Impact of Itga2-Gp6-double collagen receptor deficient mice for bone marrow megakaryocytes and platelets

Abstract

The two main collagen receptors on platelets, GPVI and integrin α2β1, play an important role for the recognition of exposed collagen at sites of vessel injury, which leads to platelet activation and subsequently stable thrombus formation. Both receptors are already expressed on megakaryocytes, the platelet forming cells within the bone marrow. Megakaryocytes are in permanent contact with collagen filaments in the marrow cavity and at the basal lamina of sinusoids without obvious preactivation. The role of both collagen receptors for megakaryocyte maturation and thrombopoiesis is still poorly understood. To investigate the function of both collagen receptors, we generated mice that are double deficient for Gp6 and Itga2 . Flow cytometric analyses revealed that the deficiency of both receptors had no impact on platelet number and led to the expected lack in GPVI responsiveness. Integrin activation and degranulation ability was comparable to wildtype mice. By immunofluorescence microscopy, we could demonstrate that both wildtype and double-deficient megakaryocytes were overall normally distributed within the bone marrow. We found megakaryocyte count and size to be normal, the localization within the bone marrow, the degree of maturation, as well as their association to sinusoids were also unaltered. However, the contact of megakaryocytes to collagen type I filaments was decreased at sinusoids compared to wildtype mice, while the interaction to type IV collagen was unaffected. Our results imply that GPVI and α2β1 have no influence on the localization of megakaryocytes within the bone marrow, their association to the sinusoids or their maturation. The decreased contact of megakaryocytes to collagen type I might at least partially explain the unaltered platelet phenotype in these mice, since proplatelet formation is mediated by these receptors and their interaction to collagen. It is rather likely that other compensatory signaling pathways and receptors play a role that needs to be elucidated.

CXCL12-Abundant Reticular (CAR) Cells Direct Megakaryocyte Protrusions across the Bone Marrow Sinusoid Wall

“…The ECM distribution pattern might contribute to MK polarization when located at the vessel and prevent ectopic platelet release into the BM cavity [ 16 , 25 ]. However, mice lacking both collagen receptors GPVI and integrin-α 2 did not show ectopic platelet release [ 26 ].…”

Role of Nitric Oxide in Megakaryocyte Function

“…Similarly, megakaryocyte glycoprotein (GP) VI-collagen I interaction has inhibited proplatelet formation [28]. However, double knockout of collagen receptors (GPVI−/− integrin α 2 β 1 −/−) shows no difference in megakaryocyte distribution, size, or blood platelet levels compared to that of wild type mice, suggesting other regulatory mechanisms may exist to suppress ectopic proplatelet generation within the osteoblastic niche [29]. In contrast, the vascular niche contains extracellular matrix proteins including collagen IV, fibronectin, fibrinogen, and von Willebrand factor, which induce proplatelet generation [30][31][32][33].…”

Megakaryocyte Cytoskeletal Proteins in Platelet Biogenesis and Diseases

“…81 In particular, megakaryocyte contact with type I collagen fibrils induces myosin light chain-2 (MLC-2) phosphorylation through the α2β1 integrin-dependent Rho-ROCK (Rho-associated protein kinase) pathway to regulate the cytoskeleton contractility, cell migration, and platelet release. [82][83][84][85] While, megakaryocyte interactions with fibrinogen, vitronectin, and fibronectin are mediated by αIIbβ3 integrin, which has a pivotal role regulating F-actin cytoskeleton during platelet biogenesis mainly through Fln-A interaction. 29,79 Mutations in the ITGA2B and ITGB3 genes cause Glanzmann thrombasthenia (GT), a bleeding disorder due to quantitative or qualitative defects of αIIbβ3 yielding reduced platelet aggregation while maintaining normal platelet count and size.…”