The differentiation capacity and vesicular content of Plekhm1 KO MSC are altered. (A) Representative images of colonies derived from WT or Plekhm1 KO MSCs, stained with crystal violet (left). Number of colonies formed by WT (red) vs Plekhm1 KO (blue) MSCs (right). The data represent results from 4 independent experiments, and the values are normalized to the WT for each independent experiment (t test). (B) Osteogenic differentiation of WT or Plekhm1 KO MSC, visualized by Von Kossa and alizarin red staining for the detection of calcium deposits (left). The scale bar depicts 40 μm. Quantification of the alizarin red staining intensity normalized by the average value of WT (right). Each dot represents a technical replicate from a total of 3 independent experiments (t test). (C) Adipogenic differentiation of WT or Plekhm1 KO MSCs, visualized by oil red O staining for the detection of intracytoplasmic lipid-rich droplets in adipocytes (left). The scale bar depicts 40 μm. Quantification of the number of adipocytes formed after differentiation of WT (red) or Plekhm1 KO (blue) MSCs (right). The data represent the results from 3 independent experiments (t test). (D) Representative immunofluorescence images of WT or Plekhm1 KO MSCs stained with an antibody to CD63 (green). The nuclei are counterstained with DAPI (blue) and the cytoplasm with phalloidin (gray) (left). The scale bar depicts 20 μm. Quantification of the CD63 fluorescence intensity of WT (red) or Plekhm1 KO (blue) MSCs (right). The values are normalized to the average value of WT. The data represent the results from 4 independent experiments (t test). DAPI, 4′,6-diamidino-2-phenylindole.