(P275) DEVELOPMENT OF A BIOSCAFFOLD BASED ON DECELLULARIZED PORCINE AORTA REPOPULATED WITH HUMAN VASCULAR SMOOTH MUSCLE CELLS

Background: Aortic aneurysm is the 2nd disease affecting the aorta and the 15th leading cause of death in individuals aged ≥55 years. The endovascular procedure involves a stent graft to exclude the aneurysm from the systemic circulation. The issues on the use of grafts are no diametral expansion under physiological pressure, insufficiency of the aortic valve, and mechanical mismatch. Compatible grafts for aortic repair are needed to improve the longevity of the grafts. This research aims to develop a novel bioscaffold using decellularized extracellular matrix (dECM) from porcine and its recellularization (rECM) through seeding Human Aortic Smooth Muscle Cells (HAoSMCs) for its potential use as an aortic graft. Herein, we report the development of decellularization and recellularization protocols along with in vitro biological and mechanical characterization.

METHODS AND RESULTS: Aortas were obtained from 5 porcine and submitted to the separation of Adventitia, Media, and Intima layer, confirmed by histology analysis. Media samples were decellularized by immersion in Sodium Dodecyl Sulfate (SDS) for 2 days followed by a DNase and non-DNase treatment for 2, 3 and 7 days. To confirm the decellularization, the samples were analyzed by DNA quantification, histology, and second harmony generation (SHG) and two-photon fluorescence (TPF) microscopy. The recellularization was carried out by seeding the HAoSMCs for 7 and 14 days. Cell proliferation, scanning electron microscopy (SEM) and confocal were used to assess the cell viability, morphology and to characterize the collagen and elastin fibers. Dynamic and static tests were carried out to determine the mechanical properties of dECM and rECM.
The DNA quantification showed complete removal of the cells with the DNase treated groups compared to the non-DNase and to the native aorta (p < 0.001). The residual DNA was significantly lower compared to the native (0.5 ng/μl v/s 129.22 ng/μl). Histology and confocal microscopy images showed no cell nuclei in the samples, while the collagen and elastin fibers maintained their fibrous morphology. The scaffolds showed high cell viability after 7 (95.41%) and 14 days (98.16%). SEM images showed cells attached on the dECM. Preliminary static and dynamic mechanical test showed similarity between native and dECM.

Conclusion: Decellularization treatment efficiently removed the cells as indicated by DNA measurement. Recellularization of dECM with HAoSMCs indicated high cell viability. SHG and TPF microscopy showed collagen and elastin fiber morphology in the dECM. The preliminary findings of this study bring new perspectives in the field of the development of a potential bioscaffold for aortic repair.