Kenichi Harada

Periductal interleukin-17 production in association with biliary innate immunity contributes to the pathogenesis of cholangiopathy in primary biliary cirrhosis

An innate immune response to bacterial components is speculated to be involved in the pathogenesis of primary biliary cirrhosis (PBC). Recently, CD4‐positive T helper type 17 (Th17) cells, characterized by the secretion of interleukin (IL)‐17, have been implicated in the pathogenesis of autoimmune diseases. Human Th17 cells are generated from Th0 cells by IL‐6 and IL‐1β and maintained by IL‐23. In this study, the role of IL‐17 in PBC and its association with biliary innate immunity were examined. Using cultured human biliary epithelial cells (BECs), the expression of Th17‐related cytokines and chemokines and changes therein on treatment with pathogen‐associated molecular patterns (PAMPs) and IL‐17 were examined. Immunohistochemistry for IL‐17 and Th17‐related cytokines was performed using tissue samples of human liver. Consequently, the expression of IL‐6, IL‐1β, IL‐23p19 and IL‐23/IL‐12p40 mRNAs, and their up‐regulation by PAMPs, were found in BECs. Moreover, BECs possessed IL‐17‐receptors and stimulation with IL‐17 induced production of IL‐6, IL‐1β, IL‐23p19 and chemokines. Several IL‐17‐positive cells had infiltrated damaged bile ducts and the expression of IL‐6 and IL‐1β was enhanced in the bile ducts of PBC patients. In conclusion, IL‐17‐positive cells are associated with the chronic inflammation of bile ducts in PBC which is associated causally with the biliary innate immune responses to PAMPs.

Incidence of and risk factors for hepatocellular carcinoma in primary biliary cirrhosis: National data from Japan

Primary biliary cirrhosis (PBC) primarily affects females and is rarely complicated by hepatocellular carcinoma (HCC). Although the HCC incidence in PBC patients is low, several characteristics and risk factors associated with its development have been reported. In this study, national data concerning the current status of carcinogenesis in PBC patients in Japan are reviewed. Using data from two national questionnaire surveys, we investigated the clinicopathological findings associated with HCC in PBC patients. According to the data of all reviewed PBC patients, the HCC incidence was 2.4% (71/2946). The HCC incidence by gender was 5.1% (19/370) in males and 2.0% (52/2576) in females, and the proportion of males was 26.7%. Prognosis was significantly poorer in the PBC patients with HCC than in those without. Multivariate analysis of risk factors associated with HCC by gender revealed histological stage at the time of PBC diagnosis as an independent risk factor associated with the development of HCC in females, but not in males. Furthermore, data from another national survey of 178 PBC patients with HCC (male/female = 49/129; proportion of males 27.5%) revealed that the duration between the diagnosis of PBC and that of HCC was significantly shorter in males than in females. In addition, histological stage at the time of HCC diagnosis was an independent risk factor for HCC in females, whereas no risk factors were identified in males. Conclusion: these data indicate that males are at risk of developing HCC at any histological stage of PBC. Therefore, male PBC patients in particular should be carefully screened for HCC from the early stages of PBC. (HEPATOLOGY 2013)

Epithelial-mesenchymal transition induced by biliary innate immunity contributes to the sclerosing cholangiopathy of biliary atresia

Infections of Reoviridae consisting of a double‐stranded RNA (dsRNA) genome and the biliary innate immune response to dsRNA are implicated in the aetiopathogenesis of biliary atresia (BA). Epithelial–mesenchymal transition (EMT) has recently been proposed as a mechanism behind the sclerosing cholangitis in BA. We hypothesized that the innate immune response to dsRNA in biliary epithelial cells plays an important role in peribiliary fibrosis via biliary EMT. Experiments using cultured human biliary epithelial cells revealed that stimulation with poly(I : C) (a synthetic analogue of viral dsRNA) increased the expression of basic fibroblast growth factor (bFGF, an EMT‐inducer), S100A4 (a mesenchymal marker) and Snail (a transcriptional factor), and decreased that of epithelial markers (biliary‐type cytokeratin 19 and E‐cadherin) and Bambi (TGF‐β1 pseudoreceptor). The expression of TGF‐β1 (EMT‐inducer) and vimentin (a mesenchymal marker) was not affected by poly(I : C). Both EMT‐inducers, bFGF and TGF‐β1, evoked a decrease and increase in the expression of the epithelial markers and of vimentin respectively, and the expression of Bambi was down‐regulated on stimulation with bFGF. Combined treatment with bFGF and TGF‐β1 quickly and completely induced a transformation of morphology as well as change from epithelial to mesenchymal features in cultured biliary epithelial cells. Immunohistochemistry revealed that biliary epithelial cells lining extrahepatic bile ducts and peribiliary glands in BA frequently show a lack of epithelial markers and an aberrant expression of vimentin. Moreover, the biliary epithelium showing sclerosing cholangitis expressed bFGF accompanied by bFGF‐positive mononuclear cells. In conclusion, the EMT may contribute to the histogenesis of sclerosing cholangiopathy, and the biliary innate immune response to dsRNA viruses induces biliary epithelial cells to undergo EMT via the production of bFGF and the increased susceptibility to TGF‐β1 caused by the down‐regulation of Bambi expression. Copyright

Evaluation of a new histologic staging and grading system for primary biliary cirrhosis in comparison with classical systems

Recently, our research team proposed a new histologic staging and grading system for primary biliary cirrhosis (PBC) that takes into account necroinflammatory activity and histologic heterogeneity. The present study aimed to confirm the usefulness of the new evaluation system. A total of 152 liver biopsy specimens and clinical data (including outcomes in patients with PBC before treatment with ursodeoxycholic acid) were analyzed with respect to the new system. Staging was evaluated on the basis of 3 histologic components (fibrosis, bile duct loss, and deposition of orcein-positive granules), and grading was assessed on the basis of chronic cholangitis activity and hepatitis activity. Concurrently, the classical systems, that is, the Scheuer and Ludwig staging systems, were also assessed and compared with our new system. PBC cases showed different distributions in each stage of the 3 systems. The new staging and grading system reflected liver dysfunctions before specific treatment. This was on a par with the results obtained using the classical systems. Development of cirrhosis-related conditions correlated well with the new staging system compared with the 2 classical staging systems, and in particular, the amount of deposition of orcein-positive granules could reflect development of cirrhosis-related conditions (scores 0-1 versus scores 2-3 groups, P < .0001). In conclusion, the new PBC staging system was demonstrated to reflect clinicolaboratory features, and its progression was associated with the development of cirrhosis-related conditions.

Biliary Innate Immunity in the Pathogenesis of Biliary Diseases

An innate immune response to bacterial and viral components is thought to be involved in the pathogenesis of cholangiopathies in case of primary biliary cirrhosis (PBC) and biliary atresia. Biliary epithelial cells possess the Toll-like receptor (TLR) family which recognizes pathogen-associated molecular patterns (PAMPs) and plays a pivotal role in the innate immune response. In PBC, disordered regulations of TLRs and a negative regulator of intracellular signaling, peroxisome proliferator-activated receptor-gamma (PPARgamma), with Th1-predominant cytokine milieu are involved in the pathogenesis of cholangitis such as chronic non-suppurative destructive cholangitis (CNSDC). Moreover, CD4-positive Th17 cells characterized by the secretion of IL-17, are implicated in the chronic inflammation of bile ducts in PBC and the induction of Th17 cells around bile ducts is causally associated with the biliary innate immune responses to PAMPs. In biliary atresia characterized by a progressive, inflammatory, and sclerosing cholangiopathy, dsRNA viruses could directly induce the expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and apoptosis in biliary epithelial cells as a result of the biliary innate immune response via dsRNA-recognizing receptors such as TLR3 and retinoic acid inducible gene I (RIG-I). Moreover, as a mechanism behind the sclerosing cholangiopathy in biliary atresia, epithelial-mesenchymal transition (EMT) has been proposed and the biliary innate immune response to dsRNA viruses is demonstrated to induce biliary epithelial cells to undergo EMT. Biliary innate immunity is associated with the pathogenesis of various cholangiopathies in biliary diseases as well as biliary defense systems.

Monocyte chemoattractant protein-1 derived from biliary innate immunity contributes to hepatic fibrogenesis

Aims Monocyte chemoattractant protein-1 (MCP-1) is a major chemotactic factor for hepatic stellate cells (HSCs) associated with hepatic fibrosis. In this study, among several fibrogenetic factors derived from biliary epithelial cells (BECs), MCP-1 produced by the biliary innate immune system was found to be most critical in the histogenesis of hepatic fibrogenesis. Methods Using cultured human BECs, the expression of five fibrogenetic factors including MCP-1 on stimulation with Toll-like receptor ligands, inflammatory cytokines or bile acids was examined. Moreover, in situ detection of MCP-1 and α-smooth muscle actin proteins was performed using sections from normal and diseased livers by immunohistochemistry. Results All fibrogenetic factors were detected in BECs, but only MCP-1 expression was upregulated, by all the Toll-like receptor ligands, IL-1β, and tumour necrosis factor-alpha. Proliferating bile ductules in interface areas expressed MCP-1 in diseased livers accompanying α-smooth muscle actin-positive activated HSCs. Conclusions Bile ductules proliferate in various hepatobiliary diseases, and its significance is still unknown. This study demonstrated that BECs in bile ductules could produce MCP-1, particularly, via biliary innate immunity, suggesting that MCP-1 derived from BECs plays an important role in the recruitment of HSCs to interface areas and the activation of HSCs resulting in the progression of periportal fibrosis.

Systemic and local expression levels of TNF‐like ligand 1A and its decoy receptor 3 are increased in primary biliary cirrhosis

Through a genome‐wide association study of a Japanese population, we recently identified TNFSF15, a gene encoding TNF‐like ligand 1A (TL1A), as a susceptibility gene for primary biliary cirrhosis (PBC). We investigated the clinical significance of TL1A and one of its receptors, decoy receptor 3 (DcR3), in PBC.

Significance of periductal Langerhans cells and biliary epithelial cell-derived macrophage inflammatory protein-3α in the pathogenesis of primary biliary cirrhosis.

Background/aims: To clarify the primary biliary cirrhosis (PBC)‐specific antigen‐presenting mechanism, we examined the distribution and phenotypic characteristics of infiltrating dendritic cells (DCs) with respect to bile ducts and the mechanism of migration in terms of the periductal cytokine milieu and biliary innate immunity.

Application and validation of a new histologic staging and grading system for primary biliary cirrhosis.

Background: We proposed a new grading and staging system for primary biliary cirrhosis (PBC), which takes into account the degree of both chronic cholangitis activity (CA) and hepatitic activity (HA) for grading disease activity and that of fibrosis, bile duct loss, and chronic cholestasis for staging. In this study, we validated our new system. Methods: Using liver biopsy specimens from 166 cases of PBC, chronic cholangitis with mild periductal lymphoplasmacytic infiltration, including chronic nonsuppurative destructive cholangitis, and the combined activity of interface hepatitis and lobular hepatitis were categorized into 4 grades on the basis of their degree and distribution (CA0-3 and HA0-3, respectively). For staging, because orcein staining was not available in this study, 2 criteria (fibrosis and bile duct loss) were independently scored from 0 to 3 on the basis of their degrees, and a final stage score was created from the sum. Results: Although there was a relatively uniform distribution of CA0/1/2/3 cases, the cases of HA0/1/2/3 were distributed as 21%, 64%, 13%, and 3%, respectively, with a prominent number of cases categorized as having none or mild HA. The distribution of stages 1 to 4 using our system was considerably different from that using the classic system and, importantly, showed a correlation with patient outcome. Conclusions: Our system revealed that the activities of chronic cholangitis and hepatitis did not correlate with each other in terms of degree and that our staging system properly reflected the outcome of PBC patients. The present study could validate the effectiveness of this new system for characterizing the pathologic condition of PBC.

Prevalence and risk factors of hepatocellular carcinoma in Japanese patients with primary biliary cirrhosis

Primary biliary cirrhosis (PBC) tends to affect females more than males. PBC selectively damages intrahepatic small bile ducts, particularly interlobular bile ducts. The clinical presentation of PBC has changed according to recent advances in clinicobiological diagnosis and improvements in therapeutic effects and prognosis. In particular, we encounter PBC patients with hepatocellular carcinoma (HCC), and the number of these patients appears to have increased. The precise reason for the increased number of PBC patients with HCC in recent decades remains unknown, but recognizing the current status of carcinogenesis in PBC patients, identifying the associated clinicopathological risk factors and understanding how the pathogenesis of PBC is directly associated with HCC, is important. In this review, we summarize the data from two nationwide surveys undertaken in Japan as well as recent data from Japanese and international studies.