Study On The Substance Basis And Mechanism On Anti-septic Acute Kidney Injury Of Tibetan Medicine Lagotis Brevituba Maxim.

Sepsis-induced acute kidney injury is a nephrotic syndrome characterized by a rapid decline in kidney function over a short period of time.The incidence and mortality of sepsis-induced acute kidney injury have been high in recent years.Furthermore,it lacks of effective and safe clinical treatment strategy.Hence,it is urgently needed to develop new medicine with high efficiency and low toxicity for treatment of sepsis-induced acute kidney injury.Tibetan medicine L.brevituba Maxim has the effect of clearing heat and detoxifying blood,and is widely used for treatment of generalized fever and nephritis.Modern pharmaceutical studies have found that L.Brevituba Maxim.is rich in phenylethanol glycosides,flavonoids,iridoids and other types of compounds,and has anti-inflammatory,antioxidant and other biological activities.In order to verify the treatment of nephritis in clinical application of L.Brevituba Maxim.,and to clarify its material basis and mechanism on anti-acute kidney injury,this study was conducted,and the results were listed as follows:1.Study on the therapeutic activity of L.Brevituba Maxim.on LPS-induced septic acute kidney injury mice:1)The weighing method was used to determine the kidney index in mice.The levels of BUN,CRE,KIM-1,TNF-α in serum and IL-1β in renal tissue were detected by ELISA kits.The histopathological changes of kidney were assessed by H&E staining.The anti-septic acute kidney injury activity of L.Brevituba Maxim.with different polarity was evaluated using LPS induced acute kidney injury model.The results showed that 30%,60% and 90% ethanol fractions possessed anti-sepsis ARF effect,and 30% ethanol fractions obtained the best therapeutic effect.2)The protein expressions of TLR4,NF-κB P65,MAPK P38 and MAPK P-P38 were detected by Western blot method to explore the therapeutic mechanism of 30%ethanol fraction on septic acute kidney injury mice,and the results showed that it could regulate the TLR4/NF-κB/MAPK signaling pathway to exert anti-septic acute kidney injury activity by down-regulating the expression of TLR4 protein,NF-κB P65 protein and phosphorylated protein of MAPK P38.Furthermore,it could reduce the contents of inflammatory factors TNF-α and IL-1β in serum and tissues.2.Study on chemical Constituents of the effective parts of L.Brevituba Maxim.:A variety of chromatographic techniques and spectroscopic methods were used to separate and identify the components from 30% ethanol fraction.The results showed that 18 compounds were obtained,which included 9 phenylethanol glycosides(2,3,5-11),2 phenylpropanoids(4,12),2 flavonoids(13-14)and 5 phenolic acid compounds(1,15-18).Among them,Lagotoside K(1),Lagotoside L(2),Lagotoside M(3)and Lagotoside N(4)were new compounds,and lycioside D(8)and Lagotisoside A(15)were isolated from L.Brevituba Maxim.for the first time.Chionoside J(9),Maxoside(10),Turrilliside A(11),Juniperoside(12),Luteolin7-O-Sophoroside(13),6-hydroxyluteolin-7-O-glucoside(14),nepetin C(16),4-hydroxybenzoic acid-β-d-glucoester(17)were isolated from the genus Lagotis.for the first time,which enriched the library of chemical composition of L.Brevituba Maxim.3.Study on anti-inflammatory mechanism of monomer compounds:The renal cytotoxicity of 18 isolated compounds was evaluated using HK-2 cell model,and the compounds with anti-inflammatory activities were screened using LPS-induced RAW264.7 cell damage model.The results showed that compounds 1,4,5,12,13,15-18 showed no obvious renal cytotoxicity at the concentration of 100μM,and all compounds showed no obvious renal cytotoxicity at the concentration of50μM.Nine phenylethanol glycosides(2,3,5-11)could significantly reduce the contcentation of NO secreted by LPS-induced RAW264.7 cells,showing good anti-inflammatory activities.In a word,our work laid a materialand experimental basis for development of new medicines for anti-septic nephritis based on Tibetan medicine L.Brevituba Maxim.