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Background. Glycoprotein-P (Pgp, АВСВ1) is a transporter protein participating in pharmacokinetics of medical drugs, and also in development of resistance of tumor cells to chemotherapy. Aim. To study the influence of progesterone on the activity of Pgp in vitro on a cell model of human small intestinal epithelium. Materials and Methods. The work was conducted on Caco-2 cells. The activity of Pgp was evaluated by transport of fexofenadine in a special transwell-system. Concentration of fexofenadine was analyzed by HPLC method. The amount of Pgp was determined by EIA method. Four series of experiments were conducted: control cells preincubated with clean transport medium without addition of any substances; influence of rifampicin on the activity and synthesis of Pgp in the concentration 10 mol/l in preincubation for 3 days (induction control); influence of progesterone on the activity of Pgp in concentrations 1, 10 and 100 mol/l in preincubation for 30 min; influence of progesterone on the activity and synthesis of Pgp in concentrations 1, 10 and 100 mol/l in preincubation for 3 days. Results. Progesterone in the concentrations 1 and 10 M in incubation with cells within 30 minutes did not show any reliable influence on the activity of Pgp, however, in concentration 100 M it reduced the activity of the transporter protein. In incubation of Caco-2 cells with progesterone in concentrations 1, 10 and 100 M within 3 days the activity of Pgp remained unchanged. Progesterone in concentration 100 M in incubation within 3 days significantly increased synthesis of Pgp in enterocytes by 114.3% as compared to control, and in other used concentrations (1 and 10 M) it produced no reliable effect. Conclusion. In in vitro experiments on Caco-2 cells progesterone in concentration 100 M produces a direct inhibiting effect on the activity of Pgp; however, in incubation within 3 days it increases synthesis of the transporter protein, which cancels out its inhibitory activity.

INTRODUCTION: Nuclear factor E2-related factor 2 (Nrf2) is a member of capncollar (CNC) family of subfamily of leucine zipper transcription factors that regulates cell protection against toxic substances and oxidants. AIM: To determine location, mechanism of activation and role of Nrf2 in conditions of oxidative stress in vitro. MATERIALS AND METHODS: The study was performed on human colon adenocarcinoma cell line (Caco-2). Oxidative stress (OS) was modeled by adding hydrogen peroxide (Н2О2) at concentrations of 0.1 М100 М to the nutritive medium and incubation for 24 and 72 hours. In assessment of Nrf2 function, its inhibitor ― AEM1 ― was added to cells at a concentration of 5 М. The extent of OS development was determined using photometric methods by the concentration of protein SH-groups and carbonyl derivatives of protein, and the activity of superoxide dismutase (SOD). Viability of cells was assessed by the results of cytotoxic test (MTT assay), the amount of Nrf2 in the cytoplasm and nucleus was determined by heterogenous ELISA method. RESULTS: Incubation of Caco-2 cells with Н2О2 resulted in decrease in the level of protein SH-groups and increase in the concentration of carbonyl derivatives of protein. In incubation with H2O2 at concentrations of 0.1 М10 М for 24 hours and 10 М for 72 hours, the activity of SOD increased. At concentrations of Н2О2 of 50 М and 100 М (24 hour and 72 hour), SOD activity and viability of cells decreased. Exposure to Н2О2 led to translocation of Nrf2 from the cytoplasm into nucleus. Direct correlation dependence was revealed between concentration of protein SH-groups and the amount of Nrf2 in the cytoplasm in incubation with H2O2 for 24 hour (r = 0.44, р = 0.03), 72 hour (r = 0.34, р = 0.05). The amount of Nrf2 in the nucleus positively correlated with SOD activity in the cytoplasm on exposure to H2O2 for 24 hour (r = 0.77, р = 0.0001) and 72 hour (r = 0.36, р = 0.06). In inhibition of Nrf2 in conditions of exposure to H2O2, the viability of cells decreased to a larger extent. CONCLUSION: Hydrogen peroxide induces the nuclear translocation of Nrf2, which promotes activation of antioxidant enzyme SOD and preserves viability of cells of OS conditions in vitro.

Рязанский государственный медицинский университет имени академика И.П. Павлова, Рязань, Российская Федерация _____________________________________________________________________________ Актуальность. Гликопротеин-Р (Pgp, АВСВ1)-белок-транспортер, обеспечивающий защиту органов и тканей от ксенобиотиков, являющихся его субстратами, выводя их из клеток во внеклеточное пространство и биологические жидкости. Цель. Изучить влияние эстрадиола на активность Pgp in vitro на линии клеток Caco-2. Материалы и методы. Исследования выполнены на линии клеток Caco-2. Активность Pgp анализировали по транспорту его маркерного субстрата-фексофенадина в трансвеллсистеме. Концентрацию фексофенадина оценивали методом высокоэффективной жидкостной хроматографии. Количество Pgp определяли методом ИФА. Эксперимент включал следующие серии: клетки, которые преинкубировали с чистой транспортной средой без добавления каких-либо веществ (контрольная серия); влияние рифампицина в концентрации 10 мкмоль/л при преинкубировании в течение 3 сут на активность и синтез Pgp (контроль индукции); влияние эстрадиола в концентрациях 1 и 10 мкмоль/л при преинкубировании в течение 30 мин на активность и синтез Pgp; влияние эстрадиола в концентрациях 1 и 10 мкмоль/л при преинкубировании в течение 3 сут на активность и синтез Pgp. Результаты. Эстрадиол в концентрациях 1 и 10 мкМ при инкубации с клетками в течение 30 мин достоверно не влиял на активность и синтез Pgp, также как и 1 мкМ эстроген при инкубации 3 сут. В то же время эстрадиол в концентрации 10 мкМ при инкубации в течение 3 сут повышал активность и синтез белка-транспортера. Заключение. Эстрадиол в эксперименте in vitro на клетках линии Caco-2 в концентрации 10 мкМ при инкубировании в течение 3 сут повышает синтез и активность белкатранспортера гликопротеина-Р.