Study On The Mechanism Of C5a Receptor Mediated Lipopolysaccharide Inducing Renal Sepsis And Renal Ischemia/Reperfusion(I/R) Injury

In the body of animals and human beings,there is a kind of biological active component which is related to the immune response and has the characteristic of enzyme activity,mainly exists in the serum or cell membrane surface.Some immune activators activate the complement system,and produce a large number of fragments,such as C3a,C4a,C5a.Studies have shown that the strongest anaphylatoxin is C5a which is the cleavage fragment of complement C5 after the activation of the complement cascade.C5a plays a variety of physiological functions in the organism mainly through the combination with C5a receptor(C5aR)to form C5a/C5aR complex.C5a receptor,also called CD88,is expressed in a variety of cell membrane glycoprotein which is a member of the large family of G protein coupled receptors.It is an important neurotransmitter of the inflammation and chemotaxis factor,plays an important role in the acute inflammatory response.C5aR is widely expressed in the kidney tissue and plays a key role in the process of renal pathology.When the kidneys are injured,C5aR can induce mononuclear cells to release TNF-α,IL-6 and IL-8,etc.Whether does C5aR mediate LPS-induced kidney damage and the mechanism of its action has not yet fully clarified.In this study,short hairpin RNA(shRNA)expression vector of C5aR gene was constructed and transfected into rat renal epithelial cell line NRK-52E with liposome 2000 and injected to rats intravenously through the tail vein.The expression of C5aR,apoptosis,and the expression of related cytokines in cells and animals were detected using fluorescence quantitative PCR method,western blot method,ELISA method,flow cytometry and other methods.To investigate the effect and mechanism of C5aR in renal ischemia/reperfusion injury and to clear the intervention effect of C5aR antagonist on the disease process,apoptosis was determined by flow cytometry,the concentration of inflammatory factors IL-6 and TNF-a in supernatant were detected by ELISA.The changes of NFAT nuclear translocation,the expression of proinflammatory cytokines and cell activity were detected by immunofluorescence assay,RT-PCR and CCK-8 method,after using C5a and C5aR antagonists stimulate kidney epithelial cells.These studies show that C5aR is an important factor in renal epithelial cell injury,blocking the C5aR can prevent kidney damage.Through in-depth study of the C5a receptor,for us to futher study the pathogenesis of kidney disease provides a new perspective;while also providing new research ideas and targets for intervention for the prevention and treatment of kidney disease,etc.By interfering with the expression of C5a receptor,it is expected to become a new method for the prevention and treatment of kidney disease.