The expression and regulation of nitric oxide synthase in human osteoarthritis-affected chondrocytes: evidence for up-regulated neuronal nitric oxide synthase.
The expression and regulation of nitric oxide synthase in human osteoarthritis-affected chondrocytes: evidence for up-regulated neuronal nitric oxide synthase.
复制标题
DOI：
10.1084/jem.182.6.2097
复制
发表时间：
1995-12-01
期刊：
影响因子：
--
通讯作者：
Abramson SB
中科院分区：
文献类型：
--
作者：
Amin AR;Di Cesare PE;Vyas P;Attur M;Tzeng E;Billiar TR;Stuchin SA;Abramson SB
传统上，骨关节炎（OA）被认为是一种非炎症性疾病。然而，在没有显著炎性细胞浸润的情况下，在骨关节炎液中检测选定的炎性介质越来越受到重视。我们试图确定人OA影响的软骨中可能参与软骨损伤/破坏的炎症组分。使用蛋白质印迹分析和一氧化氮合酶（NOS）保守区的抗体，我们已经观察到上调的NOS，炎症的“关键球员”之一，在OA影响患者的软骨细胞。值得注意的是，从正常关节检查的软骨样品中没有一个显示出可检测到的NOS量。使用相同的α-NOS抗体的Western印迹分析表明，这种NOS从OA影响的软骨（OA-NOS）是更大的大小比（和不同）转染的人肝细胞或小鼠诱导型NOS（iNOS）（150对133 kD）和类似的大小神经元组成型NOS（ncNOS）。iNOS特异性抗体显示与鼠和人iNOS结合，但不与OA-NOS、内皮组成型NOS或ncNOS结合。ncNOS特异性抗体结合ncNOS和OA-NOS，但不结合鼠或人iNOS或内皮组成型NOS。OA软骨在无血清培养基中孵育72 h后，可自发释放大量的亚硝酸盐（高达约80 μ M/100 mg湿组织），其可被各种iNOS抑制剂抑制至少80%，包括蛋白质合成和转录因子NF-κ B的抑制剂，但其（与鼠巨噬细胞iNOS不同）对氢化可的松或TGF-β不敏感。将受OA影响的软骨暴露于白细胞介素1 β、肿瘤坏死因子-α和脂多糖导致亚硝酸盐积累增加约20-50%，这对放线菌酮和吡咯烷二硫代氨基甲酸酯也敏感。因此，我们的数据表明，OA-NOS（基于免疫反应性和分子量）是类似的ncNOS，它释放一氧化氮，这可能有助于炎症和OA软骨破坏的发病机制。
Classically, osteoarthritis (OA) has been considered a noninflammatory disease. However, the detection of selected inflammatory mediators in osteoarthritic fluid, in the absence of significant inflammatory cell infiltrate, is increasingly appreciated. We sought to identify the inflammatory component in human OA-affected cartilage that may be involved in cartilage damage/destruction. Using Western blot analysis and an antibody to the conserved region of nitric oxide synthase (NOS), we have observed up-regulation of NOS, one of the "key players" of inflammation, in chondrocytes of OA-affected patients. Remarkably, none of the cartilage samples examined from normal joints demonstrated detectable amounts of this NOS. Western blot analysis using the same alpha-NOS antibody indicated that this NOS from OA-affected cartilage (OA-NOS) was larger in size than (and distinct from) transfected human hepatocyte or murine inducible NOS (iNOS) (150 versus 133 kD) and similar in size to neuronal constitutive NOS (ncNOS). Antibodies specific for iNOS showed binding to murine and human iNOS but not to OA- NOS, endothelial constitutive NOS, or ncNOS. Antibodies specific for ncNOS bound to ncNOS and also to OA-NOS, but not to murine or human iNOS or endothelial constitutive NOS. Incubation of OA cartilage in serum-free medium resulted in spontaneous release, for up to 72 h, of substantial amounts of nitrite (up to approximately 80 microM/100 mg wet tissue), which could be inhibited by at least 80% with various inhibitors of iNOS, including inhibitors of protein synthesis and transcription factor NF-kappa B, but which (unlike murine macrophage iNOS) was not sensitive to hydrocortisone or TGF-beta. Exposure of OA- affected cartilage to interleukin 1 beta, tumor necrosis factor-alpha, and lipopolysaccharide resulted in approximately 20-50% augmentation of nitrite accumulation, which was also sensitive to cycloheximide and pyrrolidine dithiocarbamate. Hence, our data indicate that OA-NOS (based on immunoreactivity and molecular weight) is similar to ncNOS and that it releases nitric oxide, which may contribute to the inflammation and pathogenesis of cartilage destruction in OA.