Sex-Specific Remodeling Phenotypes of the Tricuspid Valve Leaflets in an Ovine Model of Functional Tricuspid Regurgitation

Abstract

BackgroundModerate to severe tricuspid regurgitation (TR) affects approximately 1.6 million Americans, yet more than 90% of patients with significant TR remain untreated. Women exhibit higher TR prevalence and more rapid disease progression than men, but the valve-intrinsic mechanisms underlying these sex disparities remain unclear. We hypothesized that sex and circulating testosterone influence tricuspid leaflet remodeling during right-sided pressure overload. MethodsFemale, castrated male (C-Male), and non-castrated male (NC-Male) adult Dorset sheep (n = 45) underwent pulmonary artery banding (PAB) and were followed for 13 {+/-} 1.5 weeks. Tricuspid leaflets were evaluated using morphometry, 3D profilometry, biaxial mechanical testing, histology, and bulk RNA sequencing. Sex-stratified differential gene expression was performed, and pathway enrichment of key biological processes were compared between sexes. ResultsPAB produced a uniform hemodynamic stimulus and equivalent moderate-to-severe TR across sex groups. Despite similar TR burden, leaflet remodeling diverged substantially by sex and castration status. C-Males developed the broadest remodeling phenotype, characterized by diffuse multi-leaflet growth, thickening, increased nuclei count, and low-strain stiffening. Females demonstrated more restricted leaflet and region-specific structural and cellular changes, along with circumferential low-strain stiffening. NC-Males exhibited preferential septal remodeling characterized by growth, thickening, increased nuclei count, and radial high-strain stiffening. Transcriptomic analysis revealed that females upregulated a focused matricellular remodeling program enriched for extracellular space organization (67 DEGs; FDR=0.025), whereas C-Males activated coordinated extracellular matrix and apoptosis-regulatory programs (388 DEGs; FDR=0.009). In contrast, NC-Males exhibited broad transcriptional response (406 DEGs) without significant pathway enrichment. ConclusionsTricuspid leaflet maladaptation during pressure overload is sex-dependent and testosterone-sensitive, involving distinct structural, mechanical, and transcriptional remodeling programs. These findings identify sex and testosterone status as previously under-recognized modulators of tricuspid valve remodeling and may help explain clinical sex disparities in TR progression. NOVELTY AND SIGNIFICANCE What is known?O_LIPulmonary hypertension and right ventricular pressure overload are linked to tricuspid leaflet remodeling through leaflet thickening, enlargement, and altered mechanical properties. C_LIO_LISex and sex-steroid hormones regulate fibrosis and extracellular matrix remodeling in cardiovascular tissues, but their role in tricuspid leaflet remodeling remains poorly understood. C_LI What new information does this article contribute?O_LISex and circulating testosterone status influence the magnitude, spatial distribution, biomechanical behavior, and transcriptional organization of tricuspid leaflet remodeling during pressure overload. C_LIO_LIFemales, castrated males, and non-castrated males develop distinct remodeling programs characterized by focused matricellular remodeling, coordinated extracellular matrix/apoptosis signaling, and diffuse transcriptional activation, respectively. C_LIO_LIThese findings identify sex and hormonal status as biological regulators of tricuspid valve maladaptation during functional tricuspid regurgitation. C_LI SummarySex differences in tricuspid regurgitation progression are recognized clinically, yet the mechanobiological basis underlying these disparities remains poorly understood. Using a controlled ovine model of pressure overload-induced secondary tricuspid regurgitation, we demonstrated that tricuspid leaflet maladaptation is a sex-specific and testosterone-sensitive process spanning structural, mechanical, and transcriptional scales. Under comparable hemodynamic overload, all animals developed significant tricuspid regurgitation, but leaflet remodeling patterns diverged substantially across sexes. Castrated male sheep exhibited the broadest maladaptive phenotype, characterized by diffuse multi-leaflet growth and thickening, increased low-stretch stiffness, and coordinated extracellular matrix and apoptosis-regulatory transcriptional programs. Female sheep developed more spatially restricted remodeling accompanied by a focused matricellular and extracellular matrix secretory response, whereas non-castrated male sheep demonstrated selective leaflet remodeling with broad, but less coordinated, transcriptional activation. Different remodeling patterns emerged in females and castrated males despite comparable testosterone levels, suggesting that testosterone depletion alone does not fully explain these tricuspid valve remodeling phenotypes. These findings establish sex and testosterone status as previously underrecognized biological regulators of tricuspid leaflet maladaptation and support the emerging view that valve leaflets are active, mechanobiologically responsive, participants in functional tricuspid regurgitation progression.