Comet electrophoresis study of DNA damage in oral mucosal cells of patients with oral squamous carcinoma

Dr GS Kumar, MDS

SDM College of Dental Sciences, Dharwad, Karnataka, India.

AIM: To assess DNA damage in mucosal cells of oral cancer patients using Comet Electrophoresis. METHOD: The study group comprised of 30 patients of Oral Squamous Carcinoma (habituated to tobacco) confirmed by biopsy. The patients were assessed for TNM staging & Broders grading. The control group comprised of 30 healthy volunteers without tobacco habituation. Mucosal cells from the lesional tissue and healthy buccal mucosa from the control group were transferred to RPMI 1640, animal tissue culture medium and centrifuged. A cell suspension, using 70µ liter of low melting agarose was made. This was layered onto agarose layer (110µ liter) and overlaid with 70 µ liter of low melting agarose, on half frosted microscopic slides. These were lysed overnight in cold lysing solution and electrophoresis with alkaline buffer was carried out. Further, they were stained with ethidium bromide and DNA damage, visible as comets, was measured with an oculometer. The mean and standard deviations were calculated and statistical significance was determined. RESULTS:A high statistical significant difference was noted between control (mean 0.1842µ meter) and the study group (mean1.7469µ meter). Stepwise increase in mean DNA damage was observed from stage I to stage IV and between well, moderate and poorly differentiated grades. Statistical significance was observed between stage II and IV and also between well and moderately differentiated grades. CONCLUSIONS: The results indicate the significance of Comet Electrophoresis in assessing the DNA damage in mucosal cells of Oral Cancer patients and predicting the prognosis.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Diagnostic Markers.