Nitric oxide releasing nanofibrous Fmoc-dipeptide hydrogels for amelioration of renal ischemia/reperfusion injury

Abstract

Renal ischemia/reperfusion (I/R) injury is responsible for significant mortality and morbidity during renal procedures. Nitric oxide (NO) deficiency is known to play a crucial role in renal I/R injury; however, low stability and severe toxicity of high concentrations of NO have limited its applications. Herein, we developed an in-situ forming Fmoc-dipheylalanine hydrogel releasing s-nitroso-n-acetylpenicillamine (FmocFF-SNAP) for renal I/R injury. Fmoc-FF hydrogel comprising of β-sheet nanofibers was prepared through the pH-titration method. It was then characterized by electron microscopy, pyrene assay, and circular dichroism techniques. Mechanical properties of Fmoc-FF hydrogel (thixotropy and syringeability) were investigated by oscillatory rheology and texture analysis. To assess the therapeutic efficiency in the renal I/R injury model, expression of inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) was measured in various samples (different concentrations of free SNAP and FmocFF-SNAP, unloaded Fmoc-FF, and sham control) by real-time RT-PCR, ROS production, serum biomarkers, and histopathological evaluations. According to the results, Fmoc-FF self-assembly in physiologic conditions led to the formation of an entangled nanofibrous and shear-thinning hydrogel. FmocFF-SNAP exhibited a sustained NO release over 7 days in a concentration-dependent manner. Importantly, intralesional injection of FmocFF-SNAP caused superior recovery of renal I/R injury when compared to free SNAP in terms of histopathological scores and renal function indices (e.g. serum creatinine, and blood urea nitrogen). Compared to the I/R control group, biomarkers of oxidative stress and iNOS expression were significantly reduced possibly due to the sustained release of NO. Interestingly, the eNOS expression showed a significant enhancement reflecting the regeneration of the injured endothelial tissue. Thus, the novel FmocFF-SNAP can be recommended for the alleviation of renal I/R injury.

Keywords: Fmoc-dipeptide hydrogel; Nitric oxide; Renal ischemia/reperfusion injury; S-nitrosothiol; Self-assembly.