Epithelial Cells of Donor Origin after Allogeneic HCT: Result of Cell Fusion, Transdifferentiation (plasticity) or Transfer of Epithelial Cell Precursors with the Graft?

Hematopoietic stem cells (HSC) appear to carry a degree of developmental plasticity that allows them to differentiate across boundaries of lineage, tissue, and germ layer, as evidenced by findings of donor-type epithelial cells in recipients of hematopoietic cell transplantation (HCT). However, these claims are often subject to criticism due to either the artifacts associated with determining the recipient versus donor origin of cells using XY-FISH or cross-lineage cell fusion. Alternatively, donor-type epithelial cells post-transplant could originate from epithelial cells or their precursors transferred with the graft. Here we wished to determine whether donor nasal epithelial cells can be detected in post-transplant patients, and if so, then the underlying mechanism. We collected nasal scrapings and whole blood from 23 allo-HCT survivors, either early (2–3 months, n=5) or late (4–22 years, n=18) post-transplant. To avoid the limitations of XY-FISH, chimerism of the epithelial cells was determined as follows: Nasal cells were stained with cytokeratin (CK) and CD45 antibodies. True epithelial cells (CK+CD45−) were laser captured. DNA extracted from the captured epithelial cells and blood leukocytes was PCR amplified for a panel of 15 autosomal short tandem repeat (STR) markers and an XY-differentiating locus (ABI-Identifiler_™). In addition, a combination of immunofluorescence microscopy for CK and FISH for two autosomes was used in 8 patients to assess cross-lineage cell fusion. Magnetic separation using CD36 microbeads and staining with CK was utilized to identify epithelial cell precursors in 3 additional peripheral blood stem cell graft specimens. In all 23 HCT survivors, epithelial cells of donor origin were identified accounting for 2.4% to 12.3% of the nasal epithelial cells. The percentage of donor origin nasal epithelial cells was significantly higher (p=0.004) in late (mean=7.45%) compared to early (mean = 3.88%) post-transplant. None of the nasal epithelial cells in 8 HCT survivors exhibited the presence of cell fusion, but CD36+CK+ epithelial cells were identified in all of the three graft specimens. In conclusion, even with a method obviating the artifacts of XY-FISH; donor origin epithelial cells were identified. This appears to be due to trans-differentiation of HSC into epithelial cells or the transfer of epithelial cells (or their precursors) with the graft, but not due to cell fusion.