The Water Extract Of Phyllanthus Emblica L. Fruit Promotes Glucose Uptake And Alleviates Insulin Resistance In C2C12 Myotubes

The fruit of Phyllanthus emblica L.,a natural traditional Chinese medicine,is rich in nutritional value and pharmacological activities.More and more evidences demonstrate that the extracts of Phyllanthus emblica L.fruit can significantly reduce the blood glucose level in diabetic mice.However,the material basis and underlying mechanism of the extracts of Phyllanthus emblica L.fruit still remain unclear.In the present study,based on promoting glucose utilization and preventing disorder of glucose metabolism,we sreened out the active extract of Phyllanthus emblica L.fruit that significantly promotes glucose consumption in myotubes,and analyzed its material basis.At the meantime,our study aims to systematically investigate and clarify the molecular mechanisms of the extract of Phyllanthus emblica L.fruit on promote glucose uptake and attenuate insulin resistance in C2C12 myotubes,which will provide scientific evidence for the utilization of Phyllanthus emblica L.Skeletal muscle is the major site for disposal glucose,and plays a well-established role in maintaining glucose homeostasis.Therefore,in this study,we firstly sreened out the water extract of Phyllanthus emblica L.fruit(WEPE)based on glucose consumption in C2C12 myotubes,and analyzed its composition and content by LC-MS/MS method.To further investigate the effect of WEPE on promoting glucose utilization,2-NBDG method was used to evaluate its effect on glucose uptake,and its molecular mechanism was investigated by Western Blotting,protein target inhibitor and siRNA.The results showed that WEPE significantly promoted glucose consumption,glucose uptake and GLUT4 translocation to the plasma membrane,and increased the phosphorylation of AMPK,ACC,AS160,p38 MAPK in C2C12 myotubes.The Compound C,AMPK inhibitor,significantly decreased the phosphorylation of AMPK,impaired AMPK pathway activation,blocked GLUT4 translocation and glucose uptake induced by WEPE.Furthermore,WEPE-activated AMPK phosphorylation and promotive glucose uptake were diminished by AMPK knockown.In addition,the STO-609,CaMKKβ inhibitor,also attenuated glucose uptake,impaired translocation of GLUT4 into plasma membranes,and weakened the phosphorylation of AMPK activated by WEPE,which suggest that WEPE activates AMPK pathway through CaMKKβ to promote glucose uptake in C2C12 myotubes.Activated insulin signaling pathway and glucose uptake were impaired when C2C12 myotubes exposed to 0.5 m M Palmitate for 24 h.Moreover,Palmitate increased the expression levels of ER stress markers.In palmitate-exposed C2C12 myotubes incubated with WEPE,the nagetive effects of palmitate were prevented.Reduced glucose uptake and Akt phosphorylation induced by Palmitate were prevented when treated WEPE in the presence or absence of insulin.These protective effects of WEPE were found to be associated with increased AMPK phosphorylation and attenuated palmitate-induced ER stress.The phosphorylation of AMPK activated ACC and AS160,which act downstream of AMPK,leading to increased free fatty acid metabolism and GLUT4 translocation.Furthermore,the ability of WEPE to promote insulin-stimulated Akt phosphorylation and glucose uptake was reversed in the presence of the AMPK inhibitor Compound C,but this reversed effect was non-effect on palmitate-induced ER stress.In conclusion,these findings indicate that WEPE attenuates insulin resistance via activating AMPK pathway and preventing ER stress in palmitate-exposed C2C12 myotubes.In summary,WEPE activates AMPK pathway through CaMKKβ to promote glucose uptake in C2C12 myotubes,and attenuates insulin resistance via activating AMPK and preventing ER stress in palmitate-exposed C2C12 myotubes,which provides the experimental data to develope and utilize at the hypoglycemic activity of Phyllanthus emblica L.