| Object:Renal ischemia reperfusion injury(RIRI) is the main reason to induce acute kidney injury( AKI), such as RIRI may happen after kidney transplantation or partial resection and further induce AKI. The present study investigated the association between NICD overexpression and injury repair after renal ischemia reperfusion through constructing bone marrow mesenchymal stem cell models. These cells can overexpress NICD in vitro. NICD is one of the intracellular function fragment of Notch1 Signaling pathway. Then we further determined the role of NICD in promoting bone marrow mesenchymal stem cells to repair the injury after renal ischemia reperfusion.Methods:1. Rats bone marrow mesenchymal stem cells(MSCs) were conventionally isolated and cultured, and then the growth status of rats bone marrow mesenchymal stem cells(MSCs) was observed and recorded;2. The Mig R1-NICD plasmid was constructed by cloning technology in vitro, and then being transfected into rat bone marrow mesenchymal stem cells, the positive cells were screened by resistance and being defined as Mig R1-NICD-MSCs cells;3. The expression of NICD in Mig R1-NICD-MSCs cells was respectively detected using quality real-time polymerase chain reaction(q RT-PCR) and western blot techniques at m RNA levels and protein levels;4. The effect of NICD being exerted on the proliferation of MSCs cells was detected by CCK-8 proliferation assay kit;5. After constructing the RIRI models, rats were randomly divided into experimental group and control group. The equal volumes of Mig R1-NICD MSCs cells and Mig R1-MSCs cells were then injected into rat inferior cava vena of experimental and control group rats, respectively. Serum creatinine and blood urea nitrogen were definitively recorded before rats renal blood flow being clamped and as well as 24 h, 48 h,72h and 96 h after reperfusion;6. Rats kidneys were isolated out from the experimental and control group rats respectively. After being fast frozen, sections were stained by H&E staining techniques. The pathological difference of rats RIRI between these two groups was observed by microscope;7. statistical analysis.Results:1.The results of q RT-PCR and Western blot showed that the expression of NICD in Mig R1-NICD-MSCs cells was significantly higher than that in Mig R1-MSCs cells, which indicated that the NICD overexpression cell models were successfully constructed;2.The results of CCK-8 assay showed that the proliferation rate of Mig R1-NICD-MSCs cells was significantly faster than that of Mig R1-MSCs cells, which indicated that NICD could promote the proliferation of bone marrow mesenchymal stem cells;3. In experimental group rats, the baseline serum creatinine and blood urea nitrogen (?)±s of were 30.14±2.81 and 5.98±1.23, respectively; 24 h serum creat-inine and blood urea nitrogen (?)±s were 138.25±2.06 and 23.27±0.53, respectively; 48 h serum creatinine and blood urea nitrogen (?)±s were 60.35±2.58 and 12.17±0.33, respectively; 72 h serum creatinine and blood urea nitrogen (?)±s were 41.25±1.70 and 7.12±0.20, respectively; 96 h serum creatinine and blood urea nitrogen (?)±s were 32.16±1.636 and 23±1.14, respectively. In the control group rats, the serum creatinine and blood urea nitrogen (?)±s in the corresponding time points were 30.49±2.92 and 6.03±0.41, 146.50±2.02 and 28.45±0.12, 69.07±1.63 and 16.25±0.36, 49.12±1.41 and 8.95±0.12, 33.04±2.13 and 6.56±0.36, respectively. The renal function comparison between the two groups in the baseline level and 96 h after renal ischemia reperfusion had no significant difference(P>0.05). But, the serum creatinine and blood urea nitrogen level of experimental group rats in 24 h, 48 h and 72 h after renal ischemia reperfusion was significantly lower than the control group rats(P<0.05).4. The results of H&E staining showed that the renal tubular epithelial cell necrosis rate of experimental group rats was significantly lower than that of control group rats. The results of renal tubule score after pathological injury showed that the scores of experimental group rats were significantly lower than that of control group rats(P<0.05). Both the above results may indicate that NICD can promote bone marrow mesenchymal stem cells to repair the injury caused by renal ischemia reperfusion.Conclusions:1. NICD protein can accelerate the proliferation rate of rats bone marrow mesenchymal stem cells(MSCs);2. NICD can promote bone marrow mesenchymal stem cells to repair the injury of renal ischemia reperfusion. |
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