摘要
目的 探讨他克莫司对TNF-α刺激的HacaT细胞核因子-κB（NF-κB）表达的影响,以及对HaCaT细胞糖皮质激素受体（GR）α和β表达的影响.方法 采用Western印迹和免疫荧光-激光共聚焦显微镜技术观察：①以TNF-α（10μg/L）单独或TNF-α（10μg/L）与他克莫司（10^-8mol/L、10^-7mol/L、10^-6mol/L）同时作用HaCaT细胞,在24h和48h分别观察胞核NF-κB的表达;②以他克莫司（10^-8mol/L、10^-7mol/L、10^-6mol/L）作用HaCaT细胞,在12h、24h和36h分别观察GRα和GRβ的表达.结果 未处理组HaCaT细胞的胞核有NF-κB（p65）蛋白的表达,灰度值比值为0.4988±0.03506;TNF-α作用24h和48h后,HaCaT细胞胞核NF-κB（p65）表达随时间延长而增强,灰度值比值分别为0.73±0.0316和0.8925±0.0171,与未处理组相比较.差异均有统计学意义（P〈0.05）.TNF-α与10^-8 mol/L、10^-7 mol/L、10^-6mol/L他克莫司同时作用组HaCaT细胞胞核NF-κB表达灰度值比值分别为0.6825±0.0263、0.6200±0.0163和0.5575±0.0299,NF-κB表达随他克莫司浓度增加而减弱,后两组与TNF-α单独作用组相比,差异均有统计学意义（P〈0.05）.各浓度他克莫司作用24h和48h的NF-κB表达差异均无统计学意义（P〉0.05）.与未处理组相比,不同浓度他克莫司作用HaCaT细胞后,各时相点GRα和GRβ的表达差异均无统计学意义（P〉0.05）.结论 他克莫司能以浓度依赖方式抑制TNF-α刺激的角质形成细胞胞核NF-κB的表达,不影响角质形成细胞GRα和GRβ的表达.
Objective To investigate the effect of tacrolimus on the expression of nuclear factor-κB （NF-κB） in HaCaT cells stimulated by tumor necrosis factor-α（TNF-α）,and on the expression of glucocorticoid receptor （GR）α and β in untreated HaCaT cells in vitro.Methods Cultured Ha CaT cells were treated with TNF-α（10μg/L） only,combination of TNF-α（10μg/L） and various concentrations （10^-8mol/L, 10^-7mol/L,10^-6moL/L） of tacrolimus or tacrolimus of different concentrations only.After additional 12-,24-, 36- or 48-hour cnlture, Westem blot and immunofluorescenee-confocal laser scanning microscopy were used to detect the expressions of NF-κB,GRα and GRβ in HaCaT cells.Those untreated HaCaT cells served as the control.Results The relative protein expression level of NF-κB was increased in HaCaT cells after treatment with TNF-α for 24 and 48 hours zompared with untreated ceils （0.73±0.0316 and 0.8925±0.0171 vs 0.4988±0.03506,both P〈0.05）;however,the increase in NF-κB expression was inhibited by the combination treatment with tacrolimus,and the relative expression level of NF-κB protein was 0.6825±0.0263.0.6200±0.0163 and 0.5575±0.0299 in HaCaT cells treated with TNF-α plus tacrolimus of 10^-8mol/L 10^-7mol/L and 10^-6mol/L,respectively;the difference was significant etween TNF-α-treated cells and those dealt with the combination of NF-α and tacrolimus of 10^-7 or 10^-6 mol/L （both P〈0.05）.No significant difference was observed in the expression of NF-κB by HaCaT cells between different time oints treated with tacrolimus of 10^-8,10^-7 or 10^-6 mol/L.Also,there was no zignificant difference in the expression of GRα or GRβ between untreated HaCaT cells and those treated with tacrolimus of 10^-8, 10^-7 or 10^-6 mol/L at any time point.Conclusions Tacrolimus ould inhibit the expression of NF-κB by TNF-α-stimulated HaCaT cells,but does not affect the expression of GRα or GRβ,in untreated HaCaT cells.
出处
《中华皮肤科杂志》
CAS
CSCD
北大核心
2010年第6期385-389,共5页
Chinese Journal of Dermatology
基金
第三军医大学第二附属医院1520人才基金（2006-188）
