Inflammation and Immune Mediated Diseases Laboratory of Anhui Province, School of Pharmacy, Anhui Medical University, 81 Meishan Road, Hefei, 230032, Anhui Province, People's Republic of China.
Department of Pathology, School of Basic Medicine, Anhui Medical University, Hefei, 230032, Anhui Province, People's Republic of China.
Inflammation. 2023 Dec;46(6):2289-2305. doi: 10.1007/s10753-023-01878-3. Epub 2023 Jul 22.
Synovial hypoxia-inducible factor 1α (HIF-1α) is a prospective therapeutic target for rheumatoid arthritis (RA). AMSP-30 m, a novel HIF-1α inhibitor, was reported to have notable anti-arthritic effects in rats with adjuvant-induced arthritis. However, its roles in inhibiting the pathogenic behaviors of fibroblast-like synoviocytes (FLS) and the involved mechanisms remain unknown. Here, AMSP-30 m inhibited proliferation and induced apoptosis in hypoxia-induced RA FLS (MH7A cell line), as evidenced by decreased cell viability, reduced Ki67-positive cells, G0/G1 phase arrest, lowered C-myc and Cyclin D1 protein levels, emergence of apoptotic nuclear fragmentation, raised apoptosis rates, and activation of caspase 3. Furthermore, AMSP-30 m prevented hypoxia-induced increases in pro-inflammatory factor production, MMP-2 activity, migration index, migrated/invasive cells, and actin cytoskeletal rearrangement. In vivo, AMSP-30 m alleviated the severity of rat collagen-induced arthritis (CIA). Mechanically, AMSP-30 m reduced HIF-1α expression and blocked sonic hedgehog (Shh) pathway activation in hypoxia-induced MH7A cells and CIA rat synovium, as shown by declines in pathway-related proteins (Shh, Smo, and Gli-1). Particularly, the combination of Shh pathway inhibitor cyclopamine enhanced AMSP-30 m's inhibitory effects on the pathogenic behaviors of hypoxia-stimulated MH7A cells, whereas the combination of Shh pathway activator SAG canceled AMSP-30 m's therapeutic effects in vitro and in CIA rats, implying a close involvement of Shh pathway inhibition in its anti-arthritic effects. We likewise confirmed AMSP-30 m's anti-proliferative role in hypoxia-induced primary CIA FLS. Totally, AMSP-30 m suppressed hypoxia-induced proliferation, inflammation, migration, and invasion of MH7A cells and ameliorated the severity of rat CIA via inhibiting Shh signaling.
滑膜缺氧诱导因子 1α（HIF-1α）是类风湿关节炎（RA）的潜在治疗靶点。新型 HIF-1α 抑制剂 AMSP-30m 在佐剂诱导关节炎大鼠中表现出显著的抗关节炎作用。然而，其在抑制成纤维样滑膜细胞（FLS）的致病行为及其相关机制中的作用尚不清楚。本研究中，AMSP-30m 抑制缺氧诱导的 RA FLS（MH7A 细胞系）增殖并诱导其凋亡，表现为细胞活力降低、Ki67 阳性细胞减少、G0/G1 期阻滞、C-myc 和 Cyclin D1 蛋白水平降低、凋亡核碎片化出现、凋亡率升高和 caspase 3 激活。此外，AMSP-30m 可阻止缺氧诱导的促炎因子产生、MMP-2 活性、迁移指数、迁移/侵袭细胞和肌动蛋白细胞骨架重排增加。在体内，AMSP-30m 减轻了胶原诱导性关节炎（CIA）大鼠的严重程度。机制上，AMSP-30m 降低了缺氧诱导的 MH7A 细胞和 CIA 大鼠滑膜中 HIF-1α 的表达并阻断了 sonic hedgehog（Shh）通路的激活，表现为通路相关蛋白（Shh、Smo 和 Gli-1）的下降。特别地，Shh 通路抑制剂环巴胺增强了 AMSP-30m 对缺氧刺激的 MH7A 细胞致病行为的抑制作用，而 Shh 通路激活剂 SAG 则取消了 AMSP-30m 体外和 CIA 大鼠中的治疗作用，表明 Shh 通路抑制在其抗关节炎作用中密切相关。我们同样证实了 AMSP-30m 在缺氧诱导的原发性 CIA FLS 中的抗增殖作用。总之，AMSP-30m 通过抑制 Shh 信号通路抑制了缺氧诱导的 MH7A 细胞增殖、炎症、迁移和侵袭，并改善了大鼠 CIA 的严重程度。
