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Updated: Jul 31, 2026
Megakaryocyte Differentiation and Platelet Formation from Human Cord Blood-derived CD34+ Cells
Published on: December 27, 2017
Megakaryocyte proplatelet-like process formation in vitro is inhibited by serum prothrombin, a process which is
1Laboratory of Stem Cell Biology, Amgen Center, Thousand Oaks, CA 91320.
Abstract:
The process of platelet shedding from megakaryocytes is incompletely understood, due in part to the impossibility of studying this dynamic process in vivo. Megakaryocytes in situ and in in vitro cultures display extended cytoplasmic processes constricted at platelet-sized intervals which presumably are the structural intermediates between megakaryocytes and platelets. This study describes the establishment of a serum-free culture system of purified guinea pig megakaryocytes in which extensive cytoplasmic process formation can be observed on 21 to 29% of the cells. The addition of as little as 0.05% pooled human serum to the cultures will completely but reversibly block process development. The serum inhibitor was identified as residual prothrombin, which upon contact with megakaryocytes is converted to the serine esterase thrombin. Thrombin directly prevents the formation of new processes and also induces retraction of existing processes. When megakaryocytes are cultured on Matrigel, process formation occurs even in an excess of thrombin. This potentiation of process development in the presence of inhibitory factors is mediated by the glycosaminoglycan content of Matrigel. The physiological implications of these observations are discussed.
Insights
Platelet formation from megakaryocytes is unclear. Thrombin inhibits this process, but Matrigel
Area of Science:
- Hematology
- Cell Biology
- Biochemistry
Background:
- Platelet biogenesis from megakaryocytes is a complex process that remains poorly understood.
- Studying megakaryocyte cytoplasmic processes, the precursors to platelets, is challenging in vivo.
- In vitro models have shown megakaryocytes form platelet-sized protrusions.
Purpose of the Study:
- To establish a serum-free culture system for studying guinea pig megakaryocyte proplatelet formation.
- To identify factors in serum that inhibit megakaryocyte proplatelet formation.
- To investigate the role of Matrigel in modulating thrombin's inhibitory effects on proplatelet formation.
Main Methods:
- Developed a serum-free culture system for purified guinea pig megakaryocytes.
- Assessed the effect of pooled human serum on megakaryocyte cytoplasmic process formation.
- Identified thrombin as the serum-derived inhibitor of process formation.
- Investigated the influence of Matrigel on megakaryocyte process formation in the presence of thrombin.
Main Results:
- A serum-free culture system allowed observation of cytoplasmic process formation in 21-29% of megakaryocytes.
- As little as 0.05% pooled human serum completely blocked process development.
- Residual prothrombin in serum was converted to thrombin, which inhibited process formation.
- Culturing megakaryocytes on Matrigel promoted process formation even with excess thrombin.
- Matrigel's glycosaminoglycans mediated the potentiation of process development against inhibitory factors.
Conclusions:
- Thrombin is a key inhibitor of megakaryocyte proplatelet formation.
- Matrigel, via its glycosaminoglycan content, can overcome thrombin's inhibitory effects.
- These findings provide insights into the regulation of platelet biogenesis and the role of the extracellular matrix.
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