Noninvasive Contrast-Enhanced Ultrasound Molecular Imaging Detects Myocardial Inflammatory Response in Autoimmune Myocarditis

Background— Cardiac tests for diagnosing myocarditis lack sensitivity or specificity. We hypothesized that contrast-enhanced ultrasound molecular imaging could detect myocardial inflammation and the recruitment of specific cellular subsets of the inflammatory response in murine myocarditis. Methods and Results— Microbubbles (MB) bearing antibodies targeting lymphocyte CD4 (MB CD4 ), endothelial P-selectin (MB PSel ), or isotype control antibody (MB Iso ) and MB with a negative electric charge for targeting of leukocytes (MB Lc ) were prepared. Attachment of MB CD4 was validated in vitro using murine spleen CD4+ T cells. Twenty-eight mice were studied after the induction of autoimmune myocarditis by immunization with α-myosin-peptide; 20 mice served as controls. Contrast-enhanced ultrasound molecular imaging of the heart was performed. Left ventricular function was assessed by conventional and deformation echocardiography, and myocarditis severity graded on histology. Animals were grouped into no myocarditis, moderate myocarditis, and severe myocarditis. In vitro, attachment of MB CD4 to CD4+ T cells was significantly greater than of MB Iso . Of the left ventricular ejection fraction or strain and strain rate readouts, only longitudinal strain was significantly different from control animals in severe myocarditis. In contrast, contrast-enhanced ultrasound molecular imaging showed increased signals for all targeted MB versus MB Iso both in moderate and severe myocarditis, and MB CD4 signal correlated with CD4+ T-lymphocyte infiltration in the myocardium. Conclusions— Contrast-enhanced ultrasound molecular imaging can detect endothelial inflammation and leukocyte infiltration in myocarditis in the absence of a detectable decline in left ventricular performance by functional imaging. In particular, imaging of CD4+ T cells involved in autoimmune responses could be helpful in diagnosing myocarditis.