Effects of stimulators of protein kinases A and C and modulators of phosphorylation on plasminogen activator activity in porcine oocyte-cumulus cell complexes during in vitro maturation

1995 ◽  
Vol 40 (3) ◽  
pp. 364-370 ◽  
Author(s):  
Nam Hyung Kim ◽  
Alfred R. Menino
Keyword(s):  
Plasminogen Activator ◽  
Protein Kinases ◽  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Cell Complexes ◽  
Plasminogen Activator Activity ◽  
Porcine Oocyte

Plasminogen activator activity in cortical granules of bovine oocytes during in vitro maturation

2002 ◽  
Vol 57 (7) ◽  
pp. 1897-1905 ◽  
Author(s):  
Constantinos A. Rekkas ◽  
Urban Besenfelder ◽  
Vitezslav Havlicek ◽  
Emmanuel Vainas ◽  
Gottfried Brem
Keyword(s):  
Plasminogen Activator ◽  
In Vitro Maturation ◽  
Cortical Granules ◽  
Plasminogen Activator Activity ◽  
Bovine Oocytes

scholarly journals Exposure of Triclosan in Porcine Oocyte Leads to Superoxide Production and Mitochondrial-Mediated Apoptosis during In Vitro Maturation

2020 ◽  
Vol 21 (9) ◽  
pp. 3050 ◽  
Author(s):  
Hyo-Jin Park ◽  
Bong-Seok Song ◽  
Jin-Woo Kim ◽  
Seul-Gi Yang ◽  
Sun-Uk Kim ◽  
...  
Keyword(s):  
Antioxidant Enzymes ◽  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Superoxide Production ◽  
Mrna Levels ◽  
Female Reproduction ◽  
Mitochondrial Superoxide ◽  
Porcine Oocyte

While triclosan (TCS) exerts detrimental effects on female reproduction, the effect of TCS-derived toxins on porcine oocytes during in vitro maturation (IVM) is unclear. This study investigated the effects of TCS on mitochondrion-derived reactive oxygen species (ROS) production and apoptosis pathways during porcine oocyte maturation. Porcine oocytes were treated with TCS (1, 10, and 100 μM) and triphenylphosphonium chloride (Mito-TEMPO; 0.1 μM), and matured cumulus oocyte complexes (COCs) were stained with orcein, dichlorofluorescein diacetate (DCF-DA), and Mito-SOX. Proteins and mRNA levels of factors related to cumulus expansion and mitochondrion-mediated apoptosis and antioxidant enzymes were analyzed by western blotting and reverse-transcription polymerase chain reaction (RT-PCR), respectively. Meiotic maturation and cumulus cell expansion significantly decreased for COCs after TCS treatment along with an increase in mitochondrial superoxide levels at 44 h of IVM. Further, mitochondrion-related antioxidant enzymes and apoptosis markers were significantly elevated in porcine COCs following TCS-mediated oxidative damage. The protective effect of Mito-TEMPO as a specific superoxide scavenger from TCS toxin improved the maturation capacity of porcine COCs. Mito-TEMPO downregulated the mitochondrial apoptosis of TCS-exposed porcine COCs by reducing superoxide level. In conclusion, our data demonstrate that TCS mediates toxicity during porcine oocyte maturation through superoxide production and mitochondrion-mediated apoptosis.

265 HYALURONAN SYNTHESIS ABILITY OF PORCINE CUMULUS - OOCYTE COMPLEXES DERIVED FROM SMALL FOLLICLES

2013 ◽  
Vol 25 (1) ◽  
pp. 280
Author(s):  
M. Nakakoji ◽  
H. Funahashi
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Cumulus Expansion ◽  
Elisa Kit ◽  
Porcine Oocyte ◽  
Post Hoc ◽  
Metaphase Ii Oocytes

The degree of cumulus expansion, an important step in oocyte maturation, of porcine cumulus–oocyte complexes (COC) derived from small follicles (SF: 1 to 2 mm in diameter) is known to be lower than those derived from middle follicles (MF: 3 to 6 mm in diameter). The objective of this study was to compare the abilities of hyaluronan (HA) synthesis of COC from SF and MF. Furthermore, the effect of oestradiol during pre-incubation of COC on proliferation of the cumulus cells was examined. Cumulus–oocyte complexes from SF and MF of porcine ovaries were cultured for in vitro maturation [IVM, in modified porcine oocyte medium (Yoshioka et al. 2008 J. Reprod. Dev. 54, 208–213) supplemented with 50 µM β-mercaptoethanol, 10 IU mL–1 of eCG, 10 IU mL–1 of hCG, and 1 mM dbcAMP for 20 h and then in the fresh medium without those supplements for another 24 h]. Hyaluronan production was quantified at 20 h after the start of IVM with a commercial HA-ELISA kit (20 COC/tube × 4 times). The number of cumulus cells was assessed 0 and 20 h after the start of IVM (50 COC × 4 times). Furthermore, proliferation of cumulus cells was examined after pre-culture of COC (n = 40 COC × 5 times) in modified porcine oocyte medium with various concentrations of oestradiol (0, 0.1, 1, and 10 ng mL–1) for 6 h. Statistical analyses of results from 4 to 5 replicated trials were performed by ANOVA with a Bonferroni-Dunn post-hoc test (significance, P < 0.05). The degree of cumulus expansion of COC from MF (n = 152) was higher than that of COC from SF (n = 156). The incidence of metaphase-II oocytes was significantly lower in COC from SF (n = 133; 48.9%) than in COC from MF (n = 148; 74.7%). The HA content of COC was higher in those from MF (20.8 µg/COC) than in those from SF (10.8 µg/COC), whereas the content per cumulus cell was not different because the numbers of cumulus cells at 0 and 20 h were also higher in COC (n = 200 in each group) from MF (3.0 × 103 and 3.3 × 103 cells, respectively) than from SF (2.0 × 103 and 2.5 × 103 cells, respectively). Cumulus cells proliferated significantly in the presence of oestradiol, regardless of the concentration, during pre-incubation for 6 h (2.5 to 2.8 × 103 cells), as compared with the oestradiol-free controls (2.2 × 103 cells). These results demonstrate that the different abilities of cumulus expansion between COC (n = 200 in each group) from SF and MF may be due to the number of cumulus cells per COC. Pre-incubation in the presence of oestradiol stimulates the proliferation of cumulus cells and may improve the oocyte maturation of COC derived from SF.

In vitro maturation of blue fox oocytes and cAMP production in oocyte-cumulus cell complexes

1993 ◽  
Vol 39 (1) ◽  
pp. 250 ◽  
Author(s):  
A. Krogenæs ◽  
E. Nagyová ◽  
W. Farstad ◽  
A.L. Hafne
Keyword(s):  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Camp Production ◽  
Cell Complexes ◽  
Blue Fox

298 CYCLIC AMP AND CYCLIC GMP CONTENTS IN PORCINE OOCYTE–CUMULUS COMPLEXES, DENUDED OOCYTES, AND CUMULUS CELLS DERIVED FROM SMALL AND MIDDLE FOLLICLES DURING IN VITRO MATURATION

2015 ◽  
Vol 27 (1) ◽  
pp. 238
Author(s):  
Y. Okudaira ◽  
H. Funahashi
Keyword(s):  
Cyclic Amp ◽  
In Vitro Maturation ◽  
Critical Role ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Intracellular Camp ◽  
Camp Content ◽  
Porcine Oocyte ◽  
Camp And Cgmp

Drastic changes in intracellular cAMP and cGMP levels play a critical role in the regulation of meiotic resumption. The objective of this study was to compare cAMP and cGMP contents in cumulus-oocyte complexes (COC), denuded oocytes (DO), and cumulus cell masses (CC) derived from small (SF) and middle follicles (MF) during in vitro maturation (IVM). The COC were aspirated from SF (1–3 mm in diameter) or MF (3–6 mm in diameter) of prepubertal gilt ovaries. The COC were cultured in modified porcine oocyte medium (mPOM) with eCG, hCG, and dibutyryl cAMP for 20 h and then in fresh mPOM without those supplements for 4 h in an atmosphere of 5% CO2 in air at 39°C. At 0, 10, 20, and 24 h of IVM, COC, DO, and CC were collected. The DO were prepared by removal of cumulus cells and zona pellucida. The CC were prepared by puncturing ooplasm by using 18-gauge needle. Intracellular contents of cAMP and cGMP were determined by direct enzyme immunoassay kits. Statistical analyses of 3 to 7 replicated data were performed by ANOVA. There were no significant differences in contents of cAMP and cGMP between DO from SF and MF in all observation points (P > 0.05). Cyclic AMP contents in COC and CC derived from MF were higher than those from SF at 20 h of IVM (MF 33.0 ± 0.5 fmol/COC v. SF 28.4 ± 1.0 fmol/COC, MF 20.9 ± 0.9 fmol/CC v. SF 14.6 ± 0.8 fmol/CC; P < 0.05), whereas there were no significant differences between origins of those (SF v. MF, P > 0.05) at 0, 10, and 24 h of IVM. Furthermore, although cAMP content in CC from MF was not significantly different between 10 and 20 h of IVM (25.4 ± 1.7 and 20.9 ± 0.9 fmol/CC, respectively; P > 0.05), the content in CC from SF significantly decreased between 10 and 20 h (23.1 ± 1.2 , and 14.6 ± 0.8 fmol/CC, respectively; P < 0.05). At 0 and 10 h of IVM, cGMP contents in COC and CC from MF were significantly higher than those from SF (0 h: 81.8 ± 4.5 fmol/COC from MF v. 41.7 ± 10.6 fmol/COC from SF and 82.7 ± 7.5 fmol/CC from MF v. 10.7 ± 2.7 fmol/CC from SF; 10 h: 64.8 ± 8.4 fmol/COC from MF v. 24.8 ± 8.2 fmol/COC from SF, 49.3 ± 14.9 fmol/CC from MF v. 13.5 ± 4.8 fmol/CC from SF; P < 0.05). However, there were no significant differences in cGMP contents in COC and CC between the origins (MF v. SF) at 20 and 24 h of IVM (P > 0.05). From these results, we conclude that cAMP and cGMP contents in cumulus cells are significantly differences between the origins (MF v. SF) during IVM.

297 EFFECT OF CUMULUS CELL REMOVAL DURING IN VITRO MATURATION OF PORCINE CUMULUS–OOCYTE COMPLEXES ON THE APOPTOTIC STATUS AND MEIOTIC PROGRESSION OF THE OOCYTES

2015 ◽  
Vol 27 (1) ◽  
pp. 237
Author(s):  
P. Ferré ◽  
H. Funahashi
Keyword(s):  
In Vitro Maturation ◽  
Cumulus Cell ◽  
Annexin V ◽  
Cumulus Cells ◽  
Dapi Staining ◽  
Meiotic Progression ◽  
Viability Assay ◽  
Significant Difference ◽  
Porcine Oocyte

This study was undertaken to examine the apoptotic status and meiotic progression of oocytes from small (SF) and medium follicles (MF) when the oocytes were denuded from cumulus cells (CC) before, during and after culture for in vitro maturation (IVM). Cumulus-oocyte complexes (COC) were aspirated from SF (0.5–2 mm in diameter) or MF (3–6 mm in diameter) of slaughtered prepubertal gilt ovaries. Only COC with a good morphology of the surrounding cumulus cells were cultured for IVM in modified porcine oocyte medium supplemented with 50 µM β-mercaptoethanol, 1 mM dibutyryl c-AMP, 10 IU mL–1 of eCG, and 10 IU mL–1 of hCG for 20 h at 39°C and 5% CO2 in air and then continued culture in the absence of dibutyryl c-AMP, eCG, and hCG in the same medium for another 24 h. Before and 20 h after the start of IVM culture, some of the oocytes were denuded of CC and the oocytes continued the IVM culture. After IVM culture, oocyte viability and meiotic progression were examined by the annexin V/PI viability assay and DAPI staining. Statistical analyses of 5 replicate data were performed with a 2-way ANOVA and a Tukey's multiple comparisons test. Before IVM culture, there was no significant difference between the viability of SF and MF oocytes, but the incidence of oocytes at the GV0 stage was higher in specimens from SF than MF (24.8 v. 3.3%), and that of oocytes at the GVI stage was the opposite (57.8 in MF v. 22.7% in SF). After IVM culture, apoptotic status of oocytes was only affected by the decumulation timing. The percentage of normal live oocytes was significantly higher when CC were removed after 20 and 44 h of IVM in both SF (39.7 and 39.3 v. 17.7%) and MF (45.4 and 37 v. 22.2%). The incidence of early and late apoptotic oocytes was significantly higher when the CC were removed before IVM culture in both SF (74.3 and 7.4%) and MF (69.4 and 6.7%). The incidence of mature live oocytes was significantly affected by both the origin of COC and the decumulation timing. Although the percentage of mature oocytes was higher in MF, maturation rates were significantly higher when oocytes were denuded at 20 h of IVM culture (SF 65.4%, MF 83.1%) as compared at 0 (SF 27.9%, MF 32.3%) and 44 h (SF 41%, MF 68.5%). However, the percentage of oocytes with normal spindle morphology was significantly higher when oocytes were denuded at 44 h of IVM culture (SF 70.6%, MF 91.5%) than 20 h (SF 66.8%, MF 73%). In summary, regardless of COC from SF and MF, removal of CC at 20 h of IVM culture seems to promote meiotic progression of the oocytes to the MII stage, but factor(s) from or communication with CC during the latter half of IVM culture may be needed to obtain a normal spindle morphology in mature oocytes.

46 PLASMINOGEN ACTIVATOR ACTIVITY IN BUFFALO IN VITRO MATURED OOCYTES AFTER VITRIFICATION-WARMING

2012 ◽  
Vol 24 (1) ◽  
pp. 135
Author(s):  
M. Tsantarliotou ◽  
M. De Blasi ◽  
S. Lavrentiadou ◽  
V. Sapanidou ◽  
L. Boccia ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Proteolytic Enzymes ◽  
Sucrose Solution ◽  
Cumulus Cell ◽  
Nonparametric Test ◽  
Oocyte Cryopreservation ◽  
Control Group ◽  
Blastocyst Development ◽  
Plasminogen Activator Activity

Plasminogen activators (PA) are proteolytic enzymes that convert plasminogen into plasmin. Plasmin is involved in physiological processes such as ovulation (Liu 2004 Front. Biosci. 9, 3356–3373), cumulus cell layer expansion (Liu et al. 1986 Endocrinology 119, 1578–1587), oocyte maturation (Dow et al. 2002 Biol. Reprod. 66, 1413–1421) and fertilization (Huarte et al. 1993 Dev. Biol. 157, 539–546). Although the interest is increasing, buffalo oocyte cryopreservation is still inefficient, especially in terms of blastocyst development after IVF. The aim of the present study was to evaluate whether exposure to cryoprotectants and the vitrification procedure affect plasminogen activator activity (PAA) in buffalo in vitro-matured oocytes. A total number of 300 cumulus-oocyte complexes over 5 replicates were selected and in vitro-matured. Cumulus-oocyte complexes were mechanically stripped of their cumulus cells by gentle pipetting, washed and divided into 3 groups (20 oocytes/group, over 5 replicates). The control group consisted of fresh in vitro-matured oocytes. In the vitrification group, denuded oocytes were first exposed to 10% ethylene glycol (EG) + 10% dimethyl sulfoxide (DMSO) for 3 min, then to 20% EG + 20% DMSO and 0.5 M sucrose, loaded on cryotops and plunged into liquid nitrogen within 25 s. Subsequently, oocytes were warmed in a 1.25 M sucrose solution for 1 min and then in decreasing concentrations of sucrose (0.625 M, 0.42 M and 0.31 M) for 30 s each. In order to test cryoprotectant effects, oocytes were simply exposed to the vitrification and warming solutions (toxicity group). Surviving oocytes were extracted by a fine needle, centrifuged at 4000 rpm for 10 min and the supernatant was mixed with the reaction solution: TRIS-HCl 0.1 M, homologous plasminogen, the chromogenic substrate for plasmin S-2251 and incubated at 37°C for 30 min. The PAA levels were measured by a spectrophotometer (405 nm) expressed as Abs/20 oocytes. The data were analysed by the Kruskal-Wallis nonparametric test. Low levels of PAA were detected in the denuded oocytes of the control, toxicity and vitrification groups. No significant differences in mean PAA values were observed among the 3 experimental groups (0.017 ± 0.001, 0.018 ± 0.002 and 0.017 ± 0.001 units, in the control, toxicity and vitrification groups, respectively). In conclusion, cryoprotectants and the vitrification procedure do not affect the proteolytic activity linked to plasmin in in vitro-matured buffalo oocytes. The results show that the vitrification/warming procedure does not exert an effect on in vitro-matured buffalo oocytes in terms of PAA generation, a parameter that plays an important role in fertilization and in vitro embryo development. Further studies are needed to identify factors affecting the efficiency of oocyte cryopreservation.

scholarly journals Effect of D-Glucuronic Acid and N-acetyl-D-Glucosamine Treatment during In Vitro Maturation on Embryonic Development after Parthenogenesis and Somatic Cell Nuclear Transfer in Pigs

Animals ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 1034
Author(s):  
Joohyeong Lee ◽  
Eunhye Kim ◽  
Seon-Ung Hwang ◽  
Lian Cai ◽  
Mirae Kim ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
In Vitro Maturation ◽  
Glucuronic Acid ◽  
Cumulus Cell ◽  
Cortical Granule ◽  
Oocyte Diameter

This study aimed to examine the effects of treatment with glucuronic acid (GA) and N-acetyl-D-glucosamine (AG), which are components of hyaluronic acid (HA), during porcine oocyte in vitro maturation (IVM). We measured the diameter of the oocyte, the thickness of the perivitelline space (PVS), the reactive oxygen species (ROS) level, and the expression of cumulus cell expansion and ROS-related genes and examined the cortical granule (CG) reaction of oocytes. The addition of 0.05 mM GA and 0.05 mM AG during the first 22 h of oocyte IVM significantly increased oocyte diameter and PVS size compared with the control (non-treatment). The addition of GA and AG reduced the intra-oocyte ROS content and improved the CG of the oocyte. GA and AG treatment increased the expression of CD44 and CX43 in cumulus cells and PRDX1 and TXN2 in oocytes. In both the chemically defined and the complex medium (Medium-199 + porcine follicular fluid), oocytes derived from the GA and AG treatments presented significantly higher blastocyst rates than the control after parthenogenesis (PA) and somatic cell nuclear transfer (SCNT). In conclusion, the addition of GA and AG during IVM in pig oocytes has beneficial effects on oocyte IVM and early embryonic development after PA and SCNT.

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