Orphan nuclear receptor 4 A1 involvement in transforming growth factor beta1-induced myocardial fibrosis in diabetic mice.

We explored the involvement of orphan nuclear receptor 4 A1 (NR4A1) in myocardial fibrosis mediated by transforming growth factor-beta1 (TGF-ß1) and its response to cytosporone B (Csn-B). We developed a diabetic cardiomyopathy mouse model by administering a high-fat diet in conjunction with a low-dose streptozotocin injection. Our analysis involved monitoring alterations in blood glucose and lipid levels, cardiac function and structure, as well as profibrotic factors such as α smooth muscle actin (α-SMA), collagen I, collagen III, TGF-ß1, connective tissue growth factor, and fibronectin. These assessments were conducted using biochemical techniques, Doppler ultrasound, histopathology, and real-time quantitative polymerase chain reaction. Cardiac fibroblasts (CFs) were extracted from suckling mice and cultivated in a high-glucose medium to simulate diabetes-induced myocardial fibrosis in vitro. These CFs were then subjected to coculture experiments with TGF-ß1 or Csn-B. The proliferation and migration of CFs were assessed using cell counting kit 8 (CCK-8) assays and Transwell assays, respectively. Western blotting and immunofluorescence assays were employed to evaluate the expression levels of NR4A1, p-NR4A1, and α-SMA in CFs treated with TGF-ß1 after NR4A1 knockdown or Csn-B administration, respectively. In diabetic heart tissue, the expression of p-NR4A1 was notably elevated. Furthermore, CFs exhibited enhanced proliferative capabilities and increased p-NR4A1 expression following high glucose exposure. Interestingly, NR4A1 knockdown resulted in a significant increase in the expression of fibrosis-related proteins in CFs following treatment with TGF-ß1. Moreover, our observations revealed a marked decrease in p-NR4A1 levels and a reduction in the expression of fibrosis-related proteins after Csn-B treatment. In diabetic mice treated with Csn-B, we noted diminished NR4A1 phosphorylation and a mitigation of myocardial fibrosis. We concluded that in the mouse model, Csn-B played a pivotal role in inhibiting diabetes-induced myocardial fibrosis by activating NR4A1.

In vitro proliferation and differentiation of hepatic oval cells and their potential capacity for intrahepatic transplantation

Hepatic oval cells (HOCs) are recognized as facultative liver progenitor cells that play a role in liver regeneration after acute liver injury. Here, we investigated the in vitro proliferation and differentiation characteristics of HOCs in order to explore their potential capacity for intrahepatic transplantation. Clusters or scattered HOCs were detected in the portal area and interlobular bile duct in the liver of rats subjected to the modified 2-acetylaminofluorene and partial hepatectomy method. Isolated HOCs were positive for c-kit and CD90 staining (99.8% and 88.8%, respectively), and negative for CD34 staining (3.6%) as shown by immunostaining and flow cytometric analysis. In addition, HOCs could be differentiated into hepatocytes and bile duct epithelial cells after leukemia inhibitory factor deprivation. A two-cuff technique was used for orthotopic liver transplantation, and HOCs were subsequently transplanted into recipients. Biochemical indicators of liver function were assessed 4 weeks after transplantation. HOC transplantation significantly prolonged the median survival time and improved the liver function of rats receiving HOCs compared to controls (P=0.003, Student t-test). Administration of HOCs to rats also receiving liver transplantation significantly reduced acute allograft rejection compared to control liver transplant rats 3 weeks following transplantation (rejection activity index score: control=6.3±0.9; HOC=3.5±1.5; P=0.005). These results indicate that HOCs may be useful in therapeutic liver regeneration after orthotopic liver transplantation.