Protective Effect Of RhMG53 On Contrast-induced Acute Kidney Injury

Over the last decades,with the commonly using of vascular interventional techniques,spiral CT and new three-dimensional reconstruction techniques,the utilization of intravascular iodinated contrast media has been widespread.Consequently,the incidence of contrast-induced acute kidney injury(CI-AKI)or contrast-induced nephropathy(CIN)is rising year by year.Clinical research data show that CI-AKI has become the third major cause of iatrogenic acute renal failure,the mortality rate is as high as 34.0%.The incidence of contrast induced nephropathy in the normal population is not high,but in the population of the incidence of diabetes,chronic renal insufficiency,hypertension and other diseases,the incidence rate will be greatly increased.At present,there are still a lack of effective measures to reverse the progression of CI-AKI.How to prevent and reduce the kidney CI-AKI,has been an important issue in kidney protection,and gradually become a hot spot by clinical and scientific research.Mitsugumin 53(MG53)is a newly discovered protein,which belongs to the motif Tripartite(TRIM)family,expresses in the cardiac muscle and skeletal muscle.It is reported that in acute myocardial ischemia reperfusion injury,MG53 can play a protective role by activating RISK signaling pathway.Vivo and vitro studies showed that exogenous recombinant human MG53 protein(rhMG53)can protect renal ischemia / reperfusion injury.In recent years,our study found the expression of MG53 in kidney is abundant,and MG53 knockout mice showed renal insufficiency,indicating that MG53 may be critical in the regulation of renal function.However,the protective effect of MG53 on CI-AKI is not clear,therefore,in this study we established a rat model of acute renal injury induced by contrast media,MG53 was inserted into the externl jugular vein,and then the urine was collected to measure the renal function(BUN,Scr and CCr),the renal morphology was detected by HE staining,the renal tubular apoptosis was detected by TUNEL staining.the apoptosis protein Cleaved Caspase-3 and Totol Caspase-3 evalutes by western blot,and the signal transway of MG53 in the protective effect of CI-AKI was also tested by western blot.Research objective : To investigate the protective effect and mechanism of MG53 on acute renal injury in rats,and provide theoretical basis for clinical prevention and treatment of renal injury.Research method:Male and Female SD rats(24)(10 to 12 weeks old,weighting 225±15g)were randomly divided into four groups(n=6 each):control group,rhMG53 group,contrast medium(CM)group and contrast medium+rhMG53(CM+rhMG53)group.The rats were given indomethacin,Nw-nitro-L-Arginine methy1 ester(L-NAME)and iopromide through the external jugular vein intubation to establish contrast-induced acute kidney injury model.Then isolated subcutaneous tissue,exposed to the right external jugular vein,cut the vein,and insert the catheter to infuse the drugs.Control group:,injected normal saline at each time point.rhMG53 group: rhMG53 was inserted into the external jugular vein catheter at the dose of 5mg/kg CM group: First,10mg/kg indomethacin(dissolved in DMSO)was inserted then after 15 mins or 30 mins Nw-nitro-L-Arginine methylester(L-NAME,dissolved in normal saline)administration at the dose of 10mg/kg was inserted,at last the the low-osmolar,nonionic contrast medium agent(Iopromide)at a dose of 1600 mg I/kg was inserted.CM+rhMG53 group:Injecting rhMG53 15 min before renal injury.After the operation,all the subjects were placed in the metabolic cage to collect urine in 24 h,and then they were euthanized by inhalation of ether anesthesia.the kidney tissue and serum of the rats in each group were collected.The kidney function parameters(BUN,Scr,CCr and 24 h U-mAlb)were measured,the renal morphorogy was detected by HE staining,the renal tubular apoptosis was detected by TUNEL.The expression of Cleaved Caspase-3 、 Akt 、 GSK-3β 、 phosph-Akt 、phosph-GSK-3β by Western blot were detected,Pathological grading of renal injury was performed in three aspects,such as renal interstitial congestion,protein cast and cell necrosis.ResultsRhMG53 protected the acute renal injury caused by contrast agent.We established SD rats with contrast induced acute kidney injury model,then treated with rhMG53 to investigate the protective effects of CI-AKI.The results showed that:renal function and renal pathological score was not statistically significant between rhMG53 treatment group and Control group.Rat renal function indexes such as serum creatinine and urea nitrogen were significantly increased,creatinine clearance were significantly decreased after CI-AKI,HE staining showed that the main site of renal injury is the corticomedullary junction,because more tubular epithelial cell necrosis,inflammatory infiltration vacuolization,lumen tube,protein cast,and more severe vascular congestion and hemorrhage in renal interstitium and tubules were appeared.However,after treated with rhMG53,the severity of tubular epithelial cell necrosis and protein cast,PTC congestion and interstitial edema were significantly reduced,and renal function were improved,indicating rhMG53 has protective effects on contrast induced acute kidney injury.RhMG53 reduced the apoptosis induced by contrast induced acute kidney injury.We used TUNEL staining and Western blot to detecte the apoptosis in CI-AKI rat.Through the TUNEL test we found that control group and rhMG53 treatment group only have very few apoptosis in the kidney tissue,the TUNEL positive cells of CM group were significantly increased and mainly distributed in the renal medulla junction,TUNEL positive cells in CM+rhMG53 group were significantly reduced compared with CM group;The Western blot results show that Cleaved Caspase-3 increased significantly after CI-AKI and decreased significantly after treatment with rhMG53,which confirmed that rhMG53 can reduce the apoptosis of renal cells.RhMG53 plays a protective effect on the contrast agent of acute kidney injury via reperfusion injury salvage kinase(RISK)pathway.Previous researches found in cardiac ischemia reperfusion injury,MG53 acted as a bridge with its N-terminus interacting with CaV3 and its C-terminus interacting with p85-PI3 K,and increased the expression of phosph-Akt and phosph-GSK-3β,then activated the RISK pathway,.We found that phosph-Akt and phosph-GSK-3β decreased significantly in CI-AKI rats,but after rhMG53 treatment,the expression of phosph-Akt and phosph-GSK-3β increased.These results show that rhMG53 can play a protective effect on contrast induced acute kidney injury by activating RISK signaling pathway.Conclusion:We found that rhMG53 improved renal function,reducing the pathological injury,and renal cell apoptosis in CI-AKI via RISK signaling pathway.The protective role of rhMG53 may open an new avenues for the prevention and management of CI-AKI.