Nitric oxide induces epidermal stem cell de-adhesion by targeting integrin ß1 and Talin via the cGMP signalling pathway.

ABSTRACT

OBJECTIVE: Nitric oxide (NO) has emerged as a critical molecule in wound healing, but the mechanism underlying its activity is not well defined. Here, we explored the effect of NO on the de-adhesion of epidermal stem cells (ESCs) and the mechanism involved in this process. METHODS: The effects of NO on isolated human and mouse ESCs cultured in the presence of different concentrations of the NO donor S-nitroso-N-acetyl penicillamine (SNAP) were evaluated in cell de-adhesion assays mediated by integrin ß and collagen IV. Subsequently, changes in the expression of integrin ß1 and the phosphorylation of Talin in response to different doses of SNAP were detected by Western blot analysis and real-time PCR in vitro. Furthermore, the roles of various soluble guanylyl cyclase (sGC)- and protein kinase G (PKG)-specific inhibitors and agonists in the effects of NO on ESC de-adhesion, integrin ß1 expression and Talin phosphorylation were analysed. Moreover, the effects of NO on integrin ß1 expression and sGC/cGMP/PKG signalling-mediated wound healing were detected in vivo using 5-bromo-2-deoxyuridine (BrdU) label-retaining cells (LRCs) in a scald model and an excision wound healing model, respectively. RESULTS: SNAP promoted primary human and mouse ESC de-adhesion in a concentration-dependent manner in the integrin ß1-and collagen IV-mediated adhesion assay, and this effect was suppressed by the sGC and PKG inhibitors. Additionally, integrin ß1 expression and Talin phosphorylation at serine 425 (S425) were negatively correlated with SNAP levels, and this effect was blocked by the sGC and PKG inhibitors. Moreover, the roles of NO in integrin ß1 expression and cGMP signalling pathway-mediated wound healing were confirmed in vivo. CONCLUSION: Our data indicate that the stimulatory effects of NO on ESC de-adhesion related to integrin ß1 expression and Talin phosphorylation were mediated by the cGMP signalling pathway, which is likely involved in wound healing.