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Stem and progenitor cell subsets are affected by JAK2 signaling and can be monitored by flow cytometry.

Iida R, Welner RS, Zhao W, Alberola-lla J, Medina KL, Zhao ZJ, Kincade PW - PLoS ONE (2014)

Bottom Line: Real-Time PCR analysis of their HSC revealed that the erythropoiesis associated gene transcripts Gata1, Klf1 and Epor were particularly high.The low CD86 on HSC and multipotent progenitors paralleled the large reductions we found in lymphoid progenitors, but the few that were produced functioned normally when sorted and placed in culture.They could also serve as indicators of HSC fitness and suitability for transplantation.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology and Cancer Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, United States of America.

ABSTRACT
Although extremely rare, hematopoietic stem cells (HSCs) are divisible into subsets that differ with respect to differentiation potential and cell surface marker expression. For example, we recently found that CD86(-) CD150(+) CD48(-) HSCs have limited potential for lymphocyte production. This could be an important new tool for studying hematological abnormalities. Here, we analyzed HSC subsets with a series of stem cell markers in JAK2V617F transgenic (Tg) mice, where the mutation is sufficient to cause myeloproliferative neoplasia with lymphocyte deficiency. Total numbers of HSC were elevated 3 to 20 fold in bone marrow of JAK2V617F mice. Careful analysis suggested the accumulation involved multiple HSC subsets, but particularly those characterized as CD150(HI) CD86(-) CD18(L)°CD41(+) and excluding Hoechst dye. Real-Time PCR analysis of their HSC revealed that the erythropoiesis associated gene transcripts Gata1, Klf1 and Epor were particularly high. Flow cytometry analyses based on two differentiation schemes for multipotent progenitors (MPP) also suggested alteration by JAK2 signals. The low CD86 on HSC and multipotent progenitors paralleled the large reductions we found in lymphoid progenitors, but the few that were produced functioned normally when sorted and placed in culture. Either of two HSC subsets conferred disease when transplanted. Thus, flow cytometry can be used to observe the influence of abnormal JAK2 signaling on stem and progenitor subsets. Markers that similarly distinguish categories of human HSCs might be very valuable for monitoring such conditions. They could also serve as indicators of HSC fitness and suitability for transplantation.

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Particular HSC subsets expand in JAK2V617F Tg mice.(A) LSK were resolved with CD150 and CD34. CD34− CD150Hi, Lo, and negative HSC were gated as described by Morita [10]. The percentages of CD34− HSC subsets in BM were calculated. Closed and open circles indicate individual WT and JAK2V617F mice, respectively. The data are representative of those obtained in two independent experiments (N = 6). (B) Lower side population cells in BM (spindle-shaped gating) are thought to include myeloid biased HSC. The “side population tip” was increased in JAK2V617F mice. Representative plots are shown in the left panels, while cell percentages of lower and upper cells are given in right panels. (C) Both CD86− and CD86+ CD150+ CD48− LSK cells in BM were increased in JAK2V617F mice. CD48− LSK were resolved with CD86 and CD150. Cell percentages of CD86− and CD86+ in CD150Hi were calculated. The data are representative of those obtained in three independent experiments (N = 8) p<0.01(**), p<0.05(*).
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pone-0093643-g002: Particular HSC subsets expand in JAK2V617F Tg mice.(A) LSK were resolved with CD150 and CD34. CD34− CD150Hi, Lo, and negative HSC were gated as described by Morita [10]. The percentages of CD34− HSC subsets in BM were calculated. Closed and open circles indicate individual WT and JAK2V617F mice, respectively. The data are representative of those obtained in two independent experiments (N = 6). (B) Lower side population cells in BM (spindle-shaped gating) are thought to include myeloid biased HSC. The “side population tip” was increased in JAK2V617F mice. Representative plots are shown in the left panels, while cell percentages of lower and upper cells are given in right panels. (C) Both CD86− and CD86+ CD150+ CD48− LSK cells in BM were increased in JAK2V617F mice. CD48− LSK were resolved with CD86 and CD150. Cell percentages of CD86− and CD86+ in CD150Hi were calculated. The data are representative of those obtained in three independent experiments (N = 8) p<0.01(**), p<0.05(*).

Mentions: Previous studies revealed that HSC are heterogeneous and composed of functionally specialized subpopulations [3], [4]. In some cases, this corresponds to phenotypes [10], [39], and the JAK2V617F transgene preferentially expands CD150Hi HSC (Fig. 2A). That category of HSC was reportedly less likely to generate lymphocytes than CD150Lo/− HSC [10]. Similarly, Goodell and colleagues found lymphopoietic potential was lowest among HSC that strongly exclude Hoechst dye [40]. These “lower side population” HSC were the most increased by JAK2V617F (Fig. 2B).


Stem and progenitor cell subsets are affected by JAK2 signaling and can be monitored by flow cytometry.

Iida R, Welner RS, Zhao W, Alberola-lla J, Medina KL, Zhao ZJ, Kincade PW - PLoS ONE (2014)

Particular HSC subsets expand in JAK2V617F Tg mice.(A) LSK were resolved with CD150 and CD34. CD34− CD150Hi, Lo, and negative HSC were gated as described by Morita [10]. The percentages of CD34− HSC subsets in BM were calculated. Closed and open circles indicate individual WT and JAK2V617F mice, respectively. The data are representative of those obtained in two independent experiments (N = 6). (B) Lower side population cells in BM (spindle-shaped gating) are thought to include myeloid biased HSC. The “side population tip” was increased in JAK2V617F mice. Representative plots are shown in the left panels, while cell percentages of lower and upper cells are given in right panels. (C) Both CD86− and CD86+ CD150+ CD48− LSK cells in BM were increased in JAK2V617F mice. CD48− LSK were resolved with CD86 and CD150. Cell percentages of CD86− and CD86+ in CD150Hi were calculated. The data are representative of those obtained in three independent experiments (N = 8) p<0.01(**), p<0.05(*).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3974768&req=5

pone-0093643-g002: Particular HSC subsets expand in JAK2V617F Tg mice.(A) LSK were resolved with CD150 and CD34. CD34− CD150Hi, Lo, and negative HSC were gated as described by Morita [10]. The percentages of CD34− HSC subsets in BM were calculated. Closed and open circles indicate individual WT and JAK2V617F mice, respectively. The data are representative of those obtained in two independent experiments (N = 6). (B) Lower side population cells in BM (spindle-shaped gating) are thought to include myeloid biased HSC. The “side population tip” was increased in JAK2V617F mice. Representative plots are shown in the left panels, while cell percentages of lower and upper cells are given in right panels. (C) Both CD86− and CD86+ CD150+ CD48− LSK cells in BM were increased in JAK2V617F mice. CD48− LSK were resolved with CD86 and CD150. Cell percentages of CD86− and CD86+ in CD150Hi were calculated. The data are representative of those obtained in three independent experiments (N = 8) p<0.01(**), p<0.05(*).
Mentions: Previous studies revealed that HSC are heterogeneous and composed of functionally specialized subpopulations [3], [4]. In some cases, this corresponds to phenotypes [10], [39], and the JAK2V617F transgene preferentially expands CD150Hi HSC (Fig. 2A). That category of HSC was reportedly less likely to generate lymphocytes than CD150Lo/− HSC [10]. Similarly, Goodell and colleagues found lymphopoietic potential was lowest among HSC that strongly exclude Hoechst dye [40]. These “lower side population” HSC were the most increased by JAK2V617F (Fig. 2B).

Bottom Line: Real-Time PCR analysis of their HSC revealed that the erythropoiesis associated gene transcripts Gata1, Klf1 and Epor were particularly high.The low CD86 on HSC and multipotent progenitors paralleled the large reductions we found in lymphoid progenitors, but the few that were produced functioned normally when sorted and placed in culture.They could also serve as indicators of HSC fitness and suitability for transplantation.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology and Cancer Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, United States of America.

ABSTRACT
Although extremely rare, hematopoietic stem cells (HSCs) are divisible into subsets that differ with respect to differentiation potential and cell surface marker expression. For example, we recently found that CD86(-) CD150(+) CD48(-) HSCs have limited potential for lymphocyte production. This could be an important new tool for studying hematological abnormalities. Here, we analyzed HSC subsets with a series of stem cell markers in JAK2V617F transgenic (Tg) mice, where the mutation is sufficient to cause myeloproliferative neoplasia with lymphocyte deficiency. Total numbers of HSC were elevated 3 to 20 fold in bone marrow of JAK2V617F mice. Careful analysis suggested the accumulation involved multiple HSC subsets, but particularly those characterized as CD150(HI) CD86(-) CD18(L)°CD41(+) and excluding Hoechst dye. Real-Time PCR analysis of their HSC revealed that the erythropoiesis associated gene transcripts Gata1, Klf1 and Epor were particularly high. Flow cytometry analyses based on two differentiation schemes for multipotent progenitors (MPP) also suggested alteration by JAK2 signals. The low CD86 on HSC and multipotent progenitors paralleled the large reductions we found in lymphoid progenitors, but the few that were produced functioned normally when sorted and placed in culture. Either of two HSC subsets conferred disease when transplanted. Thus, flow cytometry can be used to observe the influence of abnormal JAK2 signaling on stem and progenitor subsets. Markers that similarly distinguish categories of human HSCs might be very valuable for monitoring such conditions. They could also serve as indicators of HSC fitness and suitability for transplantation.

Show MeSH
Related in: MedlinePlus