ABSTRACT
Polycystic ovary syndrome (PCOS) is accompanied with many disturbances in hormone synthesis and antioxidant defense. Previous reports have indicated that Vitamin D (vit.D) affects gene expression and have roles in normal follicular development. Therefore, we investigated the effects of vit.D on steroidogenesis, apoptosis, reactive oxygen species (ROS) production, and antioxidant defenses of human normal granulosa cells (N-GCs) and granulosa cells from polycystic ovaries (PCO-GCs). Ovarian GCs were obtained during oocyte retrieval procedure from 120 women with PCOS and from 100 healthy women who referred to Shiraz Fertility Center. The isolated GCs were cultured in the presence or absence of vit.D (100â¯nM), for 48â¯h. Concentration of sex steroids was measured by ELISA. Glutathione peroxidase (GPx) and superoxide dismutase (SOD) expression and activities were assessed by q-PCR and photometric methods, respectively. The amount of ROS production was estimated using chemiluminescence and fluorescence methods. Cell viability and apoptosis were detected by Annexin-V/propidium iodide detection kit. Basal estrone and progesterone secretion by N-GCs was significantly higher than that of PCO-GCs. Vit.D significantly increased aromatase and 3ß-hydroxysteroid dehydrogenase activity in N-GCs and PCO-GCs. Basal expression and activity of GPx, in PCO-GCs were significantly lower than those of N-GCs. Treatment with vit.D significantly increased genes expression and enzyme activities in both groups. Basal ROS in PCO-GCs was markedly greater than that of N-GCs, which was attenuated by vit.D treatment. Cell apoptosis was directly correlated with ROS levels. We conclude that vit.D improved N-GCs and PCO-GCs functions through affecting steroidogenesis and enzymatic antioxidant defense. Under vit.D treatment, PCO-GCs could act more similar to N-GCs.
Subject(s)
Antioxidants/metabolism , Gene Expression Regulation, Enzymologic/drug effects , Granulosa Cells/metabolism , Polycystic Ovary Syndrome/metabolism , Reactive Oxygen Species/metabolism , Steroids/biosynthesis , Vitamin D/pharmacology , Adult , Apoptosis , Case-Control Studies , Female , Granulosa Cells/drug effects , Granulosa Cells/pathology , Humans , Polycystic Ovary Syndrome/drug therapy , Polycystic Ovary Syndrome/pathology , Vitamins/pharmacologyABSTRACT
Calcium (Ca(2+)) and reactive oxygen species (ROS) constitute the most influential intracellular signaling molecules participating in the regulation of different sperm functions. Elevating intracellular Ca(2+) and ROS in physiologic range regulate capacitation, motility, acrosome reaction, and sperm-oocyte fusion; whereas cytosolic Ca(2+) overload and ROS overproduction have pathologic effects. Our aim of this study was determination of antioxidant effects of α-tocopherol on sperm motility, viability, and DNA integrity in a condition where cytosolic calcium overload was induced by A23187 (a calcium ionophore). Our results indicated that, α-tocopherol has ability to prevent sperm mortality and save sperm rapid motility after 1 hour incubation. At the same time, A23187 reduced significantly percentage of rapid sperm motility and increased sperm mortality and DNA damage. Results of sperms incubation in the medium contain a combination of A23187 and α-tocopherol showed that α-tocopherol can reduce many of the deleterious effects of A23187. In conclusion, it seems that the harmful effects of A23187 are due to excessive ROS production, and α-tocopherol neutralizes these effects.
