We compared changes in the redox status and intensity of oxidative modification of proteins in intact Jurkat tumor cells and cells cultured with buthionine sulfoximine, an inhibitor of the key enzyme of glutathione synthesis γ-glutamylcysteine synthetase. The glutathione system components play a role in modulation of the content of protein-bound glutathione, protein carbonyl derivatives, bityrosine, and oxidized tryptophan, and in dysregulation of apoptosis in Jurkat tumor cells. Inhibition of de novo synthesis of glutathione in Jurkat tumor cells was followed by accumulation of hydroxyl radical, a reduction in the level of protein-bound glutathione and oxidized tryptophan, and a rise in the concentration of protein carbonyl derivatives. These changes were accompanied by activation of programmed cell death.
Цитокинам и их рецепторам принадлежит значимая роль как в формировании, так и в последующем течении и исходе аутоиммунной патологии щитовидной железы (ЩЖ). Интерлейкин 2 (IL 2), интерлейкин 4 (IL 4) и фактор некроза опухолей альфа (TNF α)-цитокины, оказывающие многогранное влияние на различные этапы иммунного ответа: развитие воспалительного ответа, пролиферацию клеток антителопродуцентов, антителогенез и синтез белков острой фазы воспаления. Существующая ранее модель развития аутоиммунных тиреопатий рассматривала аутоиммунный тиреоидит и болезнь Грейвса (БГ) как два противоположных состояния с позиции преобладающего профиля активации Th1/Th2 лимфоцитов. В рамках исследования произведена оценка продукции цитокинов Th1 и Th2 профиля мононуклеарными лейкоцитами крови у пациентов с БГ, оценка состояния рецепторного аппарата лимфоцитов, а также определены иммунофенотипические особенности лимфоцитов крови у пациентов с БГ, влияние функционального состояния ЩЖ на изучаемые показатели. Было показано, что иммунорегуляторные цитокины, как Th1 (IL 2) , так и Th2 (IL 4) хелперных лимфоцитов, участвуют в иммунопатогенезе БГ. Уровень продукции IL 2, IL4, а также TNF α и количество презентирующих комплементарные им рецепторы лимфоцитов значимо не различались между подгруппами пациентов с БГ в фазе тиреотоксикоза или эутиреоза. Тем не менее существуют сильные корреляционные связи продукции иммунорегуляторных цитокинов (IL 2, IL 4) с увеличением продукции тиреоидных гормонов и увеличением объема ЩЖ при БГ, что подтверждает "функциональный синергизм" данных цитокинов в отношении развития аутоиммунного воспаления при БГ.
Background: Breast tumors are number one cause of cancer morbidity and mortality among women around the world, and Russia is not an exception. Many proteins that control proliferation of immortalized cells are redox-regulated, which is essential for modulating cellular proliferative activity, especially during tumor growth. Studying the role of glutaredoxin and glutathione in cell cycle phase distribution will allow not only to identify the molecular targets regulating cell proliferation, but also to develop methods of diagnosis and targeted therapy of socially sensitive diseases, including breast cancer, in the future.Aims: To evaluate the role of glutathione and glutaredoxin in the molecular mechanisms regulating MCF-7 breast cancer cell proliferation under the effects of roscovitine, a cyclin-dependent protein kinase inhibitor.Materials and methods: The MCF-7 cell line (human breast adenocarcinoma) was used in the study. The cell culture was incubated in the presence and absence of roscovitine in the final concentration of 20 µmol for 18 h. The production of reactive oxygen species, the distribution of cells between cell cycle phases and the amount of Annexin V positive cells were determined using flow cytometry. The concentrations of total, reduced and oxidized glutathione, protein SH groups and protein-bound glutathione were measured by spectrophotometry. The levels of glutaredoxin, cyclin E and cyclin-dependent protein kinases were estimated by Western blotting with monoclonal antibodies.Results: The effects of roscovitine in the MCF-7 cells resulted in cell cycle arrest in G2/М phases with the decreased levels of cyclin E and cyclin-dependent protein kinase 2. It was accompanied by activation of programmed cell death. In tumor cells incubated in the presence of roscovitine, oxidative stress was triggered, which was accompanied by the elevated generation of reactive oxygen species, the decrease in the concentration of reduced glutathione, and the rise in the level of glutaredoxin. It contributed to the increase in protein glutathionylation against the backdrop of the decreased SH group concentration.Conclusions: Breast cancer cell proliferation under the effects of roscovitine is reduced following not only the decrease in the cyclin level and cyclin-dependent protein kinase activity, but also the shift in the intracellular oxidant/antioxidant ratio. Roscovitine-induced oxidative stress in the MCF-7 cells contributed to protein glutathionylation with the changes in the protein structure and functions. It results in impaired cell cycle progression, indicating a possibility to regulate cellular proliferation through modulating functional properties of redox-dependent proteins using the glutathione/glutaredoxin system.
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