Date of Award
Open Access Thesis
Medical Doctor (MD)
Bernard Forget, Peter Tattersall
Bone marrow-derived cells (BMDCs) have significant plasticity allowing them to give rise to various non-hematopoietic cell types including epithelial cells of the lung, liver, gut, and skin. These findings offer tremendous possibilities for the use of cell therapy to treat tissue injury and disease. However, the specific populations of cells responsible for this phenomenon are still unclear with considerable controversy over the mechanism of this transformation. In this study, we sought to compare the epithelial engraftment ability of two populations enriched for hematopoietic stem cells (HSCs), specifically the fractionated, lineage-depleted, and homed subset (FLH) and the lineage-Sca-1+c-Kit+ sorted population (LSK), to whole bone marrow (WBM). Plasticity capability was assessed by examining the engraftment of type II (T2) pneumocytes in the lung following sex-mismatched bone marrow transplantation in mice. The recipient mice were knockouts of surfactant protein C (SPC), a T2-specific protein, thereby allowing detection of the transplanted wild-type cells in the lung using immunofluorescence for SPC on paraffin sections and cytospins. Additionally, quantitative PCR (qPCR) for SPC transcripts provided a sensitive method of detecting engraftment of these BMDCs. Our data suggest that the FLH and LSK populations engraft in the lung at least as well as the control WBM. Despite our effective detection techniques, only exceedingly rare donor-derived T2 cells could be found by both microscopy and qPCR. Therefore, fluorescence-activated cell sorting (FACS) of the digested lung was tested and found to be reliable in isolating and enriching for these conversion events. Vav ancestry mice were also utilized in a similar transplantation model to directly evaluate the engraftment ability of the hematopoietic and non-hematopoietic fractions of the bone marrow. Preliminarily, the hematopoietic populations showed higher levels of epithelial engraftment by both immunofluorescence and qPCR. Interestingly, a second dose of targeted lung irradiation was necessary to elicit this effect suggesting that greater levels of tissue damage may be necessary for this model's success. When taken as a whole, our results seem to implicate HSC subpopulations as enriched for highly plastic cells which are able to engraft as T2 cells in the lung. The mechanism behind this conversion still remains to be studied but we hypothesize that both cell fusion and incidental entrapment of transplanted BMDCs are responsible.
Cohen, Justin Brent, "ASSESSING PLASTICITY: THE POPULATIONS RESPONSIBLE FOR THE EPITHELIAL ENGRAFTMENT OF MARROW-DERIVED CELLS" (2010). Yale Medicine Thesis Digital Library. 131.