Abstract 4370063: Immune and Inflammatory Dysregulation Across the Single Ventricle Spectrum: A New Frontier for Therapeutic Innovation&Intervention

Abstract

Introduction: While heart failure (HF) is a leading cause of death and indication for transplant in single ventricle congenital heart disease (SV) patients, the molecular mechanisms underlying HF progression are poorly understood. Chronic inflammation is known to initiate and accelerate pathological outcomes in nearly all forms of cardiovascular disease, yet the impacts of immunologic alterations in SV patients are inadequately explored. Hypothesis: This study tests the hypothesis that SV patients display significant immunologic alterations that contribute to progressive cardiac dysfunction. Methods: Pediatric SV patients were categorized based on ventricular function and compared to healthy controls. Immune cells, sera, and plasma were isolated from whole blood using density gradient centrifugation. Bulk and single cell RNA sequencing were performed to identify dysregulated pathways and immune cell subtypes. Metabolic function was evaluated using the Seahorse Bioanalyzer (Agilent) and circulating inflammatory mediators were assessed via cytokine array, global proteomic and global lipidomic analysis. Results: RNA-seq analysis of circulating immune cells identified alterations in several key immune cell subtypes, including altered T helper cells across all SV groups, and significantly increased expression of markers for B cells, monocytes, macrophages, and proinflammatory macrophages in SV-HF. Additionally, pathways associated with immune cell activation, exhaustion, and senescence were increased across all SV groups (Figure 1A). Peripheral immune cells from both stable and SV-HF patients also exhibited dysregulated mitochondrial metabolic function, including significantly decreased basal respiration, maximal respiration, and ATP production with increased levels of extracellular acidification (Figure 1B). Moreover, several differentially expressed circulating pro-inflammatory mediators were significantly increased, such as IL-1β, CCL2, BAFF, p-selectin, CCL14, S100A8, RAGE, ICAM1, VCAM1, Cystatin C, and lactosylceramide. RNA-seq analysis of SV tissue identified immunologic alterations as a hallmark of the SV myocardium (Figure 1C). Conclusions: This study provides novel insights into the immunologic drivers of SV pathophysiology, highlighting critical pathways that could serve as targets for future therapeutic strategies to mitigate disease progression and multi-organ dysfunction in this vulnerable population.