Clinical Proteomics & Molecular Medicine, St. Marianna University Graduate School of Medicine, Kawasaki, Japan.
Clin Exp Rheumatol. 2011 Sep-Oct;29(5):768-75. Epub 2011 Oct 31.
In our previous proteomic surveillance, we found that at least 11 proteins in neutrophils were increased more than 2.5-fold by the stimulation of GM-CSF. In this paper, focusing on one of the 11 proteins, S100 calcium binding protein A8 (S100A8), we tried to elucidate the effect of S100A8 and the cooperative effect of S100A8 and GM-CSF on production and secretion of cytokines of neutrophils.
S100A8 in neutrophil was detected by western blotting, and concentrations of S100A8 in synovial fluid (SF) from patients with rheumatoid arthritis (RA) and osteoarthritis (OA) were measured by ELISA. Cytokine levels in the culture medium of neutrophils incubated with and without S100A8 were measured by an antibody array. IL-8 and IL-16 levels in the culture medium of neutrophils stimulated with S100A8, GM-CSF, and the combination of S100A8 and GM-CSF were measured by ELISA. The mRNA levels of IL-8 and IL-16 in the stimulated neutrophils were analysed by real-time PCR.
The western blotting analysis confirmed that S100A8 is up-regulated in neutrophil by the stimulation of GM-CSF. Furthermore, the ELISA analysis confirmed that S100A8 was significantly elevated in SF of patients with RA compared to SF of patients with OA. S100A8 induced mRNA expression and secretion of IL-8 and IL-16. S100A8 further enhanced production of IL-8 by GM-CSF but not that of IL-16.
These data suggest that S100A8 may be involved in the exacerbation of RA, and that S100A8 may be a therapeutic target of RA.
在我们之前的蛋白质组学监测中，我们发现粒细胞中至少有 11 种蛋白被 GM-CSF 刺激后增加了 2.5 倍以上。在本文中，我们专注于这 11 种蛋白之一，S100 钙结合蛋白 A8（S100A8），试图阐明 S100A8 的作用以及 S100A8 和 GM-CSF 对中性粒细胞细胞因子产生和分泌的协同作用。
通过 Western blot 检测中性粒细胞中的 S100A8，通过 ELISA 检测类风湿关节炎（RA）和骨关节炎（OA）患者滑液（SF）中的 S100A8 浓度。用抗体阵列检测孵育有无 S100A8 的中性粒细胞培养基中的细胞因子水平。用 ELISA 检测 S100A8、GM-CSF 及 S100A8 和 GM-CSF 联合刺激中性粒细胞后培养基中 IL-8 和 IL-16 的水平。用实时 PCR 分析刺激中性粒细胞中 IL-8 和 IL-16 的 mRNA 水平。
Western blot 分析证实 GM-CSF 刺激可使中性粒细胞中 S100A8 上调。此外，ELISA 分析证实 RA 患者的 SF 中 S100A8 明显高于 OA 患者的 SF。S100A8 诱导 IL-8 和 IL-16 的 mRNA 表达和分泌。S100A8 进一步增强 GM-CSF 诱导的 IL-8 产生，但不增强 IL-16 的产生。
这些数据表明 S100A8 可能参与 RA 的恶化，S100A8 可能是 RA 的治疗靶点。
