Engineering Dynamic 3D-Bioprinted Models of Pulmonary Vascular Disease

PROJECT SUMMARY/ABSTRACT Pulmonary arterial hypertension (PAH) is a progressive and incurable disease, characterized by elevated pulmonary blood pressure, remodeling of the pulmonary arteries, and ultimately the development of right ventricular failure. Unfortunately, the only clinically available therapeutic treatments mitigate symptoms but do

not cure the disease. Existing cell culture techniques represent a significant barrier to discovering new therapeutic targets for PAH because these systems do not adequately reproduce key aspects of human physiology, specifically, the complex 3D structure of the pulmonary vasculature and the time-dependent

changes in extracellular matrix (ECM) mechanical properties that occur during disease progression. Therefore, an urgent need remains to develop new tools and technologies that enable us to study the pathogenesis of PAH over time. We propose to develop a new class of phototunable poly(ethylene glycol) (PEG)-based

hydrogel biomaterials and biomanufacturing techniques that allow investigators to control the mechanical properties of the local microenvironment (i.e., stiffen) on-demand around patient-derived fibroblasts encapsulated within 3D-printed vascular models using focused light, with the goal of emulating PAH

pathogenesis in vitro. The advanced biomaterial platform implemented here will provide the foundation for biological models of increasing complexity comprising multiple cell types that are cultured under flow under development in the sponsor's laboratory that reveal novel mechanistic insights into reduction of human

disease. This project will bring together the applicant and a diverse mentoring team made up of bioengineers and clinician-scientists specializing in cardiovascular and pulmonary diseases to further develop the pulmonary workforce. Fellowship training will include clinical experiences through the Pulmonary Hypertension

Breakthrough Initiative, presentations at research conferences and grand rounds to aid professional development, and hands-on training in Dr. Kurt Stenmark's Cardiovascular Pulmonary Research Lab to learn experimental techniques essential to understanding PAH. The completed model will provide a platform for

testing and validating therapies for pulmonary hypertension, advancing translational research in this field. We propose two specific aims to demonstrate the feasibility of this approach. AIM I: Engineer a dynamic 3D- bioprinted cell culture platform with controllable modulus of elasticity. AIM II: Investigate the influence of

dimensionality and material modulus on fibroblast activation using patient-derived cells. The success of Aim I will be measured through rheological characterization and cell viability assays. Aim II will measure fibroblast activation through immunohistochemistry and a concise qRT-PCR array to compare phenotypic changes

among healthy patient-derived cells grown in 3D-bioprinted blood vessel mimics that emulate pathological ECM, healthy cells grown on 2D hydrogel substrates, and freshly isolated PAH patient cells.