The effect of senolytics on cardiac remodelling and repair after injury induced by isoproterenol

Abstract

Senescent cells accumulate during ageing and contribute to tissue deterioration, including in the heart. Senescent cells negatively affect an organ's microenvironment by refusing to die and producing a pro-inflammatory senescence-associated secretory phenotype (SASP). Genetic or pharmacologic clearance of senescent cells by senolytics has been shown to improve cardiac recovery and remodelling in aged mice. Moreover, senolytics have improved cardiac recovery after cardiac injury and in heart failure models in aged and young adult mice. Isoproterenol (ISO) has been widely used to induce cardiac injury in rodents, but it is unknown if ISO induces senescence and whether increased senescent cells in the heart contribute to cardiac deterioration and pathophysiology. This PhD aimed to first establish a cardiac injury model which led to increased senescence in the hearts of young adult mice by ISO. Second, to investigate the effects of the senolytics, dasatanib+quercetin (D+Q) on cardiac recovery and remodelling after ISO-injury in young adult and aged mice. To establish the dose of ISO that induced senescence with cardiac injury, ISO 150 mg/kg or 100 mg/kg was subcutaneously administered to ~12-week-old male mice (n=3 per group) for six consecutive days. Echocardiography was conducted from baseline to day 7. On day 7 after ISO, hearts were excised in order to analyse cardiac injury and cellular senescence. Results indicated that 100 mg/kg and 150 mg/kg ISO induced injury mainly in the apex of the heart. However, SA-β-gal staining was evident only in the apex of the ISO 150 mg/kg group. Therefore, ISO 150 mg/kg was chosen over the ISO 100 mg/kg dose. To establish the timeline of increased senescence after ISO-injury, 150 mg/kg ISO was subcutaneously administered to ~12-week-old male mice (n= 3-9 per group) for six consecutive days. Echocardiography was conducted at baseline, day 7 and 14 following ISO administration. Mice were sacrificed and hearts were excised for analysis of cardiac injury and senescence on days 7, 10 and 14. ISO 150 mg/kg successfully induced cardiac injury and senescence that lasted for 14 days. The peak of senescence was observed on day 10 following ISO.

Next, ISO 150 mg/kg was subcutaneously administered to ~12-week-old male mice for six consecutive days. On day 10 after ISO, D+Q (5 mg/kg, 50 mg/kg) were administered by oral gavage for five consecutive days. Echocardiography was performed at baseline, day 7 post-ISO, and day 28 after the last D+Q dose. The hearts were excised on day 28 following the last D+Q dose to investigate cardiac injury and cellular senescence. Young adult mice treated with D+Q showed better cardiac recovery and remodelling after ISO injury. D+Q enhanced cardiac function, reduced hypertrophy, and reduced senescence markers. However, levels of collagen and DNA damage were unaffected by D+Q treatment. Aged male mice had four cycles of D+Q (5 mg/kg, 50 mg/kg) via oral gavage. Each cycle was composed of 3 days on and 12 days off. After that, aged mice hearts were injured by subcutaneous injection of 50 mg/kg ISO for six consecutive days. Echocardiography was performed at baseline, 24 hours after D+Q, and 28 days after the last ISO dose. The hearts were excised on day 28 following the last ISO dose to investigate cardiac injury, cellular senescence and remodelling. D+Q did not enhance cardiac function in the aged mice before ISO injury. However, D+Q improved cardiac recovery and survival after ISO injury. In the ISO-injured aged mice, D+Q enhanced cardiac function, reduced hypertrophy, reduced fibrosis, reduced DNA damage and senescence markers, and increased cardiomyocyte DNA synthesis. Clearance of the senescent cells and their SASP factors by D+Q senolytics improved cardiac function and recovery after ISO injury in aged and young adult male mice. These findings encourage the use of senolytics as a potential adjunct therapy for cardiac injury and deterioration with ageing. Senolytics could be used to improve the microenvironment of the heart so that it is more resilient to damage and can recover more effectively.