Effect Of Ginkgo Biloba Extract On AMPK,AKT,eNOS Signal Pathways And Inflammatory Factors Of Different Uric Acid Concentrations In Glomerular Endothelium Cells

Objective:Uric acid is the result of nucleic acid catabolism and high purine food intake,internal and external interactions,and mutual influence.Humans,like most primates,have terminal metabolism of uric acid in the kidneys(90%)and intestines(10%).However,humans often suffer from hyperuricemia due to lack of uricase.When female uric acid concentration reaches 0.06g/L,male uric acid concentration reaches 0.07g/L,hyperuricemia can occur.The pathophysiological changes of the kidney caused by hyperuricemia are not all due to the deposition of urate.On the one hand,high uric acid itself can cause damage to the signaling pathways of endothelial cells,resulting in reduced nitric oxide production.On the other hand,patients with hyperuricemia can detect oxidative stress products As well as the up-regulation of inflammatory factors.How to reduce the oxidative stress and inflammatory factors of endothelial cells has become a research hotspot in recent years.Ginkgo biloba extract is to purify and separate the active ingredients of Ginkgo biloba.The active ingredients can be divided into two categories,ginkgolide and ginkgo flavonoids Ginkgolide has a high anti-platelet activation function.Ginkgo flavonoids play a role in scavenging oxygen free radicals and anti-oxidation.In recent years,Ginkgo biloba extract has played an important role in the treatment of cardiovascular and cerebrovascular diseases.This study explored the effects of Ginkgo biloba extract on the expression of AMPK,AKT and e NOS and ICAM-1 and MCP-1 inflammatory factors in glomerular endothelial cells with different uric acid concentrations.The function of renal endothelial cells in patients with hyperuricemia expands its scope of use.Methods :1.Design the research object and select human glomerular endothelial cells(HRGEC)as the experimental object.After CCK-8 cytotoxicity test,select 4mg/dl as the general concentration uric acid group,8mg/dl,16mg/d uric acid concentration It is a high-concentration uric acid group,and 0mg/dl is a control group.2.Use Ginkgo biloba(10ul/ml) 24 hours on HRGEC of different uric acid concentration,and select the best concentration of Ginkgo biloba extract by Real-time PCR method and the CCK-8 cytotoxicity test was used to detect the effect of uric acid medium and Ginkgo biloba extract on glomerular endothelial cells.3.Ginkgo biloba extract acts on HRGEC at different uric acid concentrations for 24 hours.Real-time PCR method and Western blotting method are used to detect AMPK,phosphorylated AMPK,AKT,phosphorylated AKT,e NOS m RNA and protein levels.4.The expression of ICAM-1 and MCP-1 at different uric acid concentrations in HRGEC was detected by Real-time PCR detection and Western blotting method,and the changes before and after comparison by intervention of Ginkgo biloba extract.Results:1.Observe the cell morphology and proliferation results through an inverted microscope.After 24 hours of cultivation in the general concentration uric acid group(4mg/dl),the cell morphology is normal.It can be seen that the cells are round and oval.There is no significant difference from the control group.In the 8mg/dl and 16mg/dl groups,after 24 hours of intervention,the cells were atrophic,part of the cells were necrotic,the cell culture medium was turbid,and the HRGEC growth state was poor,with fissures and even cell shedding.Ginkgo biloba extract with a concentration of 10ul/dl was used to intervene for 24 hours.Through microscopic observation,the atrophy and deformation of cells in the high-concentration uric acid group after ginkgo biloba extract intervention were improved compared with the previous ones.2.Real time PCR and Western blotting were used to detect the AMPK,AKT and e NOS m RNA and protein levels.The expression of AMPK,AKT and e NOS in the 4mg/dl uric acid group was not significantly different from that in the control group(P>0.05).The ratio of phosphorylated AMPK and total AMPK of the high concentration group(8mg/dl and 16mg/dl)was significantly lower expression than that of the control group,there was statistical significance(P<0.05);The ratio of phosphorylated AKT and total AKT of the high concentration group(8mg/dl and 16mg/dl)was significantly lower expression than that of the control group,there was statistical significance(P<0.05).The expression of e NOS in HRGEC of 8mg/dl and 16mg/dl uric acid group was significantly lower than that of the control group,Statistically significant(P<0.05).The expressions of HRGEC,AMPK,AKT and e NOS of the high concentration uric acid group(8mg/dl and 16mg/d)after Ginkgo biloba extract intervention showed an upward trend compared with before intervention,which was statistically significant(p <0.05).3.The mRNA and protein levels of AMPK,AKT and e NOS were detected by Real time PCR and Western blotting.There was no significant difference in the expression of AMPK,AKT and e NOS in the 4mg/dl uric acid group(P>0.05),and in the 8mg/dl and 16mg/dl uric acid groups Compared with HRGEC,the expression of phosphorylated AMPK and total AMPK in the control group decreased significantly,with statistical significance(P<0.05);The expression of phosphorylated AKT and total AKT decreased significantly(P <0.05).The expression of e NOS in HRGEC of 8mg/dl and 16mg/dl uric acid group was significantly lower than that of the control group(P<0.05).Ginkgo biloba extract intervened in the 8mg/dl and 16mg/dl uric acid group HRPEC,AMPK,AKT and e NOS expression showed an increasing trend compared with before intervention treatment,with statistical significance(p<0.05).Conclusion:1.High concentration of uric acid activates the AMPK,AKT,and e NOS classic inflammatory signaling pathways to reduce NO production by glomerular endothelial cells,because NO antioxidative stress improves endothelial function,and reduced NO production causes further damage to renal endothelial cells.2.Ginkgo biloba extract can effectively improve the expression of AMPK,AKT and e NOS under the condition of high concentration of uric acid,and improve the damage of glomerular endothelial cells by regulating the signaling pathway.3.Ginkgo biloba extract can effectively reduce the expression levels of ICAM-1 and MCP-1 in hyperuric acid state,and reduce the damage of inflammatory factors to glomerular endothelial cells.