Use of fluorescence in situ hybridization to detect chromosome-specific changes in exfoliated human bladder and oral mucosa cells.
Change in chromosome number, numerical aneuploidy, has been consistently linked with cancer development. Since 90% of cancers arise in epithelial tissues, techniques that measure aneuploidy in these tissues would be very useful. Here we describe methods of optimization and suggest use of fluorescent in situ hybridization (FISH) to detect aneuploidy in exfoliated epithelial cells collected from the mouth and bladder. A total of 10,383 urothelial cells and 4,691 buccal cells were scored in order to determine a baseline frequency of aneuploidy in human volunteers using a classical satellite probe for chromosome 9. Protein digestion with pepsin was found to be more efficient at removing the keratinized cell membrane and optimizing probe penetration than acid washes, detergent washes, or hypotonic treatments. A 20 min cellular digestion with 200 micrograms/ml and a 30 min digestion with 300 micrograms/ml of pepsin in 0.01 M HCl optimized probe penetration in urothelial and buccal cells, respectively. Average frequencies for 0, 1, 2, 3, and 4 hybridization regions were 10.3, 10.1, 78.4, 1.0, and 0.3% for urothelial cells and 8.8, 9.8, 79.4, 1.3, and 0.3% for buccal cells, respectively. These results are very similar to those previously described in lymphocytes. The urothelial cells of males had a lower frequency of diploid cells and a higher frequency of cells without hybridization regions than females (P < 0.02). No statistically significant variability was found between individuals or sex groups in buccal cells. Our data show that FISH is a useful tool to detect changes in frequency of aneuploidy in exfoliated epithelial cells and has good potential for monitoring human populations exposed to genotoxic agents.
Moore
L E
LE
Department of Biomedical and Environmental Health Sciences, School of Public Health, University of California, Berkeley 94720.
Titenko-Holland
N
N
Smith
M T
MT
D000328:Adult; D000782:Aneuploidy; D002899:Chromosomes, Human, Pair 9; D004847:Epithelial Cells; D004848:Epithelium / Q000648:ultrastructure; D005260:Female; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007399:Interphase; D008297:Male; D009061:Mouth Mucosa / Q000166:cytology / Q000648:ultrastructure*; D009152:Mutagenicity Tests / Q000379:methods; D015203:Reproducibility of Results; D001743:Urinary Bladder / Q000166:cytology / Q000648:ultrastructure*; D014556:Urine / Q000166:cytology