Hypoxia uniquely activated profibrotic gene programs in right ventricular fibroblasts, revealing dynamic transitions from an early stress-response phase to specialized late phenotypes.
Hypoxia-induced pulmonary hypertension drives unique, progressive profibrotic transcriptional changes specifically in right ventricular fibroblasts, identifying potential therapeutic windows for RV remodeling.
Abstract Background Pulmonary hypertension (PH) causes chronic elevation of right ventricular (RV) afterload, driving progressive remodeling associated with poor clinical outcomes. Cardiac fibroblasts regulate extracellular matrix (ECM) composition and fibrosis, yet their temporal and regional transcriptional programs during RV adaptation remain poorly defined. We hypothesized that RV fibroblasts display temporally regulated transcriptional programs during hypoxia-induced pulmonary hypertension that are not shared with fibroblasts from the left ventricle (LV) or septum. Methods C57BL/6 mice were exposed to hypobaric hypoxia (18,000 ft) for 3, 7, or 21 days (n = 2 males and females each timepoint). RV, LV, and septum were analyzed by single-cell RNA sequencing (10X Genomics Chromium Fixed RNA Profiling). Fibroblasts were examined using the SLIDE (Significant Latent Factor Interaction Discovery and Exploration) framework to identify latent transcriptional programs across three intervals: normoxia-3 days (early), 3-7 days (intermediate), and 7-21 days (chronic). Gene ontology and clustering analyses were performed to define dynamic pathways of fibroblast activation. Results Following hypoxia, fibrosis-associated genes (Postn, Col1a1, Tgfb1, Acta2, Meox1) were selectively upregulated in the RV with a 7-day peak, indicating region-specific activation despite equivalent exposure across cardiac chambers (Figure 1). From normoxia-3 days, RV fibroblasts displayed a uniform stress-response characterized by cytokine-STAT3 (Osmr, Stat3, Tlr3), ER-stress (Hspa5, Manf, Hsp90aa1), and early matrix-remodeling (Eln, Loxl2, Adamtsl2, Col1a1, Col3a1) pathways. From 3-7 days, SLIDE analysis identified two distinct fibroblast populations: one cluster enriched for 3-day fibroblasts exhibited higher expression of stress-adaptive programs consistent with acute activation of redox and proteostatic pathways, while a second cluster composed of roughly equal proportions of 3- and 7-day fibroblasts showed lower expression of antioxidant and ER-stress-response genes (Mt1, Mt2, Hspa5, Manf, Hyou1), suggesting a transition from early stress-responsive fibroblasts to a matrix-oriented, lower-stress phenotype by day 7. From 7-21 days, fibroblasts further diversified into eight transcriptionally distinct subtypes with dominant antioxidant, proteostatic, metabolic, stress-inflammatory, contractile, matrix-modulating, and biosynthetic programs. Conclusions Although all cardiac regions experienced the same hypoxic stimulus, RV fibroblasts uniquely activated profibrotic gene programs, revealing intrinsic regional susceptibility that may underlie the selective remodeling seen in PH. SLIDE analysis revealed dynamic transitions in RV fibroblasts: an early stress-response phase, a transitional phase with divergence into stress-adaptive and matrix-producing populations, and a late phase composed of specialized phenotypes. Distinct from identity-based clustering, SLIDE uncovered latent transcriptional programs that define progressive shifts in fibroblast states during hypoxia-PH RV remodeling, identifying potential therapeutic windows for selective modulation of fibroblast activation. This abstract is funded by: Janssen Pharmaceutical Companies of Johnson and Johnson ATS Early Career Investigator Award in Pulmonary Vascular Disease Program
Gu et al. (Fri,) conducted a other in Hypoxia-induced pulmonary hypertension. Hypobaric hypoxia vs. Normoxia / Left ventricle and septum was evaluated on Latent transcriptional programs and fibroblast activation pathways. Hypoxia uniquely activated profibrotic gene programs in right ventricular fibroblasts, revealing dynamic transitions from an early stress-response phase to specialized late phenotypes.