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CD9-Positive Microvesicles Mediate the Transfer of Molecules to Bovine Spermatozoa during Epididymal Maturation

Figure 7

CD26 is associated with CD9 in microvesicles and has a synergistic role with CD9 in the molecular transfer to the corpus distal sperm.

A: CD9-positive microvesicles were lysed in presence of BrijO10, CD9 (25 kDa) was immunoprecipitated and analyzed by western blot. CD26 (110 kDa) and CD224 (61 kDa) coimmunoprecipitated with CD9. Normal mouse IgG was used as the negative control. Semi-quantitative analysis of RNA expression of CD224 (B) and CD26 (C) in epididymal epithelial cells, using tubulin as the housekeeping gene and cytokeratin 8 as a marker of epithelial cells. The highest expression was considered to be 100%, results are presented as average ± s.e.m. from three different experiments, * differs significantly p<0.05. Primary cultures of epithelial cells from caput and cauda epididymal tubules show comparable amounts of total (tubulin) and epithelial cell (cytokeratin 8) transcripts. D: Relative fluorescence intensity on live corpus distal epididymal sperm after 60 min coincubation with DilC12-labeled CD9-positive microvesicles in the presence of normal IgG or anti-CD224, anti-CD26 or anti-CD9 and anti-CD26 antibody Fab fragments. The maximum fluorescence was considered to be 100%. Results are presented as average ± s.e.m. from three different experiments, * differs significantly p<0.05.

Figure 7

doi: https://doi.org/10.1371/journal.pone.0065364.g007