Combination of ADAM17 knockdown with eplerenone is more effective than single therapy in ameliorating diabetic cardiomyopathy

Abstract

Background

The renin-angiotensin-aldosterone system (RAAS) members, especially Ang II and aldosterone, play key roles in the pathogenesis of diabetic cardiomyopathy (DCM). Angiotensin-converting enzyme inhibitors or angiotensin-receptor blockers combined with aldosterone receptor antagonists (mineralocorticoid receptor antagonists) have substantially improved clinical outcomes in patients with DCM. However, the use of the combination has been limited due to its high risk of inducing hyperkalemia.

Methods

Type 1 diabetes was induced in 8-week-old male C57BL/6J mice by intraperitoneal injection of streptozotocin at a dose of 55 mg/kg for 5 consecutive days. Adeno-associated virus 9-mediated short-hairpin RNA (shRNA) was used to knock down the expression of ADAM17 in mice hearts. Eplerenone was administered via gavage at 200 mg/kg daily for 4 weeks. Primary cardiac fibroblasts were exposed to high glucose (HG) in vitro for 24 h to examine the cardiac fibroblasts to myofibroblasts transformation (CMT).

Results

Cardiac collagen deposition and CMT increased in diabetic mice, leading to cardiac fibrosis and dysfunction. In addition, ADAM17 expression and activity increased in the hearts of diabetic mice. ADAM17 inhibition and eplerenone treatment both improved diabetes-induced cardiac fibrosis, cardiac hypertrophy and cardiac dysfunction, ADAM17 deficiency combined with eplerenone further reduced the effects of cardiac fibrosis, cardiac hypertrophy and cardiac dysfunction compared with single therapy in vivo. High-glucose stimulation promotes CMT in vitro and leads to increased ADAM17 expression and activity. ADAM17 knockdown and eplerenone pretreatment can reduce the CMT of fibroblasts that is induced by high glucose levels by inhibiting TGFβ1/Smad3 activation; the combination of the two can further reduce CMT compared with single therapy in vitro.

Conclusion

Our findings indicated that ADAM17 knockout could improve diabetes-induced cardiac dysfunction and remodeling through the inhibition of RAAS overactivation when combined with eplerenone treatment, which reduced TGF-β1/Smad3 pathway activation-mediated CMT. The combined intervention of ADAM17 deficiency and eplerenone therapy provided additional cardiac protection compared with a single therapy alone without disturbing potassium level. Therefore, the combination of ADAM17 inhibition and eplerenone is a potential therapeutic strategy for human DCM.

Graphical Abstract

Mechanism of the roles of combination of ADAM17 knockdown and eplerenone treatment in DCM-induced cardiac fibrosis. The high-glucose activated RAAS increased ADAM17 activity, ACE2 shedding, and AT1R expression, thereby activating TGF-β1/Smad3 signaling in cardiac fibroblasts, which increased myofibroblast transformation and cardiac collagen deposition in DCM. The overactivated RAAS increased aldosterone expression, so aldosterone then bound to aldosterone receptors to further promote TGF-β1/Smad3 pathway-mediated myofibroblast transformation. ADAM17 deletion weakened ACE2 shedding and AT1R expression, inactivated TGF-β1/Smad3 signaling in myocardial fibroblasts, and resulted in reduced myofibroblast transformation and cardiac collagen deposition in DCM. Moreover, eplerenone acted as an aldosterone receptor antagonist to reduce the aldosterone activation of TGF-β1. Eplerenone combined with ADAM17 deficiency can further reduce the fibroblast transformation and collagen deposition caused by diabetes and improve the cardiac fibrosis and cardiac remodeling caused by DCM compared with when either of the two is used as monotherapy.