摘要
目的探讨热休克蛋白70（HSP70）对寻常性银屑病皮损中成纤维细胞白介素6（IL-6）表达的影响。方法原代培养出寻常性银屑病皮损中的成纤维细胞（PFb），然后在PFb培养基中加入5～30mg／L的HSP70以及50Ixmol／L的核因子KB特异性抑制剂吡咯啉烷二甲基硫脲（PDTC），并孵育不同时间（0～48h）。以正常生长的PFb为对照组。用酶联免疫吸附法（ELISA）检测PFb培养上清液中IL-6的含量，用半定量逆转录-聚合酶链反应（RT—PCR）检测PFbIL-6mRNA表达。运用t检验和Dunnett—t检验进行统计分析。结果HSP70呈时间依赖（0～48h）和剂量依赖（5—30mg／L）性增加PFbIL-6蛋白合成和mRNA表达。10mg／LHSP70实验组IL-6蛋白[（75．2±15．4）ng／L]和IL-6mRNA表达水平（0．439±0．093）均显著高于对照组[分别为（47．2±10．6）mg／L和0．249±0．069]，两组比较，差异均有统计学意义（P〈0．05）。30mg／LHSP70实验组培养PFb12h时IL-6蛋白开始明显增多；培养6h时IL-6mRNA开始明显增多，与对照组相比，差异均有统计学意义（P〈0．05）。随着HSP70浓度的增加和PFb培育时间的延长，HSP70实验组与对照组IL-6蛋白和IL-6mRNA的表达差异逐渐加大。30mg／LHSP70＋PDTC组IL-6蛋白（42．23±9．41n扎）和IL-6mRNA表达（0．144±0．048）均显著低于对照组（分别为68．40±14．43ng／L和0．295±0．081），两组比较，差异均有统计学意义（P〈0．05）。结论HSP70能增强核因子KB介导的PFbIL-6蛋白和IL-6mRNA表达，可能是诱导PFb高水平IL-6表达的因素之一。
Objective To evaluate the in vitro effect of heat shock protein 70 （HSP70） on interleukin-6 （IL-6） expression by cultured fibroblasts from psoriasis vulgaris lesions （PFbs）. Methods Fibroblasts were isolated from the lesions of patients with psoriasis vulgaris and subjected to a primary＇ culture. After 3 to 5 passages of culture, the fibroblasts were collected and used in the next experiment. Some PFbs were cultured with different concentrations （5, 10, 20, 30 mg/L） of HSP70 for 48 hours, or with HSP70 of 30 mg/L for different durations （3, 6, 12, 24, 48, 72 hours）; some PFbs were incubated with HSP70 of 30 mg/L for 24 hours after pretreatment with pyrrolidine dithiocarbamate （PDTC, a specific inhibitor of nuclear factor-kappa B） for 30 minutes. PFbs receiving no treatment served as the control. Enzyme-linked immunosorbent assay （ELISA） and semi-quantitative reverse transcription PCR were performed to measure the IL-6 protein expression in culture supernatant and IL-6 mRNA expression by PFbs, respectively. Differences in the expression of IL-6 protein and mRNA between PFbs receiving different treatment were analyzed by using t test and Dunnett＇s t test. Results HSP70 significantly increased both protein production and mRNA expression of IL-6 in a time （0-48 h）- and dose （5-30 mg/L）-dependent manner. The expression levels of supernatant IL-6 protein and IL-6 mRNA were significantly higher in the PFbs treated with HSP70 of 10 mg/L for 48 hours than untreated PFbs （（75.2 ± 15.4） ng/L vs. （47.2 ± 10.6） ng/L, 0.439 ± 0.093 vs. 0.249 ± 0.069, both P 〈 0.05）. A significant increase was observed as early as 6 hours in the level of IL-6 mRNA after the treatment with HSP70 of 30 mg/L, and 12 hours in the level of supernatant IL-6 protein. Decreased supernatant IL-6 protein and IL-6 mRNA were noted for PFbs treated with PDTC and HSP70 of 30 mg/L compared with untreated PFbs （ （42.23 ± 9.41） ng/L vs. （68.40 ± 14.43） ng/L, 0.144 ± 0.048 vs. 0.295 ± 0.081, both P 〈 0.05）. Conclusion HSP70 may increase the expression of IL-6 mRNA and protein by cultured PFbs via the nuclear factor-kappa B pathway.
出处
《中华皮肤科杂志》
CAS
CSCD
北大核心
2012年第11期792-795,共4页
Chinese Journal of Dermatology
基金
陕西省“13115”重大科技专项基金
