This study aimed to describe glutathione peroxidase 4 (GPx4) in rat oocytes, preimplantation embryos, and female genital organs. After copulation, Sprague Dawley female rats were euthanized with anesthetic on the first (D1), third (D3), and fifth days of pregnancy (D5). Ovaries, oviducts, and uterine horns were removed, and oocytes and preimplantation embryos were obtained. Immunohistochemical, immunofluorescent, and Western blot methods were employed. Using immunofluorescence, we detected GPx4 in both the oocytes and preimplantation embryos. Whereas in the oocytes, GPx4 was homogeneously diffused, in the blastomeres, granules were formed, and in the blastocysts, even clusters were present mainly around the cell nuclei. Employing immunohistochemistry, we detected GPx4 inside the ovary in the corpus luteum, stroma, follicles, and blood vessels. In the oviduct, the enzyme was present in the epithelium, stroma, blood vessels, and smooth muscles. In the uterus, GPx4 was found in the endometrium, myometrium, blood vessels, and stroma. Moreover, we observed GPx4 positive granules in the uterine gland epithelium on D1 and D3 and cytoplasm of fibroblasts forming in the decidua on D5. Western blot showed the highest GPx4 levels in the uterus and the lowest levels in the ovary. Our results show that the GPx4 is necessary as early as in the preimplantation development of a new individual because we detected it in an unfertilized oocyte in a blastocyst and not only after implantation, as was previously thought.

Breast cancer is the most common cancer in females. The aim of this study was to determine the effect of paclitaxel (PTX) and doxorubicin (DOX) therapy on the βIII-tubulin, carbonic anhydrase IX (CA IX), and survivin expression in chemically-induced rat mammary tumors. Animals with induced mammary carcinogenesis were randomly divided into treatment groups and an untreated group. The total proportion of tumors, the proportion of carcinoma in situ (CIS), and invasive carcinoma (IC) were evaluated. Protein expression in tumor tissue was determined using IHC. Statistical analysis of the data, evaluated by Fisher-exact test and unpaired t-test. Significantly increased levels of proteins in the tumor cells were confirmed using the IHC method for all studied proteins. The expression of βIII-tubulin, CA IX, and survivin increased significantly after treatment with both cytostatics (PTX and DOX). Depending on the type of tumor, a significant increase in all proteins was observed in IC samples after PTX treatment, and CA IX expression after DOX treatment. In CIS samples, a significant increase of βIII-tubulin and survivin expression was observed after a DOX treatment. The results suggest that βIII-tubulin, survivin, and CA IX may be significant drug resistance markers and the clinical regulation of their activity may be an effective means of reversing this resistance.

This study aimed to detect the presence of glutathione peroxidase 8 (GPx8) in rat during preimplantation period of pregnancy. Females were killed on first (D1), third (D3), and fifth (D5) day of pregnancy. The presence of GPx8 in embryos was detected under the confocal microscope, the presence of GPx8 in genital organs was confirmed immunohistochemically, and the amount of GPx8 was determined using densitometry. We found that GPx8 is dispersed in the cytoplasm of oocytes, while after fertilization, it is concentrated in granules. From 4-cell stage till blastocyst, GPx8 reaction was found in the perinuclear region. In the ovary, GPx8 was seen in granulosa-lutein cells, in plasma of blood vessels, and inside Graafian follicles. In oviduct, GPx8 was detected in the plasma and in the extracellular matrix (ECM). Moreover, epithelial cells of isthmus were positive. In uterus, GPx8 was observed in the uterine glands, in the plasma, and in ECM. On D5, the enzyme disappeared from the uterine glands and appeared in fibroblasts. Densitometry revealed that the highest amount of GPx8 was on D1 and subsequently declined. To our knowledge, this is the first paper describing GPx8 presence in the oocytes, preimplantation embryos, and female genital organs in mammals. Our results improve the understanding of antioxidant enzymes presence during pregnancy in defense against oxidative stress, which is considered to be one of the main causes of infertility.

Objectives: This study describes the presence of glutathione peroxidase 3 (GPx3) in mice oocytes, preimplantation embryos (O/PEs), and female genital organs during the preimplantation period of pregnancy. Background: Glutathione peroxidase 3, the antioxidant enzyme, is important in protecting cells against the oxidative stress. Methods: O/PEs were isolated from genital organs of superovulated females for the immunofluorescence analysis and these organs were used for the immunohistochemistry to detect GPx3. Results: GPx3 was present in all O/PEs, where it formed clusters diffusely in the cytoplasm, or under the cytoplasmic membrane. Moreover, GPx3 was likely indicated in nucleoli. GPx3 was found in the cytoplasm of granulosa-lutein cells as well as in follicular cells. The oocyte cytoplasm inside the ovary was negative. Oviductal epithelial cells before and after fertilization had different positivity patterns. Uterine luminal epithelial cells were negative before fertilization. After fertilization, a few positive luminal epithelial cells and a few cells of uterine glands had positive cytoplasm. Conclusion: The differences of the GPx3 signal intensity in nucleoli of the zygote and the 2-cell embryo, and the different localization of GPx3 in the cytoplasm of 2-cell and 4-cell embryos may be the signs of the maternal-zygotic genome transition. Moreover, GPx3 may be significant during the preimplantation period of pregnancy in O/PEs and female genital organs, where it could protect cells from oxidative stress during the hormone and protein synthesis, and during the cell metabolism. The presence of cytoplasmic granules in epithelial cells of fallopian tube may indicate their secretion into the lumen of the organ to protect gametes and preimplantation embryos passing through. To our knowledge, this is the first study investigating GPx3 occurrence in mammalian preimplantation embryos.