Maturation promoting factor destabilization mediates human chorionic gonadotropin induced meiotic resumption in rat oocytes

Chaube

2017

Dev Growth Differ

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Nitric oxides (NO) act as one of the major signal molecules and modulate various cell functions including oocyte meiosis in mammals. The present study was designed to investigate the mechanism of NO action during spontaneous meiotic exit from diplotene arrest (EDA) in rat cumulus oocytes complexes (COCs) cultured in vitro. Diplotene-arrested COCs collected from ovary of immature female rats after 20 IU pregnant mare's serum gonadotropins (PMSG) for 48 h were exposed to various concentrations of NO donor, S-nitroso-N-acetyl penicillamine (SNAP) and inducible nitric oxide synthase (iNOS) inhibitor, aminoguanidine (AG) for 3 h in vitro and downstream factors were analyzed. Our results suggest that SNAP inhibited, while AG induced EDA in a concentration-dependent manner. The iNOS-mediated total NO, cyclic nucleotides and cell division cycle 25B (Cdc25B) levels were reduced significantly. The decreased Cdc25B was associated with the increased Thr14/Tyr15 phosphorylated cyclin-dependent kinase 1 (Cdk1) level and decreased Thr161 phosphorylated Cdk1 as well as cyclin B1 levels leading to maturation promoting factor (MPF) destabilization. The destabilized MPF finally induced spontaneous EDA. Taken together, these results suggest that reduction of iNOS-mediated NO level destabilizes MPF during spontaneous EDA in rat COCs cultured in vitro.

Section: Discussionmentioning

confidence: 99%

Reduction of nitric oxide level results in maturation promoting factor destabilization during spontaneous meiotic exit from diplotene arrest in rat cumulus oocytes complexes cultured in vitro

Chaube

2017

Dev Growth Differ

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“…Immunofluorescence of iNOS, cAMP, Cdc25B, CaMKII, Wee1, specific as well as total phosphorylation of Cdk1, and cyclin B1 expressions were analyzed in oocytes using their highly specific antibodies purchased from Santa Cruz Biotechnology (Dallas, TX, USA) as per our published protocol with some modifications [38]. In brief, 12-14 oocytes from each group were fixed with 4% buffered formaldehyde (10 min) at room temperature.…”

Section: Detection Of Inos Camp Cdc25b Camkii Wee1 Specific As Wmentioning

confidence: 99%

Spontaneous Exit from Diplotene Arrest Associates with the Increase of Calcium and Reactive Oxygen Species in Rat Oocytes Cultured In Vitro

Chaube

2017

ROS

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Several signal molecules modulate the physiology of mammalian oocytes. Changes in the level of various signal molecules and their downstream impact on maturation promoting factor (MPF) during the achievement of meiotic competency remain ill understood. Therefore, the present study aimed to analyze levels of various signal molecules and MPF during the achievement of meiotic competency in rat oocytes. For this purpose, cumulus oocyte complexes (COCs) were collected from animals treated with 20 IU pregnant mare's serum gonadotropin (PMSG) for 48 h, followed by 20 IU human chorionic gonadotropin (hCG) for 14 h. The morphological changes, meiotic status and oocyte competency, cytosolic free calcium (Ca 2+), total reactive oxygen species (ROS), inducible nitric oxide synthase (iNOS), adenosine 3′,5′-cyclic monophosphate (cAMP), guanosine 3′,5′-cyclic monophosphate (cGMP), cell division cycle 25B (Cdc25B), calciumcalmodulin-dependent protein kinase II (CaMKII), Wee1, specific phosphorylation status of cyclin-dependent kinase 1 (Cdk1), and cyclin B1 expression levels were analyzed. Our data suggest that the culture of denuded diplotene-arrested oocytes resulted in spontaneous exit from diplotene arrest (EDA) in a time-dependent manner. The supplementation of hCG (2 IU) did not show any additive effect over spontaneous EDA in vitro. The hCG (20 IU) surge induced achievement of meiotic competency as most of the ovulated oocytes showed metaphase-II (M-II) arrest with extrusion of first polar body (PB-I). The spontaneous EDA was associated with a moderate increase of cytosolic free Ca 2+ as well as total ROS levels. Furthermore, iNOS, cAMP, cGMP, Cdc25B, Thr161 phosphorylated Cdk1, as well as cyclin B1 levels were significantly decreased, while CaMKII, Wee1, and Thr14/Tyr15 phosphorylated Cdk1 levels were increased significantly during spontaneous EDA. These changes were reversed in ovulated oocytes that showed M-II arrest. The above results suggest that the oscillation of signal molecules modulates MPF destabilization during the achievement of meiotic competency in rat oocytes cultured in vitro.

“…Diplotene arrested oocytes are morphologically identified by the presence of a germinal vesicle (GV) and nucleolus within the ooplasm (Pandey, Tripathi, Premkumar, Shrivastav, & Chaube, ; Tripathi, Kumar, & Chaube, ; Wassarman & Albertini, ).The diplotene arrest may last for several months to years depending on mammalian species (Tripathi et al, ). In follicular oocytes, meiotic resumption from diplotene stage is initiated by pituitary gonadotrophin surge (Chaube, ; Hao et al, ; Tiwari & Chaube, ). Removal of oocytes from follicular microenvironment and their culture in appropriate culture medium induce meiotic resumption from diplotene arrest in vitro (Hwang & Cheon, ; Tiwari & Chaube, ; Tiwari et al, ).…”

Section: Introductionmentioning

confidence: 99%

“…Molecular interactions through several pathways result in sub cellular as well as cellular changes during meiotic cell cycle progression from M‐I to M‐II stage in mammalian species. The cytostatic factors (CSFs) regulate maturation promoting factor (MPF) activity and modulate oocyte meiosis in rats (Tiwari & Chaube, ; Tiwari et al, ). MPF activity is high at M‐I stage and drops during meiotic exit from M‐I but increases at M‐II arrest in mouse oocytes (Terret et al, ).…”

Section: Introductionmentioning

confidence: 99%

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Journey of oocyte from metaphase‐I to metaphase‐II stage in mammals

Sharma

Journal Cellular Physiology

Gupta

et al. 2018

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In mammals, journey from metaphase-I (M-I) to metaphase-II (M-II) is important since oocyte extrude first polar body (PB-I) and gets converted into haploid gamete. The molecular and cellular changes associated with meiotic cell cycle progression from M-I to M-II stage and extrusion of PB-I remain ill understood. Several factors drive oocyte meiosis from M-I to M-II stage. The mitogen-activated protein kinase3/1 (MAPK3/1), signal molecules and Rho family GTPases act through various pathways to drive cell cycle progression from M-I to M-II stage. The down regulation of MOS/MEK/MAPK3/1 pathway results in the activation of anaphase-promoting complex/cyclosome (APC/C). The active APC/C destabilizes maturation promoting factor (MPF) and induces meiotic resumption. Several signal molecules such as, c-Jun N-terminal kinase (JNK2), SENP3, mitotic kinesin-like protein 2 (MKlp2), regulator of G-protein signaling (RGS2), Epsin2, polo-like kinase 1 (Plk1) are directly or indirectly involved in chromosomal segregation. Rho family GTPase is another enzyme that along with cell division cycle (Cdc42) to form actomyosin contractile ring required for chromosomal segregation. In the presence of origin recognition complex (ORC4), eccentrically localized haploid set of chromosomes trigger cortex differentiation and determine the division site for polar body formation. The actomyosin contractile activity at the site of division plane helps to form cytokinetic furrow that results in the formation and extrusion of PB-I. Indeed, oocyte journey from M-I to M-II stage is coordinated by several factors and pathways that enable oocyte to extrude PB-I. Quality of oocyte directly impact fertilization rate, early embryonic development, and reproductive outcome in mammals.