MST1过表达对棕榈酸诱导非酒精性脂肪肝细胞模型脂滴生成的影响

The Effect of MST1 Overexpression on Palmitic Acid-induced Lipid Droplets in the Cell Model of Non-alcoholic Fatty Liver Disease

目的通过构建哺乳动物STE20相关激酶(mammalian sterile 20-like kinase 1,MST1)过表达慢病毒,探讨MST1对棕榈酸孵育的HepG2细胞内脂滴生成的影响。方法构建MST1过表达载体,与慢病毒包装质粒共转染HEK293T细胞,获取慢病毒颗粒并检测其滴度。利用250μM棕榈酸诱导HepG2细胞,将MST1过表达慢病毒(LV-MST1)和对照慢病毒(LV-control)分别感染细胞。Western blot检测感染细胞中MST1的表达水平;利用油红O染色观察与检测细胞中脂滴的生成情况;利用细胞甘油三酯酶法检测细胞中甘油三酯的含量。结果成功获得MST1过表达慢病毒,其滴度达1×10~6TU/μL以上。Western blot检测显示MST1过表达的HepG2细胞中MST1的表达水平明显升高,脂滴生成和甘油三酯含量比对照组明显减少(P<0.05或<0.01)。结论成功构建MST1过表达慢病毒,MST1过表达明显抑制棕榈酸诱导的非酒精性脂肪肝细胞模型的脂滴生成。

Objective To investigate the effect of mammalian sterile 20-like kinase( MST1) on palmitic acid( PA)-induced lipid droplets synthesis in the HepG2 cells by constructing MST1 overexpressing lentivirus. Methods 393 T cells were cotransfected by lentivirus packaging vectors with constructed MST1 overexpressing lentivirus. Lentiviral particles were harvested,and lentivirus titer was measured. MST1 overexpressing lentivirus( LV-MST1) and control lentivirus( LV-control) were constructed to infect HepG2 cells induced by 250 μM palmitic acid. MST1 expression within infected the cells was detected by western blot. Lipid droplets generation of the cells was observed by Oil Red O staining. Intracellular triglycerides content was measured by triglyceride enzyme method. Results Titers of MST1 overexpressing lentivirus were up to 1 ×106TU / μL or more. Western blot showed that MST1 expression level was significantly increased in lentivirus-infected cells. Lipid droplets synthesis and triglycerides content in MST1 overexpression group were significantly lower than that in control group( P < 0. 05 or P < 0. 01). Conclusion We found by successfully constructing MST1 overexpressing lentivirus that MST1 obviously inhibited PA-induced lipid droplets synthesis in non-alcoholic fatty liver disease( NAFLD) model.