Figure 4.

Functional validation of selected compounds. (A) Germline apoptosis assay quantification. After exposure to each of the 10 least aneugenic compounds in the C. elegans assay or to each of the 10 most aneugenic compounds, we quantified apoptotic levels through the use of the Plim-7ced-1::gfp reporter DMSO was the negative control, and the DNA damaging agent camptothecin the positive control. The lower and upper edges of the box plot represent the first and third quartiles, respectively, with the median represented as a line within the box; the whiskers extend to ± 1.5 times the interquartile range. (B) DAPI staining of germ­line nuclei revealed profound germ­line defects after exposure to Maneb and TCMTB (bars = 50 µm). Assembled germ­lines show areas of reduced nuclear density (asterisks), intermixed meiotic stages (red arrowheads), and unequally spaced diakinetic nuclei (white arrows). Insets show high magnification examples of late diakinetic nuclei (bars = 4 µm); note that nuclei with chromosomes in a pachytene stage-like organization are present intermixed with diakinetic nuclei in late prophase after maneb exposure. *p < 0.001 compared with DMSO by ANOVA followed by Dunn’s comparison. The numbers of apoptotic nuclei in the 10 least and most aneugenic compounds were highly significantly different from each other (p < 0.0001; two-tailed Mann–Whitney U test, 95% CI)