Toll样受体2对人角质形成细胞增殖的影响  

Effect of Toll-like receptor 2 on the proliferation of human keratinoeytes

作　　者：陆小年[1] 张勇[1] 韩凌[1] 祝禄川[1] 郑志忠[1] 

机构地区：[1]复旦大学附属华山医院皮肤科,上海200040

出　　处：《中华皮肤科杂志》2009年第6期409-412,共4页Chinese Journal of Dermatology

摘　　要：目的探讨Toll样受体2（TLR2）对体外培养人角质形成细胞增殖的影响。方法天然配体肽聚糖（PGN）体外活化人角质形成细胞TLR2，用噻唑蓝法检测PGN对角质形成细胞体外增殖的影响，确定最适作用浓度；用实时荧光定量PCR法和Western印迹法分别检测角质形成细胞Ki67、TLR2、核因子（NF）-κBp65及转化生长因子（TGF）-α mRNA和蛋白的表达水平；采用抗体封闭实验分析封闭TLR2对PGN诱导角质形成细胞Ki67、TLR2、NF—κBp65及TGF-α表达的影响。结果不同浓度的PGN处理角质形成细胞后24h，在1．25，2．5，5μg／mL的浓度下细胞较对照组出现明显增殖（P〈0．05）。1．25、2．5和5μg／mL PGN作用于角质形成细胞后24h，其中1．25和2．5μg／mL浓度下Ki67 mRNA的表达明显增加，各组Ki67蛋白合成均有增加；各组TLR2 mRNA和蛋白的表达均增加；1．25μg／mL浓度下NF—κBp65 mRNA的表达明显增加，各组细胞核内NF—κBp65蛋白合成均增加；其中1．25和5μg／mL浓度下TGF—α mRNA和蛋白表达均明显增加（P〈0．05）。以抗人TLR2单克隆阻断性抗体封闭TLR2后，PGN诱导的角质形成细胞Ki67、TLR2、NF—κBp65及TGF—α mRNA和蛋白表达的上调均受到明显抑制（P〈0．05）。结论角质形成细胞TLR2经PGN诱导活化后，可能通过促进NF—κB活化及TGF—α表达而导致角质形成细胞的异常增殖。Objective To investigate the effect of Toll-like receptor 2 （TLR2） on the proliferation of human keratinocytes. Methods Keratinocytes were isolated from the foreskin of children, and subjected to primary culture. After 3-5 passages, the keratinocytes were incubated with various concentrations of peptidoglycan （PGN）, a TLR2 agonist. Cell proliferation was detected by MTT colorimetric assay and the suitable concentrations of PGN were determined. The mRNA and protein expressions of Ki67, TLR2, NF-κB p65 and TGF-α were detected by real-time quantitative PCR and Westem blot, respectively, in keratinocytes treated with PGN of 0, 1.25, 2.5 and 5 μg/mL. Antibody blocking test was utilized to evaluate the effect of blocking TLR2 with specific anti-TLR2 neutralizing monoclonal antibody before incubation with PGN on the expressions of Ki67, TLR2, NF-κB p65 and TGF-α by keratinocytes. Results The proliferation of keratinocytes was significantly promoted by the incubation with PGN of 1.25, 2.5 and 5 μg/mL for 24 hours （all P 〈 0.05 ）, which also increased the expression of Ki67 protein, TLR2 mRNA and protein, and NF-κB p65 protein. Further more, the mRNA expression of Ki67 in keratinocytes was elevated by PGN of 1.25 and 2.5 μg/mL, the mRNA expression of NF-κB p65 elevated by PGN of 1.25 μg/mL, and the expressions of TGF-α protein and mRNA elevated by PGN of 1.25 and 5 μg/mL （P 〈 0.05）. The mRNA and protein expressions of Ki67, TLR2, NF-κB p65 and TGF-α were all inhibited by the blocking of TLR2 before incubation with PGN （all P 〈 0.05）. Conclusion Activation of TLR2 by PGN could induce the over-proliferation of human keratinocytes, likely through promoting NF-κB activation and TGF-α expression.

关 键 词：TOLL样受体2 角蛋白细胞 细胞增殖 NF—κB 转化生长因子Α 

分 类 号：R751[医药卫生—皮肤病学与性病学]