Senolytics restore diabetes-induced aortic dysfunction in DIO mice

Background – Diabetes is closely associated with impaired vascular function and increased aortic stiffness, phenomena commonly observed in aging. Cellular senescence, a process accelerated in diabetes, contributes to vascular dysfunction through the senescence-associated secretory phenotype (SASP) consisting of a plethora of inflammatory and other activating substances. Interleukin (IL)-6, a prominent SASP factor, is known to elevate reactive oxygen species (ROS) and inhibit expression of endothelial nitric oxide synthase (eNOS), a key enzyme for vascular relaxation. This study investigates whether senescent fibroblasts contribute to IL-6-driven inflammation and oxidative stress in the diabetic aorta, thereby impairing vascular function.

Methods – Male C57Bl/6J mice (10–12 weeks old) were fed a high-fat diet for 12 weeks to induce obesity (diet-induced obesity, DIO) and a prediabetic state. Mice received either a control treatment or a senolytic combination of dasatinib (5 mg/kg) and quercetin (50 mg/kg) (DQ) for one week. Aortic senescence was assessed by measuring Cdkn1a mRNA expression using qPCR, while vascular function was evaluated in isolated aortic rings by wire myography. Histological and qPCR analysis identified senescent cell types, assessed ROS levels, and Il6 expression in aortic tissues. In vitro, human umbilical vein endothelial cells (HUVECs) were treated with recombinant IL-6 or supernatants from senescent human cardiac fibroblasts, with or without the IL-6–inhibiting antibody Tocilizumab, to assess ROS and eNOS activity. Additionally, Cdkn1a and Il6 mRNA levels were measured in aortic samples from diabetic and non-diabetic patients.

Results – DIO mice showed increased aortic senescence with elevated Cdkn1a expression, which was significantly reduced following DQ treatment. Impaired relaxation in aorta explanted from DIO mice was restored with senolytic treatment. Histological staining identified fibroblasts as the predominant senescent cell type in aortic tissue, and qPCR and histology indicated higher Il6 expression and ROS levels, respectively, in DIO mice. Treatment of HUVECs with recombinant IL-6 increased ROS formation and impaired eNOS function, effects that were also recapitulated with supernatants from senescent fibroblasts but were reversed by applying an IL-6-neutralizing antibody. Aortic tissue specimens from diabetic patients displayed significantly higher Cdkn1a and Il6 mRNA expression compared to non-diabetic control patients, supporting the link between diabetes, senescence, and inflammation. Conclusion – Senescent fibroblasts contribute to IL-6 production and oxidative stress, impairing vascular relaxation in diabetes. Senolytic treatment decreases aortic stiffness and improves vascular function in a pre-clinical diabetic model. These findings underscore the therapeutic potential of targeting senescence to address diabetes-associated vascular dysfunction and chronic inflammation.