Time-course of gelsolin and PARP processing during MK differentiation.

CD34⁺ cells were cultured during 6 days (lane 1), 11 days (lanes 2-4), and 13 days (lanes 5-7), in the presence of TPO only (lanes 1, 2, 5), TPO and Z-VAD.fmk from day 8 to day 11 (lanes 3 and 6) or TPO and STS during the last 12 hours (lanes 4 and 7). Cells were lysed and analyzed by immunoblotting with antigelsolin mAb (A) or anti-PARP antibody (B). Full-length gelsolin (A) and PARP (B) (migrating at 93 kd and 116 kd, respectively) and their cleavage products (46 kd and 86 kd, respectively) are indicated. The cleaved subunit was not detected when cells were cultured for 6 days in the presence of TPO (lane 1) and was present in mature MKs (lanes 2 and 5). This process is completely reverted by the inhibitor z-VAD.fmk (lanes 3 and 6), and the cleaved form is increased after STS treatment (lanes 4 and 7).