Activation of the iNOS/NO/cGMP pathway by Revactin® in human corporal smooth muscle cells

BACKGROUND: The combination of the nutraceuticals, Paullinia cupana, ginger rhizome, muira puama, and the amino acid L-citrulline (COMP-4) has been shown to stimulate the production of inducible nitric oxide synthase (iNOS), nitric oxide (NO), and cGMP in rat corpora cavernosa smooth muscle cells (CSMC). When administered to middle-aged rats, long-term treatment with COMP-4 resulted in both an increase in the number of CSMC and an improvement in erectile function. We, therefore, aimed to determine whether a commercial formulation of COMP-4, Revactin(®), could have a similar stimulatory effect on human CSMC. METHODS: Primary human CSMC cultures (HCSMC) were grown and incubated with Revactin® for up to 24 hours. cGMP generation and nitrite formation were determined by ELISA and Griess reaction, respectively. IBMX (1 mM), sildenafil (0.4 mM), and L-NIL (4 µM) were utilized as modulators of the NO-cGMP pathway. iNOS, endothelial NOS (eNOS), and neuronal NOS (nNOS) expressions were determined by Western blot. RESULTS: Revactin(®) up-regulated both nitrite formation and cGMP expression, achieving the highest expression at 24 hours in the HCSMC. These effects were completely blocked by L-NIL. Revactin(®) up-regulated iNOS expression, but not that of eNOS or nNOS. CONCLUSIONS: The results presented in this study confirmed that Revactin(®) activated the iNOS-NO-cGMP pathway intracellularly in HCSMC. It still needs to be determined whether the upregulation of this pathway would be an effective approach for counteracting the fibrosis and apoptosis of the corporal smooth muscle cells associated with aging.