摘要
目的探讨小白菊内酯（PAR）对关节软骨细胞蛋白聚糖合成的调节作用。方法软骨细胞取自骨关节炎（OA）患者进行膝关节置换术的股骨髁，随机分为4组：对照组、肿瘤坏死因子-α（TNF-α）组、PAR组和TNF—α＋PAR组。用3种方法测定蛋白聚糖的代谢：（1）测定蛋白聚糖的代谢产物糖胺多糖（GAG）含量。（2）用针对蛋白聚糖单克隆抗体（Mab-3B3和5D4），采用ELISA法检测培养液中蛋白聚糖的代谢片段含量。（3）半定量RT—PCR检测软骨细胞蛋白聚糖、ADAMTS-4、ADAMTS-5、基质金属蛋白酶组织抑制因子-1（TIMP-1）和丝裂原活化蛋白激酶-1（MAPK-1）的mRNA表达。结果（1）TNF—α＋PAR组培养液中GAG的百分含量明显低于TNF—α组（t=5．92，P=0．02），其降低程度与PAR呈剂量依赖性。（2）TNF—α组培养液中5134片段的含量明显高于TNF—α＋PAR组（t=7．93，P=0．016），而各组间培养液中383片段的含量无差别。（3）TNF—α组的蛋白聚糖mRNA表达明显低于其他3组（F=133．7，P=0．000）；而其ADAMTS-5及MAPK-1的mRNA表达明显高于其他3组（F=209．7，117．1；P=0．000）。结论（1）TNF—α可促进入OA关节软骨细胞蛋白聚糖的降解而PAR可抑制其降解作用。（2）PAR可下调TNF—α导致的ADAMTS-5及MAPK-1的mRNA表达。
Objective To investigate the effect of parthenolide （PAR） on the tumor necrosis factor （TNF）—α induced aggrecan catabolism of chondrocytes in ostroarthritis （OA）. Methods Human chondrocytes were obtained from the condyles of femur of OA patients undergoing knee joint replacement during operation, cultured, and randomly divided into 4 groups： control group, TNF group （cultured in medium containing 10 ng/ml TNF—α） , PAR group （cultured in medium containing 10 μmol/L PAR） , and PAR ＋ TNF group （cultured in medium containing 10 μmol/L PAR and 10 ng/ml TNF—α）. Eight days later 1, 9-dimethylmethylene blue spectrophotometric assay was used to measure the content of glycosaminoglycan （GAG） , marker of aggrecan catabolism, in the culture fluid and the cells. Using anti-aggrecan monoclonal antibodies Mab 5D4 and 3B3, ELISA was employed to detect the contents of 5D4 and 3B3, aggrecan catabolic fragments. RT-PCR was used to detect the mRNA expression of the aggrecan, ADAMTS-4 and ADAMTS-5, both aggrecanases, tissue inhibitor of metalloprotease （TIMP）-1, mitogen-activated protein kinase （MAPK）-1 and 18S, a marker. Results The GAG percentage in the culture fluid of the TNF—α was 57. 1% ±2.0%, significantly higher than those of the control and TNF—α ＋ PAR groups （P = 0. 001 and 0. 02）. The 5D4 fragment level of the TNF—α group was 509 ng/ml ± 32 ng/ml, significantly higher than that of the control group （ 166 ng/ml ± 15 ng/ml, t = 11.60,P = 0. 007 ）, and the level of 5 D4 fragment of the PAR ＋ TNF—α group was 333 ng/ml ± 15 ng/ml, significantly lower than that of the TNF—α group（ t = 7.93, P =0. 016）. There was not significant difference in the 3B3 fragment level among the 4 groups （F = 1. 316,P =0. 335）. The aggrecan mRNA expression level of the TNF—αgroup was significantly lower than those of the other 3 groups （ F = 133.7, P = 0. 000）, the mRNA expression levels of ADAMTS-5 and MAPK-1 of the TNF-α group were significantly higher than those of the other 3 groups （ F = 209. 7, 117. 1 ; P =0. 000）, the ADAMTS-5 mRNA expression level of the PAR group was significantly lower than that of the control group （ t = 11.1, P = 0. 008） , and there was not significant differences in the mRNA expression levels of ADAMTS-4and TIMP-1 among the 4 groups （ F = 1.87,0. 73 ; P 〉 0. 05 ）. Conclusion PAR inhibits TNF-α induced catabolism of aggrecan in the chondrocytes of OA and reduces the mRNA expression of ADAMTS-5 and MAPK-1. PAR may be useful in the treatment of OA and other inflammatory joint diseases.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2008年第11期764-768,共5页
National Medical Journal of China
