Substance P ameliorates tumor necrosis factor-alpha-induced endothelial cell dysfunction by regulating eNOS expression in vitro

ObjectiveThe aim of this study was to explore the beneficial effects of SP on NO production and inflammation-induced vascular endothelium cell death. MethodsTo mimic the inflammatory environment, TNF- was treated with HUVECs, and SP was added prior to TNF- to determine its protective effect. WST-1 assay was performed to detect cell viability. NO level in conditioned medium was measured by Griess Reagent System. The protein level of cleaved caspase-3, eNOS, and phosphorylated Akt was detected by Western blot analysis. ResultsTNF- declined endothelial cell viability by downregulating Akt and NO production. TNF--induced cell death was reliably restored by NO, confirming the requirement of NO for cell survival. By contrast, pretreatment of SP attenuated TNF--induced cellular apoptosis, accompanied by an increase in the phosphorylation of Akt, eNOS expression, and NO production. Blockage of NK-1R, phosphorylated Akt or eNOS by CP-96345, A6730, or L-NAME entirely eliminated the effect of SP. ConclusionsSP can protect the vascular endothelium against inflammation-induced damage through modulation of the Akt/eNOS/NO signaling pathway.