Effects of F/NAIT vs control sera on fetal/neonatal megakaryopoiesis. (A-E) Cord blood–derived CD34⁺ cells were cultured for 14 days in the presence of thrombopoietin and 10% maternal sera (F/NAIT or control). (A) The MK number for each culture was calculated by multiplying cell count and percentage of CD41⁺ cells. To quantify growth, the number of MKs generated in each culture was expressed as a percentage of the mean MK number in the corresponding control cultures (see details in supplemental Methods). Compared with control sera (n = 8), 14 out of 17 F/NAIT samples caused significant reductions in MK number at the end of culture (day 14). Samples marked with an asterisk were used for the studies shown in Figure 1F-G. (B) CD41 and CD42b surface expression in F/NAIT and control cultures. (C) MK ploidy distribution in F/NAIT and control cultures. (D) Cell numbers were evaluated on days 4, 7, 11, and 14 of culture. The cell number at each different time point was expressed as a percentage of the mean control cell count on day 14 (100%). (E) Percentages of dead cells in the cultures on days 7, 11, and 14, after staining with trypan blue. (F-G) Day 7 cells were treated with 20% of either F/NAIT (n = 6) or control sera (n = 4) for 24, 48, or 72 hours. (F) Percentage of apoptotic MKs after 24, 48, and 72 hours of exposure to F/NAIT or control sera. (G) Percentage of Ki67+ MKs after 24, 48, and 72 hours of exposure to F/NAIT or control sera. Bar graphs indicate the mean ± standard error of the mean of each group. *P < .05; ***P < .001.