| Acute kidney Injury(AKI)is a group of clinical syndrome.The pathogenesis of Acute kidney Injury is complex and unclear at present,and there is still a lack of effective treatment for Acute kidney Injury,so preventive treatment is particularly important.Traditional Chinese medicine(TCM)has been used in the treatment of a variety of diseases because of its advantages such as less toxic and side effects,many therapeutic targets and low price.It has been reported in many literatures that both ligustrazine and NO molecule can reduce Acute kidney Injury caused by ischemia reperfusion.Our research group synthesized more than ten compounds that can be degraded into ligustrazine and NO in animals.The results showed that ligustrazine furoxan complex(605-1)had an effect on the prevention and treatment of acute renal injury caused by ischemia reperfusion.In order to further study the prevention and treatment effect of ligustrazine furoxan complex on ischemia-reperfusion induced Acute kidney Injury and its possible mechanism,the experiment was divided into the following parts.1.To investigate the effect of intraperitoneal injection of ligustrazine furoxan complex on the prevention and treatment of renal ischemia-reperfusion induced Acute kidney Injury in miceMale C57BL/6 mice aged 6-8 weeks were randomly divided into 6 groups with 8mice in each group.Sham operation group(NC group)and single administration group(605-1 30mg/kg)were treated with sham operation,and 0.1ml/10 g normal saline was intraperitoneally injected 24 h and 2h before surgery.Model group(I/R group),model low,medium and high concentration groups(I/R+605-1 10mg/kg,20mg/kg,30mg/kg)were treated with surgery,and each group was intraperitoneally injected with0.1ml/10 g corresponding dose of drugs 24 h and 2h before surgery.24 h after reperfusion,blood was collected and kidney tissue was collected.In order to observe the prevention and treatment effect of ligustrazine furazan compound on acute renal injury caused by ischemia and reperfusion,pathological damage of kidney tissue in mice was detected by PAS staining,and serum levels of Cre and BUN were detected by creatinine(CRE)and urea nitrogen(BUN)kits.PAS staining showed normal renal tissue in the NC group;however,acute tubular injury was observed in the proximal tubules of the I/R group,with tubules dilatation,loss of brush boundaries,and loss of tubular cells.The renal tubular epithelium was not swollen and the brush edge was intact after the intervention of ligustrazine furazan compound,and the prevention and treatment effect was strongest when the concentration was 30 mg/kg.In addition,compared with the I/R group,the blood Scr and BUN levels of mice pretreated with different doses of ligustrazine furazan complex were decreased,and the serum Cre and BUN were the lowest at the concentration of 30 mg/kg,showing the most obvious prevention and treatment effect.Treatment with ligustrazine furazan combination alleviates acute renal injury induced by renal ischemia reperfusion in mice in a dose-dependent manner.Therefore,we chose a dose of 30 mg/kg for subsequent experiments.2.To further investigate the prevention and treatment effect of ligustrazine furoxan complex on Acute kidney Injury induced by renal ischemia reperfusion in mice by detecting the expression of KIM-1Western blot and RT-PCR were used to detect the expression of kidney injury molecule KIM-1.Immunohistochemistry and quantitative analysis of KIM-1 in renal tissues were performed.Compared with the NC group,the m RNA and protein levels of KIM-1 were increased in the I/R group.The expression of KIM-1 was decreased in the ligustrazine furoxan complex pretreatment group.With the increase of ligustrazine furoxan complex dose(10mg/kg,20mg/kg,30mg/kg),the decrease of KIM-1expression was significantly enhanced.Immunohistochemistry showed positive staining of KIM-1 in I/R group and decreased expression of KIM-1 in I/R+ligustrazine furoxan complex group.Immunohistochemical analysis of KIM-1expression was consistent with Western blot.In conclusion,it was further verified that ligustrazine furoxan complex can reduce Acute kidney Injury induced by renal ischemia reperfusion in mice.3.Explore the mechanism of ligustrazine furoxan complex ’s prevention and control effect on RIRI by detecting and analyzing the expression of NOX4,p65,p-p65 and other biomarkers.In order to explore the mechanism of ligustrazine furoxan complex alleviating RIRI,we detected the classic protein(NOX4)in oxidative stress,NF-B(p65,p-p65),and expression change of classic protein IL6,MCP-1,TNF-α in inflammatory reaction.The results showed that the protein levels of NOX4,p65 and p-p65 in renal tissue of mice with renal ischemia reperfusion were down-regulated in the ligustrazine furoxan complex preconditioning group compared with the I/R group.The RNA levels of TNF-α,IL-6 and MCP-1 were significantly decreased.The results suggested that the mechanism of ligustrazine furoxan complex alleviating Acute kidney Injury induced by renal ischemia reperfusion in mice was related to the reduction of oxidative stress and inflammatory response.4.Prevention and treatment effect of ligustrazine furazan compound on hypoxic/reoxygenated renal tubular epithelial cell(HK2)injury in vitroTo further demonstrate the efficacy of ligustrazine furoza compound in the prevention and treatment of acute kidney injury,we also tested it in vitro with renal tubular epithelial cells(HK2).First,we determined the effect of different concentrations of ligustrazine furazan complex(0,0.25,0.5,1,2,4,8,16,32μg/ m L)on HK2 cytotoxicity,i.e.survival rate,by MTT assay.The results showed that there was no cytotoxicity when the concentration of ligustrazine furazan complex was less than or equal to 4μg/ m L.By anoxia/reoxygenation stimulate HK2 cells induced acute renal tubular epithelial cells after injury,HK2 cells survival rate is measured by determined by MTT results showed that ligustrazine cefuroxime za split content can improve anoxia/reoxygenation stimulate cell survival rate,and the effect of the concentration in accordance with the lazy,when the concentration of ligustrazine cefuroxime za fitting for 4 mu g/ml control effect is best.In conclusion,the study suggested that ligustrazine furoxan complex May play a prevention and treatment role in Acute kidney Injury by inhibiting oxidative stress and inflammatory response caused by renal ischemia reperfusion in mice.ligustrazine furoxan complex can be used as a new drug for the treatment of RIRI and has a good clinical application prospect. |
PDF Full Text Request