Valacchi Giuseppe, Sticozzi Claudia, Zanardi Iacopo, Belmonte Giuseppe, Cervellati Franco, Bocci Velio, Travagli Valter
a Department of Life Sciences and Biotechnology , University of Ferrara , Ferrara , Italy ;
b Department of Biotechnology, Chemistry and Pharmacy , University of Siena , Siena , Italy.
Free Radic Res. 2016 Sep;50(9):1022-31. doi: 10.1080/10715762.2016.1219731.
The beneficial effect of low doses of ozone on wound healing has been well documented and attributed mainly to its bactericidal and pro-oxidant properties. Because ozone itself does not penetrate the cells but immediately reacts with polyunsaturated fatty acids, its effects are the results of oxidative mediators. Among the molecule produces by the interaction of ozone with biological systems, there are HNE and H2O2. At today, the cellular mechanisms accounting for the positive effects of mild ozonization on wound closure are still largely unexplored. The aim of the present study was to evaluate the effect of different non-toxic doses of ozonated saline ranging from 2 to 300 μM, in an in vitro wound scratch model by the use of human keratinocytes. The results showed that ozonated saline is able to improve in vitro wound healing by stimulating cell proliferation as measured by BrdU assay and PCNA protein levels. In order to better elucidate the molecules that play the main role in the beneficial effect of ozonated saline in wound healing, HNE and H2O2 were used alone or in combination to mimic ozonated saline effect. Surprisingly, keratinocytes treated with different doses of HNE and H2O2 did not significantly improve the wound closure, while the combination of the two compounds was able to improve wound closure. In addition, Nrf2 pathways were also activated as determined by its translocation to the nucleus and the increased HO1 gene expression. The present work suggests that ozonated saline effect on wound closure is the results of the combination of more molecules among which HNE and H2O2 play a key role.
低剂量臭氧对伤口愈合的有益作用已有充分记载，主要归因于其杀菌和促氧化特性。由于臭氧本身不会穿透细胞，而是立即与多不饱和脂肪酸发生反应，其作用是氧化介质的结果。在臭氧与生物系统相互作用产生的分子中，有HNE和H2O2。目前，关于温和臭氧处理对伤口闭合产生积极影响的细胞机制仍 largely unexplored。本研究的目的是通过使用人角质形成细胞，在体外伤口划痕模型中评估2至300μM不同无毒剂量的臭氧生理盐水的效果。结果表明，臭氧生理盐水能够通过刺激细胞增殖来改善体外伤口愈合，这通过BrdU测定法和PCNA蛋白水平来衡量。为了更好地阐明在臭氧生理盐水对伤口愈合的有益作用中起主要作用的分子，单独或联合使用HNE和H2O2来模拟臭氧生理盐水的效果。令人惊讶的是，用不同剂量的HNE和H2O2处理的角质形成细胞并没有显著改善伤口闭合，而这两种化合物的组合能够改善伤口闭合。此外，通过Nrf2易位至细胞核和HO1基因表达增加确定Nrf2途径也被激活。目前的工作表明，臭氧生理盐水对伤口闭合的作用是多种分子组合的结果，其中HNE和H2O2起关键作用。
