We studied integrins involved in the adhesion of resting and activated megakaryocytes (MK) to fibronectin (FN) and fibrinogen (FGN). Guinea pig MK were isolated and in some experiments were activated by thrombin. MK adhering to FN or FGN coated on coverslips were quantitated by a computerized image analysis program. The binding of soluble human FN to MK was detected by Western blotting. Anti-integrin antibodies, disintegrins, and cyclic RGD peptides were used to identify integrins involved in the adhesion of MK to FN or FGN. Resting MK adhered to coverslips with immobilized FN. The adhesion of MK to FN was primarily inhibited by an anti-5 antibody and EMF-10, a distintegrin highly specific for 5β1. However, the adhesion of MK to FN was not blocked by agents that inhibit ΙΙbβ3, vβ3 or 4β1. A β1 activating antibody increased the number of MK bound to FN due to the activation of 5β1. The binding of soluble FN was also primarily inhibited by agents that block 5β1. Resting MK did not adhere to FGN. However, MK activated by thrombin did adhere to FGN. This binding was mediated by ΙΙbβ3, because binding was inhibited by bitistatin, a disintegrin, and a cyclic RGD peptide that are known to block this integrin. The binding of thrombin-activated MK to FN was mediated by both 5β1 and ΙΙbβ3 based on the additive effect of agents that inhibit these integrins. The study indicates that resting MK bind to FN but not to FGN and that 5β1 is the major integrin involved in the binding of MK to FN. Activated MK bind to FGN primarily by IIbβ3. However, the binding of activated MK to FN is due to both 5β1 and IIbβ3. The demonstration that 5β1 and that IIbβ3 are involved in MK adhesion indicates that these integrins may have a role in MK maturation and platelet production.
© 1998 by The American Society of Hematology.
The Objective Assessment of Experts’ and Novices’ Suturing Skills Using An Image Analysis Program
