Limits...
Haematopoietic stem cell survival and transplantation efficacy is limited by the BH3-only proteins Bim and Bmf.

Labi V, Bertele D, Woess C, Tischner D, Bock FJ, Schwemmers S, Pahl HL, Geley S, Kunze M, Niemeyer CM, Villunger A, Erlacher M - EMBO Mol Med (2012)

Bottom Line: Thereby, we revealed a critical role for Bim and Bmf as regulators of HSPC dynamics both during early engraftment and long-term reconstitution.HSPCs derived from wild-type donors were readily displaced by Bim- or Bmf-deficient or Bcl-2-overexpressing HSPCs as early as 10 days after engraftment.Finally, we provide proof of principle that RNAi-based reduction of BIM or BMF, or overexpression of BCL-2 in human CD34(+) cord blood cells may be an attractive therapeutic option to increase stem cell survival and transplantation efficacy.

View Article: PubMed Central - PubMed

Affiliation: Division of Developmental Immunology, Biocenter, Innsbruck Medical University, Innsbruck, Austria. verena.labi@mdc-berlin.de

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Displacement of wt haematopoiesis in wt:bmf−/−, wt:bim−/− and wt:bcl-2 tg BM chimeras starts within the HSPC compartmentSixteen weeks after competitive reconstitution, recipient mice were sacrificed and the HSPC compartment was analysed by flow cytometry to determine percentages of Ly5.2+ LSK as well as LSK CD150+ cells. Significant p values (Mann–Whitney-test): wt versus bmf−/−: p = 0.03 in LSK and LSK CD150+ cells, wt versus bim−/− and wt versus bcl-2 tg cells: p = 0.01 in LSK and LSK CD150+ cells.Total numbers of Ly5.1+ and Ly5.2+ LSK cells depict the almost complete displacement of wt LSK cells in wt:bim−/− and wt:bcl-2 tg chimeras. Significant p values (Mann–Whitney-test): wt versus bim−/−: p = 0.03 in Ly5.1+ cells; wt versus bcl-2 tg cells: p = 0.01 in Ly5.1+ cells.Similarly, %Ly5.2+ LSK cells were determined 10 days after competitive reconstitution.The ratio Ly5.2+/Ly5.1+ cells within the LSK compartment changes significantly between Day 10 and week 16 in wt:bim−/− and wt:bcl-2 tg chimeras. Bars represent means of three to six animals/genotype from three independent experiments ± SEM.Representative dot plots of LSK cells derived from wt and bim−/− mice 10 days and 16 weeks after transplantation are shown.
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fig03: Displacement of wt haematopoiesis in wt:bmf−/−, wt:bim−/− and wt:bcl-2 tg BM chimeras starts within the HSPC compartmentSixteen weeks after competitive reconstitution, recipient mice were sacrificed and the HSPC compartment was analysed by flow cytometry to determine percentages of Ly5.2+ LSK as well as LSK CD150+ cells. Significant p values (Mann–Whitney-test): wt versus bmf−/−: p = 0.03 in LSK and LSK CD150+ cells, wt versus bim−/− and wt versus bcl-2 tg cells: p = 0.01 in LSK and LSK CD150+ cells.Total numbers of Ly5.1+ and Ly5.2+ LSK cells depict the almost complete displacement of wt LSK cells in wt:bim−/− and wt:bcl-2 tg chimeras. Significant p values (Mann–Whitney-test): wt versus bim−/−: p = 0.03 in Ly5.1+ cells; wt versus bcl-2 tg cells: p = 0.01 in Ly5.1+ cells.Similarly, %Ly5.2+ LSK cells were determined 10 days after competitive reconstitution.The ratio Ly5.2+/Ly5.1+ cells within the LSK compartment changes significantly between Day 10 and week 16 in wt:bim−/− and wt:bcl-2 tg chimeras. Bars represent means of three to six animals/genotype from three independent experiments ± SEM.Representative dot plots of LSK cells derived from wt and bim−/− mice 10 days and 16 weeks after transplantation are shown.

Mentions: Analysis of the erythro-myeloid lineage revealed that monocytes, granulocytes and erythroid progenitors were also derived to a great extent from the bim−/−, bmf−/− or bcl-2 tg donors 16 weeks after transplantation (Fig 2A, Supporting Information Fig 2, Table 1 and Supporting Information Table 3). Since granulocytes are short-lived cells (Pillay et al, 2010), they best mirror the composition of HSPCs at any given time point. Therefore, we concluded that bim−/− or bcl-2 tg cells account for the majority of HSPCs in BM chimeras. Indeed, around 90% of LSK or the more immature LSK CD150+ cells were derived from the Bim-deficient donor in wt:bim−/− chimeras 16 weeks after reconstitution (Fig 3A, B and E, Table 1 and Supporting Information Table III). In line with its strong mRNA induction after cytokine deprivation, but in contrast to the lack of cytoprotection upon cytokine deprivation ex vivo (Fig 1A and B), loss of Bmf also led to a significant displacement of Ly5.1+ wt LSK cells in reconstituted mice (Fig 3A and B). These data demonstrate that bim−/−, bmf−/− and bcl-2 tg LSK cells harbour a significant competitive advantage in BM reconstitution experiments and that wt haematopoiesis is suppressed or displaced completely by bim−/− and bcl-2 tg cells and to a significant degree also by bmf−/− cells. Importantly, no lymphoma or leukaemias occurred in recipient mice that were monitored in parallel over an observation period of 8 months (wt donor: n = 13, bim−/− donor: n = 14; bmf−/− donor: n = 10).


Haematopoietic stem cell survival and transplantation efficacy is limited by the BH3-only proteins Bim and Bmf.

Labi V, Bertele D, Woess C, Tischner D, Bock FJ, Schwemmers S, Pahl HL, Geley S, Kunze M, Niemeyer CM, Villunger A, Erlacher M - EMBO Mol Med (2012)

Displacement of wt haematopoiesis in wt:bmf−/−, wt:bim−/− and wt:bcl-2 tg BM chimeras starts within the HSPC compartmentSixteen weeks after competitive reconstitution, recipient mice were sacrificed and the HSPC compartment was analysed by flow cytometry to determine percentages of Ly5.2+ LSK as well as LSK CD150+ cells. Significant p values (Mann–Whitney-test): wt versus bmf−/−: p = 0.03 in LSK and LSK CD150+ cells, wt versus bim−/− and wt versus bcl-2 tg cells: p = 0.01 in LSK and LSK CD150+ cells.Total numbers of Ly5.1+ and Ly5.2+ LSK cells depict the almost complete displacement of wt LSK cells in wt:bim−/− and wt:bcl-2 tg chimeras. Significant p values (Mann–Whitney-test): wt versus bim−/−: p = 0.03 in Ly5.1+ cells; wt versus bcl-2 tg cells: p = 0.01 in Ly5.1+ cells.Similarly, %Ly5.2+ LSK cells were determined 10 days after competitive reconstitution.The ratio Ly5.2+/Ly5.1+ cells within the LSK compartment changes significantly between Day 10 and week 16 in wt:bim−/− and wt:bcl-2 tg chimeras. Bars represent means of three to six animals/genotype from three independent experiments ± SEM.Representative dot plots of LSK cells derived from wt and bim−/− mice 10 days and 16 weeks after transplantation are shown.
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fig03: Displacement of wt haematopoiesis in wt:bmf−/−, wt:bim−/− and wt:bcl-2 tg BM chimeras starts within the HSPC compartmentSixteen weeks after competitive reconstitution, recipient mice were sacrificed and the HSPC compartment was analysed by flow cytometry to determine percentages of Ly5.2+ LSK as well as LSK CD150+ cells. Significant p values (Mann–Whitney-test): wt versus bmf−/−: p = 0.03 in LSK and LSK CD150+ cells, wt versus bim−/− and wt versus bcl-2 tg cells: p = 0.01 in LSK and LSK CD150+ cells.Total numbers of Ly5.1+ and Ly5.2+ LSK cells depict the almost complete displacement of wt LSK cells in wt:bim−/− and wt:bcl-2 tg chimeras. Significant p values (Mann–Whitney-test): wt versus bim−/−: p = 0.03 in Ly5.1+ cells; wt versus bcl-2 tg cells: p = 0.01 in Ly5.1+ cells.Similarly, %Ly5.2+ LSK cells were determined 10 days after competitive reconstitution.The ratio Ly5.2+/Ly5.1+ cells within the LSK compartment changes significantly between Day 10 and week 16 in wt:bim−/− and wt:bcl-2 tg chimeras. Bars represent means of three to six animals/genotype from three independent experiments ± SEM.Representative dot plots of LSK cells derived from wt and bim−/− mice 10 days and 16 weeks after transplantation are shown.
Mentions: Analysis of the erythro-myeloid lineage revealed that monocytes, granulocytes and erythroid progenitors were also derived to a great extent from the bim−/−, bmf−/− or bcl-2 tg donors 16 weeks after transplantation (Fig 2A, Supporting Information Fig 2, Table 1 and Supporting Information Table 3). Since granulocytes are short-lived cells (Pillay et al, 2010), they best mirror the composition of HSPCs at any given time point. Therefore, we concluded that bim−/− or bcl-2 tg cells account for the majority of HSPCs in BM chimeras. Indeed, around 90% of LSK or the more immature LSK CD150+ cells were derived from the Bim-deficient donor in wt:bim−/− chimeras 16 weeks after reconstitution (Fig 3A, B and E, Table 1 and Supporting Information Table III). In line with its strong mRNA induction after cytokine deprivation, but in contrast to the lack of cytoprotection upon cytokine deprivation ex vivo (Fig 1A and B), loss of Bmf also led to a significant displacement of Ly5.1+ wt LSK cells in reconstituted mice (Fig 3A and B). These data demonstrate that bim−/−, bmf−/− and bcl-2 tg LSK cells harbour a significant competitive advantage in BM reconstitution experiments and that wt haematopoiesis is suppressed or displaced completely by bim−/− and bcl-2 tg cells and to a significant degree also by bmf−/− cells. Importantly, no lymphoma or leukaemias occurred in recipient mice that were monitored in parallel over an observation period of 8 months (wt donor: n = 13, bim−/− donor: n = 14; bmf−/− donor: n = 10).

Bottom Line: Thereby, we revealed a critical role for Bim and Bmf as regulators of HSPC dynamics both during early engraftment and long-term reconstitution.HSPCs derived from wild-type donors were readily displaced by Bim- or Bmf-deficient or Bcl-2-overexpressing HSPCs as early as 10 days after engraftment.Finally, we provide proof of principle that RNAi-based reduction of BIM or BMF, or overexpression of BCL-2 in human CD34(+) cord blood cells may be an attractive therapeutic option to increase stem cell survival and transplantation efficacy.

View Article: PubMed Central - PubMed

Affiliation: Division of Developmental Immunology, Biocenter, Innsbruck Medical University, Innsbruck, Austria. verena.labi@mdc-berlin.de

Show MeSH
Related in: MedlinePlus