| [Background] Diseases in lipid metabolism disorders,especially hypercholesterolemia,are key factors to cause cardiovascular disease such as atherosclerosis,obesity,fatty liver and so on.Oleanolic acid(OA)is a pentacyclic triterpenoid compound,which is widely distributed in olive plants and Chinese herbal medicines in the form of free fatty acids or saponins.Studies were shown that OA had many benefits,including liver protection,anti-inflammatory,glucose-lowering,anti-hypertension,anti-cancer,lipid-lowering,endothelial protection and anti-atherosclerosis.As OA derivative,difluoromethyl oleanolate,its specific biological properties have not been reported in the literature.PCSK9 was found in 2003 as a gene associated to blood cholesterol metabolism.The main physiological role of PCSK9 is to promote degradation of the low density lipoprotein receptor on the surface of liver cells,thereby increasing the level of LDL-C,resulting in reduced clearance of blood lipids,hyperlipidemia and the occurrence of atherosclerosis.Because of the important role of PCSK9 in the regulation of liver lipid metabolism,it has become an emerging target for lipid-lowering drug.[Objective] Our studies focus on the effect of difluoromethyl oleanolate in lipid metabolism in Hep G2 cells and its preliminary mechanism to reduce PCSK9 expression,receiving more experimental evident for the study of new lipid-lowering drugs targeting PCSK9.[Methods] Western blot detects PCSK9 、LDLR、SREBP2、HNF1αprotein expression after treating with difluoromethyl oleanolate in Hep G2 cells.Using cell immunofluorescence to observe the LDLR expression on the cell surface.Di I-LDL and oil red O staining was utilized to observe the effect of difluoromethyl oleanolate on lipid uptake and accumulation in HepG2 cells.[Results] With increasing concentration and time of difluoromethyl oleanolate,PCSK9 and HNF1α levels were decreased,LDLR and SREBP2 were increased;LDLR expression was increased after treating difluoromethyl oleanolate on the cell surface by using cell immunofluorescence in HepG2cells;After administration of difluoromethyl oleanolate treatment,lipid uptake was increased,and lipid accumulation was decreased,by using DiI-LDL and oil red O staining respectively.[Conclusion] Difluoromethyl oleanolate can decrease PCSK9 expression and increase LDLR levels,lowering the accumulation of intracellular lipid droplets and increasing the uptake of lipid in Hep G2 cells.The pathogenesis involves increased LDLR levels via upregulating SREBP2 expression,additionally,decreased PCSK9 expression via lowering HNF1α levels,and elevated LDLR levels ultimately. |
