Figure 2.

NOX-A12 increases CLL cell migration beneath BMSCs and induces migration of leukemic cell lines in a dose-dependent manner. (A) A representative phase-contrast photomicrograph of CLL cells migrated underneath stromal cells 9-15C and TSt-4 (control), and in comparison, increased migration of the same CLL sample after incubation of the stromal cells with 100 nM NOX-A12. (B) CLL cells that migrated underneath stromal cells were quantified by flow cytometry. The box-and-whisker diagrams represent the median including interquartile range, maximum, and minimum of migrated CLL cells from 6 different patients after pretreatment of the stromal cells with or without NOX-A12. The Wilcoxon matched-pairs signed-rank test was used for statistical analysis. (C) The T-cell leukemic cell line Jurkat, and (D) the ALL cell line Nalm-6 were labeled with Calcein AM and cocultured with murine stromal MS-5 cells preincubated with different concentrations of NOX-A12 and/or AMD3100. After incubation of 3 (Jurkat) or 6 (Nalm-6) hours, respectively, leukemia cells were washed from the stromal cell layer. Results indicate the percentages of migrated plus attached cells beneath the stromal cell layer, representing the mean ± SD values of bottom fluorescence measurement (n = 3). The one-way analysis of variance test was conducted for statistical analysis. Data are representative of 3 independent experiments. *P = .01 to .05; **P = .001 to .01; ***P < .001.