Tarr Joanna M, Winyard Paul G, Ryan Brent, Harries Lorna W, Haigh Richard, Viner Nick, Eggleton Paul
Peninsula College of Medicine and Dentistry, and University of Exeter, Exeter, UK.
Arthritis Rheum. 2010 Oct;62(10):2919-29. doi: 10.1002/art.27602.
The binding of FasL (CD95L) to its receptor, Fas (CD95), induces apoptosis. Studies have shown that in patients with rheumatoid arthritis (RA), T lymphocytes are resistant to FasL-induced apoptosis in vivo but are susceptible to FasL-induced apoptosis in vitro. Dysfunction in this mechanism may be an important contributor to the pathophysiology of RA. Thus, the present study was undertaken to determine which factors might inhibit FasL-Fas binding in vivo and those that would inhibit apoptosis of T lymphocytes in an in vitro model system.
Human Jurkat T cells rendered apoptotic by FasL exposure were analyzed by flow cytometry. Necrosis was determined according to measurement of lactate dehydrogenase release. Quantification of calreticulin in plasma and synovial fluid and of calreticulin-FasL binding was performed by enzyme-linked immunosorbent assay. Measurement of nitrite/nitrate in the plasma and synovial fluid was carried out by chemiluminescence assay.
Extracellular calreticulin was present at a significantly higher concentration in the plasma (median 10.3 ng/ml, interquartile range [IQR] 14.8 ng/ml) and synovial fluid (median 10.3 ng/ml, IQR 12.0 ng/ml) of RA patients (each P < 0.05) compared with the plasma (median 3.1 ng/ml, IQR 1.3 ng/ml) and synovial fluid (median 2.9 ng/ml, IQR 0.9 ng/ml) of patients with psoriatic arthritis and the plasma of healthy control subjects (median 2.9 ng/ml, IQR 0.9 ng/ml). Calreticulin concentrations in the synovial fluid correlated with the tender and swollen joint counts and the activity scores on the 28-joint Disease Activity Score assessment. Calreticulin also bound directly to FasL. In vitro, calreticulin (2-16 ng/ml) inhibited FasL-induced apoptosis of Jurkat T cells.
Calreticulin was present at higher concentrations in the plasma and synovial fluid of RA patients. Calreticulin had the capacity to bind directly to FasL and to inhibit FasL-mediated apoptosis of Jurkat T cells, and thus might play a role in inhibiting apoptosis of inflammatory T cells in RA.
FasL（CD95L）与其受体Fas（CD95）结合可诱导细胞凋亡。研究表明，类风湿关节炎（RA）患者体内的T淋巴细胞对FasL诱导的细胞凋亡具有抗性，但在体外对FasL诱导的细胞凋亡敏感。该机制的功能障碍可能是RA病理生理学的重要促成因素。因此，本研究旨在确定哪些因素可能在体内抑制FasL-Fas结合，以及在体外模型系统中哪些因素会抑制T淋巴细胞的凋亡。
通过流式细胞术分析经FasL处理后发生凋亡的人Jurkat T细胞。根据乳酸脱氢酶释放量测定来确定坏死情况。采用酶联免疫吸附测定法对血浆和滑液中的钙网蛋白以及钙网蛋白-FasL结合进行定量。通过化学发光测定法测量血浆和滑液中的亚硝酸盐/硝酸盐。
与银屑病关节炎患者的血浆（中位数3.1 ng/ml，四分位间距[IQR]1.3 ng/ml）、滑液（中位数2.9 ng/ml，IQR 0.9 ng/ml）以及健康对照者的血浆（中位数2.9 ng/ml，IQR 0.9 ng/ml）相比，RA患者血浆（中位数10.3 ng/ml，IQR 14.8 ng/ml）和滑液（中位数10.3 ng/ml，IQR 12.0 ng/ml）中细胞外钙网蛋白的浓度显著更高（均P < 0.05）。滑液中的钙网蛋白浓度与压痛和肿胀关节计数以及28关节疾病活动评分评估中的活动评分相关。钙网蛋白也直接与FasL结合。在体外，钙网蛋白（2 - 16 ng/ml）可抑制FasL诱导的Jurkat T细胞凋亡。
RA患者血浆和滑液中钙网蛋白浓度较高。钙网蛋白能够直接与FasL结合并抑制FasL介导的Jurkat T细胞凋亡，因此可能在抑制RA中炎性T细胞凋亡方面发挥作用。
