The Role Of MiR-663 In The Pathogenesis Of Acute Rejection Of Renal Transplantation: An In Vitro Study

BackgroundIt still remain to be solved that how to reduce the injury caused by acute rejection after renal transplantation.Immunosuppressive drugs can greatly reduce the incidence and improve the prognosis of AR,but there is a lag in the tranditional diagnostic index(eg.glomerular filtration rate,serum creatinine level,color Doppler ultrasound,renal biopsy,and so on)for AR which increase the difficulty and cost of treatment and has a great negative impact with patients.During the process of acute rejection,the molecular signal is generated earlier than pathological damage and clinical symptoms.Novel approaches of noninvasive diagnosis are blood and urine markers,proteomics,mRNA,and so on,but there are still some defects in sensitivity,specificity and stability of those diagnostic method which limit their clinical application.miRNA is a class of non-coding small molecule single stranded RNA.It is expected to be a blood marker for early diagnosis of AR because it can express diffrently in various diseases,and regulate the process of renal acute rejection,also its physical and chemical properties are stable in the peripheral blood.miRNA is involved in cell proliferation,differentiation,apoptosis,cycle regulation and other biological processes,and plays an important role in the process of biological development and pathophysiology.Studies have shown that miR-663 is involved in the process of cell apoptosis in AR of renal allograft.Therefore,this study was designed to confirm the differential expression of miR-663 in peripheral blood of AR patients and non-AR patients,to determine whether miR-663 can be used as a specific marker for the diagnosis of AR and explore the mechanism of miR-663,which provide new ideas for early diagnosis and treatment of AR in clinic.Objective1.To compare the serum miR-663 expression in renal transplant patients with and without acute rejection(AR),and then explore whether miR-663 plays a role in regulation of the pathogenesis of acute rejection of renal transplantation;2.To explore the mechanism of miR-663 in the pathogenesis of renal acute rejection at the cellular level,which might provide new direction for early diagnosis and therapy this disease.Methods1.Realtime-PCR was used to the serum miR-663 expression in renal transplant patients with and without AR.2.Four groups namely miR-663 mimic group,miR-663 inhibitor group,negative control group and blank control were set up.MTT assay and Annexin V-FITC assay were employed to evaluate the effect of miR-663 mimic/miR-663 inhibitor on the survival and apoptosis of human renal glomerular endothelial cells(HRGEC).3.ELISA was performed to evaluate the effect of miR-663 mimic/miR-663 inhibitor on the expression of inflammation related cytokines including IL-6,IFN-y,CCL-2 and TNF-?.4.Transwell assay was conducted to examine the effect of miR-663 mimic/miR-663 inhibitor on the chemotaxis capacity of macrophage.ResultsThe serum miR-663 expression level in renal transplant patients with AR was significantly higher than in patients without AR.miR-663 mimic could significantly inhibit survival of HRGEC and increase apoptosis rate,while miR-663 inhibitor could decrease the apoptosis rate.In addition,miR-663 mimic could increase the expression level of inflammation related cytokines and enhance the chemotaxis capability of macrophage.ConclusionmiR-663 might play important roles in the process of acute rejection of renal transplantation,it has potential to become a therapy target for treating renal transplantation AR.