Zhang Yuchen, Cheng Junjun, Zhang Junmeng, Wu Xiaofan, Chen Fang, Ren Xuejun, Wang Yunlong, Li Quan, Li Yu
Department of Cardiology, Beijing An Zhen Hospital of the Capital University of Medical Sciences, Beijing, 100029, China.
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100050, China.
Biochem Biophys Res Commun. 2016 Sep 2;477(4):761-767. doi: 10.1016/j.bbrc.2016.06.132. Epub 2016 Jun 27.
Apoptotic and necrotic macrophages have long been known for their existence in atherosclerotic lesions. However, the mechanisms underlying the choice of their death pattern have not been fully elucidated. Here, we report the effects of PS-341, a potent and specific proteasome inhibitor, on the cell death of primary bone marrow-derived macrophages (BMDMs) in vitro. The results showed that PS-341 could not induce macrophage apoptosis or promote TNF-induced macrophage apoptosis, on the other hand, PS-341 could significantly inhibit macrophage necroptosis induced by TNF and pan-caspase inhibitor z-VAD treatment. Remarkably, high-dose of PS-341 showed similar inhibitory effects on macrophage necroptosis comparable to that of kinase inhibition of RIP1 through specific inhibitor Nec-1 or inhibition of RIP3 via specific genetical ablation. Furthermore, the degradation of cellular inhibitor of apoptosis proteins (cIAPs) was suppressed by PS-341, which could antagonize the activation of RIP1 kinase via post-translational mechanism. Further evidences demonstrated reduced levels of both RIP1 and RIP 3 upon PS-341 treatment, concomitantly, a more strong association of RIP1 with cIAPs and less with RIP3 was found following PS-341 treatment, these findings suggested that PS-341 may disrupt the formation of RIP1-RIP3 complex (necrosome) through stabilizing cIAPs. Collectively, our results indicated that the proteasome-mediated degradation of cIAPs could be inhibited by PS-341 and followed by limited RIP1 and RIP3 kinase activities, which were indispensable for necroptosis, thus eliciting a significant necroptosis rescue in BMDMs in vitro. Overall, our study has identified a new role of PS-341 in the cell death of BMDMs and provided a novel insight into the atherosclerotic inflammation caused by proteasome-mediated macrophage necroptosis.
凋亡和坏死的巨噬细胞长期以来因其在动脉粥样硬化病变中的存在而为人所知。然而，其死亡模式选择背后的机制尚未完全阐明。在此，我们报告了一种强效且特异性的蛋白酶体抑制剂PS - 341对体外原代骨髓来源巨噬细胞（BMDMs）细胞死亡的影响。结果显示，PS - 341不能诱导巨噬细胞凋亡或促进肿瘤坏死因子（TNF）诱导的巨噬细胞凋亡，另一方面，PS - 341可显著抑制TNF和泛半胱天冬酶抑制剂z - VAD处理诱导的巨噬细胞坏死性凋亡。值得注意的是，高剂量的PS - 341对巨噬细胞坏死性凋亡显示出与通过特异性抑制剂Nec - 1抑制RIP1激酶或通过特异性基因敲除抑制RIP3相似的抑制作用。此外，PS - 341抑制了细胞凋亡蛋白抑制剂（cIAPs）的降解，其可通过翻译后机制拮抗RIP1激酶的激活。进一步的证据表明，PS - 341处理后RIP1和RIP 3的水平均降低，同时，PS - 341处理后发现RIP1与cIAPs的结合更强，与RIP3的结合更少，这些发现表明PS - 341可能通过稳定cIAPs破坏RIP1 - RIP3复合物（坏死小体）的形成。总体而言，我们的结果表明，PS - 341可抑制蛋白酶体介导的cIAPs降解，并随后限制对坏死性凋亡必不可少的RIP1和RIP3激酶活性，从而在体外BMDMs中引发显著的坏死性凋亡挽救。总的来说，我们的研究确定了PS - 341在BMDMs细胞死亡中的新作用，并为蛋白酶体介导的巨噬细胞坏死性凋亡引起的动脉粥样硬化炎症提供了新的见解。
