Cytotoxic and Apoptotic Activity of Majoranolide from Mezilaurus crassiramea on HL-60 Leukemia Cells

Figure 3

Effect of majoranolide on caspase-3 activation; expression of apoptosis-related genes and plasma membrane integrity in HL-60 cells. After cell exposure to the compound (concentrations and times indicated), the cells were stained with PE Rabbit Anti-Active Caspase-3 antibodies to quantify caspase activity or with 7-AAD to evaluate plasma membrane integrity, and then counted on a BD Accuri C6 Plus cytometer. FlowJo software was employed for data analysis. For analysis of gene expression, cDNA synthesis was performed from total RNA and data were collected using CFX Manager 3.1 software (Bio-Rad) and normalized to β-actin; untreated cells (UC) = 1. (a) Histograms depict fluorescence intensities of caspase-3 activity for treated and untreated cells after 24 h and 48 h treatments. (b) Geometric mean (GM) fluorescence intensity values. < 0.001 between treated and untreated cells; ^###p < 0.001 between 24 and 48 h treatments (one-way ANOVA followed by Tukey’s posttest). (c) mRNA relative expression by BAX, BCL2, BIRC5, and CASP8 shown as means ± SEM < 0.05; < 0.01; < 0.001 (one-way ANOVA followed by Dunnett’s posttest). (d) Percentages of viable cells according to mean fluorescence intensity are expressed as means ± SEM. < 0.001 (one-way ANOVA followed by Dunnett’s posttest).

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