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Blood platelets are assembled principally at the ends of proplatelet processes produced by differentiated megakaryocytes.

Italiano JE, Lecine P, Shivdasani RA, Hartwig JH - J. Cell Biol. (1999)

Bottom Line: We have resolved the ultrastructure of the megakaryocyte cytoskeleton at specific stages of proplatelet morphogenesis and correlated these structures with cytoplasmic remodeling events defined by video microscopy.Growth and extension of proplatelet processes is associated with repeated bending and bifurcation, which results in considerable amplification of free ends.These aspects are inhibited by cytochalasin B and, therefore, are dependent on actin.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.

ABSTRACT
Megakaryocytes release mature platelets in a complex process. Platelets are known to be released from intermediate structures, designated proplatelets, which are long, tubelike extensions of the megakaryocyte cytoplasm. We have resolved the ultrastructure of the megakaryocyte cytoskeleton at specific stages of proplatelet morphogenesis and correlated these structures with cytoplasmic remodeling events defined by video microscopy. Platelet production begins with the extension of large pseudopodia that use unique cortical bundles of microtubules to elongate and form thin proplatelet processes with bulbous ends; these contain a peripheral bundle of microtubules that loops upon itself and forms a teardrop-shaped structure. Contrary to prior observations and assumptions, time-lapse microscopy reveals proplatelet processes to be extremely dynamic structures that interconvert reversibly between spread and tubular forms. Microtubule coils similar to those observed in blood platelets are detected only at the ends of proplatelets and not within the platelet-sized beads found along the length of proplatelet extensions. Growth and extension of proplatelet processes is associated with repeated bending and bifurcation, which results in considerable amplification of free ends. These aspects are inhibited by cytochalasin B and, therefore, are dependent on actin. We propose that mature platelets are assembled de novo and released only at the ends of proplatelets, and that the complex bending and branching observed during proplatelet morphogenesis represents an elegant mechanism to increase the numbers of proplatelet ends.

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Video-enhanced light microscopy of a terminally differentiated mouse megakaryocyte forming proplatelets in vitro. During the initial stages of proplatelet formation, the megakaryocyte spreads and its cortical cytoplasm begins to unravel at one pole (this zone of erosion is labeled with an asterisk in the first panel). As the cell spreads, the cytoplasm at the erosion site is remodeled into large pseudopodia (white arrow at 2 h) that elongate and become thinner over time, forming narrow tubes of 1–4 μm diam. The proplatelet extensions frequently bend (white arrowhead at 2 h), and it is at sites of bending that the tube bifurcates to generate a new process. In this manner, the entire cytoplasmic space of the megakaryocyte is converted into anastomosing proplatelet extensions. Proplatelets also develop segmented constrictions along their length that impart a beaded appearance. The process of proplatelet elaboration culminates in a rapid retraction that separates the many strands of proplatelets from a residual nuclear mass (asterisk in last panel). Note the crawling of small cytoplasmic fragments (5–10 μm) at the ends of some of the proplatelet extensions (arrowheads at 4–7 h). Crawling begins when the end of the proplatelet adheres and flattens, forms a leading lamellipodia, and migrates away from the cell center, dragging a trail of proplatelets behind. Bar, 20 μm.
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Figure 1: Video-enhanced light microscopy of a terminally differentiated mouse megakaryocyte forming proplatelets in vitro. During the initial stages of proplatelet formation, the megakaryocyte spreads and its cortical cytoplasm begins to unravel at one pole (this zone of erosion is labeled with an asterisk in the first panel). As the cell spreads, the cytoplasm at the erosion site is remodeled into large pseudopodia (white arrow at 2 h) that elongate and become thinner over time, forming narrow tubes of 1–4 μm diam. The proplatelet extensions frequently bend (white arrowhead at 2 h), and it is at sites of bending that the tube bifurcates to generate a new process. In this manner, the entire cytoplasmic space of the megakaryocyte is converted into anastomosing proplatelet extensions. Proplatelets also develop segmented constrictions along their length that impart a beaded appearance. The process of proplatelet elaboration culminates in a rapid retraction that separates the many strands of proplatelets from a residual nuclear mass (asterisk in last panel). Note the crawling of small cytoplasmic fragments (5–10 μm) at the ends of some of the proplatelet extensions (arrowheads at 4–7 h). Crawling begins when the end of the proplatelet adheres and flattens, forms a leading lamellipodia, and migrates away from the cell center, dragging a trail of proplatelets behind. Bar, 20 μm.

Mentions: To date, proplatelets have been studied and described largely through static images, and most conclusions about mechanisms of proplatelet formation are inferred from studies conducted with only limited benefit of time-lapse cinematography. We used video-enhanced light microscopy of cultured mouse megakaryocytes to reveal a spectrum of highly reproducible morphogenetic changes and cellular movements during formation of proplatelets (Fig. 1 and accompanying video). A transformation of the entire megakaryocyte cytoplasm results in condensation of cell material into platelet-sized particles, which have the appearance of beads linked by thin cytoplasmic bridges. This process unfolds over 4–10 h, and initiates with erosion of one pole of the megakaryocyte cytoplasm. This generates a unique pseudopodial structure that elongates to yield slender tubules of uniform diameter of 2–4 μm; these in turn develop periodic densities along their length that impart the characteristic beaded appearance of proplatelets. Maturation of proplatelets ends in a rapid retraction that separates a variable portion of the proplatelets from the residual cell body, according to a process that presumably mirrors platelet release in vivo.


Blood platelets are assembled principally at the ends of proplatelet processes produced by differentiated megakaryocytes.

Italiano JE, Lecine P, Shivdasani RA, Hartwig JH - J. Cell Biol. (1999)

Video-enhanced light microscopy of a terminally differentiated mouse megakaryocyte forming proplatelets in vitro. During the initial stages of proplatelet formation, the megakaryocyte spreads and its cortical cytoplasm begins to unravel at one pole (this zone of erosion is labeled with an asterisk in the first panel). As the cell spreads, the cytoplasm at the erosion site is remodeled into large pseudopodia (white arrow at 2 h) that elongate and become thinner over time, forming narrow tubes of 1–4 μm diam. The proplatelet extensions frequently bend (white arrowhead at 2 h), and it is at sites of bending that the tube bifurcates to generate a new process. In this manner, the entire cytoplasmic space of the megakaryocyte is converted into anastomosing proplatelet extensions. Proplatelets also develop segmented constrictions along their length that impart a beaded appearance. The process of proplatelet elaboration culminates in a rapid retraction that separates the many strands of proplatelets from a residual nuclear mass (asterisk in last panel). Note the crawling of small cytoplasmic fragments (5–10 μm) at the ends of some of the proplatelet extensions (arrowheads at 4–7 h). Crawling begins when the end of the proplatelet adheres and flattens, forms a leading lamellipodia, and migrates away from the cell center, dragging a trail of proplatelets behind. Bar, 20 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2168104&req=5

Figure 1: Video-enhanced light microscopy of a terminally differentiated mouse megakaryocyte forming proplatelets in vitro. During the initial stages of proplatelet formation, the megakaryocyte spreads and its cortical cytoplasm begins to unravel at one pole (this zone of erosion is labeled with an asterisk in the first panel). As the cell spreads, the cytoplasm at the erosion site is remodeled into large pseudopodia (white arrow at 2 h) that elongate and become thinner over time, forming narrow tubes of 1–4 μm diam. The proplatelet extensions frequently bend (white arrowhead at 2 h), and it is at sites of bending that the tube bifurcates to generate a new process. In this manner, the entire cytoplasmic space of the megakaryocyte is converted into anastomosing proplatelet extensions. Proplatelets also develop segmented constrictions along their length that impart a beaded appearance. The process of proplatelet elaboration culminates in a rapid retraction that separates the many strands of proplatelets from a residual nuclear mass (asterisk in last panel). Note the crawling of small cytoplasmic fragments (5–10 μm) at the ends of some of the proplatelet extensions (arrowheads at 4–7 h). Crawling begins when the end of the proplatelet adheres and flattens, forms a leading lamellipodia, and migrates away from the cell center, dragging a trail of proplatelets behind. Bar, 20 μm.
Mentions: To date, proplatelets have been studied and described largely through static images, and most conclusions about mechanisms of proplatelet formation are inferred from studies conducted with only limited benefit of time-lapse cinematography. We used video-enhanced light microscopy of cultured mouse megakaryocytes to reveal a spectrum of highly reproducible morphogenetic changes and cellular movements during formation of proplatelets (Fig. 1 and accompanying video). A transformation of the entire megakaryocyte cytoplasm results in condensation of cell material into platelet-sized particles, which have the appearance of beads linked by thin cytoplasmic bridges. This process unfolds over 4–10 h, and initiates with erosion of one pole of the megakaryocyte cytoplasm. This generates a unique pseudopodial structure that elongates to yield slender tubules of uniform diameter of 2–4 μm; these in turn develop periodic densities along their length that impart the characteristic beaded appearance of proplatelets. Maturation of proplatelets ends in a rapid retraction that separates a variable portion of the proplatelets from the residual cell body, according to a process that presumably mirrors platelet release in vivo.

Bottom Line: We have resolved the ultrastructure of the megakaryocyte cytoskeleton at specific stages of proplatelet morphogenesis and correlated these structures with cytoplasmic remodeling events defined by video microscopy.Growth and extension of proplatelet processes is associated with repeated bending and bifurcation, which results in considerable amplification of free ends.These aspects are inhibited by cytochalasin B and, therefore, are dependent on actin.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.

ABSTRACT
Megakaryocytes release mature platelets in a complex process. Platelets are known to be released from intermediate structures, designated proplatelets, which are long, tubelike extensions of the megakaryocyte cytoplasm. We have resolved the ultrastructure of the megakaryocyte cytoskeleton at specific stages of proplatelet morphogenesis and correlated these structures with cytoplasmic remodeling events defined by video microscopy. Platelet production begins with the extension of large pseudopodia that use unique cortical bundles of microtubules to elongate and form thin proplatelet processes with bulbous ends; these contain a peripheral bundle of microtubules that loops upon itself and forms a teardrop-shaped structure. Contrary to prior observations and assumptions, time-lapse microscopy reveals proplatelet processes to be extremely dynamic structures that interconvert reversibly between spread and tubular forms. Microtubule coils similar to those observed in blood platelets are detected only at the ends of proplatelets and not within the platelet-sized beads found along the length of proplatelet extensions. Growth and extension of proplatelet processes is associated with repeated bending and bifurcation, which results in considerable amplification of free ends. These aspects are inhibited by cytochalasin B and, therefore, are dependent on actin. We propose that mature platelets are assembled de novo and released only at the ends of proplatelets, and that the complex bending and branching observed during proplatelet morphogenesis represents an elegant mechanism to increase the numbers of proplatelet ends.

Show MeSH
Related in: MedlinePlus