CAJ | 학술논문

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5～30 mg/L的HSP70以及50 μmol/L的核因子κB特异性抑制剂吡咯啉烷二甲基硫脲(PDTC),并孵育不同时间(0 ～ 48 h).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PFb IL-6 mRNA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～ 48 h)和剂量依赖(5～ 30 mg/L)性增加PFb IL-6蛋白合成和mRNA表达.10 mg/L HSP70实验组IL-6蛋白[(75.2±15.4)ng/L]和IL-6 mRNA表达水平(0.439±0.093)均显著高于对照组[分别为(47.2±10.6)ng/L和0.249±0.069],两组比较,差异均有统计学意义(P＜0.05).30 mg/LHSP70实验组培养PFb 12 h时IL-6蛋白开始明显增多;培养6h时IL-6 mRNA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-6 mRNA的表达差异逐渐加大.30 mg/L HSP70+ PDTC组IL-6蛋白(42.23±9.41 ng/L)和IL-6 mRNA表达(0.144±0.048)均显著低于对照组(分别为68.40±14.43 ng/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κB介导的PFb IL-6蛋白和IL-6 mRNA表达,可能是诱导PFb高水平IL-6表达的因素之一.
목적 탐토열휴극단백70(HSP70)대심상성은설병피손중성섬유세포백개소6(IL-6)표체적영향.방법 원대배양출심상성은설병피손중적성섬유세포(PFb),연후재PFb배양기중가입5～30 mg/L적HSP70이급50 μmol/L적핵인자κB특이성억제제필각람완이갑기류뇨(PDTC),병부육불동시간(0 ～ 48 h).이정상생장적PFb위대조조.용매련면역흡부법(ELISA)검측PFb배양상청액중IL-6적함량,용반정량역전록-취합매련반응(RT-PCR)검측PFb IL-6 mRNA표체.운용t검험화Dunnett-t검험진행통계분석.결과 HSP70정시간의뢰(0～ 48 h)화제량의뢰(5～ 30 mg/L)성증가PFb IL-6단백합성화mRNA표체.10 mg/L HSP70실험조IL-6단백[(75.2±15.4)ng/L]화IL-6 mRNA표체수평(0.439±0.093)균현저고우대조조[분별위(47.2±10.6)ng/L화0.249±0.069],량조비교,차이균유통계학의의(P＜0.05).30 mg/LHSP70실험조배양PFb 12 h시IL-6단백개시명현증다;배양6h시IL-6 mRNA개시명현증다,여대조조상비,차이균유통계학의의(P＜0.05).수착HSP70농도적증가화PFb배육시간적연장,HSP70실험조여대조조IL-6단백화IL-6 mRNA적표체차이축점가대.30 mg/L HSP70+ PDTC조IL-6단백(42.23±9.41 ng/L)화IL-6 mRNA표체(0.144±0.048)균현저저우대조조(분별위68.40±14.43 ng/L화0.295±0.081),량조비교,차이균유통계학의의(P＜0.05).결론 HSP70능증강핵인자κB개도적PFb IL-6단백화IL-6 mRNA표체,가능시유도PFb고수평IL-6표체적인소지일.
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 supematant 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 supematant 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.