Study On Mechanism Of Portulaca Oleracea L. Polysaccharides Ameliorating HepG2 Cell Insulin Resistance

Insulin resistance(IR)is a phenomenon which depicts a feature of the impairment of the utilization to insulin,resulting the attenuated glucose uptake in tissues scuh as fat,liver,and muscle,etc.IR has a close relationship with Type 2diabete mellitus(T2DM).With a result that ameliorating IR contributes to the prevention of T2 DM.Polysaccharide is one of the main functional ingredients of Portulaca oleracea L.a plant that can be used as medicine and food.Researches on anti-diabetic effects of POP mainly focus on animals with diabetic to study hyperglycemic effect,fat metabolism and antioxidation function,so far.However,the experiment on molecular mechanism of POP on anti-diabetic effects in vitro has not been reported.The liver is the main place of glucolipid metabolism and IR of liver tissue plays an important role in the pathogenesis of T2 DM.Therefore,the ameliorative effects of POP on IR and its molecular mechanism at cellular level were studied in the present study.This study will provide a theoretical reference for further development of Portulaca oleracea L.and its active ingredients as a potential therapeutic agent for IR.Firstly,in the present study,we established the model of HepG2 cells insulin resistance using insulin,glucose,palmitic acid and palmitic acid plus 30 mM glucose as inducers.Then,we choose MTT colorimetric,Neutral red uptake,and Lactate dehydrogenase to optimize the inducing condition of cell relative growth rate based on time and dose effects.The experiment results indicated that the optimum inducing conditions of the four inducers were 0.1 μM insulin,30 mM glucose,0.5 mM palmitic acid,and 0.5 mM palmitic acid plus 30 mM glucose,respectively.In the meantime,the optimum time of process was also been demonstrate to be 24 h.Conbining cell glucose absorption and the intracellular triglyceride content,insulin resistance effects of those inducers are arranged in order of difficulty: high insulin group > high sugar group > high sugar and high fat group > high-fat group.Secondly,We focus on the improvement of POP on insulin resistance ofHepG2 cell: It can be obtained that 0.1 mg/ml POP has the best effect on insulin resistance from these experiments using different concentrations of POP to separate the HepG2 cell insulin resistance model induced by high fat after 24 h,combined with cell relative growth rate,cell glucose absorption and the intracellular triglyceride content.Thirdly,we discussed the possible molecular mechanism of positive influence in POP on insulin resistance in HepG2 cells: the Western blot and RT-PCR were used to evaluate the impacts of POP on relevant protein and gene expression of insulin resistance.When POP processes HepG2 cell,the protein expression of IR,IRS-2,PI3 K,Akt as well as the gene expression of IR and GLUT-1were obvious raised.Conversely,the protein expression of GSK-3β、GLUT-2 were decreased.In conclusion,POP can reduce the content of triglyceride in liver cells and promote glucose uptake,contributing to the ameliorating of IR in liver cells,the mechanism may be related to the regulation of PI-3K/Akt/GSK-3β and GLUTs insulin signal transduction pathway.