白细胞介素-1β对类风湿关节炎患者滑膜成纤维细胞中乳腺癌耐药蛋白表达调控的研究

Impact of interleukin-1βon the expression of breast cancer resistance protein in fibroblast-like synoviocytes of patients with rheumatoid arthritis

目的探讨IL-1β对RA患者滑膜成纤维细胞(FLS)中乳腺癌耐药蛋白(BCRP/ABCG2)mRNA表达的影响,探索IL-1β对BCRP介导多药耐药性的调控作用。方法获取RA 3例和OA 3例患者FLS,建立FLS体外培养体系,培养至Ⅴ代;①采用实时荧光定量PCR(RT-qPCR)检测RA与OA的FLS中BCRP mRNA的表达水平并比较;②评估RA病情活动度与BCRP mRNA的表达水平的相关性;③RA-FLS分为4组,A组空白对照组,B组、C组、D组分别加入20、50、100 ng/ml的IL-1β,培养24、48、72 h,比较不同组间BCRP mRNA表达水平。定量资料采用M(P25,P75)表示,两独立样本比较采用Mann-Whitney U检验,组间比较采用Friedman检验。结果①RA-FLS中BCRP mRNA表达水平高于OA组[0.68(0.26,1.16)与0.06(0.05,0.53);Z=-2.136,P=0.029]。②RA-FLS的BCRP mRNA的表达量与病程、ESR、CRP及DAS28呈正相关趋势,而与发病年龄呈负相关趋势,与性别无相关性。③相同浓度IL-1β刺激不同时间RA-FLS的BCRP mRNA的表达水平比较:IL-1β浓度为0时,48 h和72 h较24 h BCRP mRNA的表达水平均升高(χ2=6.00,P=0.034);IL-1β浓度为20 ng/ml(χ2=8.00,P=0.014)或100 ng/ml(χ2=6.50,P=0.040)时,72 h组BCRP mRNA的表达水平较24 h组明显升高。④不同浓度IL-1β刺激相同时间RA-FLS的BCRP mRNA的表达水平比较:72 h时,100 ng/ml组较0 ng/ml组BCRP mRNA表达升高(χ2=12.00,P=0.006)。结论RA-FLS存在BCRP介导的耐药性,且可能与病情活动度有关。炎性细胞因子IL-1β参与调控RA-FLS的BCRP mRNA的表达,且存在时间和浓度依赖性。

Objective To explore the effects of different concentrations or time points of interleukin(IL)-1βon the expression of breast cancer resistance protein(BCRP)mRNA in fibroblast-like synoviocytes(FLS)of rheumatoid arthritis(RA)patients.Methods Synovium tissue of RA(n=3)and osteoarthritis(OA,n=3)patients was obtained through osteoarticular surgery and were used to establish FLS culture system in vitro.FLS cells in generation V were used the following purposes.①Comparing the expression levels of BCRP mRNA in RA-FLS and OA-FLS by real time quantitative polymerase chain reaction(RT-qPCR).②Analyzing the correlation between the activity of the disease and the expression level of BCRP mRNA.③FLS cells were divided into A,B,C,D four groups:Group A was the control group.Group B,C,D were cocultured with low(20 ng/ml),moderate(50 ng/ml),high(100 ng/ml)concentrations of IL-1βfor 24 h,48 h,and 72 h respectively.The expression levels of BCRP mRNA in different groups were compared.Results①The expression of BCRP mRNA in FLS of RA patients was significantly higher than that in the OA group[0.68(0.26,1.16)vs 0.06(0.05,0.53);Z=-2.136,P=0.029].②The expression of BCRP mRNA in FLS of RA patients had a positive association with the course of disease,erythrocyte sedimentation rate(ESR),C-reactive protein(CRP)and disease activity score based on 28 joint counts(DAS28),and a negative association with the age of onset,but had no correlation with gender.③While the concentration of IL-1βwas 0 ng/ml,the expression of BCRP mRNA in the 48 h and 72 h groups was higher than that in the 24 h group(χ2=6.00,P=0.034).Under the condition,when the concentration of IL-1βwas 20 ng/ml(χ2=8.00,P=0.014)or 100 ng/ml(χ2=6.50,P=0.040),the expression of BCRP mRNA in the 72 h group was significantly higher than that in the 24 hours group.④When being stimulated for 72 hours,the expression of BCRP mRNA in the 100 ng/ml group was higher than that in the 0 ng/ml group(χ2=12.00,P=0.006).Conclusion Multidrug resistance mediated by BCRP exists in RA-FLS and may be associated with disease activity.IL-1βcould regulate the expression of BCRP mRNA in FLS cells of RA patients,which may be time and concentration-dependent.