The overall goal of this work is to quantify the enzymatic activity of beta-galactosidase in stressed ovarian cancer cell monolayers. Beta-galactosidase (beta-gal)—a marker of cellular senescence—is an enzyme that catalyzes the hydrolysis of beta-glycoside bonds to yield monosaccharides. Previous studies have shown that beta-gal is upregulated in select ovarian cancers, and its expression/activity level is upregulated when cells are under oxidative stress conditions. To date, no quantitative assessment of beta-gal activity has been performed on ovarian cancer cell lines under typical and/or stressed conditions. Enzymes and other related proteins are likely targets as biomarkers for cancer diagnostics, and as a result, it is important to be able to quantify intracellular enzyme activity levels, not just presence of enzyme. Being able to detect and quantify levels of these biomarkers can aid in the early diagnosis of cancer and other diseases and in the monitoring of treatment and relapse, thereby reducing the mortality rate and improving the long-term survival of patients. Enzyme activity/expression levels in cells will be assessed using western blot analyses and a spectrometric assay with ortho-nitrophenyl-β-galactoside (ONPG) as a substrate. Immunocytochemistry has also been implemented in order to visualize the localization of the beta-gal enzyme in ovarian cancer cell monolayers. Results obtained from the spectrometric assay revealed an increasing trend in the enzymatic activity of beta-gal with each day of cell culture, while immunocytochemistry results revealed beta-gal in the nucleus and lysosomes of ovarian cancer cells. The increasing trend in beta-gal activity may be linked to senescence-associated beta-gal activity, a result of stress-induced premature senescence, which can be invoked with prolonged confluence in culture.

Quantification of beta-galactosidase activity in ovarian cancer cell monolayers

2019 Spring National Meeting

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