Targeting C/ EBP β to Suppress Myocardial Fibrosis in Hypertensive Heart Disease: Role of the ACE2 /Ang‐(1–7) Pathway

Hypertensive heart disease is characterised by ventricular remodelling and interstitial fibrosis, in which dysregulation of the renin-angiotensin system (RAS) plays a pivotal role. This study investigated whether the transcription factor CCAAT/enhancer-binding protein β (C/EBPβ) attenuates hypertensive myocardial fibrosis and remodelling by modulating the angiotensin-converting enzyme 2 (ACE2)/angiotensin-(1-7) Ang-(1-7) axis. Eight-week-old male spontaneously hypertensive rats (SHR) received tail-vein injection of lentiviral vectors encoding C/EBPβ, C/EBPβ short hairpin RNA (shRNA), or negative control (n = 6 per group), whereas Wistar-Kyoto rats injected with saline served as normotensive controls. Twelve weeks after injection, blood pressure and echocardiographic parameters were assessed, and cardiac hypertrophy and fibrosis were evaluated by heart weight, heart weight-to-body weight ratio, histology, Masson's trichrome staining with quantitative morphometry, immunohistochemistry and Western blotting. Angiotensin II (Ang II) and Ang-(1-7) levels were quantified in serum and myocardium, and circulating matrix metalloproteinase (MMP)-2/9, interleukin-6 (IL-6) and monocyte chemotactic protein-1 (MCP-1) were measured by enzyme-linked immunosorbent assay (ELISA). In parallel, primary rat cardiac fibroblasts were stimulated with Ang II and treated with lentiviral C/EBPβ overexpression in combination with ACE2-specific or control small interfering RNA (siRNA) to examine ACE2-dependent regulation of collagen I synthesis. C/EBPβ expression was markedly reduced in SHR myocardium compared with normotensive rats. C/EBPβ overexpression lowered arterial pressure, improved systolic and diastolic indices, and attenuated left ventricular hypertrophy and cardiomyocyte enlargement. Myocardial fibrosis was reduced, as shown by a smaller collagen-positive area on Masson's trichrome staining and decreased collagen I, collagen III and transforming growth factor-β1 (TGF-β1) expression. C/EBPβ overexpression shifted the renin-angiotensin system toward the ACE2/Ang-(1-7) axis, with higher ACE2 and Ang-(1-7), lower angiotensin-converting enzyme (ACE) and Ang II, and reduced IL-6 and MCP-1 levels, whereas MMP activity remained largely unchanged. In Ang II-stimulated cardiac fibroblasts, C/EBPβ upregulated ACE2 and suppressed collagen I, while ACE2 knockdown abolished these antifibrotic effects, supporting an ACE2-dependent mechanism. In contrast, C/EBPβ knockdown in vivo had no significant impact on cardiac function, remodelling, fibrosis, inflammation, or RAS components. Myocardial C/EBPβ expression is reduced in hypertensive rats, and its restoration markedly limits hypertension-induced cardiac injury. C/EBPβ overexpression lowers blood pressure, improves systolic and diastolic function and attenuates left ventricular hypertrophy, fibrosis and inflammation. These benefits are associated with a shift from the ACE/Ang II towards the ACE2/Ang-(1-7) axis and reduced collagen accumulation, supporting C/EBPβ-mediated ACE2 activation as a potential therapeutic strategy for hypertensive cardiac remodelling.