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Hematopoietic Origin of Murine Lung Fibroblasts.

McDonald LT, Mehrotra M, LaRue AC - Stem Cells Int (2015)

Bottom Line: Analysis of peripheral blood and lung tissues from engrafted mice demonstrated the ability of this population to give rise to CD45(+)/Discoidin-Domain Receptor-2(+) (DDR2) circulating fibroblast precursors (CFPs) in blood and fibroblast populations in lung.An HSC origin for lung fibroblasts was confirmed using a novel clonal cell transplantation method in which the bone marrow is reconstituted by a clonal population derived from a single HSC.Together, these findings provide evidence for an HSC contribution to lung fibroblasts and demonstrate a circulating intermediate through the CD45(+)/DDR2(+) HSC-derived CFP.

View Article: PubMed Central - PubMed

Affiliation: Research Services, Ralph H. Johnson Department of Veterans Affairs Medical Center, Charleston, SC 29401, USA ; Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29425, USA.

ABSTRACT
Multiple origins, including the bone marrow, have been suggested to contribute to fibroblast populations in the lung. Using bone marrow reconstitution strategies, the present study tested the hypothesis that the bone marrow hematopoietic stem cell (HSC) gives rise to lung tissue fibroblasts in vivo. Data demonstrate that the nonadherent bone marrow fraction is enriched for CD45(+) HSC-derived cells and was able to reconstitute hematopoiesis in lethally irradiated animals. Analysis of peripheral blood and lung tissues from engrafted mice demonstrated the ability of this population to give rise to CD45(+)/Discoidin-Domain Receptor-2(+) (DDR2) circulating fibroblast precursors (CFPs) in blood and fibroblast populations in lung. An HSC origin for lung fibroblasts was confirmed using a novel clonal cell transplantation method in which the bone marrow is reconstituted by a clonal population derived from a single HSC. Together, these findings provide evidence for an HSC contribution to lung fibroblasts and demonstrate a circulating intermediate through the CD45(+)/DDR2(+) HSC-derived CFP.

No MeSH data available.


Adherence culture enriched for the HSC-derived population in the nonadherent fraction of bone marrow. Flow cytometric analysis of CD45 expression in total bone marrow was compared to that in the 96-hour nonadherent bone marrow population. A representative analysis is shown. Panel (a) shows forward and side scatter gate and Panel (b) shows isotype control for CD45 staining gated on the live, propidium iodide (PI) negative population. A representative analysis of CD45 expression by total bone marrow is shown where Panel (c) depicts the forward and side scatter gate and Panel (d) shows CD45 staining gated on the live, PI negative population. Panel (e) depicts the forward scatter and side scatter gate for the nonadherent bone marrow fraction, and Panel (f) shows the CD45 staining gated on the live, PI negative population.
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Related In: Results  -  Collection


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fig1: Adherence culture enriched for the HSC-derived population in the nonadherent fraction of bone marrow. Flow cytometric analysis of CD45 expression in total bone marrow was compared to that in the 96-hour nonadherent bone marrow population. A representative analysis is shown. Panel (a) shows forward and side scatter gate and Panel (b) shows isotype control for CD45 staining gated on the live, propidium iodide (PI) negative population. A representative analysis of CD45 expression by total bone marrow is shown where Panel (c) depicts the forward and side scatter gate and Panel (d) shows CD45 staining gated on the live, PI negative population. Panel (e) depicts the forward scatter and side scatter gate for the nonadherent bone marrow fraction, and Panel (f) shows the CD45 staining gated on the live, PI negative population.

Mentions: Hematopoietic stem cells (HSCs) are immature cells that reside in the bone marrow and that possess the ability to give rise to colony forming cells and are capable of hematopoietic reconstitution [27–29]. These are distinct from bone marrow mesenchymal stem cells (MSCs) which are generally isolated and identified by their ability to adhere to plastic in vitro [30] and are characterized by their lack of expression of hematopoietic markers, including CD45 [31], the pan-leukocyte hematopoietic marker. In order to demonstrate that isolation based on nonadherence is enriched for the hematopoietic derived population, total bone marrow was first isolated from wild-type mice and analyzed for the expression of CD45 by flow cytometry. Approximately 68.4% ± SD 5.9% of the total, live, bone marrow population expressed CD45 (Figure 1). Total bone marrow was then plated on tissue culture treated flasks for 96 hours to allow for adherence. The nonadherent fraction was then removed and analyzed for expression of CD45 by flow cytometry. Quantification demonstrated that 93.4% ± SD 0.3% of the nonadherent fraction expressed CD45. This shows that the nonadherent bone marrow fraction obtained after 96 hours of adherence culture was significantly enriched for CD45 as compared to the total bone marrow population (93.4% ± SD 0.3% versus 68.4% ± SD 5.9%, resp.; *P = 0.002), indicating significant enrichment for the HSC-derived population (Figure 1).


Hematopoietic Origin of Murine Lung Fibroblasts.

McDonald LT, Mehrotra M, LaRue AC - Stem Cells Int (2015)

Adherence culture enriched for the HSC-derived population in the nonadherent fraction of bone marrow. Flow cytometric analysis of CD45 expression in total bone marrow was compared to that in the 96-hour nonadherent bone marrow population. A representative analysis is shown. Panel (a) shows forward and side scatter gate and Panel (b) shows isotype control for CD45 staining gated on the live, propidium iodide (PI) negative population. A representative analysis of CD45 expression by total bone marrow is shown where Panel (c) depicts the forward and side scatter gate and Panel (d) shows CD45 staining gated on the live, PI negative population. Panel (e) depicts the forward scatter and side scatter gate for the nonadherent bone marrow fraction, and Panel (f) shows the CD45 staining gated on the live, PI negative population.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4491389&req=5

fig1: Adherence culture enriched for the HSC-derived population in the nonadherent fraction of bone marrow. Flow cytometric analysis of CD45 expression in total bone marrow was compared to that in the 96-hour nonadherent bone marrow population. A representative analysis is shown. Panel (a) shows forward and side scatter gate and Panel (b) shows isotype control for CD45 staining gated on the live, propidium iodide (PI) negative population. A representative analysis of CD45 expression by total bone marrow is shown where Panel (c) depicts the forward and side scatter gate and Panel (d) shows CD45 staining gated on the live, PI negative population. Panel (e) depicts the forward scatter and side scatter gate for the nonadherent bone marrow fraction, and Panel (f) shows the CD45 staining gated on the live, PI negative population.
Mentions: Hematopoietic stem cells (HSCs) are immature cells that reside in the bone marrow and that possess the ability to give rise to colony forming cells and are capable of hematopoietic reconstitution [27–29]. These are distinct from bone marrow mesenchymal stem cells (MSCs) which are generally isolated and identified by their ability to adhere to plastic in vitro [30] and are characterized by their lack of expression of hematopoietic markers, including CD45 [31], the pan-leukocyte hematopoietic marker. In order to demonstrate that isolation based on nonadherence is enriched for the hematopoietic derived population, total bone marrow was first isolated from wild-type mice and analyzed for the expression of CD45 by flow cytometry. Approximately 68.4% ± SD 5.9% of the total, live, bone marrow population expressed CD45 (Figure 1). Total bone marrow was then plated on tissue culture treated flasks for 96 hours to allow for adherence. The nonadherent fraction was then removed and analyzed for expression of CD45 by flow cytometry. Quantification demonstrated that 93.4% ± SD 0.3% of the nonadherent fraction expressed CD45. This shows that the nonadherent bone marrow fraction obtained after 96 hours of adherence culture was significantly enriched for CD45 as compared to the total bone marrow population (93.4% ± SD 0.3% versus 68.4% ± SD 5.9%, resp.; *P = 0.002), indicating significant enrichment for the HSC-derived population (Figure 1).

Bottom Line: Analysis of peripheral blood and lung tissues from engrafted mice demonstrated the ability of this population to give rise to CD45(+)/Discoidin-Domain Receptor-2(+) (DDR2) circulating fibroblast precursors (CFPs) in blood and fibroblast populations in lung.An HSC origin for lung fibroblasts was confirmed using a novel clonal cell transplantation method in which the bone marrow is reconstituted by a clonal population derived from a single HSC.Together, these findings provide evidence for an HSC contribution to lung fibroblasts and demonstrate a circulating intermediate through the CD45(+)/DDR2(+) HSC-derived CFP.

View Article: PubMed Central - PubMed

Affiliation: Research Services, Ralph H. Johnson Department of Veterans Affairs Medical Center, Charleston, SC 29401, USA ; Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29425, USA.

ABSTRACT
Multiple origins, including the bone marrow, have been suggested to contribute to fibroblast populations in the lung. Using bone marrow reconstitution strategies, the present study tested the hypothesis that the bone marrow hematopoietic stem cell (HSC) gives rise to lung tissue fibroblasts in vivo. Data demonstrate that the nonadherent bone marrow fraction is enriched for CD45(+) HSC-derived cells and was able to reconstitute hematopoiesis in lethally irradiated animals. Analysis of peripheral blood and lung tissues from engrafted mice demonstrated the ability of this population to give rise to CD45(+)/Discoidin-Domain Receptor-2(+) (DDR2) circulating fibroblast precursors (CFPs) in blood and fibroblast populations in lung. An HSC origin for lung fibroblasts was confirmed using a novel clonal cell transplantation method in which the bone marrow is reconstituted by a clonal population derived from a single HSC. Together, these findings provide evidence for an HSC contribution to lung fibroblasts and demonstrate a circulating intermediate through the CD45(+)/DDR2(+) HSC-derived CFP.

No MeSH data available.