Analysis of chromosomal alterations induced by asbestos and ceramic fibers

Abstract

Asbestos and other mineral fibers have long been known as carcinogenic agents. However, the primary mechanisms of fiber-induced carcinogenesis still remain unclear. We have investigated mitotic disturbances caused by amosite, crocidolite, and chrysotile in Syrian hamster embryo (SHE) fibroblasts. We also analyzed micronucleus formation as a result of mitotic disturbances, and carried out a characterization of the induced micronucleus population by kinetochore staining. In addition, the spindle fiber morphology was examined. Supravital UV-microscopy was used to analyze changes in chromatin structure, impaired chromatid separation and blocked cytokinesis. All three fiber types induced micronuclei in SHE cells with a high frequency (up to 200 MN/2000 cells; dose range: 0.1–5.0 μg/cm 2) in a dose-dependent manner with a maximum between 48 and 66 h. Kinetochore staining revealed that 48% of fiber-induced micronuclei reacted positively. Furthermore, spindle deformation was observed in cells with disturbed meta- and anaphases while the spindle fiber morphology appeared unchanged. Our results show that asbestos fibers may cause both loss as well as breakage of chromosomes in the absence of direct interaction with spindle fibers. In addition, we analyzed the induction of micronuclei, hyperdiploidy and chromosome breakage in human amniotic fluid cells (AFC) in vitro by amosite, chrysotile and crocidolite asbestos and ceramic fibers. The response of human (AFC) and rodent (SHE) cells to fiber treatment was compared using the micronucleus assay. AFC were much less susceptible than SHE cells to the induction of micronuclei by mineral fibers. The application of fluorescence in situ hybridization (FISH) with tandem DNA probes yielded more detailed informations about specific structural chromosome aberrations in the 1(cen-q12) and 9(cen-q12) regions and about abnormal numbers of chromosomes in interphase AFC. Using this FISH approach we found a statistically significant increase of chromosomal breakage in the pericentric heterochromatin regions of chromosomes 1 and 9 in AFC after exposure to asbestos and ceramic fibers. The number of hyperdiploid cells was also significantly increased. These results show that asbestos as well as ceramic fibers are inducers of structural and numerical chromosomal alterations.