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目的:探讨不同细胞外基质对胚胎干细胞源性角化细胞生长、增殖及分化的影响。方法:体外培养的人类胚胎干细胞经RA、BMP4、AA诱导后,分别接种到以Matrigel(A组)、Ⅰ型胶原(B组)、Ⅳ型胶原(C组)、明胶(D组)包被的培养皿中培养,E组为无包被培养皿作为空白对照组。通过倒置显微镜观察细胞生长情况;用CCK8法检测细胞的增殖能力;并通过Western Blot检测CK14表达量,测定各组灰度值。结果:A组生长的E-KCs细胞最快到达生长对数期,增殖能力最强,且细胞集落形成率最高,与其他组比较,差异具有统计学意义(P<0.05);A组与正常口腔黏膜组(阳性对照)CK14表达量条带灰度值差异无统计学意义,但与B、C、D组比较差异具有统计学意义(P<0.01)。结论:Matrigel与其他3种细胞外基质相比能显著提高人类胚胎干细胞向角化细胞诱导分化的能力,并能促进E-KCs生长和增殖。
Objective: To investigate the effects of different extracellular matrix on the growth, proliferation and differentiation of embryonic stem cell-derived keratinocytes. Methods: Human embryonic stem cells (ESCs) cultured in vitro were induced by RA, BMP4 and AA, and then inoculated into Matrigel (group A), collagen Ⅰ (group B), collagen Ⅳ (group C) Of the culture dish, group E as a non-petri dish as a blank control group. The growth of cells was observed by inverted microscope. The proliferation of cells was detected by CCK8 method. The expression of CK14 was detected by Western Blot and the gray value of each group was measured. Results: E-KCs cultured in group A reached the logarithmic phase with the fastest proliferative ability and the highest rate of colony formation. Compared with other groups, the difference was statistically significant (P <0.05). In group A, There was no significant difference in gray value of CK14 expression in oral mucosa group (positive control), but there was significant difference compared with B, C, D group (P <0.01). Conclusion: Compared with the other three kinds of extracellular matrix, Matrigel can significantly enhance the ability of human embryonic stem cells to differentiate into keratinocytes and promote the growth and proliferation of E-KCs.