Multivesicular Bodies Are an Intermediate Stage in the Formation of Platelet α-Granules

Abstract

We have used ultrathin cryosectioning and immunogold cytochemistry to study the position of α-granules in the endocytic and biosynthetic pathways in megakaryocytes and platelets. Morphologically, we distinguished three types of granules; so-called multivesicular bodies type I (MVB I) with internal vesicles only, granules with internal vesicles and an electron dense matrix (MVB II), and the α-granules with mainly a dense content and often internal membrane vesicles at their periphery. The MVBs were prominent in cultured megakaryocytes and the megakaryoblastic cell line CHRF-288, but were less numerous in bone marrow megakaryocytes and platelets, whereas α-granules were most prominent in mature bone marrow megakaryocytes and in platelets. The internalization kinetics of bovine serum albumin-gold particles and of fibrinogen positioned the MVB subtypes and α-granules sequentially in the endocytic pathway. MVBs contained the secretory proteins von Willebrand factor (vWF) and β-thromboglobulin (β-TG), the platelet-specific membrane protein P-selectin, and the lysosomal membrane protein CD63. Within the MVBs, endocytosed fibrinogen and endogenous β-TG were restricted to the matrix, while vWF was predominantly associated with internal vesicles. CD63 was also observed in association with internal membrane vesicles in the α-granules. These observations, and the gradual morphologic transition from granules containing vesicles to granules containing predominantly dense material, suggest that MVBs represent a developmental stage in α-granule maturation.