Hypoxia-induced Abrogation of Contact-dependent Inhibition of Rheumatoid Arthritis Synovial Fibroblast Proliferation
Hypoxia-induced Abrogation of Contact-dependent Inhibition of Rheumatoid Arthritis Synovial Fibroblast Proliferation
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DOI：
10.3899/jrheum.080188
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发表时间：
2009-04-01
影响因子：
3.9
通讯作者：
Kohsaka, Hitoshi
中科院分区：
文献类型：
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作者：
Nonomura, Yoshinori;Mizoguchi, Fumitaka;Kohsaka, Hitoshi
Objective.滑膜成纤维细胞的不受控制的增殖是类风湿性关节炎（RA）的病理特征。由于类风湿关节的滑膜组织是缺氧的，我们研究了缺氧如何影响RA滑膜成纤维细胞（RASF）的增殖。在低氧（10%、3%或1%O-2）或常氧（21%O-2）条件下，以每微量滴定孔2000个细胞（低密度培养物）或5000个细胞（高密度，生长抑制汇合培养物）培养RASF。用重组人白细胞介素1受体拮抗剂（IL-1 ra）、抗肿瘤坏死因子-α（TNF-α）中和抗体、抗N-钙粘蛋白阻断抗体治疗RASF。或MG 132。H-3标记的胸苷掺入定量评估其增殖。制备总RNA和细胞裂解物用于实时聚合酶链反应和Western印迹分析。低氧对低密度培养的RASF增殖无明显影响。在高密度下，它废除了RASF的接触依赖性生长抑制，但不是人真皮成纤维细胞。添加抗TNF-α抗体或IL-1 ra不会影响结果。在常氧条件下高密度培养的细胞中观察到细胞周期蛋白依赖性激酶抑制剂p27（Kip 1）的表达上调，但在缺氧条件下则没有。缺氧降低RASE上的N-钙粘蛋白表达。抗N-钙粘蛋白阻断抗体的加入模拟了缺氧培养的作用。在常氧条件下促进高密度培养的RASF增殖。这种抗体处理也下调了p27（Kip 1）的表达。缺氧下调RASF上的N-钙粘蛋白表达，从而阻止p27（Kip 1）上调其接触抑制。类风湿性滑膜组织中的缺氧可能通过增加滑膜成纤维细胞的增殖而促成类风湿性病理。（2009年2月15日首次发布：J Rheumol 2009;36：698-705; doi：10.3899/jrheum.080188）
Objective. Uncontrolled proliferation of synovial fibroblasts is characteristic of the pathology of rheumatoid arthritis (RA). Since synovial tissues in the rheumatoid joints are hypoxic, we investigated how hypoxia affects RA synovial fibroblast (RASF) proliferation.Methods. RASF were cultured at 2000 cells (low density culture) or at 5000 cells (high density, Growth-inhibitory confluent Culture) per microtiter well under hypoxic (10%, 3%, or 1% O-2) or normoxic (21% O-2) conditions. Some RASF were treated with recombinant human interleukin 1 receptor antagonist (IL-1ra), anti-tumor necrosis factor-alpha (TNF-alpha)-neutralizing antibodies, anti-N-cadherin-blocking antibodies. or MG132. H-3-labeled thymidine incorporation was quantified to assess their proliferation. Total RNA and cell lysates were prepared for real-time polymerase chain reaction and Western blot analyses.Results. Hypoxia exerted no effect on proliferation of RASF cultured at low density. At high density, it abrogated contact-depen dent growth inhibition of RASF, but not of human dermal fibroblasts. Addition of anti-TNF-alpha antibodies or IL-1ra did not affect the results. Upregulated expression of cyclin-dependent kinase inhibitor p27(Kip1) was observed in the cells cultured at high density under normoxic conditions, but not under hypoxic conditions. Hypoxia decreased N-cadherin expression on RASE Addition of anti-N-cadherin-blocking antibodies mimicked the effects of hypoxic culture. it promoted proliferation of RASF cultured at high density under normoxic conditions. This antibody treatment also downmodulated p27(Kip1) expression.Conclusion. Hypoxia downregulates N-cadherin expression on RASF, and thus prevents p27(Kip1) upregulation for their contact inhibition. It is likely that hypoxia in rheumatoid synovial tissues contributes to rheumatoid pathology by augmenting proliferation of synovial fibroblasts. (First Release Feb 15 2009: J Rheumatol 2009;36:698-705; doi: 10.3899/jrheum.080188)