Bhardwaj N, Santhanam U, Lau L L, Tatter S B, Ghrayeb J, Rivelis M, Steinman R M, Sehgal P B, May L T
Laboratory of Cellular Physiology, Rockefeller University, New York 10021.
J Immunol. 1989 Oct 1;143(7):2153-9.
We have looked for IL-6, a cytokine that has immunomodulating and inflammation-associated activities, in joint exudates (fluid and mononuclear cells) from patients with rheumatoid arthritis and other arthritides using both biologic and biochemical assays. IL-6 was assessed by its ability to stimulate alpha 1-antichymotrypsin secretion from the human hepatoma cell line Hep3B clone 2, an activity which is blocked by an antiserum to Escherichia coli derived IL-6, and by the growth of the IL-6-dependent murine hybridoma 7TD1 cell line. IL-6 isoforms in synovial fluid were characterized by immunoaffinity chromatography followed by Western blotting. The presence of IL-1 in synovial fluids and its production by synovial fluid mononuclear cells was monitored by Western blotting and indirect immunofluorescence with polyclonal anti-IL-1 beta antisera. In an analysis of 30 effusions from 27 rheumatoid patients with acutely inflamed joints, abundant quantities of IL-6 (greater than 2 ng/ml) were detected in 23 by the alpha 1-antichymotrypsin bioassay. Several rheumatoid synovial fluids also had elevated IL-6 levels in the 7TD1 bioassay. Seven of nine nonrheumatoid effusions also contained high levels of IL-6 (greater than 2 ng/ml). No IL-1 (less than 0.25 ng/ml) could be detected by Western blotting in 10 rheumatoid effusions even though eight of these contained high levels of IL-6. The IL-6 activity could be neutralized with a rabbit antiserum to rIL-6. Multiple IL-6 isoforms (25, 30, 45 kDa) were present in two rheumatoid and one traumatic effusion studied. Fresh mononuclear cells isolated from various synovial effusions did not appear to make IL-6 constitutively, as no IL-6 could be detected in the media of cells cultured for 12 to 18 h after isolation. Similarly, there was no constitutive production of IL-1 by these cells. However, synovial fluid mononuclear cells could be induced to secrete both IL-6 and IL-1 after stimulation with LPS. The LPS-responsive cells were monocytes and not lymphocytes or dendritic cells. These findings suggest that IL-6 is involved in inflammatory joint disease. However, the primary cells synthesizing it may be located in the synovial lining instead of the joint exudate.
我们运用生物学和生化检测方法，在类风湿性关节炎及其他关节炎患者的关节渗出液（液体和单核细胞）中寻找白细胞介素-6（IL-6），这是一种具有免疫调节和炎症相关活性的细胞因子。通过检测其刺激人肝癌细胞系Hep3B克隆2分泌α1-抗糜蛋白酶的能力来评估IL-6，该活性可被抗大肠杆菌来源的IL-6抗血清阻断，同时也通过IL-6依赖性小鼠杂交瘤7TD1细胞系的生长情况来评估。通过免疫亲和层析和蛋白质印迹法对滑液中的IL-6亚型进行了鉴定。运用蛋白质印迹法和用多克隆抗IL-1β抗血清进行间接免疫荧光法监测滑液中IL-1的存在及其由滑液单核细胞的产生情况。在对27例患有急性炎症关节的类风湿患者的30份积液进行分析时，通过α1-抗糜蛋白酶生物检测法在23份积液中检测到大量的IL-6（大于2 ng/ml）。在7TD1生物检测法中，几份类风湿滑液中的IL-6水平也有所升高。9份非类风湿性积液中的7份也含有高水平的IL-6（大于2 ng/ml）。在10份类风湿性积液中，通过蛋白质印迹法未检测到IL-1（小于0.25 ng/ml），尽管其中8份含有高水平的IL-6。IL-6活性可用兔抗rIL-6抗血清中和。在所研究的两份类风湿性积液和一份创伤性积液中存在多种IL-6亚型（25、30、45 kDa）。从各种滑液积液中分离出的新鲜单核细胞似乎不会组成性地产生IL-6，因为在分离后培养12至18小时的细胞培养基中未检测到IL-6。同样，这些细胞也不会组成性地产生IL-1。然而，滑液单核细胞在用脂多糖（LPS）刺激后可被诱导分泌IL-6和IL-1。对LPS有反应的细胞是单核细胞，而非淋巴细胞或树突状细胞。这些发现表明IL-6参与了炎症性关节疾病。然而，可以合成IL-6的主要细胞可能位于滑膜衬里而非关节渗出液中。
