Micronuclei and other nuclear anomalies in buccal smears: a field test in snuff users

A revised protocol for the exfoliated cell micronucleus assay was field-tested in a population exposed to a genotoxic agent, snuff, at levels associated with a significant increase in cancer risk. The standard assay involves examination of epithelial smears to determine the prevalence of micronucleated cells, an indication of chromosome breakage or mitotic interference. The assay was revised to increase specificity and to include separate scoring of other nuclear anomalies associated with cytotoxicity and genotoxicity. The modified assay was applied to buccal smears of 38 female snuff users and 15 female nonusers recruited from a North Carolina clinic in 1987. The prevalence of micronucleation was elevated in the snuff users as compared with the nonusers (prevalence ratio = 2.4, 95% confidence interval 1.1-5.2) and, to a lesser extent, at the usual contact site as compared with a distal buccal site in the snuff users (prevalence ratio = 1.5, 95% confidence interval 0.9-2.5). The pattern of relative frequencies of several nuclear anomalies provided strong evidence of a cytotoxic effect, the prevalence ratios ranging from 2 to 13. Nuclear degenerative phenomena can be difficult to distinguish from classical micronuclei; thus, the observed association of indicators of cytotoxicity with exposure introduces the possibility of bias away from the null in micronucleus findings due to differential misclassification. Until methods to better distinguish extranuclear bodies of different origins become available, investigators should use the revised protocol and should focus on agents not thought to be cytotoxic.