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. 2011 Sep 26;208(10):2017–2031. doi: 10.1084/jem.20110750

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© 2011 Josefsson et al.

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Figure 7. — Deletion of Bak and Bax protects megakaryocytes from death signals. (a) Viability of Bak- and Bax-deficient MGKs in response to ABT-737, etoposide, and STS. Mature FLC-derived MGKs were purified from a BSA gradient and cultured in serum-free media with TPO and either ABT-737 (5 µM), Etoposide (50 µM), STS (10 µM), vehicle controls DMSO (0.05% ABT-737, 1%, STS), or ethanol (EtOH) 0.27%, or in combination with Q-VD-Oph (50 µM). Viability was measured 24 h after reseeding using the CellTiter-Glo assay system. DMSO control–treated Bak^−/−Bax^−/− levels were set as 100%. Data represent mean ± SEM. n = 3–4 independent experiments. (b) Caspase-3/7 activity in MGKs derived and treated as in a. Caspase-Glo assay was conducted 5 h after reseeding. MGK were pooled from >4 animals per genotype. Data represent mean ± SEM. n = 4 independent experiments. (c) Proplatelet formation in MGKs derived and treated as in a. Data represent mean ± SD, n = 2 technical replicates. MGKs were pooled from >2 animals per genotype. Representative of two independent experiments. (d) Platelet counts at 7 wk of age in mice lacking Bcl-x_L, Bak, and Bax in the MGK lineage. Each symbol represents 1 mouse. Data represent mean ± SEM. n = 6–10 mice per genotype. (e) Platelet counts in lethally irradiated mice reconstituted with FLC, and injected with a single IP injection of carboplatin (100 mg/kg) on day 0. Cohorts of mice (day 8; n = 13 Bak^+/+Bax^+/+ and 17 Bak^−/−Bax^−/−; day 11, n = 21 Bak^+/+Bax^+/+ and 24 Bak^−/−Bax^−/−; day 16, n = 13 Bak^+/+Bax^+/+ and n = 16 Bak^−/−Bax^−/−) were sacrificed at indicated time points. Data represent mean percentage of change to baseline ± SEM. (f) Erythrocyte counts from the experiment presented in e. Data represent mean percentage of change from baseline ± SEM. *, P < 0.05; **, P < 0.005; ***, P < 0.0001.