Extracellular Matrix Receptors and the Differentiation of Human Megakaryocytes in vitro

Abstract

We investigated the expression and functions of extracellular matrix receptors (or integrins) in the course of the differentiation of human megakaryocytes (Mks) leading to the formation of platelets. Integrins beta1 or Very Late Antigens (VLA) are specialized transmembrane receptors allowing the attachment of the cells to collagen (VLA-2), fibronectin (VLA-4 and -5) and laminin (VLA-6). A proportion of committed megakaryocytic progenitor cells (CFU-MK) adhere to fibronectin but not to collagen or laminin. The early immature Mks are retained on fibronectin (30%) and laminin (12%) but not on collagen whereas large mature Mks are still adherent to fibronectin and laminin and also acquired the capacity to adhere to collagen. The expression of the different VLA in the maturation of Mks correlates well with their adhesive properties. Hence, VLA-2 is not expressed on immature Mks but is present on the mature polyploid cells. VLA-4 is detected only on immature Mks which do not seem to bear VLA-5, while this last integrin appears on late Mks. VLA-6 showed a broad distribution from the early to late stages of Mks differentiation. Integrins beta3 of the cytoadhesin family are represented by alphaIIb beta3 that is the receptor for fibrinogen and alphaV beta3 which mediates adhesion to vitronectin. AlphaIIb beta3 is present on the CFU-MK and highly expressed throughout the Mks maturation stages while alphaV beta3 expression is much lower and seems to be detected only on the late Mks. The regulation of the expression of these receptors by cytokines and their respective roles in the maturation of Mks and the final production of platelets, are discussed. The development of efficient culture systems of human Mks in the presence of the recently cloned thrombopoietin will undoubtedly help to shed more light on the molecular mechanisms of their interactions via integrins with the BM microenvironment.

Prolonged continuous in vitro human platelet production using three-dimensional scaffolds

“…[43] The LBL deposition of nano-structured clay platelet and PDDA increased nano-scale roughness and created surface charge and stiff film, which worked in concert, along with adhered proteins, to bind and stimulate cells. We knew from the literature that megakaryocytes and CFU-MK bind to fibronectin and that TPO promotes early hematopoietic cell and megakaryocyte adherence to fibronectin [44]. We found that with the combination of both TPO and fibronectin, utilizing the SRC kinase inhibitor medium, platelet production lasted longer at a higher level vs. only one or the other or neither.…”

Delayed platelet engraftment in group O patients after autologous progenitor cell transplantation

“…38 GPIba and PECAM are later megakaryocytic markers and may contribute to growth arrest and proplatelet formation, respectively. 38,44,45 ABO antigens are also expressed by a4b1 and a5b1 integrins, where they are known to directly influence integrin function. In epithelial tumors, loss of A/B antigens, with a concomitant increase in H and Le Y , is associated with increased fibronectin binding, cell adhesion, cell motility, and resistance to apoptosis.…”

“…In epithelial tumors, loss of A/B antigens, with a concomitant increase in H and Le Y , is associated with increased fibronectin binding, cell adhesion, cell motility, and resistance to apoptosis. [46][47][48] In marrow, integrins mediate CD34+ PBPCs and megakaryocyte adhesion to ECM and stromal fibroblasts, 40,45,49 inhibiting apoptosis, 50 while promoting fibroblast growth, megakaryocyte differentiation, and proplatelet formation. 18,39,45 Finally, ABO might influence the binding and cellular response to required growth factors.…”

Matrix Mechanosensation in the Erythroid and Megakaryocytic Lineages

“…MKs express two main receptors for collagen, integrin-α2β1 and glycoprotein VI [47]; however, MK expression of leukocyte-associated immunoglobulin-like 1 (LAIR1) and discoidin domain receptor (DDR1) that also bind collagen have been described in the literature [48,49]. Early MK progenitors (CFU-MK) have a lower affinity for adhering to collagen compared to later MK progenitors [50], which is interesting as integrin-α2β1 and glycoprotein VI expression occurs early in MK development [50,51]. It has been established that collagen I inhibits proplatelet formation (PPF) in order to prevent the release of immature platelets in the bone marrow, while collagen IV and laminin support PPF in the sinusoids [52].…”

“…HSPC cultures on fibronectin resulted in an increase in mouse HSC with an increase in MK output [13]. The major fibronectin binding integrins on MKs are VLA-4 (α4β1) and VLA-5 (α5β1), which are expressed throughout MK development [50,60]. HSC cultures produced more CFU-MKs on fibronectin as stiffness increased [8].…”