Optimization of endothelialization in decellularized lung

• Grant number: 19/24185-6
• Support Opportunities: Regular Research Grants
• Start date: September 01, 2020
• End date: February 28, 2023
• Field of knowledge: Agronomical Sciences - Veterinary Medicine
• Agreement: CONFAP - National Council of State Research Support Foundations
• Principal Investigator: Maria Angelica Miglino
• Grantee: Maria Angelica Miglino
• Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated researchers: Andressa Daronco Cereta ; Renata Kelly da Palma

Abstract

Lung bioengineering may be a potential therapeutic tool for lung transplantation in patients at a very advanced stage of respiratory disease. Although bioengineering is a regenerative medical approach proposed for different organs, it has been applied in a smaller scale to the lungs, with few reports of preliminary attempts. Lung Bioengineering is particularly difficult given the variety of cells involved, the structural complexity of the bronchial tree and the pulmonary circulation circuit and the involvement of mechanical stimuli associated with breathing. However, many technical obstacles remain to achieve a viable reconstructed lung for transplantation. The inability to efficiently re-endothelialize pulmonary vasculature with a functional endothelium appears to be the main cause of failure of lung transplants with recellularized lungs to date. Therefore, the goal of this project is to develop an efficient protocol for re-endothelialization in decellularized rat lungs. For this purpose the specific objectives are divided into: To build a bioreactor that controls vascular resistance during the process of pulmonary re-endothelialization; To promote pulmonary re-endothelialization through induced pluripotent stem cells (iPSCs) and To evaluate endothelial maturation and restoration of permeability of the functional barrier of the recellularized lung vascular compartment. For this, the lungs will be collected and then the process of decellularization will be performed, followed by quantification of DNA, quantification of extracellular matrix components (ECM), scanning electron microscopy and double-photon multifocal microscopy in the decellularized lungs. The lungs will be re-endothelized with iPS, by a bioreactor which controls vascular resistance therein throughout the process of recellularization (21 days). The evaluation of the differentiation of the stem cells will be evaluated by RNA Isolation, Reverse Transcription (RT) - PCR and real time PCR analysis, Immunofluorescence and flow cytometry analysis. The vascular permeability of the recellularized lungs will be evaluated to verify the functional barrier. With this project we hope to obtain results that will allow us to better understand the basic biophysical mechanisms that participate in lung regeneration using a bioreactor. The results obtained will provide the basic scientific knowledge and may contribute in the medium term to the development and improvement of new therapeutic tools in respiratory medicine. The project will also help to enhance our knowledge of the specific technologies involved in bioreactors for regenerative medicine. (AU)