Protection of primary human macrophages by ACM is PI3K-, Ca2+-, and ERK1/2-dependent. (A) Human primary monocyte-derived macrophages remained untreated or were exposed to ACM for 16 hours. Following a medium change, apoptosis was induced for 8 hours with 10 ng/mL TNF-α and 10 μg/mL CHX. Viability was assessed by annexin V/PI staining and FACS analysis. (i) Data in histograms are presented as the mean ± SD from 3 independent experiments. Statistical differences are marked with an asterisk (P < .05). (ii) Typical FACS traces. (B) Human primary macrophages were not treated or were exposed to ACM for 16 hours in the presence or absence of LY92004, PD98059, or Bapta-AM at the indicated concentrations. Apoptosis was induced for 8 hours with 10 ng/mL TNF-α and 10 μg/mL CHX. (i) Caspase-3 activity. Data are presented as the mean ± SD from 6 independent experiments. Significant differences between samples are marked with an asterisk (P < .05). Western analysis shows expression of Bcl-2 and phospho-Ser136-Bad. One representative experiment of 3 is displayed. (ii) Quantification of Western data normalized to actin by using AIDA software. Data are presented as the mean ± SEM from 4 independent experiments.
