Presence of encircling granulosa cells protects against oxidative stress-induced apoptosis in rat eggs cultured in vitro.
Increased oxidative stress (OS) due to in vitro culture conditions can affect the quality of denuded eggs during various assisted reproductive technologies (ARTs). Presence of intact granulosa cells may protect eggs from OS damage under in vitro culture conditions. The present study was aimed to investigate whether encircling granulosa cells could protect against hydrogen peroxide (H2O2)-induced egg apoptosis in ovulated cumulus oocyte complexes (COCs) cultured in vitro. The OS was induced by exposing COCs as well as denuded eggs with various concentrations of H2O2for 3 h in vitro. The morphological changes, total reactive oxygen species (ROS) as well as catalase expression, Bax/Bcl-2, cytochrome c levels and DNA fragmentation were analysed in COCs as well as denuded eggs. Our results suggest that H2O2treatment induced morphological apoptotic features in a concentration-dependent manner in denuded eggs cultured in vitro. The 20 µM of H2O2treatment induced OS by elevating total ROS level, reduced catalase and Bcl-2 expression levels with overexpression of Bax and cytochrome c and induced DNA fragmentation in denuded eggs cultured in vitro. The presence of encircling granulosa cells protected H2O2-induced morphological apoptotic features by preventing the increase of Bax, cytochrome c expression levels and DNA fragmentation in associated egg. However, 20 µM of H2O2was sufficient to induce peripheral granulosa cell apoptosis in COCs and degeneration in few denuded eggs cultured in vitro. Taken together our data suggest that the presence of encircling granulosa cells could be beneficial to protect ovulated eggs from OS damage under in vitro culture conditions during various ART programs.
Cell Physiology Laboratory, Department of Zoology, Institute of Science, Banaras Hindu University, Varanasi, 221 005, Uttar Pradesh, India.
Meenakshi
M
Tiwari
Cell Physiology Laboratory, Department of Zoology, Institute of Science, Banaras Hindu University, Varanasi, 221 005, Uttar Pradesh, India.
Anima
A
Tripathi
Cell Physiology Laboratory, Department of Zoology, Institute of Science, Banaras Hindu University, Varanasi, 221 005, Uttar Pradesh, India. shailchaubey@gmail.com.
Shail K
SK
Chaube
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D002374:Catalase / Q000235:genetics; D002478:Cells, Cultured; D053938:DNA Fragmentation / Q000187:drug effects; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006107:Granulosa Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D009865:Oocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D027724:Reproductive Techniques, Assisted
D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D002374:Catalase
Apoptosis; Bax/Bcl-2; DNA fragmentation; Granulosa cells; Hydrogen peroxide; Rat egg