Cao Xiaopei, He Yuanzhou, Li Xiaochen, Xu Yongjian, Liu Xiansheng
Department of Pulmonary and Critical Care Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology Wuhan, China.
Key Laboratory of Pulmonary Diseases, National Ministry of Health of The People's Republic of China Wuhan, China.
Am J Transl Res. 2019 Feb 15;11(2):641-654. eCollection 2019.
Hypoxia is a common cause of pulmonary vascular remodeling and endoplasmic reticulum stress (ERS). Upon ER stress, the unfolded protein response (UPR) which activates the IRE1α, PERK and ATF6 signaling pathways is activated to cope with ERS in mammalian cells; however, the role of the three UPR arms in pulmonary vascular remodeling has not been defined. The present study showed that GRP78, a marker of ERS, was upregulated in hypoxic pulmonary artery smooth muscle cells (PASMCs). Among the three arms of the UPR, the IRE1α pathway was noticeably upregulated in hypoxic PASMCs. An inhibitor of IRE1α/XBP1 pathway, 4u8c, inhibited hypoxia-induced cell proliferation and migration and increased cell apoptosis by downregulating PCNA and MMP9 and activating mitochondrial apoptosis by enhancing the expression of BAX, activating caspase-9 and caspase-3, and eventually cleaving PARP. Quercetin affects ERS in many cell types and was shown to relieve hypoxic pulmonary hypertension (HPH) in our previous study. We demonstrated that quercetin evoked excessive GRP78 expression in hypoxic PASMCs compared with hypoxia alone by evaluating the expression of GRP78. The expression of IRE1α and XBP1s, a cleavage form of XBP1u, was upregulated by quercetin in a dose-dependent manner. Pretreatment with 4u8c reversed the apoptosis-promoting effect of quercetin by inhibiting mitochondrial apoptosis. However, 4u8c amplified the effect of quercetin on proliferation and migration in hypoxic PASMCs. In conclusion, the study demonstrated that the IRE1α-XBP1 pathway is involved in the process of hypoxia-induced pulmonary vascular remodeling; 4u8c could restrain hypoxia-induced cell proliferation and migration and reverse the hypoxia-induced apoptosis arrest, while quercetin excited excessive ERS and the IRE1α pathway in hypoxic PASMCs and promoted apoptosis. Our data suggest that intervening the IRE1α-XBP1 pathway may be useful for hypoxia-induced pulmonary arterial hypertension therapy.
缺氧是肺血管重塑和内质网应激（ERS）的常见原因。内质网应激时，激活IRE1α、PERK和ATF6信号通路的未折叠蛋白反应（UPR）被激活，以应对哺乳动物细胞中的内质网应激；然而，三条UPR分支在肺血管重塑中的作用尚未明确。本研究表明，ERS标志物GRP78在缺氧肺动脉平滑肌细胞（PASMCs）中上调。在UPR的三条分支中，IRE1α通路在缺氧PASMCs中显著上调。IRE1α/XBP1通路抑制剂4u8c通过下调PCNA和MMP9抑制缺氧诱导的细胞增殖和迁移，并通过增强BAX表达、激活caspase-9和caspase-3以及最终切割PARP来激活线粒体凋亡，从而增加细胞凋亡。槲皮素在许多细胞类型中影响内质网应激，并且在我们之前的研究中显示其可缓解缺氧性肺动脉高压（HPH）。通过评估GRP78的表达，我们证明槲皮素与单独缺氧相比，在缺氧PASMCs中引起GRP78过度表达。槲皮素以剂量依赖性方式上调IRE1α和XBP1s（XBP1u的切割形式）的表达。用4u8c预处理通过抑制线粒体凋亡逆转了槲皮素的促凋亡作用。然而，4u8c增强了槲皮素对缺氧PASMCs增殖和迁移的作用。总之，该研究表明IRE1α-XBP1通路参与缺氧诱导的肺血管重塑过程；4u8c可抑制缺氧诱导的细胞增殖和迁移并逆转缺氧诱导的凋亡停滞，而槲皮素在缺氧PASMCs中引发过度的内质网应激和IRE1α通路并促进凋亡。我们的数据表明，干预IRE1α-XBP1通路可能对缺氧诱导的肺动脉高压治疗有用。
