CAJ | 학술논문

背景：研究证实YY1主要在小鼠基底层未分化表皮细胞中表达，且随着表皮细胞向基底上层分化其表达逐渐下降，这种差异性表达方式说明YY1可能是表皮细胞分化过程中重要的调节因子之一。 　　目的：采用慢病毒-YY1感染HaCaT细胞观察YY1过表达对细胞分化的影响。 　　方法：将慢病毒-YY1感染至HaCaT细胞，经puromycin筛选，建立单克隆稳定细胞株，设对照组为慢病毒感染的HaCaT细胞和未感染的HaCaT细胞，Western blot检测分析YY1蛋白的表达水平；将慢病毒-YY1-HaCaT组和 HaCaT-YY1组细胞各分成2组，一组在低钙(0.12 mmol/L)培养基条件下培养48 h，另一组在低钙(0.12 mmol/L)培养基条件下培养24 h后在高钙(0.35 mmol/L)培养基条件下再培养24 h，用Western blot法检测YY1过表达对HaCaT细胞分化中的表皮细胞特异性分化角蛋白K1、K10、K14和晚期分化产物外皮蛋白、中间丝相关蛋白、兜甲蛋白的影响。 　　结果与结论：慢病毒-YY1成功感染HaCaT细胞，高钙条件下获得的单克隆细胞株过表达YY1蛋白，可抑制K1、外皮蛋白和兜甲蛋白的合成，从而阻止表皮细胞角质化进程并使细胞处于未分化状态。结果说明慢病毒可高效感染皮肤人永生化表皮细胞株 HaCaT 细胞，YY1可能是抑制基底表皮细胞分化并维持其未分化增殖状态的重要因子之一。
배경：연구증실YY1주요재소서기저층미분화표피세포중표체，차수착표피세포향기저상층분화기표체축점하강，저충차이성표체방식설명YY1가능시표피세포분화과정중중요적조절인자지일。 　　목적：채용만병독-YY1감염HaCaT세포관찰YY1과표체대세포분화적영향。 　　방법：장만병독-YY1감염지HaCaT세포，경puromycin사선，건립단극륭은정세포주，설대조조위만병독감염적HaCaT세포화미감염적HaCaT세포，Western blot검측분석YY1단백적표체수평；장만병독-YY1-HaCaT조화 HaCaT-YY1조세포각분성2조，일조재저개(0.12 mmol/L)배양기조건하배양48 h，령일조재저개(0.12 mmol/L)배양기조건하배양24 h후재고개(0.35 mmol/L)배양기조건하재배양24 h，용Western blot법검측YY1과표체대HaCaT세포분화중적표피세포특이성분화각단백K1、K10、K14화만기분화산물외피단백、중간사상관단백、두갑단백적영향。 　　결과여결론：만병독-YY1성공감염HaCaT세포，고개조건하획득적단극륭세포주과표체YY1단백，가억제K1、외피단백화두갑단백적합성，종이조지표피세포각질화진정병사세포처우미분화상태。결과설명만병독가고효감염피부인영생화표피세포주 HaCaT 세포，YY1가능시억제기저표피세포분화병유지기미분화증식상태적중요인자지일。
BACKGROUND:YY1 is mainly expressed in the undifferentiated epidermic cells in mouse basal lamina, and the expression level is gradual y down-regulated as the differentiation towards suprabasal lamina. The differential expression indicates that, YY1 is one of the regulators in the process of epidermic cells differentiation. OBJECTIVE:To observe the effects of YY1 over-expression on the differentiation of HaCaT cells infected with lentivirus. METHODS:Lentivirus-YY1 was transferred into the HaCaT cells by using Lipofectamine 2000. After selection of the puromycin, monoclonal celllines were established, and the control group were lentivirus-infected HaCaT cells and uninfected HaCaT cells. The expression of YY1 was detected by using western blot analysis. cells in Lentivirus-YY1-HaCaT group and HaCaT-YY1 group were further divided into two subgroups according to the calcium concentration in culture medium, cells were either cultured in low-calcium medium (0.12 mmol/L) for 48 hours, or cultured in low-calcium medium (0.12 mmol/L) for 24 hours and in high-calcium medium (0.35 mmol/L) for additional 24 hours. Keratin K1, K10, K14, and involucrin, filaggrin and loricrin after over-expression of YY1 were detected with western blot analysis. RESULTS AND CONCLUSION:The HaCaT cells were successful y infected with lentivirus-YY1, and we obtained over-expression of YY1 protein in monoclonal celllines under high-calcium concentrations, the over-expressed YY1 could decrease the expression of K1, involucrin and loricrin, thereby preventing the process of epidermal keratinocytes and maintaining the cells in an undifferentiated state. Lentivirus can efficiently infect human immortalized epidermal cellHaCaT, and YY1 may the important factor of inhibiting the differentiation of basal epidermal cells and maintaining the undifferentiated proliferation status.