Proplatelet formation defects in gm/gm MKs. (A) Proplatelet formation is reduced approximately 10-fold in gm/gm MKs. Mouse fetal liver cells were cultured in thrombopoietin for 4 days before isolation of MKs that were analyzed for proplatelet formation the following day by phase-contrast microscopy. The percentage of proplatelet (PP)-producing MKs is expressed as mean ± SD. (B) Phase-contrast micrographs of representative +/gm and gm/gm cells on culture day 5. The +/gm MKs elaborate rich proplatelet networks, whereas gm/gm MKs only form a few blunt pseudopodia. (C-D) Immunofluorescent detection of β1 tubulin in cultured +/gm (C) and gm/gm (D) MKs. The increased sensitivity of this assay reveals that gm/gm MKs initiate proplatelet formation but the cytoplasmic extensions develop and elongate poorly. Original magnifications, × 400. (E) Immunoblot analysis of cultured +/gm and gm/gm MKs showing normal levels of β1 tubulin and p45 NF-E2. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is used as a loading control.