16 research outputs found
Dual Effects of Hydrogen Sulfide Donor on Meiosis and Cumulus Expansion of Porcine Cumulus-Oocyte Complexes
<div><p>Hydrogen sulfide (H<sub>2</sub>S) has been revealed to be a signal molecule with second messenger action in the somatic cells of many tissues, including the reproductive tract. The aim of this study was to address how exogenous H<sub>2</sub>S acts on the meiotic maturation of porcine oocytes, including key maturation factors such as MPF and MAPK, and cumulus expansion intensity of cumulus-oocyte complexes. We observed that the H<sub>2</sub>S donor, Na<sub>2</sub>S, accelerated oocyte <i>in vitro</i> maturation in a dose-dependent manner, following an increase of MPF activity around germinal vesicle breakdown. Concurrently, the H<sub>2</sub>S donor affected cumulus expansion, monitored by hyaluronic acid production. Our results suggest that the H<sub>2</sub>S donor influences oocyte maturation and thus also participates in the regulation of cumulus expansion. The exogenous H<sub>2</sub>S donor apparently affects key signal pathways of oocyte maturation and cumulus expansion, resulting in faster oocyte maturation with little need of cumulus expansion.</p></div
Effect of Na<sub>2</sub>S on meiosis resumption and transition to meiosis II in DOs.
<p>Proportion of GVBD (A) and meiosis I to II transition (B) during <i>in vitro</i> cultivation after 20 and 30 h <i>in vitro</i> cultivation, respectively. H<sub>2</sub>S: 300 µM Na<sub>2</sub>S. <sup>a,b,c</sup>Statistically significant differences among experimental groups (P<0.05).</p
Effect of Na<sub>2</sub>S on meiotic resumption and transition to meiosis II during oocyte cultivation.
<p>Proportion of GVBD (A) and meiosis I to II transition (B) in oocytes during <i>in vitro</i> cultivation over 2 h time scale. H<sub>2</sub>S: 300 µM Na<sub>2</sub>S. *Statistically significant differences between control and H<sub>2</sub>S groups (P<0.05).</p
Effect of different Na<sub>2</sub>S concentrations on meiosis resumption and transition to meiosis II in oocytes.
<p>Proportion of GVBD (A) and meiosis I to II transition (B) during <i>in vitro</i> cultivation after 20 and 30 h <i>in vitro</i> cultivation, respectively. <sup>a,b,c</sup>Statistically significant differences among experimental groups (P<0.05).</p
Effect of Na<sub>2</sub>S on HA content in expanded cumulus.
<p>(A) Total and retained HA content in COCs cultivated with 150–900 µM Na<sub>2</sub>S for 48 hs, total HA is related to the control group. (B) Total and retained HA content in COCs during <i>in vitro</i> cultivation with 300 µM Na<sub>2</sub>S over 12 h time scale, total HA is related to the control group after 48 h cultivation. (C) Total and retained HA content in COCs and OOXs cultivated with or without H<sub>2</sub>S donor, total HA is related to the control group of COCs. H<sub>2</sub>S: 300 µM Na<sub>2</sub>S. <sup>a,b,c</sup>Statistically significant differences among experimental groups in total HA, <sup>1,2</sup>statistically significant differences among experimental groups in retained HA, *statistically significant differences in total HA between control and H<sub>2</sub>S groups (P<0.05).</p
Effect of Na<sub>2</sub>S on partenogenetic development of porcine oocytes.
<p>Oocytes were matured with or without Na<sub>2</sub>S and partenogenetically activated using calcium ionophore. Pronucleus formation after 24 h zygote culture, cleavage rate after 2 days and blastocyst achievement after 7 days presumptive embryos culture were evaluated (%±SE).</p><p>H<sub>2</sub>S: 300 µM Na<sub>2</sub>S during oocyte maturation.</p><p>*Statistically significant differences between control and H<sub>2</sub>S group – in column (P<0.05).</p
Effect of Na<sub>2</sub>S on MPF and MAPK activities during oocyte cultivation.
<p>Representative autoradiograms and signal quantifications of phosphorylated histone H1 (A) and MBP (B) reflecting MPF and MAPK activity, respectively. Kinase activity was measured in oocytes cultivated with or without Na<sub>2</sub>S over 2 h time scale. The kinase activity was related to oocytes cultivated for 24 hs. C: control; H<sub>2</sub>S: 300 µM Na<sub>2</sub>S. *Statistically significant differences between control and H<sub>2</sub>S groups (P<0.05).</p
Hydrogen Sulfide Donor Protects Porcine Oocytes against Aging and Improves the Developmental Potential of Aged Porcine Oocytes
<div><p>Porcine oocytes that have matured in in vitro conditions undergo the process of aging during prolonged cultivation, which is manifested by spontaneous parthenogenetic activation, lysis or fragmentation of aged oocytes. This study focused on the role of hydrogen sulfide (H<sub>2</sub>S) in the process of porcine oocyte aging. H<sub>2</sub>S is a gaseous signaling molecule and is produced endogenously by the enzymes cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (MPST). We demonstrated that H<sub>2</sub>S-producing enzymes are active in porcine oocytes and that a statistically significant decline in endogenous H<sub>2</sub>S production occurs during the first day of aging. Inhibition of these enzymes accelerates signs of aging in oocytes and significantly increases the ratio of fragmented oocytes. The presence of exogenous H<sub>2</sub>S from a donor (Na<sub>2</sub>S.9H<sub>2</sub>O) significantly suppressed the manifestations of aging, reversed the effects of inhibitors and resulted in the complete suppression of oocyte fragmentation. Cultivation of aging oocytes in the presence of H<sub>2</sub>S donor positively affected their subsequent embryonic development following parthenogenetic activation. Although no unambiguous effects of exogenous H<sub>2</sub>S on MPF and MAPK activities were detected and the intracellular mechanism underlying H<sub>2</sub>S activity remains unclear, our study clearly demonstrates the role of H<sub>2</sub>S in the regulation of porcine oocyte aging.</p></div
Parthenogenetic activation of oocytes aged under the effect of the H<sub>2</sub>S donor.
<p>At Oocytes were cultivated 48 hours to the metaphase II and then divided into 4 groups (see table). Control group (MII) was parthenogenetically activated immediately (without any exposure to prolonged cultivation). Other groups were exposed to prolonged cultivation (aging) for 24 hours in modified M199 medium supplemented with a H2S donor (Na<sub>2</sub>S.9H<sub>2</sub>O; 0μM, 150μM, and 300μM) and then parthenogenetically activated with calcium ionophore (25μM, 5 min) combined with 6-dimethyl aminopurine (2mM, 2 h). Subsequently, oocytes were cultured in NCSU 23 medium for the following 24 hours.</p><p><sup>a,b,c</sup> Statistically significant differences in the ratio of activated oocytes between individual treatments (in columns) are indicated with different superscripts (P<0.05).</p><p>Parthenogenetic activation of oocytes aged under the effect of the H<sub>2</sub>S donor.</p
Effect of H<sub>2</sub>S donor on MPF and MAPK activity. 6A
<p>Histone H1 kinase assay was carried out to determine the activity of MPF by measurement of MPF capacity to phosphorylate its substrate (histone H1). <b>6B</b>: MBP kinase assay was carried out to determine the activity of MAPK by measurement of MAPK capacity to phosphorylate its substrate (MBP – Myelin basic protein). MPF and MAPK activities were determined in the MII oocytes (C – control, white column), the oocytes aged 12h and 24h in modified M199 medium, the oocytes aged 12h and 24h in modified M199 medium supplemented with a H<sub>2</sub>S donor (Na<sub>2</sub>S, black column), and the oocytes aged 12h and 24h in modified M199 medium supplemented with triple combination of inhibitors (3Ki, grey column). The results are presented as a ratio relative to the group of oocytes at metaphase II. <i>(GV – germinal vesicle stage; MII – oocytes at metaphase II; A12–12 hours of aging; A24–24 hours of aging; C – control, white column; Na<sub>2</sub>S—Na<sub>2</sub>S.9H<sub>2</sub>O, 300 μM, black column; 3Ki - 1mM oxamic acid + 1mM beta-kyano-L-alanine + 5mM alpha-ketoglutaric acid disodium salt dihydrate). <sup>a,b,</sup> Statistically significant differences in activity (MPF or MAPK) between individual treatments at the same time are indicated with different superscripts (P<0.05)</i>.</p