Placental stem cells as a treatment for lung disease

The use of cells isolated from human placenta for regenerative medicine represents a field of investigation that is still in its infancy, but holds great promise on several fronts. Specifically, their plasticity, immune characteristics, lack of tumorigenicity and the absence of ethical barriers to their procurement, make them ideal candidates for the basis of further research into disease treatment. This thesis aimed to investigate the potential of human amnion epithelial cells (hAECs) to be used as a cell therapy for the treatment of lung disease. The ability to isolate, characterize and store hAECs, under conditions suitable for human use will be essential to facilitate application of these cells as a future cell therapy. I have developed methods that enable the isolation, characterization and cryopreservation of hAECs that are suitable for future clinical applications. These methods produce an average yield of 120 million viable hAECs per placenta. I also assessed important “safety” attributes of hAECs that make them attractive as a cell therapy, demonstrating that hAECs maintain a normal karyotype in culture, have conserved long telomere lengths and, unlike embryonic stem cells (ESCs), do not form teratomas in vivo. I characterized physiological changes that occur during hAEC culture, performing extensive analysis of the cell surface marker profiles of freshly isolated as well as passaged hAECs. Since our group has shown that hAECs possess low immunogenicity, I investigated the ability of hAECs to reduce inflammation and fibrosis and improve pulmonary function in a xenogenic stetting utilizing a mouse model of lung disease. I demonstrated that hAEC transplantation reduces bleomycin-induced pulmonary fibrosis, showing, for the first time, that hAECs can mitigate the inflammatory cytokine response in an immune competent mouse and, again for the first time, that these effects result in improved lung function. Interestingly, I found no evidence of hAEC engraftment in the lung, or their differentiation into functional lung epithelium in vivo. The conclusions drawn from this study suggest that, rather than restoring function to the damaged lung by integrating and differentiating into functional lung epithelial cells, hAECs exert their effect by interacting with the host immune system. I then explored the mechanisms by which hAECs are able to abrogate pulmonary inflammation and fibrosis. To facilitate this, I utilized surfactant protein C deficient (Sftpc-/-) mice and their wild-type counterparts. Sftpc-/- mice are known to have a heightened susceptibility to lung injury, secondary to impaired macrophage function, including reduced phagocytic activity and increased metalloproteinase activity. This allowed us to investigate the role of macrophages in hAEC-mediated immunemodulation in the bleomcyin injury model. In this study I confirmed that the effect of hAEC therapy was independent of engraftment and differentiation in vivo, and that hAEC therapy was not effective in the Sftpc-/- strain. By the investigating the key differences in the response to hAEC administration between the responsive wild-type strain and the non-responsive Sftpc-/- strain, I was able to determine the importance of the hAEC effect on macrophage migration and polarization to the alternatively activated phenotype. Finally, with the aim of developing a cell source for the treatment of cystic fibrosis, I demonstrated the ability of differentiated hAECs to express functional cystic fibrosis transmembrane conductance regulator (CFTR). I demonstrated hAECs can be induced to express CFTR at both the gene and protein level. Using high resolution synchrotron imaging, I showed that CFTR was located on the periphery of 3-dimensional cellular structures formed by hAEC in vitro and its expression was polarized, consistent with its function as an ion channel. Finally I demonstrated that differentiated hAECs formed ion channels whose activity is regulated by the CFTR activator, forskolin. We believe that hAECs will be a valuable tool for exploring novel therapies for cystic fibrosis. hAECs hold great promise as a tool for regenerative medicine. The questions explored in this thesis progresses the development of hAECs as a potential cell therapy to restore function to damaged or dysfunctional tissues