Date of Award

January 2026

Document Type

Thesis

Degree Name

Medical Doctor (MD)

Department

Medicine

First Advisor

Peter J. Gruber

Abstract

Background: Currently, pediatric valve replacements are limited by size and in many cases the need for anticoagulation and reoperation. Other than pulmonary autografts for the semilunar positions, there are no long-term studies of growth-adaptive pediatric valve replacements for atrioventricular valves. Acellular extracellular matrix (ECM) derived from a porcine small intestinal submucosa (SIS) have been routinely used in vessel repair and complex congenital cardiac reconstruction. More recently, SIS-ECMs have been used for tricuspid valve replacements. Despite document success in animals and human, SIS-ECM for tricuspid valve replacements have had variable reported outcomes in clinical case reports. Current data has been primarily limited to clinical outcomes and histology on primarily failed SIS-ECM valves, fundamentally limiting our knowledge of the biology of healthy SIS-ECM valve cellularization.

Aim: Our objective is to define the functional and molecular process of valvular cellularization and functional remodeling of SIS-ECM tricuspid valve replacements.

Methods: Sheep underwent SIS-ECM TV replacement with sacrifice at 3 and 4 months (n = 3; Early time point group) and 5 and 10 months (n=3; Late time point group). Valves were analyzed for longitudinal function using Transthoracic Echocardiogram (TTE) and subsequently harvested and divided for histological analysis using light microscopy and single-nuclei RNA sequencing.

Results: Functionally, TTEs demonstrated thin pliable leaflets with excellent coaptation and trace tricuspid regurgitation. In the early time point group, left ventricular end diastolic (LVEDV) and end systolic volumes (LVESV) significantly increased from baseline to terminal measurements (LVEDV: 96.9 ± 17.1 to 178.0 ± 15.1 mL, p = 0.008; LVESV: 44.6 ± 6.9 to 107.7 ± 27.6 mL, p = 0.036). Cardiac output also significantly increased from baseline to terminal measurement in the early cohort (5.81 ± 2.25 to 8.04 ± 2.02 L/min, p = 0.028). However, in the late time point group, baseline to terminal measurements were not significantly different for any functional measurement. Gross examination showed intact suture lines, well-healed annular and ventricular attachments, and no evidence of dehiscence or adverse chamber remodeling. Histology demonstrated reduced inflammatory cell infiltration, improved collagen organization, and progressive endothelialization from 3 to 10 months. Late explants showed significantly greater endothelialization than early explants (2.33 ± 0.91 vs 1.50 ± 0.71; p = 0.007) and significantly less intramural fibrin deposition (0.67 ± 1.03 vs 1.44 ± 1.15; p = 0.036). Spatial analysis suggested regional heterogeneity, with septal-associated regions showing a less favorable remodeling profile. Single-nucleus sequencing identified major cell populations, including macrophages, fibroblasts, valvular interstitial cells (VICs), valvular endothelial cells (VECs), osteoclast-like cells, T cells, B cells, neuronal cells, and proliferating cell populations. Early timepoints were enriched for macrophages and fibroblasts, whereas later timepoints showed increased endothelial and lymphocyte populations, supporting a staged process of inflammatory remodeling, scaffold recellularization, and tissue maturation. Transcriptional heterogeneity within macrophage, VIC, and VEC populations suggests dynamic shifts in cellular function during valve remodeling, highlighting the roles of balanced pro- and anti-inflammatory macrophage signaling and endothelial-to-mesenchymal transition in SIS-ECM valvular maturation.

Conclusion: SIS-ECM tricuspid valve replacements demonstrate preserved function and recapitulation of native valve features. Valvular remodeling is a progressive host-mediated process characterized by increasing endothelialization, resolution of early inflammatory features, and dynamic cellular recellularization of the scaffold. These findings support the potential of SIS-ECM as a biologically integrated, growth-adaptive valve replacement platform, and provide biological targets for future studies to investigate manipulating the material and environment to standardize outcomes, enhance valve maturation, and better emulate TV function and morphology to provide a reliable growth-adaptive valve replacement.

Comments

This thesis is restricted to Yale network users only. It will be made publicly available on 07/14/2028

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