HGF/c-Met反应轴对脂肪干细胞修复烧伤创面的调控

HGF/c-Met axis regulates human adipose tissue-derived stem cells in burn wound healing

背景:研究发现,肝细胞生长因子(hepatocyte growth factor,HGF)能促进脂肪干细胞增殖迁移,进而促进创面愈合。目的:观察HGF/c-Met调节脂肪干细胞增殖和迁移的作用机制及其对烧伤创面愈合的影响。方法:将提取的人脂肪干细胞按实验要求分为空白对照组(未处理的脂肪干细胞)、HGF组(10μg/L,24 h)、c-Met si RNA+HGF组(10μg/L,24 h);采用Western blot技术检测c-Met、AKT、p-AKT的蛋白表达,CCK-8和Transwell法分别检测细胞增殖和迁移情况。将12只裸鼠随机分为2组,用99℃烫头接触裸鼠背部皮肤5 s,构建直径为1 cm的皮肤Ⅲ度烫伤模型。实验组创面周围皮肤内注射1×106个10μg/L HGF处理24 h的脂肪干细胞,对照组注射等量未处理的脂肪干细胞。所有裸鼠在实验第7天处死,取新生创面全层皮肤组织,进行苏木精-伊红染色,计算创面愈合率和再上皮化厚度。结果与结论:(1)HGF能够诱导c-Met表达(P<0.05);HGF促进脂肪干细胞的增殖和迁移,c-Met si RNA对脂肪干细胞的增殖与迁移起抑制作用(P<0.05);HGF促进p-AKT蛋白表达,c-Met si RNA抑制p-AKT蛋白表达;(2)实验组创面再上皮化厚度大于对照组(P<0.05),创面愈合率高于对照组(P<0.05);(3)结果表明,HGF干预的脂肪干细胞能通过诱导c-Met基因的表达,激活PI3K/AKT信号通路,提高脂肪间充质干细胞的增殖与迁徙能力;HGF干预的脂肪干细胞能促进裸鼠烫伤创面的愈合。

BACKGROUND:Studies have found that hepatocyte growth factor(HGF)can promote proliferation and migration of adipose-derived stem cells(ADSCs)and accelerate wound healing.OBJECTIVE:To investigate the mechanism of HGF/c-Met on the proliferation and migration of ADSCs and its effects on burn wound healing.METHODS:ADSCs were separated from subcutaneous adipose tissues and primary cultured.ADSCs untreated were considered as control group.Cells in experimental groups were treated with HGF(10μg/L,24 hours)or c-Met siRNA combined with HGF(10μg/L,24 hours).Then,western blot assay was used to detect expressions of c-Met,p-AKT,and AKT in the cells.The proliferation and migration abilities of ADSCs were observed by cell counting kit-8 and Transwell assay.Twelve nude mice were burned by copper bar at 99 oC for 5 seconds in order to establish animal models of skin III degree burns with the diameter of 1 cm,and then these model mice were randomly divided into experimental and control group(n=6 per group).In control group,only 1×106 ADSCs were injected around the wound,while in experimental group,1×106 ADSCs transfected with Ad-HGF were injected around the wound.All the mice were killed at 7 days of the experiment,and fresh full-thickness skin wound tissues were taken.Wound healing rate was calculated and the thickness between dermis and epidermis was measured by hematoxylin-eosin staining.RESULTS AND CONCLUSION:Significant expression of c-Met was induced by HGF(P<0.05).The proliferation and migration of ADSCs were promoted by HGF but suppressed by c-Met siRNA(P<0.05).p-AKT expression could be promoted by HGF but inhibited by c-Met siRNA.Compared with the control group,the experimental group showed higher wound healing rate and thickened re-epithelium(P<0.05).To conclude,the HGF/c-Met axis can promote the proliferation and migration of ADSCs by activating PI3K/AKT pathway,and HGF can accelerate wound healing of burn nude mice.