocol. Here a scalable ion exchange chromatography purification method was developed, and purified CVB1 was inactivated with UV-C or formalin. Virus morphology and concentration remained unchanged, when the UV (2 min) or formalin (5 days) inactivation were performed in the presence of tween80 detergent. The concentration of the native and UV inactivated CVB1 remained constant at 4 °C during a six months stability study, whereas the concentration of the formalin inactivated vaccine decreased 29% during this time. UV treatment decreased, whereas formalin treatment increased the thermal stability of the capsid. The formalin inactivated CVB1 vaccine was more immunogenic than the UV inactivated vaccine; the protective neutralizing antibody levels were higher in mice immunized with formalin inactivated vaccine. High levels of CVB1 neutralizing antibodies as well as IgG1 antibodies were detected in mice that were protected against viremia induced by experimental CVB1 infection. In conclusion, this study describes a scalable ion exchange chromatography purification method and optimized 5-day long formalin inactivation method that preserves CVB1 capsid structure and immunogenicity. Formalin treatment stabilizes the virus particle at elevated temperatures, and the formalin inactivated vaccine induces high levels of serum IgG1 antibodies (Th2 type response) and protective levels of neutralizing antibodies. Formalin inactivated CVB vaccines are promising candidates for human clinical trials.

Formalin treatment increases the stability and immunogenicity of coxsackievirus B1 VLP vaccine.

Type B Coxsackieviruses (CVBs) are a common cause of acute and chronic myocarditis, dilated cardiomyopathy and aseptic meningitis. However, no CVB-vaccines are available for human use. We have previously produced virus-like particles (VLPs) for CVB3 with a baculovirus-insect cell production system. Here we have explored the potential of a VLP-based vaccine targeting CVB1 and describe the production of CVB1-VLPs with a scalable VLP purification method. The developed purification method consisting of tangential flow filtration and ion exchange chromatography is compatible with industrial scale production. CVB1-VLP vaccine was treated with UV-C or formalin to study whether stability and immunogenicity was affected. Untreated, UV treated and formalin treated VLPs remained morphologically intact for 12  months  at 4 °C. Formalin treatment increased, whereas UV treatment decreased the thermostability of the VLP-vaccine. High neutralising and total IgG antibody levels, the latter predominantly of a Th2 type (IgG1) phenotype, were detected in female BALB/c mice immunised with non-adjuvanted, untreated CVB1-VLP vaccine. The immunogenicity of the differently treated CVB1-VLPs (non-adjuvanted) were compared in C57BL/6 J mice and animals vaccinated with formalin treated CVB1-VLPs mounted the strongest neutralising and, CVB1-specific IgG and IgG1 antibody responses. This study demonstrates that formalin treatment increases the stability and immunogenicity of CVB1-VLP vaccine and may offer a universal tool for the stabilisation of VLPs in the production of more efficient vaccines.

Coxsackievirus B Persistence Modifies the Proteome and the Secretome of Pancreatic Ductal Cells.

The group B Coxsackieviruses (CVB), belonging to the Enterovirus genus, can establish persistent infections in human cells. These persistent infections have been linked to chronic diseases including type 1 diabetes. Still, the outcomes of persistent CVB infections in human pancreas are largely unknown. We established persistent CVB infections in a human pancreatic ductal-like cell line PANC-1 using two distinct CVB1 strains and profiled infection-induced changes in cellular protein expression and secretion using mass spectrometry-based proteomics. Persistent infections, showing characteristics of carrier-state persistence, were associated with a broad spectrum of changes, including changes in mitochondrial network morphology and energy metabolism and in the regulated secretory pathway. Interestingly, the expression of antiviral immune response proteins, and also several other proteins, differed clearly between the two persistent infections. Our results provide extensive information about the protein-level changes induced by persistent CVB infection and the potential virus-associated variability in the outcomes of these infections.

Complications of type 2 diabetes in Iranian population: An updated systematic review and meta-analysis.

Complications among patients with type 2 diabetes mellitus (T2DM) have increased dramatically through two past decades. Thus, the aim of this updated systematic review and meta-analysis was to estimate the pooled prevalence of T2DM complications in Iranian patients. Using Medical Subject Headings terms, Emtree, and related equal Persian key words, international databases including PubMed, ISI/WOS, Scopus, Iran Medex, SID, Magiran, Irandoc, Medlib, domestic databases were searched from January 1990 till January 2018 reporting prevalence of any complications of type 2 diabetes in Iran. All the keywords were searched electronically by two Boolean operators through the explained search strategy, separately. Relevant additional articles were identified from the lists of the retrieved articles. Random and fixed effect meta-analysis was used to estimate the pooled prevalence of complications in Iranian patients with T2DM. Through searching steps, among 1238 publications retrieved from literature search, finally 45 studies met the inclusion criteria for meta-analysis, with number of 30679 participants. According to random effect, the estimated pooled prevalence of diabetic foot ulcer, cardiovascular disease, retinopathy, neuropathy and nephropathy in Iranian patients with T2DM were 3%(95% CI: 1-5%), 33%(95% CI: 16-49%), 36%(95% CI: 27-45%), 38% (95% CI: 14-63%), and 43% (95% CI: 27-60%), respectively. This updated meta-analysis shows that prevalence of major microvascular complications of T2DM in Iran is high. Our findings provide practical evidence for better planning and clinical decision making.

The effect of diabetes on the risk of endometrial Cancer: an updated a systematic review and meta-analysis.

BACKGROUND: Previous studies conducted on the association between diabetes and the risk of endometrial cancer have reported controversial results that have raised a variety of questions about the association between diabetes and the incidence of this cancer. Thus, the aim of this systematic review and meta-analysis was to more precisely estimate the effect of diabetes on the risk of endometrial cancer incidence. METHODS: All original articles were searched in international databases, including Medline (PubMed), Web of sciences, Scopus, EMBASE, and CINHAL. Search was done from January 1990 to January 2018 without language limitations. Also, logarithm and standard error logarithm relative risk (RR) were used for meta-analysis. RESULTS: A total of 22 cohort and case-control studies were included in this meta-analysis, of which 14 showed statistically significant associations between diabetes and risk of endometrial cancer. Diabetes was associated with increased risk of endometrial cancer (RR = 1.72, 95% CI 1.48-2.01). The summary of RR for all 9 cohort studies was 1.56 (95% CI 1.21-2.01), and it was 1.85 (95% CI 1.53-2.23) for 13 case control studies. The summary of RR in hospital-based studies was higher than other studies. Thirteen of the primary studies-controlled BMI as a confounding variable, and the combined risk of their results was 1.62 (95% CI 1.34-1.97). CONCLUSIONS: Diabetes seems to increases the risk of endometrial cancer in women, and this finding can be useful in developing endometrial cancer prevention plans for women having diabetes.

Molecular epidemiology of enteroviruses in young children at increased risk of type 1 diabetes.

BACKGROUND: Young children are susceptible to enterovirus (EV) infections, which cause significant morbidity in this age group. However, the current knowledge regarding the epidemiology of EVs and the circulating virus strains is mostly based on viruses detected in children with severe diseases leading to contact with the health care system, while the vast reservoir of EVs that circulate in the general population is less characterized. METHODOLOGY: The present study investigates the types and the prevalence of EVs circulating in the young children of the background population in Georgia, Colorado, and Washington State in the USA, and Germany, Sweden, and Finland in Europe. A total of 4018 stool samples, collected monthly from 300 healthy and non-hospitalized children at the age of 3-18 months in 2005-2009, were analyzed for the presence of EVs using RT-PCR, followed by sequencing of the VP1-2A region of the viral genome to type the EV(s) present. All of the children carried type HLA-DQ2 or -DQ8 alleles associated with type 1 diabetes. PRINCIPAL FINDINGS: Altogether 201 children (67%) were found to be EV positive. The prevalence was much lower in Finnish children (26%) than in the children of the other counties combined (75%). Infections increased by age and showed a nadir during the winter months. Children who carried both the HLA-DQ2 and -DQ8 alleles had less infections than children who were homozygous for these alleles. Coxsackieviruses type A were the most frequently detected viruses in all geographical regions. Coxsackievirus type A4, Echovirus type 18, and Echovirus type 25 were shed for longer time periods than the other EV types. CONCLUSIONS: Compared to prevalence data from symptomatic patients requiring medical attention, this study provides a better view of EVs circulating in young children in the USA and in Europe. The observations may prove useful for the selection of strategies for designing EV vaccines in the future. The study also confirms our previous serological findings suggesting that EV infections are relatively rare in Finland.