mAKAPβ targeted gene therapy prevents cerebral arteriole endothelial dysfunction and vascular stiffness in female Ossabaw swine with cardiometabolic heart failure

Heart failure with preserved ejection fraction (HFpEF) is more prevalent in females compared to males, and ~75% of HFpEF patients experience cognitive impairment, thus increasing their risk of hospitalization and mortality. We previously demonstrated cerebrovascular insufficiency in female Ossabaw swine with cardiometabolic HFpEF (HF) and improved cardiac function and ventricular-vascular interactions by a novel mAKAPβ targeted gene therapy (AAV9sc.AKAP6). AAV9sc.AKAP6 produces a peptide (AKAP6 1694-1833) which prevents RSK3 and CAMKII binding to mAKAPβ, ultimately reducing cardiac fibrosis and infiltration of inflammatory CD3+ T-cells. The goal of this study was to examine the effects of AAV9sc.AKAP6 on endothelial function, vascular stiffness, transcriptomic signaling, and protein expression in cerebral arterioles of an Ossabaw swine model of HF. Female Ossabaws were assigned to HF (HF; n=4-5), HF+AAV9sc.AKAP6 (HF+AKAP6; n=4-7), and Lean control (LC; n=5). LC (12 mo. old) were fed standard chow while HF and HF+AKAP6 were fed a Western diet (3 mo. old) and aortic banded (6 mo. old) prior to terminal experiments (12 mo. old). HF+AKAP6 underwent infusion of intravenous saline containing 3 x 1014 viral genomes of AAV9sc.AKAP6 biologic immediately after aortic banding. AAV9sc.AKAP6 decreased wet lung weight (283±19 vs 423±64g, unpaired t-test, p< 0.05), prevented diastolic dysfunction (EDPVR: 0.022±0.003 vs 0.039±0.004mmHg/mL; unpaired t-test, p< 0.05) and increased stroke volume (50±2 vs 40±2mL; unpaired t-test, p< 0.05), compared to HF. In second order cerebral arterioles isolated from the middle cerebral artery, reductions in bradykinin-induced endothelial-dependent dilation in HF vs LC (53.5 vs 75.0% max dilation) were prevented by AAV9sc.AKAP6 (84.6%; Two-way ANOVA, Group x Dose interaction, p< 0.05). Further, increased vascular stiffness (incremental modulus of elasticity) in HF vs LC (15 ± 2 vs 6 ± 1 x106 dyn/cm2), was prevented by AAV9sc.AKAP6 (6±1 x106 dyn/cm2; One-way ANOVA, p< 0.05). However, reductions in cerebral blood flow in HF vs LC (22.9±4.48 vs 41.0±4.59mL/min/100g), were unaltered by AAV9sc.AKAP6 (27.7±3.44mL/min/100g; One-way ANOVA, p< 0.05). RNA sequencing of cerebral arterioles identified 893 differentially expressed genes (DEGs) in HF vs LC, 1,535 DEGs in HF+AKAP6 vs LC, and 26 DEGs in HF+AKAP6 vs HF. Ingenuity pathway analysis (IPA) identified decreased endothelial cell movement and cell junction organization, and increased immune cell infiltration in HF vs LC, decreased immune cell infiltration in HF+AKAP6 vs LC, and inhibition of POR, an upstream regulator of fibrosis, in HF+ AKAP6 vs HF (z-score < -2 or > 2). While immunofluorescence demonstrated increased expression of POR in the prefrontal cortex (PFC) of HF (64.81±14.31%) vs LC (6.60±3.32%), this increase was unaltered by AAV9sc.AKAP6 (69.25±20.47%; One-way ANOVA, p< 0.05) and similarly unchanged in the hippocampus (HC; HF vs HF+AKAP6: 0.46±0.11 vs 0.31±0.11%). Total and perivascular fibrosis were also unaltered in the PFC and HC in HF vs HF+AKAP6 (0.83±0.12 vs 1.06±0.63; 1.54±0.63 vs 2.39±0.36%). These findings demonstrate AAV9sc.AKAP6-induced improvement in cerebral arteriole endothelial function is linked with reduced vascular stiffness, but unrelated to alterations in cerebral blood flow, POR expression, and collagen deposition. Future studies will investigate the role of endothelial cell junction integrity and inflammatory immune cell infiltration in AAV9sc.AKAP6 mediated improvements in cerebral vascular function. Funding: MU Tier 1 Sequencing Grant and Department of Defense (DOD) Grant W81XWH-18-1-0179 This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.