Hepatic stellate cells (HSC) and hydrogen sulfide (H2S) both play important roles in the development of hepatocellar carcinoma (HCC). Whereas, in the microenvironment of HCC, whether HSC participate in regulating the biological process of HCC cells by releasing H2S remains elusive. In vitro, Flow cytometry (FCM), CCK-8, RNA-sequencing, Western blotting, RT-qPCR, immunofluorescence and ChIP assays were carried out in the HCC cells to investigate the effect of H2S on biological functions and JNK/JunB-TNFSF14 signaling pathway. Specimens from HCC patients were analyzed by RT-qPCR and Western blotting assays for evaluating the expression of TNFSF14 and CSE. Statistical analysis was used to analyze the correlation between TNFSF14 expression and clinical data of HCC patients. Based on the FCM and CCK-8 results, we found the LX-2 cells were able to induce HCC cells apoptosis through releasing H2S. RNA-sequencing, RT-qPCR, and Western blotting results showed that TNFSF14 gene was upregulated in both LX-2 and NaHS group. NaHS treated in HCC cells led to JNK/JunB signaling pathway activating and greater binding of p-JunB to its responsive elements on TNFSF14 promoter. Impairment of TNFSF14 induction alleviated LX-2 and NaHS induced apoptosis of HepG2 and PLC/PRF/5 cells. Furthermore, TNFSF14 expression in HCC tissues was lower than the adjacent tissue. HCC patients with low expression of TNFSF14 had higher malignant degree and poor prognosis. In summary, demonstration of the involvement of HSC-derived H2S in JNK/JunB mediated expression of TNFSF14 gene strongly indicates H2S palys an important role in the regulation of HCC apoptosis.
Background LCSCs is a key factor in the occurrence and development of HCC. Hydrogen sulphide (H2S) is the third gas signalling molecule after carbon monoxide (CO) and nitrogen monoxide (NO). Exogenous H2S has been shown to inhibit the progression of HCC by increasing apoptosis and autophagy. However, whether H2S can affect LCSCs in HCC microenvironment still remains poorly understand. Methods In vitro, Flow cytometry (FCM), imaging quantitive flow cytometry, RNA-sequencing, and Tumor sphere-forming assay were carried out in the HCC cells and LCSCs to investigate the effect of H2S on biological functions and Wnt/β-catenin signaling pathway. Data was analyzed using unpaired Student's t-test and nonlinear regression. P < 0.05 was considered to indicate a statistically significant difference. Results Based on the FCM, we found that exogenous H2S were able to induce HCC cells apoptosis and promote stemness in HCC cells. RNA-sequencing, imaging quantitive flow cytometry showed that 23 genes was regulated in NaHS group, and NaHS can activate the Wnt/β-catenin signaling pathway in HCC cell lines. Treatment with the Wnt/β-catenin signaling inhibitor ICRT3 alleviated the NaHS-induced stemness increase in HepG2 and Hep3B cells. Furthermore, the tumor sphere-forming assay show that the number of sphere-forming cells was significantly increased in NaHS treated group and was inhibited significantly in ICRT3 treated group. Conclusion Exogenous H2S increased the expression of CD133, CD44 and β-catenin and promoted the stemness in LCSCs by activating the Wnt/β-catenin signalling, which strongly clarify the relationship between Exogenous H2S and LCSC stemness and may provide theoretical guidance and potential therapeutic approaches for liver cancer.
This study sought to investigate risk factors for 6-week mortality of patients with decompensated liver cirrhosis associated esophagogastric variceal bleeding (EGVB) and clinical characteristics of myocardial injury in cirrhotic patients with EGVB. This retrospective cohort study included 249 patients with decompensated liver cirrhosis associated EGVB in the Department of Emergency. Patients were divided into two groups including liver cirrhosis associated EGVB without myocardial injury and liver cirrhosis associated EGVB with myocardial injury. Myocardial injury, recurrent bleeding, total bilirubin (TBIL) level and dyslipidemia are independent risk factors for 6-week mortality in liver cirrhosis associated EGVB. Among all patients with liver cirrhosis associated EGVB, 90 (36.2%) had myocardial injury and 159 individuals (63.8%) not. The 6-week mortality in the group with myocardial injury was 21%, which was significantly higher than that of 7% in the group without myocardial injury. More patients in the myocardial injury group smoked, had moderate to severe esophageal varices, liver failure, and Child–Pugh C liver function compared to the non-myocardial injury group. Myocardial injury, recurrent bleeding, TBIL level and dyslipidemia are independent risk factors for death within 6 weeks in liver cirrhosis associated EGVB. The 6-week mortality is considerably higher in patients with myocardial injury in liver cirrhosis associated EGVB than those without myocardial injury.
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