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. 2020 Dec 8;21(24):9342. doi: 10.3390/ijms21249342

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© 2020 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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78-kDa glucose-regulated protein (GRP78) mRNA and protein expression before and after oocyte in vitro maturation, as well as its immunolocalisation in cumulus–oocyte complexes (COCs). COCs (50/group/experiment) were selected after morphological examination of material collected from the ovaries taken from 100 pigs (15 follicles/ovary). COCs were collected before (0 h) and after (44 h) in vitro maturation and oocytes were denuded by gently pipetting in hyaluronidase, then real-time PCR and Western blot analysis were performed to determine mRNA (A) and protein (B) level of GRP78 in oocytes and cumulus cells. Immunolocalisation of GRP78 was studied by immunofluorescence in COCs (C). Gene expression level was normalised to the geometric mean of actin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and protein to actin. GRP78 immunostaining shown by arrows. Experiments were performed independently and repeated three times (n = 3). The data are plotted as the mean ± SEM of three independent experiments. Significance between groups before and after maturation is indicated by ** p < 0.01 and *** p < 0.001; Cumulus cells (Cc), oocyte (Oo), Immunoglobulin G (IgG).

78-kDa glucose-regulated protein (GRP78) mRNA and protein expression before and after oocyte in vitro maturation, as well as its immunolocalisation in cumulus–oocyte complexes (COCs). COCs (50/group/experiment) were selected after morphological examination of material collected from the ovaries taken from 100 pigs (15 follicles/ovary). COCs were collected before (0 h) and after (44 h) in vitro maturation and oocytes were denuded by gently pipetting in hyaluronidase, then real-time PCR and Western blot analysis were performed to determine mRNA (A) and protein (B) level of GRP78 in oocytes and cumulus cells. Immunolocalisation of GRP78 was studied by immunofluorescence in COCs (C). Gene expression level was normalised to the geometric mean of actin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and protein to actin. GRP78 immunostaining shown by arrows. Experiments were performed independently and repeated three times (n = 3). The data are plotted as the mean ± SEM of three independent experiments. Significance between groups before and after maturation is indicated by ** p < 0.01 and *** p < 0.001; Cumulus cells (Cc), oocyte (Oo), Immunoglobulin G (IgG).