Knockout of ACSL3 increasing the sensitivity of HCC cells to glucose deficiency

Objective:To investigate the effect of long-chain acyl-CoA synthetase 3 (ACSL3) on the survival of Huh7 cells of hepatocellular carcinoma (HCC) under metabolic stress.Methods:The RNA expression profile and clinical data of HCC patients were downloaded from the TCGA-LICH database to analyze the expression of ACSL3 in HCC patients and draw Kaplan-Meier survival curve.From the liver cancer cohort of the First Affiliated Hospital of Guangxi Medical University, 44 primary liver cancer tissues and paracancerous tissues in HCC patients were collected, and the expression of ACSL3 was detected by immunohistochemistry.ACSL3 expression in Huh7 cells was silenced/knocked-out by siRNA and CRISPR/CAS9 technology; the effect of ACSL3 knockout on the proliferation and invasion of Huh7 cells were detected by cell counting kit (CCK-8) assay, clonal formation assay and Transwell invasion assay.Under full medium and glucose deficiency culture conditions, cell death in control group and ACSL3 silencing or knockout group was measured by flow cytometry; and level changes of ATP and β-hydroxybutyrate in cells were detected.Results:Compared with normal liver tissues, the expression of ACSL3 was higher in HCC tissues (P< 0.05); and patients with high ACSL3 exhibited shorter survival time(P< 0.05).ACSL3 knockout did not affect cell proliferation, clonogenicity and invasion of Huh7 cells(P> 0.05).Under the condition of glucose deficiency, ACSL3 silencing or knockout significantly decreased the survival rate of Huh7 cells (P< 0.05), decreased ATPproduction and inhibited β-hydroxybutyrate production (P< 0.05).Conclusion:Knockout of ACSL3 promotes the HCC cell death from glucose deficiency by inhibiting β-oxidation.