## 4 Discussion  
The cardiac toxicity associated with anthracyclines is characterized by oxidative stress and apoptotic processes. Numerous studies conducted both in vitro and in vivo have suggested that the death of cardiomyocytes due to apoptosis plays a central role in the development of cardiomyopathy caused by anthracyclines [1]. In the current research, the cardiotoxic effects of epirubicin were identified through histological assessments of heart tissue. We noted disturbances in the intermediate filament network and cytoplasmic vacuolization, along with the loss of myofibrils, disorganization of fibers, and a slight increase in interstitial collagen fibers. These findings are consistent with previous observations made for other anthracyclines such as doxorubicin [19] and mitoxantrone [20]. Similar disruptions in cellular organization within the heart have been previously reported in rodent studies following epirubicin treatment [8,21]. Our results showed that co-treatment with hesperetin offers significant preservation of cardiac cell morphology, as evident from the histological analyses. Mitochondrial degeneration has also been recognized as a key aspect of subcellular damage caused by epirubicin [8,22]. Cardiac mitochondria possess an array of enzymatic defenses such as manganese-superoxide dismutase (Mn-SOD), glutathione reductase, and glutathione peroxidase, which modulate the redox balance in response to drug-induced changes [22]. Additionally, mitochondrial outer membrane permeabilization, a conduit for the release of pro-apoptotic factors like Bax and BAK, has been identified [23]. The released cytochrome c forms the apoptosome complex in the cytosol, triggering the caspase cascade, leading to target cleavage and apoptosis [24]. Conversely, Bax and Bak's effect on mitochondrial outer membrane permeabilization is inhibited by antiapoptotic Bcl-2 proteins, including Bcl-2 and Bcl-xL [25]. In this study, we investigated the influence of hesperetin (HSP) on Bcl-2 family activities in the cardiac tissue of mice administered with a cumulative dose of 12 mg/kg of epirubicin. Our findings indicate that a 100 mg/kg dose of HSP significantly reduced the activity of the pro-apoptotic protein Bax and simultaneously enhanced the activity of the anti-apoptotic protein Bcl-2. These observations suggest that HSP's inhibition of cardiomyocyte apoptosis by epirubicin might be linked to the mitochondrial signaling pathway, a conclusion that aligns with prior research indicating that cardiomyocyte death by epirubicin is mitochondally induced [26]. The HSP concentration used in our study aligns with doses documented in prior investigations, such as those by [PERSON] et al. in 2013,  
Figure 6: (**A**). TUNEL staining of cardiac sections: (**a**) control; (**b**) epirubicin group—most cells exhibit dark-stained TUNEL + nuclei (**c**); epirubicin + hesperetin group—TUNEL-positive cells were significantly decreased and exhibit less darkly stained nuclei than the Epi group; (**d**) hesperetin group. (**B**). Tunel (+) cells/field. A value of \(p<0.01\) was considered significant; *** as compared to control; *** as compared to Epi group. Magnification \(20\times\). which have been shown to attenuate oxidative stress in rodent models [27]. The reduction in apoptosis and the preservation of mitochondrial integrity observed in our histopathological study upon HSP administration could potentially be attributed to its antioxidant capabilities, suggesting that these effects might underlie the cardioprotective benefits of HSP. This is underscored by the well-known link between mitochondrial dysfunction and oxidative stress. Anthracyclines trigger cardiomyocyte apoptosis through both extrinsic and intrinsic pathways [28; 29]. Our research indicates that HSP markedly inhibits caspase-3 activation, which is typically upregulated by Epi toxicity, suggesting that HSP interferes with the intrinsic pathway of apoptosis.