一氧化氮合酶抑制剂对炎性刺激下软骨细胞增殖和基质代谢的影响

Effects of NO synthase inhibitors on proliferation and matrix metabolism of chondrocytes and cartilage entities induced by the inflammatory environment

目的 用白细胞介素 1β(IL 1β)和脂多糖 (LPS)模拟软骨细胞和软骨的炎性环境 ,探讨一氧化氮合酶 (NOS)抑制剂对软骨细胞增殖和基质代谢的影响。方法 实验共分 5组 :对照组、IL 1β和LPS刺激组、IL 1β和LPS +S 甲基异硫脲 (SMT)组、IL 1β和LPS +L NIL组、IL 1β和LPS +L NMA组。清洗干净的软骨块和传至第二代的软骨细胞依次加入上述药物。采用MTT法和BrdU掺入法分别检测软骨细胞和软骨的增殖活性 ;化学比色法检测NO释放量、NOS活性 ;3 5S掺入法检测蛋白多糖合成率。结果 ①外源性IL 1β和LPS可增加软骨细胞和软骨NO释放 [(2 0 4 5± 14 )μmol/L ,(2 0 9 8± 18) μmol/L],诱导iNOSmRNA表达 ,增加NOS活性 [(2 0 1 7± 17)U/ml,(2 98 7±2 0 )U/ml) ],抑制软骨蛋白多糖合成和软骨细胞增殖 (P <0 0 5 ) ;②不同浓度NOS抑制剂均能减少NO释放和降低NOS活性 ,并与药物剂量正相关 ;③ 1mmol/L的NOS抑制剂可显著逆转软骨细胞和软骨蛋白多糖合成抑制 (P <0 0 5 ) ,增加软骨细胞增殖活性。结论 NOS抑制剂能有效抑制炎性因子 (IL 1β和LPS)诱发的软骨代谢改变 ,对关节软骨损害具有潜在的保护作用。

Objective To induce inflammatory environment with IL 1β and Lipopolysaccharides (LPS) in rabbit chondrocytes and cartilage entities, and study effects of NO synthase(NOS) inhibitors on proliferation and matrix metabolism of chondrocytes and cartilage entities. Methods The experiments were performed in 5 groups: control group; IL 1β and LPS group; IL 1β and LPS+SMT group; IL 1β and LPS+ L NIL group; IL 1β and LPS+ L NMA group. The release of NO and the activity of NOS were measured by spectrophotometric methods. MTT and BrdU assay was employed to observe the proliferation of chondrocytes. The proteoglycan synthesis was assessed by incorporation of radiolabelled sodium sulphate Na 35 2SO 4. Results ①IL 1β and LPS induced iNOS mRNA expression in rabbit chondrocytes and cartilage entities,and increased NO release[(204 5±14) μmol/L, (209 8±18) μmol/L] and NOS activity[(201 7±17) U/ml, (298 7±20) U/ml)],and inhibit cell proliferation and proteoglycan synthesis. ②NOS inhibitors in different concentration all reduced NO release and NOS activity in chondrocytes and cartilage entities.③ 1 mmol/L NOS inhibitors resulted in a partial recovery of proteoglycan synthesis and cell proliferation in chondrocytes and cartilage entities. Conclusion NOS inhibitors can reduce NO release, inhibit NOS activity, and restore proteoglycan synthesis, so it can protect chondrocytes and cartilage entities.