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Aging of the microenvironment influences clonality in hematopoiesis.

Vas V, Senger K, Dörr K, Niebel A, Geiger H - PLoS ONE (2012)

Bottom Line: Based on a retroviral-insertion mutagenesis approach to generate primitive hematopoietic cells with an intrinsic potential for clonal expansion, we determined clonality of transduced hematopoietic progenitor cells (HPCs) exposed to a young or aged microenvironment in vivo.While HPCs displayed primarily oligo-clonality within a young microenvironment, aged animals transplanted with identical pool of cells displayed reduced clonality within transduced HPCs.Our data show that an aged niche exerts a distinct selection pressure on dominant HPC-clones thus facilitating the transition to mono-clonality, which might be one underlying cause for the increased age-associated incidence of leukemia.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology and Allergic Diseases, University of Ulm, Ulm, Germany.

ABSTRACT
The mechanisms of the age-associated exponential increase in the incidence of leukemia are not known in detail. Leukemia as well as aging are initiated and regulated in multi-factorial fashion by cell-intrinsic and extrinsic factors. The role of aging of the microenvironment for leukemia initiation/progression has not been investigated in great detail so far. Clonality in hematopoiesis is tightly linked to the initiation of leukemia. Based on a retroviral-insertion mutagenesis approach to generate primitive hematopoietic cells with an intrinsic potential for clonal expansion, we determined clonality of transduced hematopoietic progenitor cells (HPCs) exposed to a young or aged microenvironment in vivo. While HPCs displayed primarily oligo-clonality within a young microenvironment, aged animals transplanted with identical pool of cells displayed reduced clonality within transduced HPCs. Our data show that an aged niche exerts a distinct selection pressure on dominant HPC-clones thus facilitating the transition to mono-clonality, which might be one underlying cause for the increased age-associated incidence of leukemia.

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The clonal composition of the hematopoiesis is different in old compared to young microenvironment. (A)Schematic representation of the gammaretroviral SF91/IRES-eGFP vector used for insertional mutagenesis and the experimental setup (LTR: long terminal repeat with strong enhancer element, wPRE: woodchuck hepatitis virus posttranscriptional regulatory element). Lineage depleted (lin-) BM cells from young C57BL/6 mice were pre-stimulated with a cytokine cocktail and transduced. Cells were subsequently transplanted in equal amounts into young and aged recipient mice. (The graft contained 51.6%, 30.7%, 32.4% GFP+ cells in the 3 transduction/transplantation.) 24–26 week post-transplantation recipients were sacrificed and PB, spleen and BM analyzed by flow cytometry for lineage differentiation markers as well as the number of primitive L-S+K+ cells. At the same time CFC-assays on BM cells were performed and DNA from GFP+ colonies (representative pictures) isolated for LM-PCR. GFP expression among BM cells of young (B) and aged (C) recipient mice. (young mice n = 6, aged mice n = 5, from 3 independent experiments).
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pone-0042080-g001: The clonal composition of the hematopoiesis is different in old compared to young microenvironment. (A)Schematic representation of the gammaretroviral SF91/IRES-eGFP vector used for insertional mutagenesis and the experimental setup (LTR: long terminal repeat with strong enhancer element, wPRE: woodchuck hepatitis virus posttranscriptional regulatory element). Lineage depleted (lin-) BM cells from young C57BL/6 mice were pre-stimulated with a cytokine cocktail and transduced. Cells were subsequently transplanted in equal amounts into young and aged recipient mice. (The graft contained 51.6%, 30.7%, 32.4% GFP+ cells in the 3 transduction/transplantation.) 24–26 week post-transplantation recipients were sacrificed and PB, spleen and BM analyzed by flow cytometry for lineage differentiation markers as well as the number of primitive L-S+K+ cells. At the same time CFC-assays on BM cells were performed and DNA from GFP+ colonies (representative pictures) isolated for LM-PCR. GFP expression among BM cells of young (B) and aged (C) recipient mice. (young mice n = 6, aged mice n = 5, from 3 independent experiments).

Mentions: Pools of cells from the same transduction procedure were subsequently transplanted into young (2 month) and aged (18–19 months) recipient mice and GFP chimerism in peripheral blood was monitored by flow cytometry to follow the fate of the transduced cell population in vivo. 24–26 weeks post transplantation, when hematopoiesis is driven by the transplanted hematopoietic stem cells, GFP+ (transduced) cells in PB, spleen and BM were analyzed in detail (Figure 1A). This experimental set-up combined two goals: (1) The determination of clonality was possible as HSCs were individually marked by the random nature of the genomic site of the retroviral-integration. (2) Integration of this vector elicits retroviral insertional mutagenesis, and the transduced cells are prone to form dominant clones, serving as a model system for the initiation of leukemogenesis.


Aging of the microenvironment influences clonality in hematopoiesis.

Vas V, Senger K, Dörr K, Niebel A, Geiger H - PLoS ONE (2012)

The clonal composition of the hematopoiesis is different in old compared to young microenvironment. (A)Schematic representation of the gammaretroviral SF91/IRES-eGFP vector used for insertional mutagenesis and the experimental setup (LTR: long terminal repeat with strong enhancer element, wPRE: woodchuck hepatitis virus posttranscriptional regulatory element). Lineage depleted (lin-) BM cells from young C57BL/6 mice were pre-stimulated with a cytokine cocktail and transduced. Cells were subsequently transplanted in equal amounts into young and aged recipient mice. (The graft contained 51.6%, 30.7%, 32.4% GFP+ cells in the 3 transduction/transplantation.) 24–26 week post-transplantation recipients were sacrificed and PB, spleen and BM analyzed by flow cytometry for lineage differentiation markers as well as the number of primitive L-S+K+ cells. At the same time CFC-assays on BM cells were performed and DNA from GFP+ colonies (representative pictures) isolated for LM-PCR. GFP expression among BM cells of young (B) and aged (C) recipient mice. (young mice n = 6, aged mice n = 5, from 3 independent experiments).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3412859&req=5

pone-0042080-g001: The clonal composition of the hematopoiesis is different in old compared to young microenvironment. (A)Schematic representation of the gammaretroviral SF91/IRES-eGFP vector used for insertional mutagenesis and the experimental setup (LTR: long terminal repeat with strong enhancer element, wPRE: woodchuck hepatitis virus posttranscriptional regulatory element). Lineage depleted (lin-) BM cells from young C57BL/6 mice were pre-stimulated with a cytokine cocktail and transduced. Cells were subsequently transplanted in equal amounts into young and aged recipient mice. (The graft contained 51.6%, 30.7%, 32.4% GFP+ cells in the 3 transduction/transplantation.) 24–26 week post-transplantation recipients were sacrificed and PB, spleen and BM analyzed by flow cytometry for lineage differentiation markers as well as the number of primitive L-S+K+ cells. At the same time CFC-assays on BM cells were performed and DNA from GFP+ colonies (representative pictures) isolated for LM-PCR. GFP expression among BM cells of young (B) and aged (C) recipient mice. (young mice n = 6, aged mice n = 5, from 3 independent experiments).
Mentions: Pools of cells from the same transduction procedure were subsequently transplanted into young (2 month) and aged (18–19 months) recipient mice and GFP chimerism in peripheral blood was monitored by flow cytometry to follow the fate of the transduced cell population in vivo. 24–26 weeks post transplantation, when hematopoiesis is driven by the transplanted hematopoietic stem cells, GFP+ (transduced) cells in PB, spleen and BM were analyzed in detail (Figure 1A). This experimental set-up combined two goals: (1) The determination of clonality was possible as HSCs were individually marked by the random nature of the genomic site of the retroviral-integration. (2) Integration of this vector elicits retroviral insertional mutagenesis, and the transduced cells are prone to form dominant clones, serving as a model system for the initiation of leukemogenesis.

Bottom Line: Based on a retroviral-insertion mutagenesis approach to generate primitive hematopoietic cells with an intrinsic potential for clonal expansion, we determined clonality of transduced hematopoietic progenitor cells (HPCs) exposed to a young or aged microenvironment in vivo.While HPCs displayed primarily oligo-clonality within a young microenvironment, aged animals transplanted with identical pool of cells displayed reduced clonality within transduced HPCs.Our data show that an aged niche exerts a distinct selection pressure on dominant HPC-clones thus facilitating the transition to mono-clonality, which might be one underlying cause for the increased age-associated incidence of leukemia.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology and Allergic Diseases, University of Ulm, Ulm, Germany.

ABSTRACT
The mechanisms of the age-associated exponential increase in the incidence of leukemia are not known in detail. Leukemia as well as aging are initiated and regulated in multi-factorial fashion by cell-intrinsic and extrinsic factors. The role of aging of the microenvironment for leukemia initiation/progression has not been investigated in great detail so far. Clonality in hematopoiesis is tightly linked to the initiation of leukemia. Based on a retroviral-insertion mutagenesis approach to generate primitive hematopoietic cells with an intrinsic potential for clonal expansion, we determined clonality of transduced hematopoietic progenitor cells (HPCs) exposed to a young or aged microenvironment in vivo. While HPCs displayed primarily oligo-clonality within a young microenvironment, aged animals transplanted with identical pool of cells displayed reduced clonality within transduced HPCs. Our data show that an aged niche exerts a distinct selection pressure on dominant HPC-clones thus facilitating the transition to mono-clonality, which might be one underlying cause for the increased age-associated incidence of leukemia.

Show MeSH
Related in: MedlinePlus