cAMP increases CD34⁺ progenitor cell survival by PKCζ-activated pathway while inhibiting their proliferation. (A) Increased survival of CB CD34⁺ cells cultured for 3 days under cytokine-deprived conditions in the presence of 500 μM dbcAMP (cAMP) as analyzed by flow cytometry. Data represented as fold change (mean ± SD of 6 independent experiments each in triplicate) in the number of PI-positive (Dead cells) in cAMP-treated compared with untreated (CTRL) samples. Inset shows representative changes in the proportion of CD34⁺ cells with fragmented DNA (dead cells [%]) cultured for the indicated time periods untreated (CTRL), treated with cAMP, or treated with cAMP and 10 μM PKCζ PS (cAMP+PSζ). (B) Comparison of the percentage of dividing CD34⁺ cells (mean ± SD of 4 independent experiments) cultured for 3 days in serum-supplemented medium with 500 μM cAMP or without (CTRL), fixed, and PI labeled for cell cycle analysis. (C) Ki-67 expression in G2 cells cultured in full growth medium for 3 days either untreated (CTRL) or with 500 μM cAMP. Data are shown as mean ± SD of 3 independent experiments. Representative flow cytometry analyses are shown on the right; numbers indicate percentages of Ki-67-positive cells. (D) Immunoblot analysis of ERK1/2 phosphorylation (P-ERK1/2) status compared with the total ERK1/2 level in G2 cells incubated for 24 hours with 500 μM cAMP or left untreated (CTRL). Representative of 1 of 3 independent experiments with similar results.