Project description:We performed RNA-seq of livers obtained from mice fed either a control or the NASH-inducing FPC diet (Wang et al., 2016) for 24wk. In some FPC-fed mice the NASH-activated transcription factors, Elf3 or Glis2, had been deleted specifically in hepatocytes, when mice reached a steatotic state.
Project description:Comparison between livers of FLS mice and livers of DS (DD shionogi) mice We used FLS mice as model animals of human NASH, while DS mice as control animals. FLS mice develops NASH spontaneously. DS mouse strain is a sister strain of the FLS mouse strain. We compared RNA from pooled livers of three FLS mice and three DS mice at 19 weeks. NASH in livers from FLS mice was confirmied pathologically while simple steatosis of DS mouse livers confirmed.
Project description:Non-alcoholic steatohepatitis (NASH) is a life-threatening liver disease for which no drug has been approved. We have previously shown that human-derived hepatic in vitro models can be used to mimic key cellular mechanisms involved in the progression of NASH. In the present study, we first assess the predictive capacity of different in vitro models and then investigate how the reduction of NASH-specific parameters upon treatment with elafibranor, a PPAR-α/δ agonist, correlates with clinical NASH-resolution obtained through bariatric surgery. Whole genome transcriptomics analyses revealed that in vitro NASH models based on primary human hepatocytes (PHH), HepaRG and human skin stem cell-derived hepatic progenitors (hSKP-HPC) exhibit up to 35% overlap with publicly available datasets of liver biopsies of 4 cohorts of NASH patients. Exposure of the in vitro NASH models to elafibranor partially reverses these human-specific transcriptional NASH signatures, with the hSKP-HPC-derived NASH model showing the most sensitive response. NASH-specific transcriptomic changes observed in patients that underwent bariatric surgery correlated with the changes observed in the in vitro NASH models exposed to the PPAR-α/δ agonist. PPARGC1A, PPARA and SIRT1 are shared upstream regulators in the PHH-, HepaRG- and hSKP-HPC NASH models exposed to elafibranor. Activation of these upstream regulators increases the expression of ANGPTL4, PDK4 and PLIN2, while this does not occur in patients that underwent bariatric surgery, suggesting an adverse effect on lipid metabolism. In conclusion, pathologic and therapeutic (anti-)NASH-specific transcriptional responses can be mimicked in PHH, HepaRG and hSKP-HPC, while the latter most sensitively responds to drug testing. PPAR-α/δ agonism adversely modulates pro-steatogenic genes which deserves attention in further studies.
Project description:BACKGROUND & AIMS: Recent studies revealed that hemoglobin is expressed in some non-erythrocytes and it suppresses oxidative stress when overexpressed. Oxidative stress plays a critical role in the pathogenesis of non-alcoholic steatohepatitis (NASH). This study was to investigate whether hemoglobin is expressed in hepatocytes and how it is related to oxidative stress in NASH patients. METHODS: Microarray was performed to identify differentially expressed genes in NASH. Quantitative real time PCR (qRT-PCR) was used to examine gene expression levels. Western blotting and immunofluorescence staining were employed to examine hemoglobin proteins. Flow cytometry was used to measure intracellular oxidative stress. RESULTS: Analysis of microarray gene expression data has revealed a significant increase in the expression of hemoglobin alpha (HBA1) and beta (HBB) in liver biopspies from NASH patients. Increased hemoglobin expression in NASH was validated by qRT-PCR. However, the expression of erythrocyte specific marker genes such as SPTA, SPTB, GYPA, GATA1, and ALAS2 did not change, indicating that increased hemoglobin expression in NASH was not from erythropoiesis, but could result from increased expression in hepatocytes. Immunofluorescence staining demonstrated positive HBA1 and HBB expression in the hepatocytes of NASH livers. Hemoglobin expression was also observed in human hepatocellular carcinoma HepG2 cell line. Furthermore, treatment with hydrogen peroxide, a known oxidative stress inducer, induced a dose dependent increase in HBA1 expression in HepG2 cells. Intriguingly, forced hemoglobin expression suppressed oxidative stress. CONCLUSIONS: Oxidative stress upregulates hemoglobin expression in hepatocytes. Suppression of oxidative stress by hemoglobin could be a mechanism to protect hepatocytes from oxidative damage. These findings suggest that hemoglobin is an inducible antioxidant in hepatocytes in response to increased oxidative stress as found in NASH livers. Twelve biopsy diagnosed NASH patients were included in this study. For control groups, total RNA from 5 different subjects were purchased from ADMET. These subjects are free from liver disease.
Project description:Transcriptional profiling by bulk RNA sequencing of murine livers or sorted immune cells, from DEN ALIOS NASH-HCC mouse model. Mice treated with CXCR2i (AZD5069) monotherapy, anti-PD1 monotherapy, or combination CXCR2i, anti-PD1 treatment. Published in doi: 10.1136/gutjnl-2021-326259
Project description:Comparison between livers of FLS mice and livers of DS (DD shionogi) mice We used FLS mice as model animals of human NASH, while DS mice as control animals. FLS mice develops NASH spontaneously. DS mouse strain is a sister strain of the FLS mouse strain.
Project description:To investigate the mechanism of hepatic Activin A and Gpnmb in NAFLD/NASH, we studied C57BL/6J mice on a FPC NASH diet and sugar water for 16 weeks, compared with standard chow diet, and used adeno-associated viral vectors with a liver-specific thyroxine binding globulin (TBG) promoter to express Activin A or GFP (control), or AAV8-H1-shRNA to knockdown of Gpnmb or scramble control in NAFLD/NASH liver. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different livers in each group.
