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Normal Hematopoietic Progenitor Subsets Have Distinct Reactive Oxygen Species, BCL2 and Cell-Cycle Profiles That Are Decoupled from Maturation in Acute Myeloid Leukemia

View Article: PubMed Central - PubMed

ABSTRACT

In acute myeloid leukemia (AML) quiescence and low oxidative state, linked to BCL2 mitochondrial regulation, endow leukemic stem cells (LSC) with treatment-resistance. LSC in CD34+ and more mature CD34− AML have heterogeneous immunophenotypes overlapping with normal stem/progenitor cells (SPC) but may be differentiated by functional markers. We therefore investigated the oxidative/reactive oxygen species (ROS) profile, its relationship with cell-cycle/BCL2 for normal SPC, and whether altered in AML and myelodysplasia (MDS). In control BM (n = 24), ROS levels were highest in granulocyte-macrophage progenitors (GMP) and CD34− myeloid precursors but megakaryocyte-erythroid progenitors had equivalent levels to CD34+CD38low immature-SPC although they were ki67high. BCL2 upregulation was specific to GMPs. This profile was also observed for CD34+SPC in MDS-without-excess-blasts (MDS-noEB, n = 12). Erythroid CD34− precursors were, however, abnormally ROS-high in MDS-noEB, potentially linking oxidative stress to cell loss. In pre-treatment AML (n = 93) and MDS-with-excess-blasts (MDS-RAEB) (n = 14), immunophenotypic mature-SPC had similar ROS levels to co-existing immature-SPC. However ROS levels varied between AMLs; Flt3ITD+/NPM1wild-type CD34+SPC had higher ROS than NPM1mutated CD34+ or CD34− SPC. An aberrant ki67lowBCL2high immunophenotype was observed in CD34+AML (most prominent in Flt3ITD AMLs) but also in CD34− AMLs and MDS-RAEB, suggesting a shared redox/pro-survival adaptation. Some patients had BCL2 overexpression in CD34+ ROS-high as well as ROS-low fractions which may be indicative of poor early response to standard chemotherapy. Thus normal SPC subsets have distinct ROS, cell-cycle, BCL2 profiles that in AML /MDS-RAEB are decoupled from maturation. The combined profile of these functional properties in AML subpopulations may be relevant to differential treatment resistance.

No MeSH data available.


Related in: MedlinePlus

ROS levels in blast cells of different AML sub-groups.DCF levels in AML blasts (gated by CD45/SSC/CD117/CD34). Data is shown for total CD34+ blasts in CD34+AMLs (n = 70) (A) and total CD34− CD117+ blasts in CD34− AMLs (n = 23) (B) compared to the equivalent blast subset of control BMs (n = 24). AML samples, except for CBF-AMLs (CD34+, n = 12; CD34−, n = 1), were grouped according to Flt3ITD+/NPM1 mutational status: ITD−/NPM1wt, (CD34+, n = 37; CD34−, n = 4), ITD+/NPM1wt (all CD34+, n = 6), ITD-–/NPM1mut (CD34+, n = 7; CD34−, n = 4), ITD+/NPM1mut, (all CD34−, n = 9). Unk = represents patient samples lacking mutational data.
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pone.0163291.g004: ROS levels in blast cells of different AML sub-groups.DCF levels in AML blasts (gated by CD45/SSC/CD117/CD34). Data is shown for total CD34+ blasts in CD34+AMLs (n = 70) (A) and total CD34− CD117+ blasts in CD34− AMLs (n = 23) (B) compared to the equivalent blast subset of control BMs (n = 24). AML samples, except for CBF-AMLs (CD34+, n = 12; CD34−, n = 1), were grouped according to Flt3ITD+/NPM1 mutational status: ITD−/NPM1wt, (CD34+, n = 37; CD34−, n = 4), ITD+/NPM1wt (all CD34+, n = 6), ITD-–/NPM1mut (CD34+, n = 7; CD34−, n = 4), ITD+/NPM1mut, (all CD34−, n = 9). Unk = represents patient samples lacking mutational data.

Mentions: In CD34+ AMLs (Fig 4A), CBF-AMLs (n = 4) had globally higher ROS levels than Flt3ITD−/NPM1wild type (wt) patients (MFI 4.61 vs 2.28, p = 0.03). Flt3ITD+/NPM1wt cases (n = 6) overall also displayed significantly higher ROS than Flt3ITD–/NPM1wt AMLs (n = 44) (MFI 4.48 vs 2.28, p = 0.04) in total CD34+ cells and a trend to higher than Flt3ITD–/NPM1mutated CD34+AMLs (n = 7) (MFI 4.48 vs 2.86; p = 0.07). In 3/6 of Flt3ITD+/NPM1wt patients, all SPC types were ROS-high, with DCF MFI of >5.0, the 75% percentile for all patients (S2 Fig).


Normal Hematopoietic Progenitor Subsets Have Distinct Reactive Oxygen Species, BCL2 and Cell-Cycle Profiles That Are Decoupled from Maturation in Acute Myeloid Leukemia
ROS levels in blast cells of different AML sub-groups.DCF levels in AML blasts (gated by CD45/SSC/CD117/CD34). Data is shown for total CD34+ blasts in CD34+AMLs (n = 70) (A) and total CD34− CD117+ blasts in CD34− AMLs (n = 23) (B) compared to the equivalent blast subset of control BMs (n = 24). AML samples, except for CBF-AMLs (CD34+, n = 12; CD34−, n = 1), were grouped according to Flt3ITD+/NPM1 mutational status: ITD−/NPM1wt, (CD34+, n = 37; CD34−, n = 4), ITD+/NPM1wt (all CD34+, n = 6), ITD-–/NPM1mut (CD34+, n = 7; CD34−, n = 4), ITD+/NPM1mut, (all CD34−, n = 9). Unk = represents patient samples lacking mutational data.
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pone.0163291.g004: ROS levels in blast cells of different AML sub-groups.DCF levels in AML blasts (gated by CD45/SSC/CD117/CD34). Data is shown for total CD34+ blasts in CD34+AMLs (n = 70) (A) and total CD34− CD117+ blasts in CD34− AMLs (n = 23) (B) compared to the equivalent blast subset of control BMs (n = 24). AML samples, except for CBF-AMLs (CD34+, n = 12; CD34−, n = 1), were grouped according to Flt3ITD+/NPM1 mutational status: ITD−/NPM1wt, (CD34+, n = 37; CD34−, n = 4), ITD+/NPM1wt (all CD34+, n = 6), ITD-–/NPM1mut (CD34+, n = 7; CD34−, n = 4), ITD+/NPM1mut, (all CD34−, n = 9). Unk = represents patient samples lacking mutational data.
Mentions: In CD34+ AMLs (Fig 4A), CBF-AMLs (n = 4) had globally higher ROS levels than Flt3ITD−/NPM1wild type (wt) patients (MFI 4.61 vs 2.28, p = 0.03). Flt3ITD+/NPM1wt cases (n = 6) overall also displayed significantly higher ROS than Flt3ITD–/NPM1wt AMLs (n = 44) (MFI 4.48 vs 2.28, p = 0.04) in total CD34+ cells and a trend to higher than Flt3ITD–/NPM1mutated CD34+AMLs (n = 7) (MFI 4.48 vs 2.86; p = 0.07). In 3/6 of Flt3ITD+/NPM1wt patients, all SPC types were ROS-high, with DCF MFI of >5.0, the 75% percentile for all patients (S2 Fig).

View Article: PubMed Central - PubMed

ABSTRACT

In acute myeloid leukemia (AML) quiescence and low oxidative state, linked to BCL2 mitochondrial regulation, endow leukemic stem cells (LSC) with treatment-resistance. LSC in CD34+ and more mature CD34− AML have heterogeneous immunophenotypes overlapping with normal stem/progenitor cells (SPC) but may be differentiated by functional markers. We therefore investigated the oxidative/reactive oxygen species (ROS) profile, its relationship with cell-cycle/BCL2 for normal SPC, and whether altered in AML and myelodysplasia (MDS). In control BM (n = 24), ROS levels were highest in granulocyte-macrophage progenitors (GMP) and CD34− myeloid precursors but megakaryocyte-erythroid progenitors had equivalent levels to CD34+CD38low immature-SPC although they were ki67high. BCL2 upregulation was specific to GMPs. This profile was also observed for CD34+SPC in MDS-without-excess-blasts (MDS-noEB, n = 12). Erythroid CD34− precursors were, however, abnormally ROS-high in MDS-noEB, potentially linking oxidative stress to cell loss. In pre-treatment AML (n = 93) and MDS-with-excess-blasts (MDS-RAEB) (n = 14), immunophenotypic mature-SPC had similar ROS levels to co-existing immature-SPC. However ROS levels varied between AMLs; Flt3ITD+/NPM1wild-type CD34+SPC had higher ROS than NPM1mutated CD34+ or CD34− SPC. An aberrant ki67lowBCL2high immunophenotype was observed in CD34+AML (most prominent in Flt3ITD AMLs) but also in CD34− AMLs and MDS-RAEB, suggesting a shared redox/pro-survival adaptation. Some patients had BCL2 overexpression in CD34+ ROS-high as well as ROS-low fractions which may be indicative of poor early response to standard chemotherapy. Thus normal SPC subsets have distinct ROS, cell-cycle, BCL2 profiles that in AML /MDS-RAEB are decoupled from maturation. The combined profile of these functional properties in AML subpopulations may be relevant to differential treatment resistance.

No MeSH data available.


Related in: MedlinePlus