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The subendocardium is highly vulnerable to damage and is thus affected even in subclinical disease stages. Therefore, methods reflecting subendocardial status are of great clinical relevance for the early detection of cardiac damage and the prevention of functional impairment. The aim of this study was to investigate the potential ability of myocardial strain parameters to evaluate changes within the subendocardium.Male 129/Sv mice were injected with isoproterenol (ISO; n = 32) to induce isolated subendocardial fibrotic lesions or saline as appropriate control (n = 15). Transthoracic echocardiography was performed using a 30-MHz linear-frequency transducer coupled to a high-resolution imaging system, and acquired images were analyzed for conventional and strain parameters. The degree of collagen content within the different cardiac layers was quantified by histologic analysis and serum levels of tissue inhibitor of metalloproteinase–1, a biomarker for fibrosis, were assessed.ISO treatment induced a marked increase in subendocardial collagen content in response to cell loss (control vs ISO, 0.6 ± 0.3% vs 5.8 ± 0.9%; P < .001) and resulted in a moderate increase in left ventricular wall thickness with preserved systolic function. Global longitudinal peak strain (LS) and longitudinal strain rate were significantly decreased in ISO-treated animals (LS, −15.49% vs −11.49% [P = .001]; longitudinal strain rate, −4.81 vs −3.88 sec−1 [P < .05]), whereas radial and circumferential strain values remained unchanged. Global LS was associated with subendocardial collagen content (r = 0.46, P = .01) and tissue inhibitor of metalloproteinase–1 serum level (r = 0.52, P < .05). Further statistical analyses identified global LS as a superior predictor for the presence of subendocardial fibrosis (sensitivity, 84%; specificity, 80%; cutoff value, −14.4%).Assessment of LS may provide a noninvasive method for the detection of subendocardial damage and may consequently improve early diagnosis of cardiac diseases.STE was performed in an experimental model of isolated subendocardial fibrosis, and comprehensive morphologic-functional correlation analyses were performed.Functional impairment was present only in decreased global LS and global LSR values, whereas global systolic function and other strain parameters remained unaffected.Global LS was associated with subendocardial collagen content and serum level of TIMP-1.Global LS was identified as a superior predictor for the presence of subendocardial damage.