Novel X gene point mutations in chronic hepatitis B and HBV related cirrhotic patients.

INTRODUCTION: HBx is a multifunctional modulator viral protein with key roles in various biological processes such as signal transduction, transcription, proliferation, and cell apoptosis. Also, HBx has an important role in the progression of cirrhosis and hepatocellular carcinoma (HCC). This study aimed to determine mutations in X gene, enhancer II (EnhII), and basal core promoter (BCP) of genotype D of Hepatitis B Virus (HBV) in cirrhotic and chronic HBV patients. MATERIAL AND METHODS: This cross-sectional study was performed on 68 cases with chronic HBV (cHBV) and 50 cases with HBV related cirrhosis. Serum samples were obtained for genomic DNA extraction. Semi-nested PCR was used to amplify the HBx region. Point mutations in the HBx region were detected by sequencing. RESULT: Novel mutations were detected, including C1491G, C1500T, G1613T, and G1658T in the N-terminal of the X gene. The frequency of C1481T/G1479A, T1498C, C1500T, G1512A, A1635T, C1678T, A1727T, and A1762T/ G1764A/ C1773T was significantly higher in cirrhotic patients compared to chronically HBV infected ones. A higher rate of A1635T, C1678T, A1727T, A1762T, G1764A, and C1773T was observed in cirrhotic patients. CONCLUSION: Our findings showed that the frequency of mutations in the basal-core promoter, enhancer II, and regulatory region of the HBx gene was more seen in cirrhotic patients than in chronic HBV cases. Novel mutations were detected in the HBx gene, causing amino acid substitutions; however, the clinical impact of these novel mutations is yet to be cleared.

A single-cell transcriptomic landscape of the lungs of patients with COVID-19.

The lung is the primary organ targeted by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), making respiratory failure a leading coronavirus disease 2019 (COVID-19)-related mortality. However, our cellular and molecular understanding of how SARS-CoV-2 infection drives lung pathology is limited. Here we constructed multi-omics and single-nucleus transcriptomic atlases of the lungs of patients with COVID-19, which integrate histological, transcriptomic and proteomic analyses. Our work reveals the molecular basis of pathological hallmarks associated with SARS-CoV-2 infection in different lung and infiltrating immune cell populations. We report molecular fingerprints of hyperinflammation, alveolar epithelial cell exhaustion, vascular changes and fibrosis, and identify parenchymal lung senescence as a molecular state of COVID-19 pathology. Moreover, our data suggest that FOXO3A suppression is a potential mechanism underlying the fibroblast-to-myofibroblast transition associated with COVID-19 pulmonary fibrosis. Our work depicts a comprehensive cellular and molecular atlas of the lungs of patients with COVID-19 and provides insights into SARS-CoV-2-related pulmonary injury, facilitating the identification of biomarkers and development of symptomatic treatments.

Role of the CXCR3­mediated TLRs/MyD88 signaling pathway in promoting the development of hepatitis B into cirrhosis and liver cancer.

Chronic hepatitis B can lead to liver cirrhosis and primary hepatocellular carcinoma. The present study aimed to investigate whether C­X­C motif chemokine receptor 3 (CXCR3) regulates the genes in Toll­like receptors (TLRs)/myeloid differentiation primary response protein 88 (MyD88) signaling pathway in the development of hepatitis B into cirrhosis and liver cancer in vitro. A hepatitis B virus (HBV) overexpression lentivirus was constructed and infected into a LX­2 cell line to obtain stable HBV­overexpressing cells (named HBV­LX­2 cells). The CXCR3 gene was knocked down using small interfering RNA in HBV­LX­2 cells. Cell Counting Kit­8 assays, cell scratch tests and flow cytometry were used to detect cell proliferation, migration and apoptosis, respectively. The levels of IL­1ß and IL­6 in serum samples of patients with liver cancer were measured via ELISA, and the collagen content in liver cancer tissues was detected using Masson staining. Western blotting was used to detect the expression levels of proteins in the TLRs/MyD88 signaling pathway. Excessive fibrosis was identified in the liver cancer tissues, and the serum levels of IL­6 and IL­1ß were abnormally increased in patients with liver cancer. It was found that interfering with CXCR3 inhibited cell proliferation and migration, as well as promoted the apoptosis of HBV­LX­2 cells. Moreover, interfering with CXCR3 inhibited the expression levels of collagen type I α 1 chain and the proteins in the TLRs/MyD88 pathway. In conclusion, CXCR3 knockdown could inhibit the expression levels of proteins in the TLR4/MyD88 signaling pathway, decrease cell proliferation and migration, and promote cell apoptosis, thus inhibiting the development of liver cirrhosis to liver cancer.

αV ß6 Integrin: An Intriguing Target for COVID-19 and Related Diseases.

The outbreak of SARS-CoV-2 has been an extraordinary event that constituted a global health emergency. As the novel coronavirus is continuing to spread over the world, the need for therapeutic agents to control this pandemic is increasing. αV ß6 Integrin may be an intriguing target not only for the inhibition of SARS-CoV-2 entry, but also for the diagnosis/treatment of COVID-19 related fibrosis, an emerging type of fibrotic disease which will probably affect a significant part of the recovered patients. In this short article, the possible role of this integrin for fighting COVID-19 is discussed on the basis of recently published evidence, showing how its underestimated involvement may be interesting for the development of novel pharmacological tools.

Viral fibrotic scoring and drug screen based on MAPK activity uncovers EGFR as a key regulator of COVID-19 fibrosis.

Understanding the molecular basis of fibrosis, the lethal complication of COVID-19, is urgent. By the analysis of RNA-sequencing data of SARS-CoV-2-infected cells combined with data mining we identified genes involved in COVID-19 progression. To characterize their implication in the fibrosis development we established a correlation matrix based on the transcriptomic data of patients with idiopathic pulmonary fibrosis. With this method, we have identified a cluster of genes responsible for SARS-CoV-2-fibrosis including its entry receptor ACE2 and epidermal growth factor EGF. Then, we developed Vi-Fi scoring-a novel drug repurposing approach and simultaneously quantified antiviral and antifibrotic activities of the drugs based on their transcriptomic signatures. We revealed the strong dual antifibrotic and antiviral activity of EGFR/ErbB inhibitors. Before the in vitro validation, we have clustered 277 cell lines and revealed distinct COVID-19 transcriptomic signatures of the cells with similar phenotypes that defines their suitability for COVID-19 research. By ERK activity monitoring in living lung cells, we show that the drugs with predicted antifibrotic activity downregulate ERK in the host lung cells. Overall, our study provides novel insights on SARS-CoV-2 dependence on EGFR/ERK signaling and demonstrates the utility of EGFR/ErbB inhibitors for COVID-19 treatment.

A molecular single-cell lung atlas of lethal COVID-19.

Respiratory failure is the leading cause of death in patients with severe SARS-CoV-2 infection1,2, but the host response at the lung tissue level is poorly understood. Here we performed single-nucleus RNA sequencing of about 116,000 nuclei from the lungs of nineteen individuals who died of COVID-19 and underwent rapid autopsy and seven control individuals. Integrated analyses identified substantial alterations in cellular composition, transcriptional cell states, and cell-to-cell interactions, thereby providing insight into the biology of lethal COVID-19. The lungs from individuals with COVID-19 were highly inflamed, with dense infiltration of aberrantly activated monocyte-derived macrophages and alveolar macrophages, but had impaired T cell responses. Monocyte/macrophage-derived interleukin-1ß and epithelial cell-derived interleukin-6 were unique features of SARS-CoV-2 infection compared to other viral and bacterial causes of pneumonia. Alveolar type 2 cells adopted an inflammation-associated transient progenitor cell state and failed to undergo full transition into alveolar type 1 cells, resulting in impaired lung regeneration. Furthermore, we identified expansion of recently described CTHRC1+ pathological fibroblasts3 contributing to rapidly ensuing pulmonary fibrosis in COVID-19. Inference of protein activity and ligand-receptor interactions identified putative drug targets to disrupt deleterious circuits. This atlas enables the dissection of lethal COVID-19, may inform our understanding of long-term complications of COVID-19 survivors, and provides an important resource for therapeutic development.

[Histopathological features due to the SARS-CoV-2]. / Les lésions histologiques associées à l'infection par le SARS-CoV-2.

The infection due to the SARS-CoV-2 leads lesions mainly observed at the respiratory tract level, but not exclusively. The analyses of these lesions benefited from different autopsy studies. Thus, these lesions were observed in different organs, tissues and cells. These observations allowed us to rapidly improve the knowledge of the pathophysiological mechanisms associated with this emergent infectious disease. The virus can be detected in formalin fixed paraffin embedded tissues using immunohistochemistry, in situ hybridization, molecular biology and/or electron microscopy approaches. However, many uncertainties are still present concerning the direct role of the SARS-CoV-2 on the different lesions observed in different organs, outside the lung, such as the heart, the brain, the liver, the gastrointestinal tract, the kidney and the skin. In this context, it is pivotal to keep going to increase the different tissue and cellular studies in the COVID-19 positive patients aiming to better understanding the consequences of this new infectious disease, notably considering different epidemiological and co-morbidities associated factors. This could participate to the development of new therapeutic strategies too. The purpose of this review is to describe the main histological and cellular lesions associated with the infection due to the SARS-CoV-2.

Fibrotic progression and radiologic correlation in matched lung samples from COVID-19 post-mortems.

Data on the pathology of COVID-19 are scarce; available studies show diffuse alveolar damage; however, there is scarce information on the chronologic evolution of COVID-19 lung lesions. The primary aim of the study is to describe the chronology of lung pathologic changes in COVID-19 by using a post-mortem transbronchial lung cryobiopsy approach. Our secondary aim is to correlate the histologic findings with computed tomography patterns. SARS-CoV-2-positive patients, who died while intubated and mechanically ventilated, were enrolled. The procedure was performed 30 min after death, and all lung lobes sampled. Histopathologic analysis was performed on thirty-nine adequate samples from eight patients: two patients (illness duration < 14 days) showed early/exudative phase diffuse alveolar damage, while the remaining 6 patients (median illness duration-32 days) showed progressive histologic patterns (3 with mid/proliferative phase; 3 with late/fibrotic phase diffuse alveolar damage, one of which with honeycombing). Immunohistochemistry for SARS-CoV-2 nucleocapsid protein was positive predominantly in early-phase lesions. Histologic patterns and tomography categories were correlated: early/exudative phase was associated with ground-glass opacity, mid/proliferative lesions with crazy paving, while late/fibrous phase correlated with the consolidation pattern, more frequently seen in the lower/middle lobes. This study uses an innovative cryobiopsy approach for the post-mortem sampling of lung tissues from COVID-19 patients demonstrating the progression of fibrosis in time and correlation with computed tomography features. These findings may prove to be useful in the correct staging of disease, and this could have implications for treatment and patient follow-up.

Impact of Human Cytomegalovirus and Human Herpesvirus 6 Infection on the Expression of Factors Associated with Cell Fibrosis and Apoptosis: Clues for Implication in Systemic Sclerosis Development.

Systemic sclerosis (SSc) is a severe autoimmune disorder characterized by vasculopathy and multi-organ fibrosis; its etiology and pathogenesis are still largely unknown. Herpesvirus infections, particularly by human cytomegalovirus (HCMV) and human herpesvirus 6 (HHV-6), have been suggested among triggers of the disease based on virological and immunological observations. However, the direct impact of HCMV and/or HHV-6 infection on cell fibrosis and apoptosis at the cell microenvironment level has not yet been clarified. Thus, this study aimed to investigate the effects of HCMV and HHV-6 infection on the induction of pro-fibrosis or pro-apoptosis conditions in primary human dermal fibroblasts, one of the relevant SSc target cells. The analysis, performed by microarray in in vitro HCMV- or HHV-6-infected vs. uninfected cells, using specific panels for the detection of the main cellular factors associated with fibrosis or apoptosis, showed that both viruses significantly modified the expression of at least 30 pro-fibrotic and 20 pro-apoptotic factors. Notably, several recognized pro-fibrotic factors were highly induced, and most of them were reported to be involved in vivo in the multifactorial and multistep pathogenic process of SSc, thus suggesting a potential role of both HCMV and HHV-6.

Identification of a new strain of mouse kidney parvovirus associated with inclusion body nephropathy in immunocompromised laboratory mice.

Inclusion body nephropathy (IBN) and kidney fibrosis in aged immunodeficient mice and, to lesser extent, in immunocompetent mice have been recently linked to infection of mouse kidney parvovirus (MKPV), also known as murine chapparvovirus (MuCPV). Knowledge about its prevalence and the complete genome sequence of more MKPV strains is essential for understanding phylogenetic relationships and pathogenicity among MKPV strains. In the present study using PCR and genome walking, we determined the complete 4440-nucleotide genome of a new MKPV strain, namely MIT-WI1, which was identified in IBN-affected Il2rg-/-Rag2-/- c-Kit W-sh/W-sh mice housed in the vivarium at Whitehead Institute for Biomedical Research (WI). The overall nucleotide (>94%) and deduced amino acid sequences (>98%) of p10, p15, NS1 (replicase), NS2 and VP1 (capsid protein) within the MIT-WI1 genome, are closely related to MKPV/MuCPV strains described in laboratory and wild Mus musculus mice. In addition, PCR and qPCR assays using newly designed primers conserved among the known MKPV/MuCPV genomes were developed and utilized to assess MKPV status in selected laboratory mice. MKPV was also detected in immunodeficient (NSG) and immunocompetent (Crl:CD1(ICR), UTXflox) mouse strains/stocks. The abundance of the MKPV genome copies was significantly correlated with the severity of IBN. Our data indicate that MKPV is present in selected mouse strains/stocks, and provides new insights into the genome evolution of MKPV.

Heart Dysfunction Following Long-Term Murine Cytomegalovirus Infection: Fibrosis, Hypertrophy, and Tachycardia.

Human cytomegalovirus (HCMV) is associated with increased risk of chronic diseases of the heart and vasculature, including myocarditis, atherosclerosis, and transplant vasculopathy. To investigate CMV infection of the heart, murine cytomegalovirus (MCMV) was used to evaluate both acute and latent infection and the subsequent phenotypic and functional consequences of infection. Female BALB/c mice were intraperitoneally (i.p.) inoculated with 1 × 106 pfu of MCMV and evaluated at 14 and 50 days postinfection (dpi). At each time point, echocardiography was used to evaluate cardiac function and histology was conducted for phenotypic evaluation. MCMV replication in the heart was detected as early as 3 dpi and was no longer detectable at 14 dpi. Infected animals had significant cardiac pathology at 14 and 50 dpi when compared to uninfected controls. Histology revealed fibrosis of the heart as early as 14 dpi and the presence of white fibrous deposits on the surface of the heart. Functional evaluation showed significantly increased heart rate and muscle thickening in the latently infected animals when compared to the control animals. At 50 dpi, latent virus was measured by explant reactivation assay, demonstrating that MCMV establishes latency and is capable of reactivation from the heart, similar to other tissues such as spleen and salivary glands. Collectively, these studies illustrate that MCMV infection results in phenotypic alterations within the heart as early as 14 dpi, which progress to functional abnormalities during latency. These findings are similar to sinus tachycardia and hypertrophy of the heart muscle observed in cases of HCMV-induced acute myocarditis.

Immune Correlates of Diffuse Myocardial Fibrosis and Diastolic Dysfunction Among Aging Women With Human Immunodeficiency Virus.

Human immunodeficiency virus (HIV) imparts increased heart failure risk to women. Among women with HIV (WHIV), immune pathways relating to heart failure precursors may intimate targets for heart failure prevention strategies. Twenty asymptomatic, antiretroviral-treated WHIV and 14 non-HIV-infected women matched on age and body mass index underwent cardiac magnetic resonance imaging and immune phenotyping. WHIV (vs non-HIV-infected women) exhibited increased myocardial fibrosis (extracellular volume fraction, 0.34 ± 0.06 vs 0.29 ± 0.04; P = .002), reduced diastolic function (diastolic strain rate, 1.10 ± 0.23 s-1 vs 1.39 ± 0.27 s-1; P = .003), and heightened systemic monocyte activation. Among WHIV, soluble CD163 levels correlated with myocardial fibrosis (r = 0.53; P = .02), while circulating inflammatory CD14+CD16+ monocyte CCR2 expression related directly to myocardial fibrosis (r = 0.48; P = .04) and inversely to diastolic function (r = -0.49; P = .03). Clinical Trials Registration. NCT02874703.

Molecular epidemiology, phylogenetic analysis and genotype distribution of hepatitis B virus in Saudi Arabia: Predominance of genotype D1.

Despite the implementation of various vaccination programs, hepatitis B virus (HBV) poses a considerable health problem in Saudi Arabia. Insight on HBV evolutionary history in the region is limited. We performed a comprehensive epidemiological and phylogenetic reconstruction based on a large cohort of HBV infected patients. Three hundred and nineteen HBV-infected patients with different clinical manifestations, including inactive and active chronic carriers and patients with cirrhosis and hepatocellular carcinoma (HCC), were enrolled in this study. The full-length large S gene was amplified and sequenced. Phylogenetic analysis was performed to determine the genotype and subgenotypes of the isolates. Phylogenetic tree analysis revealed that genotype D is the most dominant genotype among patients. Moreover, this analysis identified two strains with genotype E isolated from active carriers. Detailed phylogenetic analyses confirmed the presence of four HBV D subgenotypes, D1 (93%, n = 296), D2 (0.02%, n = 5), D3 (0.003%, n = 1), and D4 (0.003%, n = 1). In addition, six genotype D strains were not assigned to any existing HBV D subgenotype. The large S gene of eight strains showed signatures of genotype recombination between the genotypes D and A and between D and E. Several strains harbored medically important point mutations at the protein level. Along with the dominance of the HBV genotype D, isolation of the E genotype and several recombinant strains from patients with Saudi Arabian origin is an essential result for decisions involving therapeutic measures for patients. Development of vaccines and detection of diagnostic escape mutations at antigenic epitopes on the HBsAg will be valuable to public health authorities. Furthermore, the diversity at the nucleotide and amino acid levels and different proportions of dN/dS at the PreS1, PreS2, and HBsAg reveal the selective pressure trend from inactive status towards advanced liver diseases.

The effects of losartan on cytomegalovirus infection in human trabecular meshwork cells.

BACKGROUND: Human cytomegalovirus (CMV) has been emerged as one of the causes of acute recurrent or chronic hypertensive anterior uveitis in immunocompetent. In hypertensive anterior uveitis, human trabecular meshwork (TM) cells are considered a focus of inflammation. We investigated the effects of losartan, a selective angiotensin II receptor antagonist, on CMV infection in human TM cells. METHODS: Human TM cells were infected with CMV AD169. Virus infected and mock-infected cells were treated with losartan or dexamethasone or ganciclovir with or without transforming growth factor (TGF)-ß1. Viral DNA accumulation and host cell response were analyzed using real-time PCR. Levels of secreted TGF-ß1 were measured by determining its concentration in conditioned medium using a commercially available sandwich enzyme-linked immunosorbent assay (ELISA) kits. RESULTS: CMV infection significantly increased the concentrations of the secreted TGF-ß1 at 3, 5, and 7 day post infection in TM cells. Treatment with dexamethasone or losartan significantly decreased the levels of TGF-ß1, whereas treatment with ganciclovir did not affect TGF-ß1 levels. TM cells treated with TGF-ß1 along with the presence of losartan for 48 hours showed marked decrease in the expression of α-smooth muscle actin (SMA), lysyl oxidase (LOX), connective tissue growth factor (CTGF), fibronectin and collagen-1A, compared with cells treated with TGF-ß1 alone. CMV-infected TM cells stimulated by TGF-ß1 significantly increased the expression of α-SMA and CTGF, which were attenuated by additional treatment with losartan. CONCLUSION: Losartan inhibited the expression of TGF-ß1 and fibrogenic molecules in human TM cells. Thus, losartan has the potential to decrease TM fibrosis in patients with CMV-induced hypertensive anterior uveitis.

Perspectives on the expansion of human precision oncology and genomic approaches to sea turtle fibropapillomatosis.

Our recent Communications Biology research article revealed the genomic drivers and therapeutic vulnerabilities of sea turtle fibropapillomatosis tumors. Fibropapillomatosis is a debilitating tumorous disease afflicting populations of green sea turtles globally. While a virus is involved in the development of this disease, it is increasingly understood that the key trigger is linked to anthropogenic disturbances of the environment. The specific environmental co-trigger(s) has yet to be functionally confirmed. Here we outline the next steps required to advance our understanding of this enigmatic disease, to enable us to more effectively clinically combat it and to ultimately tackle its environmental co-trigger to halt and hopefully reverse the spread of fibropapillomatosis.

Glecaprevir/pibrentasvir for the treatment of chronic hepatitis C virus infection.

In recent years, management of chronic hepatitis C virus (HCV) infection has been revolutionized by the availability of oral direct-acting antivirals (DAAs), which have significantly better efficacy and safety profiles than interferon-containing regimens. Simple, short-duration DAA therapies will facilitate expansion of HCV treatment to nonspecialist providers, which will be vital to achieve the WHO target of eliminating chronic HCV as a major public health threat by 2030. Coformulated glecaprevir/pibrentasvir is the only 8-week, pan-genotypic, 2-DAA regimen recommended by international guidelines as a first-line regimen in treatment-naive, noncirrhotic HCV genotype 1-6 patients. This review provides a comprehensive summary of the pharmacodynamic and pharmacokinetic parameters, efficacy, safety and place in the HCV treatment paradigm for glecaprevir/pibrentasvir.

Identification of abnormal fucosylated-glycans recognized by LTL in saliva of HBV-induced chronic hepatitis, cirrhosis, and hepatocellular carcinoma.

The hepatitis B virus (HBV)-induced chronic liver diseases are serious health threats worldwide. There is evidence to display the alterations of salivary N-linked glycans related to the development of HBV-infected liver diseases. Here, we further investigated the alterations of fucosylated N/O-glycans recognized by LTL in saliva from 120 subjects (30 healthy volunteers (HV), 30 patients with hepatitis B (HB), 30 patients with hepatic cirrhosis (HC), and 30 patients with hepatocellular carcinoma (HCC)) using salivary microarrys and MALDI-TOF/TOF-MS. The results showed that the expression level of fucosylated glycans recognized by LTL was significantly increased in HCC compared with other subjects (P < 0.0001). Besides, the fucosylated glycoproteins were isolated from pooled saliva of HV, HB, HC, and HCC by LTL-magnetic particle conjugates. Then, N/O- glycans were released from the isolated glycoproteins with PNGase F and NaClO, and were identified by MALDI-TOF-MS, respectively. Totally, there were 21/20, 25/18, 29/19, and 28/24 N/O-glycan peaks that were identified and annotated with proposed structures in saliva of HV, HB, HC, and HCC. Among the total, there were 8 N-glycan peaks (e.g., m/z 1905.634, 2158.777 and 2905.036) and 15 O-glycan peaks (e.g., 1177.407, 1308.444 and 1322.444) that only presented in patients with HBV-induced liver diseases. One N-glycan peak (m/z 2205.766) was unique in HC, and 9 O-glycan peaks (e.g., m/z 1157.420, 1163.417 and 1193.402) were unique in HCC. This study could facilitate the discovery of biomarkers for HC and HCC based on precise alterations of fucosylated N/O-glycans in saliva.

Transcriptional Regulatory Networks in Hepatitis C Virus-induced Hepatocellular Carcinoma.

Understanding the transcriptional regulatory elements that influence the progression of liver disease in the presence of hepatitis C virus (HCV) infection is critical for the development of diagnostic and therapeutic approaches. Systems biology provides a roadmap by which these elements may be integrated. In this study, a previously published dataset of 124 microarray samples was analyzed in order to determine differentially expressed genes across four tissue types/conditions (normal, cirrhosis, cirrhosis HCC, and HCC). Differentially expressed genes were assessed for their functional clustering and those genes were annotated with their potential transcription factors and miRNAs. Transcriptional regulatory networks were constructed for each pairwise comparison between the 4 tissue types/conditions. Based on our analysis, it is predicted that the disruption in the regulation of transcription factors such as AP-1, PPARγ, and NF-κB could contribute to the liver progression from cirrhosis to steatosis and eventually to HCC. Whereas the condition of the liver digresses, the downregulation of miRNAs' (such as miR-27, Let-7, and miR-106a) expression makes the transition of the liver through each pathological stage more apparent. This preliminary data can be used to guide future experimental work. An understanding of the transcriptional regulatory attributes acts as a road map to help design interference strategies in order to target the key regulators of progression of HCV induced HCC.

An Atypical Parvovirus Drives Chronic Tubulointerstitial Nephropathy and Kidney Fibrosis.

The occurrence of a spontaneous nephropathy with intranuclear inclusions in laboratory mice has puzzled pathologists for over 4 decades, because its etiology remains elusive. The condition is more severe in immunodeficient animals, suggesting an infectious cause. Using metagenomics, we identify the causative agent as an atypical virus, termed "mouse kidney parvovirus" (MKPV), belonging to a divergent genus of Parvoviridae. MKPV was identified in animal facilities in Australia and North America, is transmitted via a fecal-oral or urinary-oral route, and is controlled by the adaptive immune system. Detailed analysis of the clinical course and histopathological features demonstrated a stepwise progression of pathology ranging from sporadic tubular inclusions to tubular degeneration and interstitial fibrosis and culminating in renal failure. In summary, we identify a widely distributed pathogen in laboratory mice and establish MKPV-induced nephropathy as a new tool for elucidating mechanisms of tubulointerstitial fibrosis that shares molecular features with chronic kidney disease in humans.

The role of the NLRP3 inflammasome and the activation of IL-1ß in the pathogenesis of chronic viral hepatic inflammation.

BACKGROUND AND AIMS: Chronic viral hepatitis is a prevalent disease with major health implications. Its underlying pathophysiological mechanisms are not fully understood. IL-1ß and the NLRP3 inflammasome involvement has been suggested in recent years, from in vitro data and data from peripheral blood samples. Therefore, we investigated IL-1ß and the NLRP3 inflammasome in liver tissues in an effort to clarify their role in the pathophysiology of chronic viral hepatitis. METHODS: We studied liver biopsies from patients with a new diagnosis of either chronic hepatitis B (CHB) and chronic hepatitis C (CHC) or patients with chronic hepatitis B in remission (CHB-rem). The biopsies were separated in two parts. The first part was sent to histology to determine the grade of inflammation and fibrosis. From the second part, RNA was extracted and converted to cDNA used in semi-quantitative Real-Time PCR to measure the levels of IL1B, CASP1, NLRP3, ASC and IL1RA. The cell lines used in the in vitro experiments were Huh7.5, LX2 and THP-1 in variety of combinations of monocultures, co-cultures and triple cultures with one of the cell lines infected with the JFH-1 HCV clone. From the cell cultures RNA was extracted and converted to cDNA. For cell lines, we focused in the expression of IL1B and NLRP3. RESULTS: The expression of IL1B, CASP1 and NLRP3 were found significantly different between our groups (p = 0.001, p = 0.001 and p = 0.038, respectively). CHB patients displayed significantly higher IL1B and CASP1 mRNA levels compared to both CHB-rem and CHC patients. IL1B expression significantly correlates with liver biochemical data in CHB patients (AST: p = 0.006, r = 0.457; ALT p = 0.002, r = 0.497). Finally, mRNA levels of IL1B in CHB patients significantly correlate with the degree of inflammation (p = 0.016) but not the stage of fibrosis (p = 0.362). Interestingly, the relative expression of IL1B in triple culture experiments in vitro was below of 1.5-fold, suggesting no activation of IL1B. Moreover, no activation of NLRP3 was demonstrated in all investigated in vitro conditions. CONCLUSION: IL-1ß might play an important role in the pathogenesis of chronic hepatic inflammation from HBV, but not from HCV.