Myocardial ischemia-reperfusion injury (MIRI) induces oxidative stress and inflammatory signaling that drive fibroblast activation, myocardial fibrosis, and progressive cardiac dysfunction, for which effective targeted therapies remain limited. The electrophilic fatty acid nitroalkene 10-nitro-octadec-9-enoic acid (NO₂-FA) exhibits anti-inflammatory and antifibrotic properties but pharmacological actions have been limited by extents of myocardial delivery. Ultrasound-targeted cavitation (UTC) using lipid-shelled gas-filled nanoparticles (LNPs) enables spatially controlled drug release and represents a promising theranostic strategy. NO₂-FA were incorporated in LNPs and delivered focally to the myocardium using UTC. Therapeutic efficacy was evaluated in rodent models of MIRI and myocardial fibrosis. In the MIRI model, animals received UTC + NO₂-FA LNPs, intravenous NO₂-FA, or sham treatment. Cardiac structure, function, and molecular remodeling were assessed using echocardiography, histological staining, polymerase chain reaction, and enzyme-linked immunosorbent assay. In the myocardial fibrosis model, additional analyses included immunohistochemistry and cardiovascular magnetic resonance imaging. UTC-mediated delivery of NO₂-FA LNPs significantly reduced myocardial fibrosis compared with intravenous NO₂-FA (p = 0.03) and sham treatment (p = 0.001), as demonstrated by histological quantification and reduced late gadolinium enhancement. Targeted NO₂-FA delivery also improved cardiac output and favorably modulated molecular markers associated with fibrosis and inflammation. Across both experimental models, UTC-facilitated NO₂-FA delivery consistently demonstrated superior therapeutic efficacy relative to non-targeted administration. Spatially targeted delivery of NO₂-FA using ultrasound-mediated cavitation enhances cardioprotective and antifibrotic effects in experimental models of MIRI and myocardial fibrosis. This platform integrates targeted therapy with imaging-based assessment and supports further development of UTC-enabled theranostic approaches for ischemic heart disease.

Background: Interstitial myocardial fibrosis is a crucial pathological feature of many cardiovascular disorders. Myocardial fibrosis resulting in extracellular volume (ECV) expansion can be quantified via cardiac MRI (CMR) with T₁ mapping before and after minimally invasive gadolinium (Gd) contrast agent administration. However, longitudinal repetitive ECV measurements are challenging in rodents due to the prolonged scan time with cardiac and respiratory gating that is required for conventional T₁ mapping and the invasive nature of the rodent intravenous lines. Methods: To address these challenges, the objective of this study is to establish a fast, free-breathing, and gating-free ECV procedure using a minimally invasive subcutaneous catheter for in-scanner Gd administration that can allow longitudinal repetitive ECV evaluations in rodent models. This is achieved by the (1) IntraGate sequence for free-breathing, gating-free cardiac imaging; (2) minimally invasive subcutaneous in-scanner Gd administration; and (3) fast T₁ mapping with a varied flip angle (VFA) in conjunction with (4) triple jugular vein blood T₁ normalization. Additionally, full cine CMR (multi-slice short-axis, long-axis 2-chamber, and long-axis 4-chamber) was acquired during the waiting period to assess comprehensive cardiac function and strain. Results: We successfully established a minimally invasive fast ECV quantification protocol to enable longitudinal repetitive ECV quantifications in rodents. Minimally invasive subcutaneous Gd bolus administration induced a reasonable dynamic contrast enhancement (DCE) time course, reaching a steady state in ~20 min for stable T₁ quantification. The free-breathing gating-free VFA T₁ quantification scheme allows for rapid cardiac (~2.5 min) and jugular vein (49 s) T₁ quantification with no motion artifacts. The triple jugular vein T₁ acquisitions (1 pre-contrast and 2 post-contrast) immediately flanking the heart T₁ acquisitions enable accurate myocardial ECV quantification. Our data demonstrated that left-ventricular myocardial ECV quantification was highly reproducible with repeated scans, and the ECV values (0.25) are comparable to reported ranges among humans and rodents. This protocol was successfully applied to the ischemia–reperfusion injury model to detect myocardial fibrosis, which was validated by histopathology. Conclusions: We established a simple, fast, minimally invasive, and robust CMR protocol in rodents that can enable longitudinal repetitive ECV quantification for cardiovascular disease progression. It can be used to monitor disease regression with interventions.