Effect Of Hyperosmotic Stress On The Proliferation And Apoptosis Of Hepatocellular Carcinoma Huh7Cells

BackgroundIt is critical to understand the underlying molecular mechanisms of the development of hepatocellular carcinoma (HCC) which is a malignant seriously threatening people’s life. Toni city-responsive enhancer binding protein (TonEBP/nuclear factor of activated T-cells5,NFAT5) is a Rel homologytranscription factor classically known for its osmosensitive role in regulating cellular homeostasis during states of hypo-and hypertonic stress.Apoptosis is characterized by cell shrinkage,nuclear condensation, DNA fragmentation and apoptoticbody formation. These features distinguish apoptosis from other types of cell death, such as necrosis.Whereas some signs of apoptosis, such as externalization of phosphatidylserine, altered mitochondrial function or activation of caspases are cell type-and death signal-dependent, apoptotic cell volume decrease (AVD) is an early and ubiquitous event. Hyperosmotic hepatocyte shrinkage could decrease cell volume and induce cell apoptosis. Since there is no documentation of carcinoma tissue osmolality,it is unknown if cancerous tumors possess an osmolality vastly different from that of the surrounding tissues or blood. Since there is no documentation of carcinoma tissue osmolality,it is unknown if cancerous tumors possess an osmolality vastly different from that of the surrounding tissues or blood.In conclusion, the aim of this research is to find the influence of osmotic stress in tumor microenvironment on hepatoma cell apoptosis and proliferation to provide a new molecular targeted therapy in HCC. Continuing research will provide a better understanding as to how these and other alternative TonEBP stimuli regulate gene expression in both health and disease.ObjectiveTo investigate the effect of hyperosmotic stress on the proliferation and apoptosis of hepatocellular carcinoma Huh7cells,and preliminarily explore the mechanism. Methods1. The expression of mammalian osmotic regulator Nuclear factor of activated T-cells5(NFAT5) was detected in64cases of hepatocellular carcinoma by immunohistochemistry staining method.2. The NFAT5mRNA level was analyzed by RT-PCR.3. NaCl was used as a chemical hyperosmotic-inducible reagent to mimic tumor hyperosmotic microenvironment. The expression of NFAT5protein was analyzed by Western blotting Successfully construct the NFAT2overexpressed lentiviral vector and package the lentivirus.4. Cell apoptosis was examined by flow cytometry.5. Cell growth was measured by Methyl-Thiazol-Tetrazolium (MTT) assay.Results1. NFAT5positive expression rate was low(9.38%) in hepatocellular carcinoma tissue but was high in the corresponding peritumoral tissue(68.75%)(P<0.01).2. NFAT5mRNA level was low in hepatocellular carcinoma tissue but was high in the corresponding peritumoral tissue(P<0.01).3. Under hyperosmotic stress(100mmol/LNaCl) for24h,the Huh7cell protein expression levels of NFAT5were up-regulated(p<0.01).4. The apoptosis rate in experimental group (26.0±3.2%) was significantly higher than that in blank control group (0.8±0.2%)(P<0.01).5. Hyperosmotic stress(100mmol/LNaCl) also inhibit growth of Huh7cells, cell growth inhibitory rate was31.35±2.09%(P<0.01).