1. Inhibition of stromal cell–induced survival of primary B-CLL cells by cyclopamine in vitro.
Inhibition of stromal cell–induced survival of primary B-CLL cells by cyclopamine in vitro. Primary B-CLL cells were cultured in control medium or cocultured with OMA-AD stromal cells or OMA-AD cells with 5 μmol/L cyclopamine for 72 h. The proportion of B-CLL cells at different stages of apoptosis was determined by flow cytometry using the Annexin-PI assay. A to C. A representative culture of primary B-CLL cells on the OMA-AD stromal cell layer. D to F. Representative flow cytometric profile of B-CLL cells cultured in the presence of control medium or OMA-AD cells or OMA-AD cells with cyclopamine as determined by the Annexin-PI assay. There was a decrease in apoptotic B-CLL cells in the presence of coculture with OMA-AD stromal cells compared with controls, and this influence was reversed in B-CLL cells cocultured with OMA-AD cells and cyclopamine. G. Expression of Hh ligand in OMA-AD cells using mouse brain as positive control. H to K. Primary B-CLL cells were cultured in control medium or stromal-CM or stromal-CM with 5 μmol/L cyclopamine for 24 h, and GLI1 expression was determined by real-time PCR. The proportion of B-CLL cells at different stages of apoptosis was determined after 72 h by flow cytometry using the Annexin-PI assay. There was a significantly increased GLI1 expression (fold change) and decreased apoptotic cell population in the presence of stromal-CM compared with control. The effect was reversed by cyclopamine. Results are obtained from B-CLL cells of three different patients.
Ganapati V. Hegde et al. Mol Cancer Res 2008;6:
©2008 by American Association for Cancer Research