Simultaneous induction of differentiation and senescence in cardiac fibroblasts by TGF-β1: effects of senotherapeutics drugs

IntroductionAging is a major risk factor for cardiovascular diseases (CVD), partly due to the accumulation of senescent cells. Senescence is characterized by irreversible cell cycle arrest and the acquisition of a senescence associated secretory phenotype (SASP), which promotes inflammation and tissue remodeling, thereby contributing to cardiac dysfunction. Cardiac fibroblasts (CFs), key regulators of cardiac repair, differentiate into myofibroblasts (CMFs) in response to pathological stimuli such as mechanical stiffness and TGF-β1. CMFs secrete abundant extracellular matrix (ECM) proteins, and while their senescence may transiently restrict fibrosis in acute cardiac injury, persistent senescence promotes chronic remodeling through SASP activity. Given that TGF-β1 is a central mediator of CF-to-CMF differentiation and can induce senescence in various cell lines, we investigated whether it simultaneously triggers both processes in neonatal rat CFs, and whether the senotherapeutics Navitoclax and Dasatinib + Quercetin modulate the viability of senescent CMFs.MethodsCF from neonatal rats were incubated in DMEM-F12 (10% FBS), and treated with TGF-β1 (10 46 ng/mL, 7 days). Proteins were analyzed by Western blot and immunocytochemistry. Cytokines were analyzed by Milliplex. After 7 days cells were treated with senolytics drugs.ResultsCFs treated with TGF-β1 exhibited senescence markers (p15-p16, p21), increased SA-β-gal activity, and reduced Ki-67 and p-Rb expression. CMFs displayed enhanced secretion of IL-1β, IL-6, IL-5, and IL-10, together with elevated collagen and VCAM-1 levels, indicating a pro-fibrotic and low-grade inflammatory phenotype. Treatment with Navitoclax or Dasatinib + Quercetin decreased cell viability and reduced the proportion of SA-β-gal -positive cells.ConclusionTGF-β1 simultaneously promotes CF differentiation into CMFs and induces senescence, resulting in a sustained pro-fibrotic and inflammatory phenotype. Senotherapeutics treatment attenuates senescent CMFs, supporting its potential as a therapeutic strategy to mitigate cardiac fibrosis.