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Effect of Rab32 on lipid metabolism in mouse hepatic AML12 cells



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Effect of Rab32 on lipid metabolism in mouse hepatic AML12 cells


ZHOU Wei MA Li WAN Ying LI Fuxiang

Graduate School, Biomedical Analysis Center, Army Medical University (Third Military Medical University), Chongqing, 400038; Laboratory of Lipid & Glucose Metabolism, the First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016; Department of Intensive Care Medicine, General Hospital of Chengdu Military Command, Chengdu, Sichuan Province, 610083, China


nonalcoholic fatty liver disease AML12 cells Rab32 lipid metabolism

Q591.5; R322.47; R329.26

Objective    To determine the effect of regulation of Rab32 on the lipid metabolism in murine hepatic alpha mouse liver 12 (AML12) cells. Methods    Lentivirus expressing FCD-EYFP-Rab32 was transfected into AML12 cells for overexpression of Rab32, and CRISPR/Cas9 gene knockout technology was used to knockout Rab32 in the cells. Untreated AML12 cells were used as normal control. The protein level of Rab32 was determined by Western blotting. The intracellular lipid droplets were observed by confocal microscopy after Nile red fluorescent staining. The contents of intracellular triglyceride (TG) and total cholesterol (TC) were measured by corresponding quantitation kits. Cell apoptosis was assayed by flow cytometry. Results    Compared with the normal control cells, the protein level of Rab32 was significantly increased in the overexpressed cells but decreased in the knockout cells (P<0.01). The number (21.91±14.32 vs 33.89±17.31, P<0.01) and size (16.82±14.37 vs 22.27±14.10 μm2, P<0.01) of lipid droplets per cell were significantly decreased in the overexpressed cells when compared with the untreated cells, while the intracellular contents of TG (104.03±12.28 vs 130.94±4.21 nmol, P<0.01) and TC (46.92±1.26 vs 81.11±0.65 μg, P<0.01) were decreased. Whereas, the knockout cells had larger lipid droplet number (58.23±42.28, P<0.01) and total area (53.31±36.33 μm2, P<0.01), and higher intracellular TG (159.03±8.85 nmol, P<0.01) and TC (93.38±3.33 μg, P<0.01). No obvious difference was found in cell apoptosis among the 3 groups of cells (P>0.05). Conclusion    Rab32 promotes the lipid metabolism in the AML12 cells, and the molecule may be regarded as a new target in treatment of nonalcoholic fatty


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Last Update: 2018-06-13