Growth and maturation of megakaryocytes is regulated by Lnk/Sh2b3 adaptor protein through crosstalk between cytokine- and integrin-mediated signals

Exp Hematol. 2008 Jul;36(7):897-906. doi: 10.1016/j.exphem.2008.02.004. Epub 2008 May 5.

Abstract

Objective: Various cytokines and growth factors control the differentiation and maturation of megakaryocytes (MKs). However, the mechanism regulating platelet release from MKs is not well understood. Here, we investigated a role of Lnk/Sh2b3, an intracellular adaptor protein, in megakaryopoiesis.

Materials and methods: Number of MK progenitor in bone marrow (BM) of wild-type or Lnk(-/-) mice and their sensitivity to thrombopoietin (TPO) were determined in colony-forming unit assay. Using BM-derived wild-type or Lnk(-/-) MKs stimulated with TPO, activation of the signaling molecules was biochemically analyzed and effect of integrin stimulation on TPO signals was studied by addition of vascular cell adhesion molecule (VCAM-1). Platelet production from MKs in the presence of VCAM-1 was counted by flow cytometry and their morphological change was observed by time-lapse microscopy.

Results: Lnk(-/-) mice showed elevated platelets and mature MKs due to enhanced sensitivity of progenitors to TPO. Erk1/2 phosphorylation induced by TPO was augmented and prolonged in Lnk(-/-) MKs while activation of signal transducers and activators of transcription (Stat)3, Stat5, and Akt was normal. Wild-type MKs, but not in Lnk(-/-) MKs on VCAM-1 showed reduced Stat5 phosphorylation and mitogen-activated protein kinases activation upon stimulation with TPO. Additionally, the presence of VCAM in culture accelerated spontaneous platelet release from mature wild-type MKs, but not from Lnk(-/-) MKs.

Conclusions: Results suggest that contact of MKs with adhesion molecules via integrins might contribute to platelet release, which is under Lnk-mediated regulation of Stat-5 activation and show that Lnk functions in responses controlled by cell adhesion and in crosstalk between integrin- and cytokine-mediated signaling.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blood Platelets / metabolism
  • Enzyme Activation / drug effects
  • Enzyme Activation / physiology
  • Humans
  • Integrins / metabolism*
  • Intracellular Signaling Peptides and Proteins
  • MAP Kinase Signaling System / drug effects
  • MAP Kinase Signaling System / physiology*
  • Megakaryocytes / cytology
  • Megakaryocytes / metabolism*
  • Membrane Proteins
  • Mice
  • Mice, Knockout
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Mitogen-Activated Protein Kinase 3 / metabolism
  • Phosphorylation / drug effects
  • Proteins / genetics
  • Proteins / metabolism*
  • Proto-Oncogene Proteins c-akt / metabolism
  • STAT3 Transcription Factor / metabolism
  • STAT5 Transcription Factor / metabolism
  • Thrombopoietin / metabolism
  • Thrombopoietin / pharmacology*
  • Vascular Cell Adhesion Molecule-1 / metabolism*

Substances

  • Integrins
  • Intracellular Signaling Peptides and Proteins
  • Lnk protein, mouse
  • Membrane Proteins
  • Proteins
  • STAT3 Transcription Factor
  • STAT5 Transcription Factor
  • Stat3 protein, mouse
  • Vascular Cell Adhesion Molecule-1
  • Thrombopoietin
  • Proto-Oncogene Proteins c-akt
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3