Abstract Body (Do not enter title and authors here): Background:
Thoracic aortic aneurism and dissection (TAAD) is a severe macrovascular disease that lead to aortic rupture and sudden death. To date, there are no effective clinical agents to prevent the formation or control the progression of TAAD.
Objectives:
Investigate the mechanism of empagliflozin improves aortic pathological remodeling in TAAD mice.
Methods:
Four-week-old male mice were given BAPN (β-aminopropionitrile monofumarate; 1 g/kg/d) in drinking water for 4 weeks for TAD modeling. RNA sequencing analysis was performed to recapitulate transcriptome profile changes.TRIM59 with USP7 and NLRP3 in VSMCs was tested by coimmunoprecipitation assay.
Results:
1. Compared with BAPN group or MFS group, the degree of elastic fiber rupture and collagen fiber deposition in TAAD mice with two kinds of NLRP3 knockout were significantly improved (all P<0.05). Transcriptional sequencing showed that the effect of NLRP3 knockout on the aorta of TAAD mice was mainly focused on extracellular matrix remodeling.
2. Empagliflozin was confirmed to significantly ameliorate pathological remodeling of aortic ECM in TAAD mice (all P<0.05). GO/KEGG enrichment analysis revealed that empagliflozin primarily impacts ECM structure and function, as well as inflammation-related signaling pathways. Compared with the vehicle group, BAPN group displayed increased mRNA and protein levels of NLRP3, while empagliflozin decreased NLRP3 protein levels. These findings suggest that empagliflozin may regulate the post-transcriptional level of NLRP3.
3. In vitro studies have shown that empagliflozin promotes the degradation of NLRP3 protein in a dose- and time-dependent manner. MG132 offset the effect of empagliflozin on NLRP3 degradation; coimmunoprecipitation assay confirmed that empagliflozin promotes the degradation of NLRP3 through the ubiquitin-proteasome system (all P<0.05). In addition, TRIM59 knockdown increased NLRP3 ubiquitination levels, whereas the presence of TRIM59 increased NLRP3 deubiquitination levels. Further through the Ubibrowser 2.0 website found that TRIM59 has a strong interaction with USP7. Finally, compared with DMSO group, the interaction between USP7 and NLPR3 was weakened and the ubiquitination level of NLRP3 was increased in the empagliflozin-treated group. Notably, overexpression of TRIM59 reversed this effects of empagliflozin.
Conclusion:
Empagliflozin reduces TRIM59 expression reduces the binding of USP7 to NLRP3 and promotes ubiquitination and degradation of NLRP3.