Abstract P233: Single-cell Transcriptomic Characterization Of Aortic Lymphocyte Heterogeneity During Hypertension

Vascular inflammation is a hallmark of hypertension. T and B lymphocytes accumulate in the aortic wall during hypertension; however, it is unclear which subtypes or functions contribute to disease progression. Thus, we aimed to characterize T and B cell subtypes, activation states and phenotypes in hypertension. Angiotensin II infusion (0.7 mg/kg/day; s.c. for 28 days, n = 15) was used to induce hypertension in 12-week-old male C57BL/6 mice. Age-matched normotensive controls received vehicle infusion (0.5% NaCl, 0.1% acetic acid; s.c. for 28 days, n = 11). Systolic blood pressure (SBP) was measured weekly by tail-cuff plethysmography. Aortic stiffness and fibrosis were assessed at endpoint via high resolution ultrasound and picrosirius red-staining, respectively. Whole aortas (n = 3 per treatment group) were isolated and digested into single-cell suspensions and enriched for live, nucleated, metabolically active cells. Single-cell RNA sequencing was performed using Chromium 10x and NovaSeq genomics platforms. Bioinformatic analyses identified cell types, acquired molecular and biological activities and inferred gene regulatory networks. At endpoint, angiotensin II-treated mice had elevated SBP (179 ± 4 vs. 124 ± 2 mmHg; P < 0.0001), with increased aortic pulse wave velocity (1.5 ± 0.2 vs 3.1 ± 0.3 m/s; P = 0.001) and collagen content (51 ± 2% vs. 60 ± 2% wall area; P = 0.02), compared to vehicle controls. Total aortic T cells and B cells increased by 16% (184 vs. 215 cells) and 74% (433 vs. 771 cells) in hypertension, respectively. These increases were attributed to the expansion of CD8+, regulatory T cells (Tregs), gamma-delta (GD) T cells, immature transitional B2 cells, naïve B2, memory B2, B1 B cells and plasma cells, with a reduction in regulatory B2 B cells (Bregs). Gene ontology analyses revealed both pro- and anti-inflammatory B and T cells were activated during hypertension. However, antigen receptor activity pathways were only enriched in GD T cells, B1 B cells and effector Bregs (running enrichment scores > 0.2). Overall, these findings identify GD T cells, B1 B cells and effector Bregs as key lymphocyte subtypes that drive vascular inflammation during experimental hypertension and aortic stiffening.