Culture Into Perfusion-Assisted Bioreactor Promotes Valve-Like Tissue Maturation of Recellularized Pericardial Membrane

Abstract

Derivation of tissue-engineered valve replacements is a strategy to overcome the limitations of the current valve prostheses, mechanical, or biological. In an effort to set living pericardial material for aortic valve reconstruction, we have previously assessed the efficiency of a recellularization strategy based on a perfusion system enabling mass transport and homogenous distribution of aortic valve-derived "interstitial" cells inside decellularized pericardial material. In the present report, we show that alternate perfusion promoted a rapid growth of valve cells inside the pericardial material and the activity of a proliferation-supporting pathway, likely controlled by the YAP transcription factor, a crucial component of the Hippo-dependent signaling cascade, especially between 3 and 14 days of culture. Quantitative mass spectrometry analysis of protein content in the tissue constructs showed deposition of valve proteins in the decellularized pericardium with a high variability at day 14 and a reproducible tissue maturation at 21 days. These results represent a step forward in the definition of strategies to produce a fully engineered tissue for replacing the calcified leaflets of failing aortic valves.

A novel approach for the endothelialization of xenogeneic decellularized vascular tissues by human cells utilizing surface modification and dynamic culture

“…The combination of coating with the E8 fragment of human laminin-411 and dynamic ow culture enables su cient endothelial cell coverage on the decellularized vascular tissue Dynamic ow culture has been reported to improve the recellularization e ciency of decellularized pericardial tissues 17 , possibly owing to the reproduction of the physical conditions in the living body. We prepared a custom-made bioreactor that could confer dynamic ow of the culture medium and incubated gelatin-coated slices of porcine aorta endothelium seeded with human umbilical venous endothelial cells (HUVECs) under dynamic ow conditions (Fig.…”

“…Our results indicate that this combination was necessary for su cient recellularization of human endothelial cells as a single cell layer of the endothelium. Dynamic ow culture has been reported to improve the recellularization e ciency in various kinds of decellularized tissues 11,17,28 . In conventional methods of recellularization aiming for ex vivo tissue reconstruction before in vivo implantation, seeding cells by circulating a cell suspension (cells + culture medium) requires a relatively long duration of recellularization incubation, and results in a relatively low e ciency of recellularization 29,30 .…”

Lithotripsy of Calcified Aortic Valve Leaflets by a Novel Ultrasound Transcatheter-Based Device

“…Since application of shockwaves to tissues can determine mechanical damages to cells, we were prompted to assess the effects on cellular survival and tissue ruptures using our experimental setup. This was done by measuring the vitality of living pericardial membranes from pigs, a tissue that we already employed for engineering valve tissues after recellularization (20) and is the golden standard material for manufacturing valve replacements. As shown in Figures 2A,B, the samples, stained with 3-(4.5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) and treated with the device, exhibited no differences in the overall vitality of the treated vs. the non-treated areas on both sides of the tissue.…”

Engineering Efforts to Refine Compatibility and Duration of Aortic Valve Replacements: An Overview of Previous Expectations and New Promises

“…Interestingly, when valve cells were statically cultured on the surface of the decellularized porcine pericardium, only minimal penetration was observed ( 98 ). In contrast, the increase in tissue permeability following decellularization rendered the tissue perfusable with an oscillating perfusion bioreactor ( 88 ), which led to a homogeneous distribution of the cells throughout the entire depth of the construct thereby effecting valve-like tissue maturation ( 99 ). Despite the lack, as yet, of confirmation of total biological compatibility of the recellularized tissue or the scalability of the method, the approach seems amenable to generate a fully engineered, personally tailored (for example, using recipient mesenchymal cells), living valve for selected classes of patients (e.g., infants/children and young adults).…”