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目的探讨人羊膜上皮细胞(HAEC)损伤后能否在体外构建的纤维蛋白支架上修复,以及表皮生长因子(EGF)、碱性成纤维细胞生长因子(bFGF)和转化生长因子β1(TGF-β1)对HAEC增殖的影响。方法采用环钻钻切培养板上的HAEC建立定量损伤模型,用制备的纤维蛋白块覆盖环钻钻切范围,分别加入不同浓度的培养液进行培养。其中加入EGF为EGF组、加入bFGF为bFGF组、加入TGF-β1为TGF-β1组,不加任何细胞生长因子作为对照组。显微镜下观察各组HAEC生长移行情况,5-溴脱氧尿嘧啶掺入法检测HAEC的增殖效应。结果(1)各组HAEC均能向环钻缺损处移行并向内生长,EGF组、bFGF组的HAEC移行速度快,细胞数量多,对照组次之,TGF-β1组最少。(2)EGF组不同浓度的EGF(1·0、5·0、10·0、20·0、40·0、80·0及160·0ng/ml)培养下,HAEC细胞增殖率分别为17·8%、28·0%、35·3%、51·6%、34·1%、34·2%及26·0%,EGF组10~80ng/ml的HAEC增殖效应均明显大于对照组(17·1%,P<0·05)。(3)bFGF组不同浓度的bFGF(1·0、5·0、10·0、20·0、40·0、80·0及160·0ng/ml)培养下,HAEC细胞增殖率分别为18·0%、35·7%、43·0%、52·7%、67·4%、43·6%及30·5%。bFGF组5~80ng/ml的HAEC增殖效应均明显大于对照组(P<0·05)。其中40ng/ml时的细胞增殖率最高(P<0·05)。(4)TGF-β1组不同浓度的TGF-β1(0·1、0·2、0·4、0·8、1·6、3·2、6·4及12·8ng/ml)培养下,HAEC细胞增殖率分别为17·1%、15·1%、9·3%、6·2%、4·8%、3·6%、2·0%、1·2%。TGF-β1组0·8~12·8ng/ml的HAEC增殖效应明显小于对照组(P<0·05)。结论通过纤维蛋白支架,HAEC能移行修复缺损部位。EGF、bFGF能促进HAEC的增殖,而TGF-β1则抑制HAEC的增殖。
Objective To investigate whether human epithelial cells (HAEC) can repair fibrin scaffolds in vitro and the effects of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and transforming growth factor β1 (TGF-β1) ) On the proliferation of HAEC. METHODS: HAEC was used to establish the model of quantitative injury by drilling and cutting the HAEC. The prepared fibrin block was used to cover the range of drilling and drilling. Different concentrations of culture medium were added for culture. EGF was added to EGF group, bFGF group was added to bFGF group, TGF-β1 was added to TGF-β1 group, without any cell growth factor as control group. The growth and migration of HAEC in each group were observed under the microscope, and the proliferative effect of HAEC was detected by 5-bromodeoxyuridine incorporation method. Results (1) HAECs in each group migrated to ingot defects and grew inwards. HAEC in EGF group and bFGF group migrated faster and had more cells than control group, followed by TGF-β1 group. (2) The proliferation rates of HAEC cells cultured in different concentrations of EGF (1.0, 5.0, 10.0, 20.0, 40.0, 80.0 and 160.0 ng / ml) in EGF group were 17 The proliferative effects of HAECs of 8-80%, 28.0%, 35.3%, 51.6%, 34.1%, 34.2% and 26.0% in EGF group were significantly higher than those in control group (17.1%, P <0.05). (3) Under the different concentrations of bFGF (1.0, 5.0, 10.0, 20.0, 40.0, 80.0 and 160.0 ng / ml) in bFGF group, the proliferation rates of HAEC cells were 18 · 0%, 35.7%, 43.0%, 52.7%, 67.4%, 43.6% and 30.5% respectively. The proliferative effects of 5 ~ 80ng / ml HAEC in bFGF group were significantly greater than those in control group (P <0.05). Among them, the cell proliferation rate was highest at 40ng / ml (P <0.05). (4) The TGF-β1 group at different concentrations of TGF-β1 (0 · 1,0 · 2,0 · 4,0 · 8,1 · 6,3 · 2,6 · 4 and 12 · 8ng / ml) The proliferation rates of HAEC cells were 17.1%, 15.1%, 9.3%, 6.2%, 4.8%, 3.6%, 2.0%, 1.2% respectively. The proliferative effect of HAEC from 0.8 to 12.8 ng / ml in TGF-β1 group was significantly less than that in control group (P <0.05). Conclusion HAEC can migrate to repair the defect by fibrin scaffold. EGF, bFGF can promote the proliferation of HAEC, while TGF-β1 inhibits the proliferation of HAEC.