Protective Effects Of Polysaccharide From Lentinus Edodes Mycelium On Islet ? Cells Damage Induced By Glucose Toxicity

Diabetes mellitus is one of the common metabolic diseases with a high mortality rate and an increasing incidence year by year.It has become the third most serious disease that endangers human health after cardiovascular and cerebrovascular diseases and cancer.About 90% of them are type 2 diabetes mellitus(type 2 diabetes mellitus,T2DM).Studies have shown that glucose toxicity caused by significant and sustained hyperglycemia is one of the important causes of aggravation of islet ? cells dysfunction.Glucose toxicity induced oxidative stress under pathological conditions of diabetes is the direct cause of islet ? cells apoptosis and dysfunction.Therefore,in the prevention and treatment of diabetes mellitus,how to effectively inhibit islet ? cells oxidative stress damage has become a key target.Natural products have many advantages such as wide source and small side effects.The use of natural products to protect islet ? cells from oxidative stress and explore its possible protective pathways and mechanisms have become new trends and important research topics in biology,medicine and food science.In this study,we used the polysaccharide from Lentinus edodes mycelium prepared in the early stage of the laboratory to further investigate the protective effect on glucose toxicity induced islet ? cells damage and related mechanisms.It provided experimental basis for the application of polysaccharide from Lentinus edodes mycelium in the prevention and treatment of diabetes mellitus.In this study,mouse islet cells MIN6 and rat islet cells INS-1 were selected as the research objects;the safe concentration range of polysaccharide from Lentinus edodes mycelium was determined by MTT colorimetry,and the appropriate high glucose damage concentration was selected.Simultaneously,the effects of 0.0125,0.025,and 0.05 mM polysaccharide from Lentinus edodes mycelium on glucose toxicity induced cell viability were detected.The effects of polysaccharide from Lentinus edodes mycelium on the morphology of glucose toxicity injured cells were observed under microscope.The effects of lactate dehydrogenase(LDH)release on the damage of cell membrane induced by glucose toxicity were detected.The effects of polysaccharide from Lentinus edodes mycelium on glucose toxicity induced apoptosis were detected by Hoechst33258 staining test;The levels of reactive oxygen species(ROS),superoxide dismutase(SOD)and malondialdehyde(MDA)were detected to determine the effects of polysaccharide from Lentinus edodes mycelium on glucose toxicity induced cellular oxidative stress.Finally,the effects of polysaccharide from Lentinus edodes mycelium on the expression of apoptosis-related proteins,such as Bax,Bcl-2,caspase-3 and caspase-1 induced by high glucose were detected by western blot.The expression of key proteins in p38 MAPK,JNK and NF-?B signaling pathways were detected,and the translocation of nuclear Nrf2 and pancreatic duodenal homeobox factor-1(PDX-1)were detected.The results showed that 40 mM high glucose could reduce cell viability,damage cell morphology,increase LDH release in cell supernatant,and increase cell apoptosis.0.0125,0.025,and 0.05 mM polysaccharide from Lentinus edodes mycelium could effectively increase the cell viability in a concentration-dependent manner,which could improve cell morphology,reduce the release of LDH in cell supernatant,and reduce cell apoptosis.Under the action of glucose toxicity,intracellular ROS levels increased,SOD activity decreased,MDA content increased,and intracellular oxidative stress levels were enhanced.With the increasing concentration of polysaccharide from Lentinus edodes mycelium,the activity of SOD increased,and the levels of ROS and MDA decreased in the cells.It indicated that the polysaccharide from Lentinus edodes mycelium could effectively inhibit the oxidative stress induced by glucose toxicity.Western Blot assay showed that the expression of Bax and apoptosis-related proteins cleaved-caspase-3 and cleaved-caspase-1 were up-regulated by glucose toxicity,the expression of apoptosis-inhibiting protein Bcl-2 was down-regulated,p38 MAPK and JNK signaling pathways were activated and the expression of related phosphorylated were increased,enhanced nuclear transfer of NF-?B p65,activated NF-?B signaling pathway,and reduced the translocation of Nrf2 and PDX-1 proteins.The group of polysaccharide from Lentinus edodes mycelium could effectively inhibit the above trend and inhibit apoptosis by reducing the expression of Bax,cleaved-caspase-3,cleaved-caspase-1 and enhancing the expression of Bcl-2 protein.At the same time,it inhibited the activation of p38 MAPK and JNK signaling pathway,reduced the level of oxidative stress in cells,reduced the translocation of NF-?B p65,inhibited the activation of NF-?B signaling pathway,increased the translocation of Nrf2,enhanced the ability of cells to resist oxidative stress,and enhanced the translocation of PDX-1 to maintain transcription and regulation of islet cell properties and function-related genes.The above results demonstrated that the polysaccharide from Lentinus edodes mycelium could effectively inhibit the apoptosis and oxidative stress damage induced by glucose toxicity in MIN6 and INS-1 cells,providing theoretical and experimental basis for the development of Lentinus edodes resources and the study of Lentinus edodes polysaccharide function.