Comparative decellularization and recellularization of normal versus emphysematous human lungs
Introduction
Devastating lung diseases, such as pulmonary fibrosis and chronic obstructive pulmonary diseases (COPD), are increasing in prevalence and remain without a cure except for lung transplantation. However, there are not enough donor lungs to match the current clinical need and transplantation efficacy is further inhibited by acute and chronic rejection and by complications from immunosuppressive drugs. Alternative options need to be explored to increase the potential supply of donor lungs and the subsequent efficacy of transplantation. A rapidly growing body of literature suggests that decellularized (acellular) whole lung scaffolds recellularized with autologous stem or progenitor cells obtained from the intended transplant recipient may provide a potential means of utilizing failed donor or even cadaveric lungs in clinical transplantation approaches [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15], [16], [17], [18], [19], [20], [21], [22]. However, lungs available for producing acellular lung scaffolds may come from older individuals with a history of pre-existing pulmonary disorders, such as emphysema or pulmonary fibrosis. It is currently unknown whether these lungs could be used to produce suitable acellular scaffolds for ex vivo tissue regeneration. In previous studies, we found that murine alveolar epithelial cells (C10) had limited survival when inoculated into acellular mouse lung scaffolds obtained from mice with experimentally-induced emphysema compared to normal mice [13]. To assess whether these results translated to repopulation of acellular emphysematous human lung tissue, acellular scaffolds were generated from cadaveric lungs obtained from emphysematous and healthy individuals. Lung architecture and residual protein content were assessed by histology, immunohistochemistry, and mass spectrometry as well as the ability of the acellular scaffolds to support short and long term recellularization with a variety of human cell types including human bronchial epithelial cells (HBEs), endothelial progenitor cells (CBFs), lung fibroblasts (HLFs), and bone marrow-derived MSCs (hMSCs). Potential scaffold toxicity and the ability of matrix components to support cell growth was also assessed on tissue culture plates coated with solubilized extracellular matrix from decellularized normal and emphysematous lungs.
Section snippets
Human lungs
Human lungs were obtained from autopsy services at Fletcher Allen Hospital in Burlington, Vermont. Classification of normal versus COPD (emphysematous) was based on review of available clinical records including known previous COPD or other lung diseases, smoking history, chest radiographs, pulmonary function testing, and use of respiratory medications. Subsequent confirmation of emphysematous changes on histologic sections were utilized to corroborate clinical data. A total of 18 lungs (7
Histologic and proteomic characterization of acellular emphysematous and normal lungs
Whole cadaveric lungs or isolated pulmonary lobes were decellularized using a protocol we have developed for lungs from larger animals and humans (Fig. 1A) [22]. The lungs were predominantly from an aged population (mean age at death 65.7 years, range 38–89 years) with no statistically significant differences (p < 0.015) between time from death until autopsy, cause of death, or additional findings at autopsy. Relevant clinical characteristics are detailed in Table 1.
Histological assessment
Discussion
Ex vivo lung bioengineering using decellularized whole lungs as scaffolds represents one potential new strategy for lung transplantation and in particular addresses the shortage of available donor lungs [1]. The acellular donor or cadaveric lungs could be inoculated with autologous stem or progenitor cells obtained from the transplant recipient and then clinically implanted after a suitable ex vivo regeneration strategy. Our group and others have demonstrated in a range of animal and human lung
Conclusions
Our data suggests that emphysematous lungs support initial engraftment and proliferation of a variety of human cell lines, but for reasons presently unknown, do not support prolonged viability. However, COPD, including emphysema, is a heterogenous collection of disorders and more extensive exploration of these findings needs to be performed in a wider range of diseased human lungs. Further, other acute pulmonary pathologies at the time of lung harvest from autopsy may also conceivably impact
Acknowledgments
The authors would like to thank the autopsy staff at Fletcher Allen Hospital, particularly Dr. Nick Hardin, Dr. Barbara Waters from the UVM Department of Pathology for technical assistance in methodologic development for identifying and securing small airways and blood vessels for cell inoculations, Dr. Rachael Oldinski from UVM College of Engineering for assistance with the calcium alginate coating, Dr. Mervin Yoder from Indiana University – Purdue University Indianapolis for supplying the
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