Abstract
OBJECTIVE: Adipocyte Fatty Acid-Binding Protein 4 (FABP4) exerts a direct negative inotropic effect on cardiac muscle, but the underlying cellular mechanisms remain elusive. This study aimed to dissect the specific effects of FABP4 on the contractility and calcium (Ca(2+)) homeostasis of isolated mouse ventricular myocytes and to characterize the functional role and critical residues of its N-terminal domain. METHODS: Contractility and intracellular Ca(2+) transients were simultaneously measured in isolated adult mouse ventricular myocytes using an IonOptix system following acute application of recombinant human FABP4 or its synthetic N-terminal peptide (FABP4(aa1-20)). L-type Ca(2+) current was assessed via the whole-cell patch-clamp technique. Dose-response curves were analyzed using non-linear regression, and site-directed mutagenesis (E15K) was performed to evaluate the functional importance of a key amino acid residue. RESULTS: FABP4 inhibited myocyte contraction in a biphasic, dose-dependent manner, with a high-affinity (EC(50) = 0.010 pM) and a low-affinity (EC(50) = 0.120 nM) component. This inhibition was largely independent of Ca(2+) handling, as Ca(2+) transient amplitude was only weakly attenuated at higher concentrations (EC(50) = 0.412 nM), and L-type Ca(2+) current was unaffected. In stark contrast, the FABP4(aa1-20) peptide also inhibited contraction (EC(50) = 0.110 nM) but did so via a Ca(2+)-dependent pathway, robustly suppressing Ca(2+) transients. Mutation of glutamic acid at position 15 (E15K) significantly attenuated the peptide's inhibitory activity. CONCLUSION: Full-length FABP4 suppresses cardiomyocyte contractility primarily through a Ca(2+)-independent pathway, likely by reducing myofilament Ca(2+) sensitivity. Conversely, its isolated N-terminal domain operates via a distinct, Ca(2+)-dependent mechanism. These findings reveal a complex dual-pathway regulation of cardiac function by FABP4 and identify its N-terminal region as a potential therapeutic target for mitigating obesity-related cardiac dysfunction.