Two potential clinical applications of the alkaline single-cell gel electrophoresis assay: (1). Human bladder washings and transitional cell carcinoma of the bladder; and (2). Human sperm and male infertility.
Part 1: The alkaline single-cell gel electrophoresis (comet) assay was used to analyse the integrity and DNA content of exfoliated cells extracted from bladder washing specimens from 9 transitional cell carcinoma patients and 15 control patients. DNA damage, as expressed by % tail DNA and tail moment values, was observed to occur in cells from both control and bladder cancer samples. The extent of the damage was, however, found to be significantly greater in the cancer group than in the control group. Comet optical density values were also recorded for each cell analysed in the comet assay and although differences observed between tumour grades were not found to be statistically significant, the mean comet optical density value was observed to be greater in the cancer group than in the control population studied. These preliminary results suggest that the comet assay may have potential as a diagnostic tool and as a prognostic indicator in transitional cell carcinoma. Part 2: Baseline DNA damage in sperm cells from 13 normozoospermic fertile males, 17 normozoospermic infertile males and 11 asthenozoospermic infertile males were compared using a modified alkaline comet assay technique. No significant difference in the level of baseline DNA damage was observed between the 3 categories of sperm studied; however the untreated sperm cells were observed to display approximately 20% tail DNA. This is notably higher than the background DNA damage observed in somatic cells where the % tail DNA is normally less than 5%. Sperm from the 3 groups of men studied were also compared for sensitivity to DNA breakage, using the modified alkaline comet assay, following X-ray irradiations (5, 10 and 30 Gy) and hydrogen peroxide treatments (40, 100 and 200 microM). Significant levels of X-ray-induced damage were found relative to untreated control sperm in the two infertile groups following 30 Gy irradiation. Significant damage in hydrogen peroxide-treated sperm was observed in sperm from fertile samples, at 200 microM and in infertile samples at 100- and 200-microM doses relative to controls. These results therefore indicate that fertile sperm samples are more resistant to X-ray- and hydrogen peroxide-induced DNA breakage than infertile samples. Further studies involving greater numbers of individuals are currently in progress to confirm these findings.
McKelvey-Martin
V J
VJ
Cancer and Ageing Research Group, School of Biomedical Sciences, University of Ulster, Coleraine, UK. V.McKelvey@ulst.ac.uk
Melia
N
N
Walsh
I K
IK
Johnston
S R
SR
Hughes
C M
CM
Lewis
S E
SE
Thompson
W
W
D000368:Aged; D001706:Biopsy; D002295:Carcinoma, Transitional Cell / Q000175:diagnosis* / Q000235:genetics; D004249:DNA Damage; D004252:DNA Mutational Analysis / Q000379:methods*; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007248:Infertility, Male / Q000175:diagnosis / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D009152:Mutagenicity Tests / Q000379:methods*; D013094:Spermatozoa / Q000737:chemistry; D001749:Urinary Bladder Neoplasms / Q000175:diagnosis* / Q000235:genetics
D004273:DNA, Neoplasm