Hematopoietic Stem Cell Origin of Human Fibroblasts: Cell Culture Studies of Female Recipients of Gender Mismatch Stem Cell Transplantation and Patients with Chronic Myelogenous Leukemia

Objectives— Our series of studies using transplantation of single hematopoietic stem cells (HSCs) demonstrated that mouse fibroblasts/myofibroblasts are derived from HSCs. In order to determine the origin of human fibroblasts, we established a method for culturing fibroblasts from human peripheral blood (PB) mononuclear cells and studied fibroblasts from gender mismatch HSC transplant recipients and patients with untreated Philadelphia chromosome-positive chronic myelogenous leukemia (CML). Methods— We cultured PB cells from three female subjects who showed near complete hematopoietic reconstitution from transplantation of granulocyte-colony stimulating factor (G-CSF)-mobilized male PB cells and examined the resulting fibroblasts using fluorescent in situ hybridization (FISH) for Y chromosome. Because the mobilized PB cells may contain mesenchymal stem cells (MSCs), we could not determine the HSC or MSC origin of the fibroblasts seen in culture. To further document the HSC origin of human fibroblasts, we next examined fibroblasts from two patients with untreated CML, a known clonal disorder of HSCs. Results— All cultured fibroblasts from female recipients of male cells showed the presence of Y-chromosome, indicating the donor origin of fibroblasts. Cultured fibroblasts from the CML patients revealed the presence of BCR-ABL translocation. This demonstration provided strong evidence Conclusions— These studies strongly suggest that human fibroblasts are derived from HSCs. In addition, the results suggest that fibrosis seen in patients with CML may be a part of the clonal process. Real-time RT-PCR was conducted on total RNA extracted from PB MNCs, cultured fibroblasts and a human foreskin fibroblast cell line (HFF-1). Analysis showed that cultured human fibroblasts expressed mRNA for pro-collagen I α 1, vimentin and fibronectin at levels similar to a HFF-1 and far higher than those by PB MNCs. These findings provide biochemical support for the identity of the cultured cells as fibroblasts.

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