Liu Hongtao, Wang Zhongxiao, Yu Shujie, Xu Jian
Section of Endocrinology and Diabetes, Department of Medicine, Harold Hamm Diabetes Center, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
Department of Physiology, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA.
Cardiovasc Res. 2014 Jul 1;103(1):131-9. doi: 10.1093/cvr/cvu116. Epub 2014 Apr 29.
Hypoxia induces vascular inflammation by a mechanism not fully understood. Emerging evidence implicates O-GlcNAc transferase (OGT) in inflammation. This study explored the role of OGT in hypoxia-induced vascular endothelial inflammatory response.
Hypoxia was either induced (1% O2 chamber) or mimicked by exposure to hypoxia-mimetic agents in cultured endothelial cells. Hypoxia increased hypoxia-inducible factor (HIF-1α) and inflammatory response (gene and protein expression of interleukin (IL)-6, IL-8, monocyte chemoattractant protein-1, and E-selectin) but, surprisingly, reduced OGT protein (not mRNA) levels. Hypoxia-mimetic CoCl2 failed to reduce OGT when proteasome inhibitors were present, suggesting proteasome involvement. Indeed, CoCl2 enhanced 26S proteasome functionality evidenced by diminished reporter (Ub(G76V)-GFP) proteins in proteasome reporter cells, likely due to increased chymotrypsin-like activities. Mechanistically, β-TrCP1 mediated OGT degradation, since siRNA ablation of this E3 ubiquitin ligase stabilized OGT. Administration of the oxidative stress inhibitors reversed both proteasome activation and OGT degradation. Furthermore, up-regulation of OGT by stabilization, overexpression, or activation mitigated CoCl2-elicited inflammatory response. These observations were recapitulated in a mouse (C57BL/6J) model mimicking hypoxia, in which lung tissues presented higher levels of HIF-1α, proteasome activity, and inflammatory response, but lower levels of OGT (n = 5/group, hypoxia vs. normoxia, P < 0.05). However, administration of an activator of OGT (glucosamine: 1 mg/g/day, vehicle: saline, ip, 5 days) abolished the up-regulation of proteasome activity and inflammatory response (n = 5/group, the treated vs. untreated hypoxia groups, P < 0.05).
26S proteasome-mediated OGT reduction contributed to hypoxia-induced vascular endothelial inflammatory response. Modulation of OGT may represent a new approach to treat diseases characterized by hypoxic inflammation.
缺氧通过一种尚未完全了解的机制诱导血管炎症。新出现的证据表明O-连接N-乙酰葡糖胺转移酶（OGT）与炎症有关。本研究探讨了OGT在缺氧诱导的血管内皮炎症反应中的作用。
在培养的内皮细胞中，通过1%氧气舱诱导缺氧或用缺氧模拟剂模拟缺氧。缺氧增加了缺氧诱导因子（HIF-1α）和炎症反应（白细胞介素（IL）-6、IL-8、单核细胞趋化蛋白-1和E-选择素的基因和蛋白表达），但令人惊讶的是，降低了OGT蛋白（而非mRNA）水平。当存在蛋白酶体抑制剂时，缺氧模拟剂CoCl2未能降低OGT，提示蛋白酶体参与其中。事实上，CoCl2增强了26S蛋白酶体功能，蛋白酶体报告细胞中报告蛋白（Ub(G76V)-GFP）减少证明了这一点，这可能是由于胰凝乳蛋白酶样活性增加所致。从机制上讲，β-TrCP1介导OGT降解，因为该E3泛素连接酶的siRNA敲除使OGT稳定。给予氧化应激抑制剂可逆转蛋白酶体激活和OGT降解。此外，通过稳定、过表达或激活上调OGT可减轻CoCl2引发的炎症反应。在模拟缺氧的小鼠（C57BL/6J）模型中也观察到了这些现象，其中肺组织中HIF-1α、蛋白酶体活性和炎症反应水平较高，但OGT水平较低（每组n = 5，缺氧组与常氧组比较，P < 0.05）。然而，给予OGT激活剂（氨基葡萄糖：1 mg/g/天，载体：生理盐水，腹腔注射，5天）可消除蛋白酶体活性和炎症反应的上调（每组n = 5，治疗组与未治疗的缺氧组比较，P < 0.05）。
26S蛋白酶体介导的OGT减少导致缺氧诱导的血管内皮炎症反应。调节OGT可能代表一种治疗以缺氧炎症为特征的疾病的新方法。
