Enhanced survival and proliferation of gp130^Y757F/Y757F hematopoietic cells. (A) Survival of bone marrow and spleen cells from each genotype in response to IL-6. Equivalent cell numbers were cultured over 5 days, and the number of viable cells was counted. Data from 3 separate experiments were pooled and are presented as the mean ± SD. ^*P < .05 versus data from the corresponding gp130^+/+ cultures. □ indicates gp130^+/+; ▪, gp130^Y757F/Y757F; and ▦, gp130^Y757F/Y757F:Stat3^+/- mice. (B) Cells from IL-6–treated bone marrow and spleen cultures were stained with FITC-labeled annexin-V and subjected to flow cytometric analysis. The x-axis represents FITC intensity, and y-axis represents relative cell number. The numbers indicate the percent of annexin-V^- (M1 region) or annexin-V⁺ (M2 region) cells. (C,D) Bone marrow cells were cultured in the presence of IL-6 for 3 hours and 24 hours, after which whole-cell lysates were immunoblotted with the indicated antibodies (C), and semiquantitative RT-PCR analysis of Socs3 and Bcl-xL gene expression was performed on cDNA derived from total RNA (D). Also shown is the RT-PCR for β-actin as an internal control for sample normalization and integrity. Proliferation of unfractionated (E) and Gr1⁺ (F) bone marrow cells from gp130^+/+, gp130^Y757F/Y757F, and gp130^Y757F/Y757F:Stat3^+/- mice with or without IL-6 treatment over 48 hours prior to being pulsed with ³H-thymidine for 12 hours. Data are presented as the mean ± SD for triplicate cultures. ^*P < .05 versus data from the corresponding gp130^+/+ cultures. No significant differences in ³H-thymidine incorporation were observed in cultures of Gr1⁺ bone marrow cells from each genotype treated with IL-3 (gp130^+/+, 3740 ± 1101 counts per minute [cpm]; gp130^Y757F/Y757F, 3612 ± 472 cpm; gp130^Y757F/Y757F:Stat3^+/-, 4227 ± 372 cpm).