东南大学刘乃丰课题组发现，恩格列净通过抑制依赖Bhlhe40的NLRP3炎性小体活化并改善糖尿病小鼠的血管钙化状况。相关论文于2024年1月3日在线发表在《中国药理学报》杂志上。

研究人员表示，2型糖尿病（T2DM）患者更容易发生血管钙化（VC），导致死亡和残疾的风险更高。然而，目前还没有治疗血管钙化的特效药物。NLRP3炎性小体激活是内侧钙化的标志性事件，会导致动脉僵化，造成T2DM患者的血管收缩功能障碍。恩格列净（Empagliflozin，EMPA）是一种钠-葡萄糖协同转运体2抑制剂（SGLT2i），可抑制高血糖并对心血管有明显益处。

鉴于EMPA的抗炎活性，研究人员揭示了EMPA是否能通过抑制NLRP3炎性小体的活化来防止T2DM小鼠主动脉中的VC。由于接受正常饮食的db/db小鼠在大约20周大时就会出现VC，因此研究人员给8周大的db/db小鼠注射了12周的EMPA（5、10、20 mg-kg-1-d-1，i.g）。研究人员发现，EMPA干预剂量依赖性地改善了钙沉积，同时降低了主动脉中RUNX2和BMP2蛋白的表达。

研究人员发现，EMPA（10 mg-kg-1-d-1，持续6周）还能保护维生素D3过量的小鼠免受VC的影响，这表明其保护作用与新陈代谢无关。研究人员发现，EMPA（10 mg-kg-1-d-1）抑制了NLRP3炎性小体在db/db小鼠主动脉平滑肌层的异常激活。敲除（KO）NLRP3能显著缓解STZ诱导的糖尿病小鼠的VC。EMPA的保护作用在高糖（HG）处理的小鼠主动脉平滑肌细胞（MOVAS）中得到了验证。在经HG处理的NLRP3 KO MOVAS中，EMPA（1 μM）并不能进一步改善其功能。

生物信息学和Western印迹分析表明，EMPA能显著提高碱性螺旋环-螺旋家族转录因子e40（Bhlhe40）在HG处理的MOVAS中的表达水平，而e40是直接与Nlrp3启动子结合的负性转录因子。研究人员认为，EMPA通过抑制Bhlhe40依赖的NLRP3炎性小体的激活来改善VC。这些结果可能为EMPA治疗T2DM患者的VC提供了潜在的意义。

附：英文原文

Title: Empagliflozin ameliorates vascular calcification in diabetic mice through inhibiting Bhlhe40-dependent NLRP3 inflammasome activation

Author: Li, Xiao-xue, Chen, Zheng-dong, Sun, Xue-jiao, Yang, Yi-qing, Jin, Hong, Liu, Nai-feng

Issue&Volume: 2024-01-03

Abstract: Type 2 diabetes mellitus (T2DM) patients exhibit greater susceptibility to vascular calcification (VC), which has a higher risk of death and disability. However, there is no specific drug for VC therapy. NLRP3 inflammasome activation as a hallmark event of medial calcification leads to arterial stiffness, causing vasoconstrictive dysfunction in T2DM. Empagliflozin (EMPA), a sodium-glucose co-transporter 2 inhibitor (SGLT2i), restrains hyperglycemia with definite cardiovascular benefits. Given the anti-inflammatory activity of EMPA, herein we investigated whether EMPA protected against VC in the aorta of T2DM mice by inhibiting NLRP3 inflammasome activation. Since db/db mice receiving a normal diet developed VC at the age of about 20 weeks, we administered EMPA (5, 10, 20mg·kg1·d1, i.g) to 8 week-old db/db mice for 12 weeks. We showed that EMPA intervention dose-dependently ameliorated the calcium deposition, accompanied by reduced expression of RUNX2 and BMP2 proteins in the aortas. We found that EMPA (10mg·kg1·d1 for 6 weeks) also protected against VC in vitamin D3-overloaded mice, suggesting the protective effects independent of metabolism. We showed that EMPA (10mg·kg1·d1) inhibited the abnormal activation of NLRP3 inflammasome in aortic smooth muscle layer of db/db mice. Knockout (KO) of NLRP3 significantly alleviated VC in STZ-induced diabetic mice. The protective effects of EMPA were verified in high glucose (HG)-treated mouse aortic smooth muscle cells (MOVASs). In HG-treated NLRP3 KO MOVASs, EMPA (1μM) did not cause further improvement. Bioinformatics and Western blot analysis revealed that EMPA significantly increased the expression levels of basic helix-loop-helix family transcription factor e40 (Bhlhe40) in HG-treated MOVASs, which served as a negative transcription factor directly binding to the promotor of Nlrp3. We conclude that EMPA ameliorates VC by inhibiting Bhlhe40-dpendent NLRP3 inflammasome activation. These results might provide potential significance for EMPA in VC therapy of T2DM patients.

DOI: 10.1038/s41401-023-01217-0

Source: https://www.nature.com/articles/s41401-023-01217-0