摘要:
Objective To characterize the pigment epithelium-derived factor (PEDF) expression in the mechanically expanding skin tissues.Methods The rat PEDF expression was identified by immunofluorescence staining on skin tissue sections and by quantitative characterization using Western blotting on tissue lysates originated from the mechanically expanding skin samples.To further elucidate the effects of expansion-relevant tissue microenvironment stimuli on the cellular PEDF expression,the PEDF gene transcription levels were determined respectively on epidermal cells that were incubated in vitro under either hypoxia or mechanical stretching conditions.Results For normal oxygen group,epidermal cell line HaCaT cells were cultured and collected at 1,3,6 and 9 h,and the PEDF mRNA espression levels were 0.392 ±0.174,0.385 ±0.207,0.491 ±0.134,0.122 ±0.040 respectively at 1,3,6 and 9 h.For hypoxia group,PEDF mRNA expression levels were markedly upregulated correspondingly [2.271 ± 0.781 (P <0.05),3.419 ±0.774 (P<0.05),2.886 ±0.019 (P<0.05),and 2.673 ±0.219 (P<0.05) respectively at 1,3,6 and 9 h].For stretch group,HaCaT cells were applied to mechanical stretch for the same period under the normal oxygen.It was found that the PEDF mRNA expression levels were also upregulated correspondingly [1.535 ±0.199,1.078 ±0.167,1.295 ±0.012 and 1.500 ±0.104 respectively at 1,3,6 and 9 h].However,the difference in the mRNA expression levels between stretch group and normal oxygen group was just significant at 1 h after the onset of mechanical stretching.Conclusion The microenvironmental hypoxia and mechanical strain significantly upregulate epidermal cellular PEDF expression respectively.%目的 观察皮肤软组织扩张器施加扩张应力条件下,皮肤组织表达色素上皮衍生因子(PEDF)的特征.方法 通过免疫荧光染色和Western blot检测大鼠扩张期皮肤样品中PEDF的表达、分布及PEDF蛋白表达水平;利用体外细胞牵拉装置和低氧培养箱分别对表皮细胞株HaCaT细胞进行处理,以模拟扩张皮肤中缺氧和机械应力微环境,探讨影响表皮细胞PEDF基因转录水平的主要组织微环境因素,此两组实验组分别为缺氧组和牵拉组,无任何干预组为对照组.结果 在缺氧处理后,取1、3、6、9h的细胞,检测到细胞PEDF mRNA表达水平显著上调并维持在较高水平,1h时缺氧组PEDF mRNA (2.271±0.781)高于对照组(0.392±0.174,P<0.05),3h时缺氧组PEDF mRNA(3.419±0.774)高于对照组(0.385±0.207,P<0.05),6h时缺氧组PEDF mRNA(2.886±0.019)高于对照组(0.491±0.134,P<0.05),9h时缺氧组PEDF mRNA(2.673±0.219)高于对照组(0.122±0.040,P<0.05).常氧条件下对HaCaT细胞施加牵拉应力培养相同时间后,检测到1h细胞PEDF mRNA表达水平(1.535±0.199)高于对照组(0.392±0.174,P<0.05),3h时牵拉组PEDF mRNA(1.078±0.167)与对照组(0.385±0.207)差异无统计学意义(P>0.05),6h时牵拉组PEDF mRNA(1.295±0.012)与对照组(0.491±0.134)差异无统计学意义(P>0.05),9h时牵拉组PEDF mRNA(1.500±0.104)与对照组(0.122±0.040)差异无统计学意义(P>0.05).结论 采用皮肤软组织扩张器实施扩张时,组织微环境内缺氧及机械应力因素可分别显著上调皮肤表皮细胞PEDF的表达.