An In Vivo Analysis of Hematopoietic Stem Cell Potential

Abstract

Recent studies evaluating hematopoietic stem cell (HSC) potential raise the possibility that, in addition to embryonic sources, adult valve fibroblasts may be derived from HSCs. To test this hypothesis, we used methods that allow the potential of a single HSC to be evaluated in vivo. This was achieved by isolation and clonal expansion of single lineage-negative (Lin-), c-kit(+), Sca-1(+), CD34- cells from the bone marrow of mice that ubiquitously express enhanced green fluorescent protein (EGFP) combined with transplantation of individual clonal populations derived from these candidate HSCs into a lethally irradiated congenic non-EGFP mouse. Histological analyses of valve tissue from clonally engrafted recipient mice revealed the presence of numerous EGFP+ cells within host valves. A subpopulation of these cells exhibited synthetic properties characteristic of fibroblasts, as evidenced by their expression of mRNA for procollagen 1alpha1. Further, we show by Y-chromosome-specific fluorescence in situ hybridization analysis of female-to-male transplanted mice that the EGFP+ valve cells are the result of HSC-derived cell differentiation and not the fusion of EGFP+ donor cells with host somatic cells. Together, these findings demonstrate HSC contribution to the adult valve fibroblast population.

Identification and Characterization of Aortic Valve Mesenchymal Progenitor Cells with Robust Osteogenic Calcification Potential

“…40 While some of the mechanisms involved in valve disease are similar to those in atherosclerosis, recent observations suggest anatomical, cellular, and genetic mechanisms that are unique to the aortic valve. 12,40,41 Accordingly, an improved understanding of the pathogenesis of calcific aortic valve disease requires better definition of these unique features, including the characteristics of the cell populations involved in the disease. In the present study, we identified for the first time a mesenchymal progenitor cell population in the aortic valve using rigorous quantitative assays established in the mesenchymal stem cell field.…”

Congenital Heart Disease: In Search of Remedial Etiologies

“…In quantitative terms, bone marrow-derived cells accounted for 20-30 % of fibroblasts in normal adult myocardial tissue [36,37,39]. In contrast, we found GFP+ label in myocytes at exceedingly low frequency [36] making this approach suitable for a targeted assessment of non-myocyte contributions to cardiac structure and function. It is important to recall, in this context, that not all lineage markers of hematopoietic cell sources (e.g., CD34) are applicable across species.…”

“…Recent single cell engraftment experiments indicate that in postnatal and adult life, cardiac fibroblasts are also derived from bone marrow or from pericytes that express the hematopoietic stem cell marker -CD45 [33][34][35][36][37][38]. In these experiments, adult mice were lethally irradiated and a single (or clone) of a rigorously isolated wild-type CD45+ hematopoietic stem cell (HSC) carrying a green fluorescent protein (GFP) marker was injected into their tail vein.…”

“…We found that CD45+/GFP+ cells migrate and engraft in several organs including the heart. Specifically they engrafted into the inlet and outlet valves, ventricular fibrous interstitium, and as pericytes surrounding coronary microvasculature [36,37]. In addition to the CD45 marker and GFP, they also expressed fibroblast markers, e.g., collagen-1, HSP47, vimentin, discoid domain receptor 2 (DDR2), and, importantly, periostin [33,37].…”

“…Importantly, their numbers increase in the valves and ventricular interstitium significantly if the heart was injured by coronary ligation or cryoablation, indicating that they are also a population of fibrogenic precursors that can respond dynamically to injury or inflammatory signals [38,40]. In quantitative terms, bone marrow-derived cells accounted for 20-30 % of fibroblasts in normal adult myocardial tissue [36,37,39]. In contrast, we found GFP+ label in myocytes at exceedingly low frequency [36] making this approach suitable for a targeted assessment of non-myocyte contributions to cardiac structure and function.…”