Contributions of hepatocytes and bile ductular cells in ductular reactions and remodeling of the biliary system after chronic liver injury.

Mature hepatocytes are suggested to possess a capacity for bile ductular transdifferentiation, but whether and how hepatocytes contribute to ductular reaction in chronic liver diseases has not been elucidated. We examined whether mouse hepatocytes can transdifferentiate into bile ductular cells in vitro, using a three-dimensional collagen gel culture method, and in vivo, using a liver repopulation model in which ß-galactosidase-positive hepatocytes from Alb-Cre × ROSA26R mice were transplanted into the liver of wild-type mice. We further examined the relative contribution of intrinsic hepatocytes in ductular reaction in a hepatocyte lineage-tracing model using Mx1-Cre × ROSA26R mice treated with polyinosinic-polycytidylic acid. Within collagen gels, hepatocytes exhibited branching morphogenesis associated with the emergence of bile duct-like phenotype. In the liver repopulation model, many ß-galactosidase-positive, hepatocyte-derived bile ductular structures were identified; these markedly increased after liver injury. In Mx1-Cre × ROSA26R mice, relatively minor but significant contributions of hepatocyte-derived bile ductules were observed in both periportal and centrilobular ductular reaction. As the centrilobular ductular reaction progressed, the portal ducts or ductules migrated toward the injured area and joined with hepatocyte-derived ductules, leaving the portal tract without biliary structures. We conclude that hepatocytes and bile ducts or ductules are important sources of ductular reaction and that the intrahepatic biliary system undergoes remarkable remodeling in response to chronic liver injury.

Tumor necrosis factor-α promotes bile ductular transdifferentiation of mature rat hepatocytes in vitro.

We previously showed that mature hepatocytes could transdifferentiate into bile ductular cells when placed in a collagen-rich microenvironment. To explore the mechanism of transdifferentiation, we examined whether inflammatory cytokines affected the phenotype of hepatocytes in a three-dimensional culture system. Spheroidal aggregates of rat hepatocytes were embedded within a type I collagen gel matrix and cultured in the presence of various cytokines. In the control, hepatocytes gradually lost expression of albumin, tyrosine aminotransferase, and hepatocyte nuclear factor (HNF)-4α, while aberrantly expressed bile ductular markers, including cytokeratin 19 (CK 19) and spermatogenic immunoglobulin superfamily (SgIGSF). Among the cytokines examined, tumor necrosis factor (TNF)-α inhibited expression of albumin and HNF-4α, both at mRNA and protein levels. After culturing for 2 weeks with TNF-α, hepatocytic spheroids were transformed into extensively branching tubular structures composed of CK 19- and SgIGSF-positive small cuboidal cells. These cells responded to secretin with an increase in secretion and expressed functional bile duct markers. TNF-α also induced the phosphorylation of Jun N-terminal kinase (JNK) and c-Jun, and the morphogenesis was inhibited by SP600125, a specific JNK inhibitor. Furthermore, in chronic rat liver injury induced by CCl(4) , ductular reaction in the centrilobular area demonstrated strong nuclear staining of phosphorylated c-Jun. Our results demonstrate that TNF-α promotes the ductular transdifferentiation of hepatocytes and suggest a role of TNF-α in the pathogenesis of ductular reaction.

Recovery of mature hepatocytic phenotype following bile ductular transdifferentiation of rat hepatocytes in vitro.

We previously demonstrated that mature rat hepatocytes transdifferentiate to bile ductular cells when cultured in a three-dimensional collagen-rich matrix. Here, we show that the phenotype of transdifferentiated hepatocytes can be reversed by modulating culture conditions. Spheroidal aggregates of hepatocytes were cultured within a collagen gel matrix in the presence of serum and tumor necrosis factor-α. Spheroids transformed into ductular structures composed of small cuboidal cells, lost the expression of hepatocytic markers, whereas aberrantly expressed bile ductular markers. The transdifferentiated cells were then retrieved from the gels, plated on surfaces coated with a basement membrane-like material, and cultured in serum-free media. Cells spontaneously formed spheroidal aggregates and recovered hepatocytic phenotype. Dexamethasone (Dex), which suppressed the phosphorylation of ERK and Jun N-terminal kinase, facilitated the recovery, and the combination with interleukin-6 or oncostatin M resulted in the recovery of hepatocyte nuclear factor 4 α protein expression and the typical hepatocytic morphology, and a decrease in the expression of bile ductular markers. A cDNA microarray analysis revealed that the hepatocyte-specific mRNA expression profile was recovered in these cells. Our results demonstrate that hepatocytes are able to recover their phenotypes following bile ductular transdifferentiation, suggesting that hepatocytic and bile ductular phenotypes may be mutually reversible.

IgA vasculitis with transient glomerular hematuria, diarrhea, and pericarditis following COVID-19 mRNA vaccination in a young patient with possible pre-existing ulcerative colitis.

Exacerbations or de novo autoimmune/autoinflammatory disease have been reported after COVID-19 vaccination. A young male presented with cutaneous IgA vasculitis with glomerular hematuria, diarrhea and pericarditis following his second COVID-19 mRNA vaccination. He also showed positivity for proteinase 3 anti-neutrophil cytoplasmic antibody (PR3-ANCA) and anti-cardiolipin antibody. Skin biopsy was compatible to IgA vasculitis. His purpura subsided and hematuria spontaneously disappeared. Treatment with anti-inflammatory medications and prednisolone resolved the pericarditis. He had a history of persistent diarrhea, and colonic biopsies showed possible ulcerative colitis without vasculitis. Kidney biopsy after prednisolone therapy revealed minor glomerular abnormalities without any immune reactants and did not show vasculitis. After prednisolone treatment, PR3-ANCA decreased in a medium degree despite of improvement of symptoms and inflammatory data, suggesting that his PR3-ANCA may be associated with ulcerative colitis. The cause of the transient glomerular hematuria was unclear, however, it might be caused by focal glomerular active lesions (glomerular vasculitis) due to vaccine-induced IgA vasculitis with nephritis. This case highlights that COVID-19 mRNA vaccination can activate multiple autoimmune/autoinflammatory systems. The conditions might help us better understand the mutual mechanisms of the relevant disorders.