Adhesion to different collagens modulates megakaryocyte development

Megakaryopoiesis occurs in a complex microenvironment within the bone marrow. The first events occur in the osteoblastic niche and include commitment of the hemopoietic progenitor cell to megakaryopoiesis. The second step is megakaryocyte (MK) maturation and is associated with rapid cytoplasm expansion and intense synthesis of proteins. Finally MKs, which migrate to the vascular niche, convert the bulk of their cytoplasm into multiple long processes called proplatelets that protrude through the vascular endothelium into the sinusoid lumen, where the platelets are released. Growing evidence indicates that a complex regulatory mechanism, involving MK-matrix interactions, may contribute to the quiescent or permissive microenvironment related to platelet release within bone marrow. To address this hypothesis, in this work we have investigated the role of type I, IV and VI collagens in regulating MK function. Fibrillar type I collagen is the most abundant extracellular protein of the osteoblastic niche, while microfibrillar type IV and VI collagens are primary subendothelial extracellular matrix components. Human MKS (hMKs) were differentiated from cord blood derived CD34+ cells for 12 days. Mature hMKs were plated onto glass coverslips coated with type I, IV or VI collagen. Proplatelet formation (PPF) was evaluated by phase contrast and fluorescence microscopy upon cell staining with anti-tubulin and CD41 antibodies. Type I, but not type IV or type VI collagen suppressed PPF. This process was triggered by the engagement of integrin alpha2beta1 through activation of Rho/ROCK pathway and myosin-IIA. Moreover, after a short incubation hMKs were spread on all collagens, while, prolonging incubation, hMKs on type IV and VI collagens returned round and started to extend proplatelets, while MKs on type I collagen remained spread and did not proceed on maturation. Overall our data represent the first evidence that hMK function on different collagens may depend on peculiar structural properties of the collagens, as well as on differences in receptor engagement.