MAL expression in MKs. CD34⁺ cells from cord blood or cytapheresis were cultured with TPO. (A) At day 9 of culture, mRNA levels of MAL and MAL16 were evaluated by quantitative RT-PCR. Their expression was normalized with respect to HPRT mRNA. (B) To investigate MAL transcription level during MK differentiation, cells were sorted at day 6 of culture in 4 populations: for cord blood, CD34⁺CD41⁻, CD34⁻CD41⁺CD42⁻, CD34⁻CD41⁺CD42^low, and CD34⁻CD41⁺CD42^high; and for cytapheresis, CD34⁺CD41⁻, CD34⁺CD41⁺CD42⁻, CD34⁺CD41⁺CD42⁺, and CD34⁻CD41⁺CD42⁺. MAL mRNA level was evaluated as in panel A. Error bars in panels A and B represent the SD of the mean of 3 repeated experiments each performed in triplicate wells. (C) To investigate protein levels by Western blot analysis during MK differentiation, total cell populations were harvested on day 3 (D3), day 6 (D6), day 9 (D9), and day 12 (D12) for cord blood-derived MK and at day 0 (D0), day 3 (D3), day 6 (D6), and day 9 (D9) for cytapheresis-derived MKs. HDAC-1 and HSC70 were used as internal control. The figure illustrates representative data of 2 experiments with similar results.