Liu Mengguo, Yang Ji, Xing Xiaojing, Cui Xiangxiang, Li Ming
Arthritis Res Ther. 2014 Dec 31;16(6):4223. doi: 10.1186/s13075-014-0512-2.
Dermal vascular smooth muscle cells (DVSMCs) are important for vascular wall fibrosis in microangiopathy of systemic sclerosis (SSc). T helper 17 cell-associated cytokines, particularly interleukin-17A (IL-17A), have been demonstrated to play a role in the pathogenesis of SSc. However, the effect of IL-17A on the DVSMCs in microangiopathy of SSc has not been established. In the present study, we investigated the effect of IL-17A on the SSc patient-derived DVSMCs.
DVSMCs from patients with SSc and healthy subjects were incubated using IL-17A or serum derived from patients with SSc. Subsequently, the proliferation, collagen synthesis and secretion, and migration of DVSMCs were analysed using a cell counting kit-8 (CCK-8), dual-luciferase reporter assay, real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blot, enzyme-linked immunosorbent assay (ELISA) and transwell assay. The protein phosphorylation of signalling pathways in the process of IL-17A-mediated DVSMC activation was investigated and validated by specific signalling pathway inhibitor.
IL-17A and serum from patients with SSc could promote the proliferation, collagen synthesis and secretion, and migration of DVSMCs. IL-17A neutralising antibody could inhibit the IL-17A-induced activation of DVSMCs. Additionally, IL-17A induced the activation of extracellular-regulated protein kinases 1/2 (ERK1/2) in DVSMCs, and ERK1/2 inhibitor could block the IL-17A-elicited activation of DVSMCs.
Our results suggested that IL-17A derived from patients with SSc might induce the proliferation, collagen synthesis and secretion, and migration of DVSMCs via ERK1/2 signalling pathway, raising the likelihood that IL-17A and ERK1/2 might be promising therapeutic targets for the treatment of SSc-related vasculopathy.
真皮血管平滑肌细胞(DVSMC)在系统性硬化症(SSc)微血管病变的血管壁纤维化过程中起重要作用。辅助性T细胞17相关细胞因子,尤其是白细胞介素-17A(IL-17A),已被证明在SSc发病机制中发挥作用。然而,IL-17A对SSc微血管病变中DVSMC的影响尚未明确。在本研究中,我们调查了IL-17A对SSc患者来源的DVSMC的影响。
使用IL-17A或SSc患者来源的血清培养SSc患者和健康受试者的DVSMC。随后,使用细胞计数试剂盒-8(CCK-8)、双荧光素酶报告基因检测、实时逆转录-聚合酶链反应(RT-PCR)、蛋白质印迹法、酶联免疫吸附测定(ELISA)和Transwell检测分析DVSMC的增殖、胶原合成与分泌以及迁移情况。通过特异性信号通路抑制剂研究并验证IL-17A介导的DVSMC激活过程中信号通路的蛋白磷酸化情况。
IL-17A和SSc患者的血清可促进DVSMC的增殖、胶原合成与分泌以及迁移。IL-17A中和抗体可抑制IL-17A诱导的DVSMC激活。此外,IL-17A可诱导DVSMC中细胞外调节蛋白激酶1/2(ERK1/2)的激活,ERK1/2抑制剂可阻断IL-17A引发的DVSMC激活。
我们的结果表明,SSc患者来源的IL-17A可能通过ERK1/2信号通路诱导DVSMC的增殖、胶原合成与分泌以及迁移,这增加了IL-17A和ERK1/2可能成为治疗SSc相关血管病变的有前景治疗靶点的可能性。