SS-31 peptide attenuates radiation-induced cardiomyocyte senescence and mitochondrial dysfunction in vitro

Exposure to ionizing radiation, whether from radiation therapy for cancers like lung or breast cancer, or accidental exposure, can cause significant damage to healthy tissues, particularly the heart. A critical consequence of this exposure is cardiomyocyte senescence, a state of irreversible cell cycle arrest associated with inflammation and tissue dysfunction. Currently, there are no established strategies to prevent this radiation-induced cardiac injury. The mitochondrial-targeted peptide SS-31 (elamipretide) is being investigated for its protective effects against mitochondrial dysfunction, a key driver of cellular senescence and apoptosis.

This study investigated SS-31's effects on radiation-induced senescence in two in vitro models: cardiomyoblast H9C2 cells and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Cells were exposed to γ-radiation at doses of 2, 5, and 10 Gy. The intervention involved treating cells with SS-31 (1 μM) for 7 days following radiation. Primary endpoints included senescence-associated beta-galactosidase (SA-β-gal) staining, expression levels of senescence markers p16 and p21, and quantification of pro-inflammatory senescence-associated secretory phenotype (SASP) markers like TNF-α, IL-6, and IL-1β. Mitochondrial health was assessed by BAX/bcl-2 ratio, reactive oxygen species (ROS) production, and mitochondrial respiration.

Exposure to γ-radiation inhibited H9C2 cell proliferation in a dose-dependent manner and significantly induced SA-β-gal staining. Treatment with SS-31 (1 μM) for 7 days effectively reduced SA-β-gal positive staining from 67% to 38% in H9C2 cells under 10 Gy radiation. SS-31 also prevented the radiation-induced increases in the expression of p16 and p21, two critical senescence markers, in both irradiated H9C2 cells and hiPSC-CMs.

SS-31 significantly decreased the canonical senescence-associated secretory phenotype (SASP) markers TNF-α, IL-6, and IL-1β, indicating a reduction in pro-inflammatory signaling. Furthermore, SS-31 reversed the elevated BAX/bcl-2 ratio, a marker of mitochondrial-related apoptosis, and mitigated mitochondrial production of reactive oxygen species (ROS). Interestingly, 10 Gy radiation increased mitochondrial respiration, and SS-31 successfully reversed this elevation, suggesting a restoration of mitochondrial function.