Peripheral Blood Monocytes as Adult Stem Cells: Molecular Characterization and Improvements in Culture Conditions to Enhance Stem Cell Features and Proliferative Potential
Coculture with lymphocytes increases the fraction of mitotically active PCMO. (a) Adherently growing peripheral blood monocytes isolated by elutriation were indirectly cocultured with increasing numbers of autologous lymphocytes in transwell inserts (top, pore size 0.4 μm). Ratios of monocytes (M) : lymphocytes (L) are indicated below the images. After 4 days, cultured cells were fixed and double-stained with an antibody to Ki67 (magenta) and DAPI (blue) as control. The procedure of elutriation was described previously . (b) Detection of expression in elutriated monocytes cocultured with lymphocytes at different ratios. After coculture, monocytes were lysed and analyzed for expression of by immunoblotting. The housekeeping protein α-tubulin served as a loading control. (c) As in (b), except that the immunoblots were probed with antibodies to phospho-Smad2 (p-Smad2) and phospho-Smad3 (p-Smad3). Signal strengths of Smad3 and Smad2 should be assessed relative to those for α-tubulin.