Effects of cGMP phosphodiesterase inhibitor on expression of eNOS and VEGF in rats with cyclosporine A induced nephrotoxicity

Abstract

[한글]

[영문]Background: Cyclosporine-A (CsA) inhibits the differentiation of T cells by suppressing calcineurin and thus blocks the activation of the IL-2 promotor gene which blocks the rejection of kidney transplants and is an essential immune suppressor used after kidney transplantation. However, when CsA is used long term, nephrotoxicity develops. Hence, long term CsA use must be limited. The mechanism of CsA induced nephrotoxicity have been suggested vasoconstriction due to the reduction of nitric oxide (NO), and tissue fibrosis caused by the elevation of transforming growth factor (TGF)-beta, vascular endothelial growth factor (VEGF). The cyclic guanosine monophosphate (cGMP) phosphodiesterase (PDE) inhibitor (Udenafil, Zydena®), active in the NO/cGMP pathway, with its physiological function induced by cGMP, has been reported to ameliorate renal injury by the increase of cGMP through the suppression of cGMP-specific PDE-5. In this study, in a rat model of CsA-induced nephrotoxicity, the administration of the PDE-5 inhibitor udenafil was studied to determine whether it ameliorated kidney dysfunction and altered the expression of eNOS and VEGF.Methods: A right nephrectomy was performed in Sprague-Dawley rats (N=30, 200-250g, male). The Ischemia group (N=6), had ligation of the left renal artery for 45 minutes (IR) and was maintained for 28 days. The Udenafil group (N=6), after ischemia, was treated with 10 mg/kg udenafil orally for 28 days. The CsA group (N=6), after IR and then 15 mg/kg cyclosporine-A was injected subcutaneously for 28 days. The CsA with udenafil group (N=6), after IR received 15 mg/kg cyclosporine-A injected subcutaneously together with the oral administration of 10 mg/kg udenafil for 28 days. A low salt diet was provided to all groups. After the collection of blood on day 28, the remaining kidney was resected surgically and the injury was assessed by hematoxylin-eosin (H&E) staining, immunohistochemical staining, western blot, real time-PCR analysis was used to assess degree of renal injury and the expression of eNOS and VEGF.Results: The comparison with the Ischemia group and the CsA group, after the administration of udenafil, showed that the creatinine was significantly decreased (p=0.002, p=0.002, respectively). The H&E staining, comparing the Ischemia group and the CsA group, showed that the level of loss of the nuclei in the proximal tubules was significantly decreased after the administration of udenafil (p=0.004, p=0.002, respectively). The immunohistochemical staining showed that eNOS stained strongly in the Udenafil group and the CsA with udenafil group compared to the other groups. The results of the eNOS western blot showed that it was not changed in the Ischemia group; however, the expression of the protein was decreased in the CsA group, and increased in the Udenafil group. For the VEGF western blot, the expression of the protein was reduced only in the CsA with udenafil group. The eNOS mRNA assessed by real time-PCR was decreased in the CsA group compared to the other groups (p=0.000). VEGF mRNA was decreased in the CsA group, and after the administration of udenafil, it showed a tendency to decrease more (p=0.003, p=0.005, respectively).Conclusion: cGMP phosphodiesterase inhibitor ameliorated kidney injury in a rat model of CsA-induced nephrotoxicity. The mechanism appears to be associated with an increase of eNOS and reduction of VEGF.