1. Introduction

Oocytes and eggs of the African clawed frog Xenopus laevis provide the most common model for studying oogenesis, fertilization, meiotic and mitotic cell cycle progression, and apoptosis because of their large size and high biochemical tractability. The term “eggs” is generally used in the frog model for mature ovulated oocytes arrested in metaphase of the second meiotic division by high activity of the key meiotic regulators, such as the maturation promoting factor (MPF) and the cytostatic factor (CSF) [1]. The meiotic metaphase arrest prevents cell cycle progression and parthenogenesis prior to fertilization. Meiotically arrested eggs awaiting fertilization can experience various injuries leading to the loss of their quality. The stress- and age-triggered damage leads to decreased rates of fertilization, polyspermy, parthenogenesis, and abnormal development of embryos. Poor quality of oocytes and eggs is considered to be a cause of infertility and abnormal embryo development in different animals, including mammals [2, 3]. In addition, spontaneous egg activation and exit from the meiotic metaphase arrest make successful fertilization impossible [4, 5]. It was reported that unfertilized Xenopus eggs spontaneously activate, exit the meiotic arrest, and degrade by a robust apoptotic process within 48 hours after ovulation [6, 7].

The intracellular pathways involved in spontaneous egg activation are poorly investigated. It has been suggested that this process might engage a calcium-dependent mechanism in mammalian eggs [4, 8–10]. Indeed, artificial elevation of intracellular calcium concentration is known to initiate parthenogenetic activation of eggs in various species. However, spontaneous activation may also utilize calcium-independent mechanisms. In aging unfertilized sea urchin eggs, apoptosis was shown to be triggered by progressive inactivation of MAPK [11, 12]. Also, it was demonstrated that aged mouse and pig eggs have decreased activities of major CSF and MPF components [13, 14]. The gradual decrease in the content and/or activity of the key meiotic regulators below a threshold level necessary to maintain the meiotic metaphase arrest was hypothesized to cause meiotic exit in the absence of intracellular calcium signal [5].

At present, physiological inducers of spontaneous egg activation remain unidentified. It was suggested that oxidative stress might act as the initiator for a cascade of events that lead to expedited aging and deterioration of postovulatory oocytes [15]. Using Xenopus eggs, it was demonstrated previously that hydrogen peroxide initiates tyrosine phosphorylation and elevates intracellular calcium, resulting in Src kinase-dependent egg activation [16]. The study also reported that prolonged treatment with hydrogen peroxide led to excessive cortical contraction, egg swelling, and overactivation with a very distinctive egg phenotype. Studies of overactivated eggs are important because they can expand our understanding of cell death by unveiling alternative physiological mechanisms. So far, the intracellular events that occur in eggs upon overactivation have not been investigated in detail. Considering that hydrogen peroxide can easily diffuse through the cell plasma and subcellular compartment membranes to directly inflict oxidative damage [17], it could be expected that the drug might interfere with various intracellular processes and severely damage egg homeostasis.

In this work, we investigated oxidative stress-induced overactivation of Xenopus eggs using light and fluorescent microscopy, histochemical staining, protein, and bioluminescent assays. It was found that lipofuscin accumulation, decrease of soluble cytoplasmic protein content, and depletion of intracellular ATP occur in the eggs overactivated by strong oxidative stress. To the best of our knowledge, this is the first report concerning the changes of cellular homeostasis in overactivated eggs.