Abstract 2626


The mechanisms by which megakaryocytes (Mks) proliferate, differentiate, and release platelets into circulation are not well understood. Mk maturation and platelet generation occur in the bone marrow and is consequent to Mk migration from the osteoblastic to the vascular niche, where Mks extend proplatelets and newly generated platelets can be released into the bloodstream. Growing evidence indicate that a complex regulatory mechanism, involving megakaryocyte-matrix interactions, may contribute to the quiescent or permissive microenvironment related to Mk differentiation and maturation within the bone marrow. It has been demonstrated that interactions of primary human Mks with matrices supposed to fill the vascular niche, such as fibrinogen or von Willebrand factor, is able to sustain Mk maturation and proplatelet formation, while type I collagen, in the osteoblastic niche, totally suppresses these events and prevents premature platelet release. The negative regulation of proplatelet formation by type I collagen is mediated by the interaction with integrin alpha2beta1, and involves the Rho/ROCK pathway.


The dynamic interaction of Mks with different extra-cellular matrices, that fill the bone marrow spaces, may orchestrate their maturation in specific sites. Despite the improvement in knowledge of biochemical niche, little is known about the mechanical force that regulate Mk-niche interactions. Therefore, in this work, we correlated activation of signaling cascade with generation of contractile force to understand the influences of bone marrow environment on Mk function.

Methodology/Principal Findings.

To address this hypothesis, we first demonstrated that human Mks express and synthesize cellular fibronectin (cFN), with a predominance of the EDA isoform, and transglutaminase FXIII-A. Thereafter, we proposed that these two molecules are involved in a new regulatory mechanism of Mk-type I collagen interaction in the osteoblastic niche. We propose that Mk adhesion on type I collagen promotes Mk spreading through a mechanism that involves FN, membrane receptors and FXIII-A activity. This mechanism seemed to be mediated by the exposure of cFN to the cell membrane and maintained by FN polymerization catalyzed by FXIII-A. These data address a new role to FN that, upon specific activation, could be released and thereby modulate Mk interaction with extracellular matrices. In this context FXIII-A catalyzes FN cross-linking at cellular sites, stabilizes FN assembly and promotes the organization of extracellular matrix. Consistently, the same mechanism regulated the assembly of plasma FN (pFN) by adherent Mks to type I collagen. Most importantly, our results demonstrated that only Mks adherent to type I collagen, and not to fibrinogen, were able to promote FN assembly. As a result, we observed that Mk adhesion to type I collagen promoted Mk spreading overtime, while Mks on fibrinogen showed a shortened spreading that was replaced by proplatelet formation in sixteen hours of adhesion. Thus, FN assembly regulate the anchoring of Mks to type I collagen with consequent activation of biochemical signalling and generation of contractile force that may prevent proplatelet formation.


In conclusion, this study provides important new elements in the understanding of the regulatory pathways for Mk-matrix interactions within bone marrow environment. In particular, our results demonstrate that fibronectins and FXIII-A modulate Mk spreading on type I collagen by promoting matrix assembly. This work opens new prospective in the study of illnesses, such as primary myelofibrosis or MYH9-related thrombocytopenia, related to defect of Mk-matrix interactions within the bone marrow environment, whose origin is still matter of debate.


No relevant conflicts of interest to declare.

Author notes


Asterisk with author names denotes non-ASH members.