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Quantitative trait analysis reveals transforming growth factor-beta2 as a positive regulator of early hematopoietic progenitor and stem cell function.

Langer JC, Henckaerts E, Orenstein J, Snoeck HW - J. Exp. Med. (2004)

Bottom Line: A quantitative trait locus (QTL) for the effect of TGF-beta2 was identified on chromosome 4 overlapping with a QTL regulating the frequency of LSK cells.These data suggest that TGF-beta2 can act cell autonomously and is important for HSCs that have undergone replicative stress.Thus, TGF-beta2 is a novel, genetically determined positive regulator of adult HSCs.

View Article: PubMed Central - PubMed

Affiliation: The Carl C. Icahn Center for Gene Therapy and Molecular Medicine, Mount Sinai School of Medicine, Box 1496, Gustave L. Levy Place, New York, NY 10029, USA.

ABSTRACT
Elucidation of pathways involved in mouse strain-dependent variation in the hematopoietic stem cell (HSC) compartment may reveal novel mechanisms relevant in vivo. Here, we demonstrate genetically determined variation in the proliferation of lin-Sca1++kit+ (LSK) primitive hematopoietic progenitor cells in response to transforming growth factor-beta (TGF-beta) 2, the dose response of which was biphasic with a stimulatory effect at low concentrations. In contrast, the dose responses of TGF-beta1 or -beta3 were inhibitory and did not show mouse strain-dependent variation. A quantitative trait locus (QTL) for the effect of TGF-beta2 was identified on chromosome 4 overlapping with a QTL regulating the frequency of LSK cells. These overlapping QTL were corroborated by the observation that the frequency of LSK cells is lower in adult Tgfb2+/- mice than in wild-type littermates, indicating that TGF-beta2 is a genetically determined positive regulator LSK number in vivo. Furthermore, adult Tgfb2+/- mice have a defect in competitive repopulation potential that becomes more pronounced upon serial transplantation. In fetal TGF-beta2-deficient HSCs, a defect only appears after serial reconstitution. These data suggest that TGF-beta2 can act cell autonomously and is important for HSCs that have undergone replicative stress. Thus, TGF-beta2 is a novel, genetically determined positive regulator of adult HSCs.

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Linkage analysis in BXD RI strains. (a) Strain distribution pattern of the effect of 0.1 ng/ml TGF-β2 on the KL, flt3L, and TPO-supported proliferation of LSK cells in BXD mice (mean ± SEM; n = 2–4). (b) Pattern of likelihood ratio statistic (y axis) values along chromosome 4 for the absolute number of lin−Sca1++ cells (reference 9) and their responsiveness to TGF-β2. The y axis scale is relative to normalize the size of both peaks (the maximum likelihood ratio statistic value is 15.9 for the number of lin−Sca1++ cells, and 13.7 of the effect of TGF-β2). (c) Correlation between the effect of TGF-β2 on the proliferation of LSK cells (a) and the frequency of lin−Sca1++ cells (reference 9).
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fig2: Linkage analysis in BXD RI strains. (a) Strain distribution pattern of the effect of 0.1 ng/ml TGF-β2 on the KL, flt3L, and TPO-supported proliferation of LSK cells in BXD mice (mean ± SEM; n = 2–4). (b) Pattern of likelihood ratio statistic (y axis) values along chromosome 4 for the absolute number of lin−Sca1++ cells (reference 9) and their responsiveness to TGF-β2. The y axis scale is relative to normalize the size of both peaks (the maximum likelihood ratio statistic value is 15.9 for the number of lin−Sca1++ cells, and 13.7 of the effect of TGF-β2). (c) Correlation between the effect of TGF-β2 on the proliferation of LSK cells (a) and the frequency of lin−Sca1++ cells (reference 9).

Mentions: To further investigate mouse strain–dependent variation in the dose response of TGF-β2, we performed linkage analysis using BXD RI strains. Although the TGF-β2 dose response in LSK cells from C57BL/6 and DBA/2 mice (the progenitors of the BXD RI set) was very similar, linkage analysis was attempted using these strains because a relatively large number are readily available in this set. If a trait is multigenic, phenotypic variation can be more pronounced among RI strains than between their progenitor strains because the balance between positive-acting and negative-acting alleles present in the progenitor strains may be completely unbalanced in some of the RI strains (25). This may allow quantitative trait locus (QTL) identification, even when phenotypic variation between the progenitor strains is not significant. The effect of TGF-β2 at 0.1 ng/ml on the proliferation of LSK cells from 30 BXD RI strains was measured because this concentration revealed the most pronounced difference in the TGF-β2 dose response between the inbred strains (Fig. 1 a). The individual BXD strains showed wide variation in this assay (Fig. 2 a), indicating that the responsiveness to TGF-β2 is indeed a multigenic trait. One QTL contributing to the effect of TGF-β2 was identified on chromosome 4 between 67 (D4Mit158) and 79 (D4Mit33) cM (maximum likelihood ratio statistic 13.7; P = 0.00021). The level of significance was suggestive according to permutation analysis (27, 28). Although suggestive QTL require confirmation (28), the location of this QTL was compelling because this interval overlaps with the interval containing a previously identified QTL contributing to genetic variation in the pool size of lin−Sca1++ and of LSK cells in vivo (Fig. 2 b; reference 9). Furthermore, a significant correlation was found between the effect of TGF-β2 and the number of lin−Sca1++ cells in BXD RI strains (Fig. 2 c). One interpretation of these findings is that TGF-β2 signaling regulates the size of the hematopoietic stem and progenitor cell compartment as defined by phenotype in vivo.


Quantitative trait analysis reveals transforming growth factor-beta2 as a positive regulator of early hematopoietic progenitor and stem cell function.

Langer JC, Henckaerts E, Orenstein J, Snoeck HW - J. Exp. Med. (2004)

Linkage analysis in BXD RI strains. (a) Strain distribution pattern of the effect of 0.1 ng/ml TGF-β2 on the KL, flt3L, and TPO-supported proliferation of LSK cells in BXD mice (mean ± SEM; n = 2–4). (b) Pattern of likelihood ratio statistic (y axis) values along chromosome 4 for the absolute number of lin−Sca1++ cells (reference 9) and their responsiveness to TGF-β2. The y axis scale is relative to normalize the size of both peaks (the maximum likelihood ratio statistic value is 15.9 for the number of lin−Sca1++ cells, and 13.7 of the effect of TGF-β2). (c) Correlation between the effect of TGF-β2 on the proliferation of LSK cells (a) and the frequency of lin−Sca1++ cells (reference 9).
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Related In: Results  -  Collection

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fig2: Linkage analysis in BXD RI strains. (a) Strain distribution pattern of the effect of 0.1 ng/ml TGF-β2 on the KL, flt3L, and TPO-supported proliferation of LSK cells in BXD mice (mean ± SEM; n = 2–4). (b) Pattern of likelihood ratio statistic (y axis) values along chromosome 4 for the absolute number of lin−Sca1++ cells (reference 9) and their responsiveness to TGF-β2. The y axis scale is relative to normalize the size of both peaks (the maximum likelihood ratio statistic value is 15.9 for the number of lin−Sca1++ cells, and 13.7 of the effect of TGF-β2). (c) Correlation between the effect of TGF-β2 on the proliferation of LSK cells (a) and the frequency of lin−Sca1++ cells (reference 9).
Mentions: To further investigate mouse strain–dependent variation in the dose response of TGF-β2, we performed linkage analysis using BXD RI strains. Although the TGF-β2 dose response in LSK cells from C57BL/6 and DBA/2 mice (the progenitors of the BXD RI set) was very similar, linkage analysis was attempted using these strains because a relatively large number are readily available in this set. If a trait is multigenic, phenotypic variation can be more pronounced among RI strains than between their progenitor strains because the balance between positive-acting and negative-acting alleles present in the progenitor strains may be completely unbalanced in some of the RI strains (25). This may allow quantitative trait locus (QTL) identification, even when phenotypic variation between the progenitor strains is not significant. The effect of TGF-β2 at 0.1 ng/ml on the proliferation of LSK cells from 30 BXD RI strains was measured because this concentration revealed the most pronounced difference in the TGF-β2 dose response between the inbred strains (Fig. 1 a). The individual BXD strains showed wide variation in this assay (Fig. 2 a), indicating that the responsiveness to TGF-β2 is indeed a multigenic trait. One QTL contributing to the effect of TGF-β2 was identified on chromosome 4 between 67 (D4Mit158) and 79 (D4Mit33) cM (maximum likelihood ratio statistic 13.7; P = 0.00021). The level of significance was suggestive according to permutation analysis (27, 28). Although suggestive QTL require confirmation (28), the location of this QTL was compelling because this interval overlaps with the interval containing a previously identified QTL contributing to genetic variation in the pool size of lin−Sca1++ and of LSK cells in vivo (Fig. 2 b; reference 9). Furthermore, a significant correlation was found between the effect of TGF-β2 and the number of lin−Sca1++ cells in BXD RI strains (Fig. 2 c). One interpretation of these findings is that TGF-β2 signaling regulates the size of the hematopoietic stem and progenitor cell compartment as defined by phenotype in vivo.

Bottom Line: A quantitative trait locus (QTL) for the effect of TGF-beta2 was identified on chromosome 4 overlapping with a QTL regulating the frequency of LSK cells.These data suggest that TGF-beta2 can act cell autonomously and is important for HSCs that have undergone replicative stress.Thus, TGF-beta2 is a novel, genetically determined positive regulator of adult HSCs.

View Article: PubMed Central - PubMed

Affiliation: The Carl C. Icahn Center for Gene Therapy and Molecular Medicine, Mount Sinai School of Medicine, Box 1496, Gustave L. Levy Place, New York, NY 10029, USA.

ABSTRACT
Elucidation of pathways involved in mouse strain-dependent variation in the hematopoietic stem cell (HSC) compartment may reveal novel mechanisms relevant in vivo. Here, we demonstrate genetically determined variation in the proliferation of lin-Sca1++kit+ (LSK) primitive hematopoietic progenitor cells in response to transforming growth factor-beta (TGF-beta) 2, the dose response of which was biphasic with a stimulatory effect at low concentrations. In contrast, the dose responses of TGF-beta1 or -beta3 were inhibitory and did not show mouse strain-dependent variation. A quantitative trait locus (QTL) for the effect of TGF-beta2 was identified on chromosome 4 overlapping with a QTL regulating the frequency of LSK cells. These overlapping QTL were corroborated by the observation that the frequency of LSK cells is lower in adult Tgfb2+/- mice than in wild-type littermates, indicating that TGF-beta2 is a genetically determined positive regulator LSK number in vivo. Furthermore, adult Tgfb2+/- mice have a defect in competitive repopulation potential that becomes more pronounced upon serial transplantation. In fetal TGF-beta2-deficient HSCs, a defect only appears after serial reconstitution. These data suggest that TGF-beta2 can act cell autonomously and is important for HSCs that have undergone replicative stress. Thus, TGF-beta2 is a novel, genetically determined positive regulator of adult HSCs.

Show MeSH
Related in: MedlinePlus