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Pan-myeloid Differentiation of Human Cord Blood Derived CD34+ Hematopoietic Stem and Progenitor Cells

Pan-myeloid Differentiation of Human Cord Blood Derived CD34⁺ Hematopoietic Stem and Progenitor Cells

Published on: August 9, 2019

CD34+ cells derived from fetal liver contained a high proportion of immature megakaryocytic progenitor cells

D C Ma¹, Y H Sun, W Zuo

¹Department of Experimental Medicine, Northern Hospital, Shenyang, Liaoning, China.

European Journal of Haematology

|June 23, 2000

Abstract:

Endoreplication and maturation of the megakaryocyte (MK) may be retarded or delayed during ontogenesis. In this study, CD34+ cells were isolated from both human fetal liver and adult bone marrow and incubated with thrombopoietin (TPO). The cell number, morphological characteristics, platelet-associated antigen phenotype, maturation stage and DNA ploidy of CD41+ cells were examined from day 0 to day 12 in culture. 1) TPO stimulated the proliferation of fetal liver (FL)-derived CD34+ cells with a mean 73.14-fold increase of CD41+ cells after 12 d in culture. Adult BM-derived CD34+ cells increased only slightly, with a mean 8.18-fold increase of CD41+ cells. 2) Although the membrane phenotype of both FL CD34+-derived MKs and BM CD34+ -derived MKs analyzed with CD41a, CD42a, CD61 and CD34 were similar, all FL CD34+-derived MKs were in maturation stage I and II and in low ploidy (<4N) class. By comparison, BM CD34+ MKs possessed 15% MKs in maturation stage III and IV and with 23% MKs in high ploidy class ( > 4N). 3) Most of cultured FL-derived CD34+ cells did not have a well developed demarcation system (DM) and numerous alpha-granules after 12 d incubation. von Willebrand factor (vWF) appeared earlier on the cultured BM-derived CD34+ cells than on FL-derived CD34+ cells. 4) The expression of both cyclin E and cyclin B1 progressively increased in FL CD34+ cells induced by TPO during 12 d in culture. 5) The expression of cyclin D1 gradually decreased in FL CD34+ cells induced by TPO over 12 d incubation. 6) Immunocytochemical analysis showed that cyclin D3 was detected only in cytoplasm of cultured FL-derived CD34+ cells, whereas in both cytoplasm and nuclei of cultured BM-derived CD34+ cells. These data suggest that FL-derived CD34+ cells contain a high proportion of immature megakaryocytic progenitor cells. It further suggests that TPO can push these progenitor cells into proliferation by upregulating the expression of cyclins B1 and E, and drive a high proportion of cells into megakaryocytic lineage.

Insights

Thrombopoietin (TPO) significantly boosts fetal liver CD34+ cell proliferation into megakaryocytes (MKs), unlike adult bone marrow cells. Fetal cells show delayed maturation and lower ploidy, indicating immature megakaryocytic progenitors responsive to TPO.

Area of Science:

Hematology
Developmental Biology
Cell Biology

Background:

Megakaryocyte (MK) endoreplication and maturation can be delayed during development.
Understanding the differences between fetal and adult hematopoietic stem cells is crucial for regenerative medicine.

Purpose of the Study:

To investigate the proliferative and differentiation potential of CD34+ cells from human fetal liver (FL) and adult bone marrow (BM) in response to thrombopoietin (TPO).
To compare the maturation stages, ploidy, and expression of key cell cycle regulators in MKs derived from FL and BM sources.

Main Methods:

Isolation of CD34+ cells from human fetal liver and adult bone marrow.
Culture of CD34+ cells with TPO for 12 days, monitoring cell proliferation and CD41+ cell expansion.
Analysis of cell morphology, platelet-associated antigen phenotype (CD41a, CD42a, CD61, CD34), maturation stage, and DNA ploidy.
Assessment of demarcation system (DM) development and alpha-granule presence.
Evaluation of von Willebrand factor (vWF) expression.
Immunocytochemical analysis of cyclin E, cyclin B1, cyclin D1, and cyclin D3 expression.

Main Results:

TPO induced a substantial increase (73.14-fold) in CD41+ cells from FL CD34+ progenitors, compared to a modest increase (8.18-fold) from adult BM CD34+ cells.
FL-derived MKs exhibited predominantly early maturation stages (I and II) and low ploidy (<4N), while BM-derived MKs showed later maturation (15% in stages III/IV) and higher ploidy (23% >4N).
FL-derived MKs displayed less developed DM and fewer alpha-granules, with delayed vWF expression compared to BM-derived MKs.
TPO upregulated cyclin E and cyclin B1 expression in FL CD34+ cells, while cyclin D1 expression decreased over 12 days.
Cyclin D3 was detected only in the cytoplasm of FL-derived MKs, but in both cytoplasm and nuclei of BM-derived MKs.

Conclusions:

Human fetal liver CD34+ cells contain a higher proportion of immature megakaryocytic progenitor cells compared to adult bone marrow.
TPO promotes proliferation of these immature progenitors by upregulating cyclins B1 and E, driving them towards the megakaryocytic lineage.
The distinct patterns of cell cycle regulation and maturation suggest differential developmental pathways for MKs originating from fetal versus adult sources.

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