β₂‑Adrenergic Receptor Disruption Exacerbates Cardiac Injury and Fibroblast Activation in a Female Mouse Model of Takotsubo Cardiomyopathy

Takotsubo cardiomyopathy (TTC) is an acute stress‑induced cardiac syndrome that predominantly affects women and is driven by surges in catecholamines that excessively activate β‑adrenergic receptors (βARs). Although β₁AR signaling mediates much of the injury, β₂ARs have recognized cytoprotective roles in other cardiac settings, yet their contribution to TTC‑associated remodeling remains unclear. To address this gap, we induced a TTC‑like phenotype in female wild‑type (WT) and β₂AR‑deficient (β2AR-/-) mice using a single high dose of isoproterenol (ISO). Following ISO injection, β2AR-/- mice treated with ISO exhibited exacerbated myocardial injury, characterized by greater hypertrophy, higher levels of apoptosis and necrosis compared with WT/ISO mice. This heightened injury was accompanied by a more robust inflammatory response, including increased inflammatory score, enhanced CD68⁺ macrophage infiltration, and marked recruitment of CCR2+MHC-IIlow monocytes. β₂AR-/-/ISO hearts also displayed more extensive interstitial fibrosis. Because fibrosis is a key driver of long-term functional decline, we isolated cardiac fibroblasts (CFs) and characterized their activation state. CFs from β2AR-/-/ISO hearts displayed a significantly higher percentage of α-SMA+ cells, increased collagen 3 and MMP-2 staining, along with upregulation of pro‑fibrotic genes (Col1a1, Col3a1 and Fap). Functionally, β₂AR-/-/ISO CFs exhibited an activated molecular signature enriched in cytokines and growth factors, and their conditioned media induced greater hypertrophy in neonatal cardiomyocytes, revealing a potent paracrine contribution to the remodeling process. These findings demonstrate that the female heart relies on β₂AR signaling to limit acute catecholamine‑induced injury, underscoring the potential of β₂AR-targeted interventions as therapeutic strategies in a Takotsubo‑like setting.