Rixing Zhan1, Fan Wang2, Ying Wu3, Ying Wang4, Wei Qian5, Menglong Liu6, Tengfei Liu7, Weifeng He8, Hui Ren9, Gaoxing Luo10. 1. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China; School of Nursing, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: zhanrixing@sina.com. 2. Department of Plastic and Reconstructive Surgery, Southwestern Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: 693199983@qq.com. 3. The Institute of Hepatobiliary Surgery, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing, China. Electronic address: 26130501@qq.com. 4. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: yingwang0528@163.com. 5. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: weiqian87@126.com. 6. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: lml1390@126.com. 7. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: liutengfei93@sina.com. 8. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: whe761211@aliyun.com. 9. School of Nursing, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: renhui_tmmu@163.com. 10. Institute of Burn Research, State Key Laboratory of Trauma, Burn and Combined Injury, Key Laboratory of Proteomics of Chongqing, Southwest Hospital, The Third Military Medical University (Army Medical University), Chongqing 400038, China. Electronic address: logxw@yahoo.com.
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 β and collagen IV. Subsequently, changes in the expression of integrin β1 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 β1 expression and Talin phosphorylation were analysed. Moreover, the effects of NO on integrin β1 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 β1-and collagen IV-mediated adhesion assay, and this effect was suppressed by the sGC and PKG inhibitors. Additionally, integrin β1 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 β1 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 β1 expression and Talin phosphorylation were mediated by the cGMP signalling pathway, which is likely involved in wound healing.
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 donorS-nitroso-N-acetyl penicillamine (SNAP) were evaluated in cell de-adhesion assays mediated by integrin β and collagen IV. Subsequently, changes in the expression of integrin β1 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 β1 expression and Talin phosphorylation were analysed. Moreover, the effects of NO on integrin β1 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 β1-and collagen IV-mediated adhesion assay, and this effect was suppressed by the sGC and PKG inhibitors. Additionally, integrin β1 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 β1 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 β1 expression and Talin phosphorylation were mediated by the cGMP signalling pathway, which is likely involved in wound healing.