Studies On The Clinical Trial And Mechanism Of Sancai Lianmei Partical In The Treatment Of Type 2 Diabetes Mellitus Combined With Non-Alcoholic Fatty Liver Disease

Background: Studies have shown that T2 DM is closely associated with the incidence of NAFLD,and patients with NAFLD have an increased risk of T2 DM.NAFLD has been recognized as an early lesion and independent risk factor for T2 DM.Sancai Lianmei Particle is derived from classic prescription and long-term clinical practice.Previous studies have confirmed that Sancai Lianmei Particle can improve insulin resistance(IR),control blood glucose and regulate blood lipid,alleviate inflammatory response and oxidative stress.However,its clinical efficacy and mechanism of action in T2 DM patients with NAFLD are still uncertain.Objective: To observe primarily the therapeutic effect of Sancai Lianmei Particle on T2 DM patients with NAFLD through clinical trial.To screen the key target proteins and key signaling pathways of Sancai Lianmei Particle by network pharmacological method.This study focused on the actions of Sancai Lianmei Particle on improving hepatic insulin resistance and alleviating hepatic oxidative stress.To elucidate the mechanism of actions of Sancai Lianmei Particle by regulating ROS-JNK signaling pathway to improve hepatic insulin resistance and reduce oxidative stress in vivo study.Methods:(1)A clinical trial was conducted in patients with T2 DM combined with NAFLD,Sancai Lianmei Particle was applied as test product,to clarify the therapeutic action and safety of Sancai Lianmei Particle on T2 DM patients with NAFLD.(2)Using network pharmacology as research method to analyze the chemical composition,potential therapeutic targets and related signaling pathways of Sancai Lianmei Particle.(3)High fat and sugar + STZ induced diabetic with fatty liver rats used as models,intervention with Sancai Lianmei Particle,intraperitoneal glucose tolerance test(IPGTT)to assess insulin resistance,ELISA method to detect the mice serum biochemistry,insulin levels;ELISA method to detect inflammatory factors,hepatic fibrosis levels in liver homogenate;HE staining was conducted to observe the liver tissue;SOD and MDA levels were monitored;Real-time PCR was used to detect the JNK/AP-1/NF-?B m RNA expression in the liver tissue;Western blot was used to detected the expression of apoptotic and autophagy-related proteins;and the ultrastructure of liver tissue was observed by transmission electron microscopy(TEM).Results:(1)Sancai Lianmei Particle can reduce the levels of blood glucose,blood lipid and Hb A1 c in T2 DM patients with NAFLD,significantly improve IR,protect the pancreatic ? cell function,reduce liver fibrosis levels,remove AGEs in patients,and reduce TCM syndrome integral in T2 DM patients with NAFLD.(2)A total of 65 compounds related to Sancai Lianmei Particle were identified by network pharmacological analysis,involving 140 signaling pathways.Among the top 20 pathways with P values,the pathways related to this study included the MAPK signaling pathway,advanced glycation end products(AGEs)signaling pathway associated with diabetic complications,and NAFLD signaling pathway.(3)Sancai Lianmei Particle can obviously reduce the body weight of T2 DM with NAFLD model rats;reduce the levels of GHb,INS,TC,TG,LDL-C,ALT,AST,IL-1?,IL-6,TNF-?,P?P,C?,HA,LN,MDA and decline HOMA-IR index;improve glucose tolerance;elevate HDL-C and SOD.HE staining of liver tissue showed that Sancai Lianmei Particle could alleviate the vacuolar degeneration of liver and the deposition of lipid droplets.Sancai Lianmei Particle can effectively down-regulate the expressions of JNK m RNA,AP-1m RNA and NF-?B m RNA in liver tissues of model rats.Western Blot results exhibited that Sancai Lianmei Particle could significantly regulate the expression of apoptotic proteins of Bax,Caspase-3,and Bcl-2.Simultaneously,Sancai Lianmei Particle could obviously down-regulate the expression of autophagy proteins of Beclin1,LC3?,LC3?.The results of TEM displayed that Sancai Lianmei Particle could inhibit the apoptosis and excessive autophagy of hepatocytes.Conclusions:(1)Clinical trial proved that Sancai Lianmei Particle can prevent and treat fatty liver disease in patients with type 2 diabetes by reducing blood glucose and lipid levels,improving IR,reducing liver fibrosis,clearing AGEs in patients,and improving clinical symptoms.Meanwhile Sancai Lianmei Particle has exact effect and security.(2)The network pharmacological analysis showed that the signaling pathways involved in Sancai Lianmei Particle mainly included the MAPK signaling pathway,AGEs signaling pathway associated with diabetes complications and NAFLD signaling pathway.(3)The experiment proved that Sancai Lianmei Particle can improve hepatic insulin resistance and oxidative stress,slow the progression of NAFLD by inhibiting the ROS-JNK signaling pathway,up-regulating the expression of Bcl-2 and down-regulating the expression of Bax,Caspase-3,Beclin1,LC3?,LC3?.