Inhibition of platelet formation by Bcl-2 overexpression.

Bone marrow CD34⁺ cells were cultured in the presence of TPO. At day 6, cells were cocultured for 2 days with cells producing either the MFG-GFP or MFG-BIG virus. MKs transduced by the retroviral vectors were sorted on the expression of CD41 (R-PE) and GFP. GFP⁺ CD41⁺ and CFP⁻CD41⁺ cells were subsequently cultured with TPO and proplatelet formation was studied. Panels A and B: analysis by flow cytometry of the correlation between GFP and Bcl-2 expression in MFG-GFP–transduced cells (A) and MFG-BIG–transduced cells (B). In these cells a linear relationship between GFP and Bcl-2 is observed. Panels C and D: representative MKs obtained after MFG-GFP and MFG-BIG transduction. Proplatelet-forming MKs expressing GFP are found after transduction with the MFG-GFP virus (C), while MKs expressing GFP remain round after transduction with the MFG-BIG virus (D). Original magnification, × 150. Panel E: Kinetics of proplatelet formation in Bcl-2–overexpressing MKs. The graph shows a significant inhibition of proplatelet formation by Bcl-2–overexpressing MKs (one experiment made in triplicate). Experiments were repeated 3 times and yielded similar results each time.