Effector Role of Neonatal Hepatic CD8+ Lymphocytes in Epithelial Injury and Autoimmunity in Experimental Biliary Atresia

University of Cincinnati, Cincinnati, Ohio, United States
Gastroenterology (Impact Factor: 16.72). 07/2007; 133(1):268-77. DOI: 10.1053/j.gastro.2007.04.031
Source: PubMed


Lymphocytes populate the livers of infants with biliary atresia, but it is unknown whether neonatal lymphocytes regulate pathogenesis of disease. Here, we investigate this question by examining the role of T lymphocytes in the destruction of extrahepatic bile ducts of neonatal mice using an experimental model of biliary atresia.
Inoculation of neonatal mice with rhesus rotavirus followed by multistaining flow cytometry to quantify expression of interferon-gamma by hepatic lymphocytes, and real-time polymerase chain reaction for mRNA expression of pro-inflammatory cytokines. This was followed by determining the consequences of antibody-mediated depletion of lymphocyte subtypes on the development of biliary obstruction, and coculture and cell transfer experiments to investigate the effector role of lymphocyte subtypes on neonatal biliary disease.
Rotavirus infection results in overexpression of interferon-gamma by neonatal hepatic T cells. Among these cells, depletion of CD4(+) cells did not change the course of inflammatory injury and obstruction of neonatal bile ducts. In contrast, loss of CD8(+) cells remarkably suppressed duct injury, prevented luminal obstruction, and restored bile flow. Coculture experiments showed that rotavirus-primed, but not naïve, CD8(+) cells were cytotoxic to cholangiocytes. In adoptive transfer experiments, we found that primed CD8(+) cells preferentially homed to extrahepatic bile ducts of neonatal mice and invaded their epithelial lining.
Primed neonatal CD8(+) cells can activate a pro-inflammatory program, target diseased and healthy duct epithelium, and drive the phenotypic expression of biliary atresia, thus constituting a potential therapeutic target to halt disease progression.

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Available from: Pranavkumar Shivakumar
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    • "Previously, we used a similar approach to survey the gene expression signature in bile ducts in search of genes or gene-groups with potential roles in pathogenesis of biliary atresia [5]. The strength of the approach was validated in subsequent studies showing that the suppression of gene groups induced by the loss of CD8+ lymphocytes [12] and natural killer [33] and dendritic cells [34] substantially decreased epithelial injury, prevented bile duct obstruction, and improved cholestasis. "
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    ABSTRACT: Biliary atresia is a fibroinflammatory obstruction of extrahepatic bile duct that leads to end-stage liver disease in children. Despite advances in understanding the pathogenesis of biliary atresia, very little is known about the role of microRNAs (miRNAs) in onset and progression of the disease. In this study, we aimed to investigate the entire biliary transcriptome to identify miRNAs with potential role in the pathogenesis of bile duct obstruction. By profiling the expression levels of miRNA in extrahepatic bile ducts and gallbladder (EHBDs) from a murine model of biliary atresia, we identified 14 miRNAs whose expression was suppressed at the times of duct obstruction and atresia (>=2 fold suppression, P < 0.05, FDR 5%). Next, we obtained 2,216 putative target genes of the 14 miRNAs using in silico target prediction algorithms. By integrating this result with a genome-wide gene expression analysis of the same tissue (>=2 fold increase, P < 0.05, FDR 5%), we identified 26 potential target genes with coordinate expression by the 14 miRNAs. Functional analysis of these target genes revealed a significant relevance of miR-30b/c, -133a/b, -195, -200a, -320 and -365 based on increases in expression of at least 3 target genes in the same tissue and 1st-to-3rd tier links with genes and gene-groups regulating organogenesis and immune response. These miRNAs showed higher expression in EHBDs above livers, a unique expression in cholangiocytes and the subepithelial compartment, and were downregulated in a cholangiocyte cell line after RRV infection. Integrative genomics reveals functional relevance of miR-30b/c, -133a/b, -195, -200a, -320 and -365. The coordinate expression of miRNAs and target genes in a temporal-spatial fashion suggests a regulatory role of these miRNAs in pathogenesis of experimental biliary atresia.
    Full-text · Article · Oct 2013 · BMC Systems Biology
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    • "Depletion of CD8+ cells, as well as loss of IFN-γ, reduces the incidence of experimental BA. IFN-type II regulated chemokines such as Mig, IP-10 and I-Tac can be shown to peak at day 7 only in BA-developing Balb/c but not in IFN-γ–/–mice [110]. Thus in these knock-out mice, early lobular and portal inflammation with periductal infiltration by neutrophils is still seen at day 3 and is followed by bile duct proliferation. "
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    ABSTRACT: Biliary atresia (BA) is a rare disease of unknown etiology and unpredictable outcome, even when there has been timely diagnosis and exemplary surgery. It has been the commonest indication for liver transplantation during childhood for the past 20 years. Hence much clinical and basic research has been directed at elucidating the origin and pathology of BA. This review summarizes the current clinical variations of BA in humans, its occasional appearance in animals and its various manifestations in the laboratory as an experimental model.
    Full-text · Article · Aug 2013 · Orphanet Journal of Rare Diseases
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    • "In order to perform mechanistic studies, the Rhesus rotavirus (RRV)-induced mouse model of BA has been employed by many investigators [11–14]. In this model, the bile duct injury is associated with Th1-mediated inflammation and specifically with bile duct epithelial autoreactive T cells [15,16]. Much less is known about the role of B cells in the pathogenesis of BA. "
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    ABSTRACT: A leading theory regarding the pathogenesis of biliary atresia (BA) is that bile duct injury is initiated by a virus infection, followed by an autoimmune response targeting bile ducts. In experimental models of autoimmune diseases, B cells have been shown to play an important role. The aim of this study was to determine the role of B cells in the development of biliary obstruction in the Rhesus rotavirus (RRV)-induced mouse model of BA. Wild-type (WT) and B cell-deficient (Ig-α(-/-)) mice received RRV shortly after birth. Ig-α(-/-) RRV-infected mice had significantly increased disease-free survival rate compared to WT RRV-infected BA mice (76.8% vs. 17.5%). In stark contrast to the RRV-infected BA mice, the RRV-infected Ig-α(-/-) mice did not have hyperbilirubinemia or bile duct obstruction. The RRV-infected Ig-α(-/-) mice had significantly less liver inflammation and Th1 cytokine production compared to RRV-infected WT mice. In addition, Ig-α(-/-) mice had significantly increased numbers of regulatory T cells (Tregs) at baseline and after RRV infection compared to WT mice. However, depletion of Tregs in Ig-α(-/-) mice did not induce biliary obstruction, indicating that the expanded Tregs in the Ig-α(-/-) mice were not the sole reason for protection from disease. Conclusion : B cell deficient Ig-α(-/-) mice are protected from biliary obstruction in the RRV-induced mouse model of BA, indicating a primary role of B cells in mediating disease pathology. The mechanism of protection may involve lack of B cell antigen presentation, which impairs T-cell activation and Th1 inflammation. Immune modulators that inhibit B cell function may be a new strategy for treatment of BA.
    Preview · Article · Aug 2013 · PLoS ONE
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