Department of Immunology, Medical College of Nanchang University, Nanchang, China; Department of Clinical Laboratory, People's Hospital of Deyang City, Deyang, China.
Department of Immunology, Medical College of Nanchang University, Nanchang, China.
Cytokine. 2020 Sep;133:155146. doi: 10.1016/j.cyto.2020.155146. Epub 2020 Jun 3.
Rheumatoid arthritis (RA) is a chronic, symmetric, systemic autoimmune disease. Because insufficient apoptosis of fibroblast-like synoviocytes (FLS) is an important characteristic of RA, promoting apoptosis is considered a potential therapeutic tool for treating RA. We have previously found that daphnetin (7,8-dihydroxycoumarin, DAP) has a pro-apoptotic effect on fibroblast-like synoviocytes from collagen-induced arthritis (CIA) rats. In the present study, we further investigated the mechanisms of DAP-induced apoptosis in CIA-FLS. CIA-FLS were incubated with DAP for 48 h in the presence or absence of caspase inhibitors, including inhibitors of caspase-3, caspase-8, or caspase-9 or a pan-caspase inhibitor; then, a series of experiments were performed to evaluate the mechanisms of DAP-induced apoptosis. Our results showed that DAP markedly decreased cell viability and induced the apoptosis of CIA-FLS along with typical morphological and ultrastructural changes; moreover, DAP increased FasL, cytochrome c (Cyt-c), Bax, caspase-3, caspase-8, and caspase-9 mRNA expression and Bax, caspase-3, caspase-8, and caspase-9 protein expression. In contrast, DAP decreased Bcl-2 mRNA and protein expression and promoted the release of Cyt-c from the mitochondria into the cytosol; these effects were attenuated to varying degrees by pre-treatment with caspase inhibitors, especially with caspase-3 or caspase-9 inhibitors or a pan-caspase inhibitor. In conclusion, the current findings demonstrate that the DAP-induced apoptosis of CIA-FLS occurred mainly via a caspase-dependent pathway, in particular the mitochondrial pathway, and that the Bax/Bcl-2 ratio was involved in this process. Thus, DAP may be a potential therapeutic agent for RA.
类风湿关节炎（RA）是一种慢性、对称、系统性自身免疫性疾病。由于成纤维样滑膜细胞（FLS）凋亡不足是 RA 的一个重要特征，因此促进凋亡被认为是治疗 RA 的一种潜在治疗手段。我们之前发现，瑞香素（7,8-二羟基香豆素，DAP）对胶原诱导关节炎（CIA）大鼠的成纤维样滑膜细胞具有促凋亡作用。在本研究中，我们进一步研究了 DAP 诱导 CIA-FLS 凋亡的机制。将 CIA-FLS 与 DAP 孵育 48 小时，存在或不存在半胱天冬酶抑制剂，包括 caspase-3、caspase-8 或 caspase-9 抑制剂或泛半胱天冬酶抑制剂；然后，进行了一系列实验来评估 DAP 诱导凋亡的机制。我们的结果表明，DAP 显著降低细胞活力并诱导 CIA-FLS 凋亡，同时伴有典型的形态学和超微结构变化；此外，DAP 增加 FasL、细胞色素 c（Cyt-c）、Bax、caspase-3、caspase-8 和 caspase-9 mRNA 表达以及 Bax、caspase-3、caspase-8 和 caspase-9 蛋白表达。相反，DAP 降低了 Bcl-2 mRNA 和蛋白表达，并促进 Cyt-c 从线粒体释放到细胞质；这些作用被 caspase 抑制剂预处理不同程度地减弱，尤其是 caspase-3 或 caspase-9 抑制剂或泛半胱天冬酶抑制剂。总之，目前的研究结果表明，DAP 诱导 CIA-FLS 凋亡主要通过 caspase 依赖性途径发生，特别是线粒体途径，并且 Bax/Bcl-2 比值参与了这一过程。因此，DAP 可能是 RA 的一种潜在治疗药物。
