MC-cultured WT MKs display in situ–like DMS development and morphology. (A) Confocal microscopy images of MKs from either 3-day in vitro cultures or in situ BM section, showing the various morphologies of the DMS. The plasma membrane and DMS are labeled with an anti-GPIbβ antibody (green) and the nucleus with 4′,6-diamidino-2-phenylindole (DAPI; white). Images are representative of at least 3 different experiments. Scale bar = 5 µm. (B) Impact of the MC medium on MK morphology following 3-day culture. In the upper panel, the DMS appears in green and the nucleus appears in white as in (A). Scale bar = 5 µm. In the lower panel, the mean diameter of MKs (left) and their proportion in each class (right) are shown for liquid and MC hydrogel (0.8, 1.3, 2 and 2.5%) cultures. Results are expressed as the mean ± SEM in 3 independent cultures, with a total of at least 100 MKs examined. ***P < .0001 using 1-way analysis of variance (ANOVA) with Bartlett’s test comparing the 2.5% MC condition to all other conditions. (C) Bar graphs represent the total cell fluorescence for GPIbβ staining in liquid cultures and 2% or 2.5% MC hydrogels. Data are representative of 3 different cultures for each condition, with a total number of 137, 134, and 120 MKs analyzed for liquid cultures and 2% and 2.5% MC gels, respectively. Results are expressed as the mean ± SEM: ***P < .0001 using a 1-way ANOVA analysis with a Newman-Keuls posttest.