Preservation of micro-architecture and angiogenic potential in a pulmonary acellular matrix obtained using intermittent intra-tracheal flow of detergent enzymatic treatment

Panagiotis Maghsoudlou,Thomas W Gilbert,Panicos Shangaris,Angelo Cenedese,John F Alcorn,Agostino Pierro,Athanasios Tyraskis,Paolo De Coppi,Alan J Burns,Martin A Birchall,Mark Turmaine,Neil J Sebire,Martin J Elliott,Simon Eaton,Anthony Atala,Giorgia Totonelli,Fanourios Georgiades,Giuseppe Orlando,Peggy Lange,Jean Marie Delalande ,Brogan Guest ,Stavros Loukogeorgakis

doi:10.1016/j.biomaterials.2013.05.015

Abstract

Tissue engineering of autologous lung tissue aims to become a therapeutic alternative to transplantation. Efforts published so far in creating scaffolds have used harsh decellularization techniques that damage the extracellular matrix (ECM), deplete its components and take up to 5 weeks to perform. The aim of this study was to create a lung natural acellular scaffold using a method that will reduce the time of production and better preserve scaffold architecture and ECM components. Decellularization of rat lungs via the intratracheal route removed most of the nuclear material when compared to the other entry points. An intermittent inflation approach that mimics lung respiration yielded an acellular scaffold in a shorter time with an improved preservation of pulmonary micro-architecture. Electron microscopy demonstrated the maintenance of an intact alveolar network, with no evidence of collapse or tearing. Pulsatile dye injection via the vasculature indicated an intact capillary network in the scaffold. Morphometry analysis demonstrated a significant increase in alveolar fractional volume, with alveolar size analysis confirming that alveolar dimensions were maintained. Biomechanical testing of the scaffolds indicated an increase in resistance and elastance when compared to fresh lungs. Staining and quantification for ECM components showed a presence of collagen, elastin, GAG and laminin. The intratracheal intermittent decellularization methodology could be translated to sheep lungs, demonstrating a preservation of ECM components, alveolar and vascular architecture. Decellularization treatment and methodology preserves lung architecture and ECM whilst reducing the production time to 3 h. Cell seeding and in vivo experiments are necessary to proceed towards clinical translation.

Highlights

Chronic obstructive pulmonary disease (COPD) is the fifth largest cause of death with a worldwide mortality of 3 million people annually
Decellularization using an intravascular approach from the pulmonary artery led to a reduction in DNA from 602 Æ 145 ng/mg to 114 Æ 22 ng/mg (p < 0.05), following 1 cycle of treatment (Fig. 1A)
The incomplete decellularization that was evident by the presence of nuclear and cytoplasmic material in hematoxylin and eosin (H&E) and Masson’s trichrome (MT) staining (Fig. 1D, G), was not improved with further cycles of treatment

Summary

Introduction

Chronic obstructive pulmonary disease (COPD) is the fifth largest cause of death with a worldwide mortality of 3 million people annually. P. Maghsoudlou et al / Biomaterials 34 (2013) 6638e6648 indicate that by 2030, COPD will become the third leading cause of death [1]. Due to the poor regenerative capability of the lung, the only definitive treatment is lung transplantation. Acute rejection, graft failure and the need for immunosuppression lead to a survival of 79% at 1 year, 53% at 5 years, and 30% at 10 years [2]. The development of a tissue-engineered lung that could be transplanted without the need for immunosuppression would provide a therapeutic alternative

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