Protective Effects And Mechanism Investigation Of Salvianolic Acid A On Diabetic Peripheral Neuropathy

To study the protective effects of Salvianolic acid A on diabetic peripheral neuropathy and to investigate the underlying mechanism.Spontaneous type 2 diabetic KK-Ay mice were orally administered with Salvianolic acid A(0.3,1 mg/kg)for 8 weeks.The body weight,24 h food intake and water intake,blood glucose and blood lipids were observed.Thermal pain threshold,mechanical pain threshold and sciatic nerve conduction velocity were evaluated and scanning electron microscopy was used to detect the ultrastructural changes of sciatic nerve myelin.H&E staining and immunohistochemistiy staining were used to detect the tissue structure and changes of myelin protein zero(MPZ/PO)and nerve growth factor(NGF)protein expression in sciatic nerve.The rat Schwann cell(RSC96)was used to establish the diabetic periphery neuropathy screening model under high glucose stimulation and Salvianolic acid was used to incubate the high glucose-injured RSC96 cell to observe the changes of oxidative stress and mitochondrial function.The inflammatory factors mRNA expression was examined by RT-PCR.The reporter gene,RT-PCR combined with high content screening,Western blot and molecular docking were used to evaluate Nrf2 promoter activity,transcriptional level,nuclear translocation,effects on the Keapl/Nrf2 signaling pathway and predict the potential interaction of SalA and Nrf2 protein active sites.As a result,Sal A could effectively improve the abnormal glucose and lipid metabolism in KK-Ay mice,increase the mechanical pain threshold and sciatic nerve conduction velocity,reduce the ultrastructural changes and upregulate the expression of P0 and NGF protein in the injured sciatic nerve myelin.In vitro,SalA can significantly protect Schwann cells against high glucose injury,reduce reactive oxygen species(ROS)production,oxidized glutathione(GSSG)and malondialdehyde(MDA)content,and markedly downregulate the inflammatory factors mRNA expression such as Mcp1,Icam1,116,Tnfa and Cox2.Salvianolic acid A increased the mitochondrial number,promoted mitochondrial ATP production,restored mitochondrial membrane potential,reduced mitochondrial ROS production and thus improved the mitochondrial function.More importantly,Salvianolic acid A can significantly inhibit Nrf2 promoter activity,downregulate Nrf2 mRNA expression,but promote Nrf2 nuclear translocation.SalA directly bound with strong affinity to Nrf2 at multiple active sites.In summary,Salvianolic acid A can significantly improve the diabetic peripheral neuropathy and its underlying mechanism may be related to anti-oxidative stress,anti-inflammation and improvement of mitochondrial function mediated by Nrf2.Salvianolic acid A might be a promising clinical therapeutics for the treatment of diabetic peripheral neuropathy.