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      Effect of Melatonin on the In Vitro Maturation of Porcine Oocytes, Development of Parthenogenetically Activated Embryos, and Expression of Genes Related to the Oocyte Developmental Capability

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          Abstract

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          Exogenous melatonin has beneficial effects on improving cumulus oophorus expansion; mitochondrial distribution; intracellular level of glutathione; and first polar body extrusion rate of porcine oocytes derived from in vitro maturation. Moreover; melatonin supplementation increases relative abundances of BMP15 and CAT mRNA; and decreases intracellular levels of reactive oxygen species; and expression values of P53 and BAX genes; which are related to in vitro development of porcine oocytes.

          Abstract

          Melatonin treatment can improve quality and in vitro development of porcine oocytes, but the mechanism of improving quality and developmental competence is not fully understood. In this study, porcine cumulus–oocyte complexes were cultured in TCM199 medium with non-treated (control), 10 −5 M luzindole (melatonin receptor antagonist), 10 −5 M melatonin, and melatonin + luzindole during in vitro maturation, and parthenogenetically activated (PA) embryos were treated with nothing (control), or 10 −5 M melatonin. Cumulus oophorus expansion, oocyte survival rate, first polar body extrusion rate, mitochondrial distribution, and intracellular levels of reactive oxygen species (ROS) and glutathione of oocytes, and cleavage rate and blastocyst rate of the PA embryos were assessed. In addition, expression of growth differentiation factor 9 (GDF9), tumor protein p53 (P53), BCL2 associated X protein (BAX), catalase (CAT), and bone morphogenetic protein 15 (BMP15) were analyzed by real-time quantitative PCR. The results revealed that melatonin treatment not only improved the first polar body extrusion rate and cumulus expansion of oocytes via melatonin receptors, but also enhanced the rates of cleavage and blastocyst formation of PA embryos. Additionally, melatonin treatment significantly increased intraooplasmic level of glutathione independently of melatonin receptors. Furthermore, melatonin supplementation not only significantly enhanced mitochondrial distribution and relative abundances of BMP15 and CAT mRNA, but also decreased intracellular level of ROS and relative abundances of P53 and BAX mRNA of the oocytes. In conclusion, melatonin enhanced the quality and in vitro development of porcine oocytes, which may be related to antioxidant and anti-apoptotic mechanisms.

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          Most cited references52

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          Growth differentiation factor-9 is required during early ovarian folliculogenesis.

          Growth factors synthesized by ovarian somatic cells directly affect oocyte growth and function, but it is unclear whether oocyte-secreted factors play a reciprocal role in modulating somatic cell functions in vivo. During the functional analysis of members of the transforming growth factor-beta superfamily in mouse development, we have uncovered a new family member, growth differentiation factor-9 (GDF-9), which is required for ovarian folliculogenesis. GDF-9 messenger RNA is synthesized only in the oocyte from the primary one-layer follicle stage until after ovulation. Here we analyse ovaries from GDF-9-deficient female mice and demonstrate that primordial and primary one-layer follicles can be formed, but there is a block in follicular development beyond the primary one-layer follicle stage which leads to complete infertility. Oocyte growth and zona pellucida formation proceed normally, but other aspects of oocyte differentiation are compromised. Thus, GDF-9 is the first oocyte-derived growth factor required for somatic cell function in vivo.
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            Mitochondria and calcium: from cell signalling to cell death.

            M R Duchen (2000)
            While a pathway for Ca2+ accumulation into mitochondria has long been established, its functional significance is only now becoming clear in relation to cell physiology and pathophysiology. The observation that mitochondria take up Ca2+ during physiological Ca2+ signalling in a variety of cell types leads to four questions: (i) 'What is the impact of mitochondrial Ca2+ uptake on mitochondrial function?' (ii) 'What is the impact of mitochondrial Ca2+ uptake on Ca2+ signalling?' (iii) 'What are the consequences of impaired mitochondrial Ca2+ uptake for cell function?' and finally (iv) 'What are the consequences of pathological [Ca2+]c signalling for mitochondrial function?' These will be addressed in turn. Thus: (i) accumulation of Ca2+ into mitochondria regulates mitochondrial metabolism and causes a transient depolarisation of mitochondrial membrane potential. (ii) Mitochondria may act as a spatial Ca2+ buffer in many cells, regulating the local Ca2+ concentration in cellular microdomains. This process regulates processes dependent on local cytoplasmic Ca2+ concentration ([Ca2+]c), particularly the flux of Ca2+ through IP3-gated channels of the endoplasmic reticulum (ER) and the channels mediating capacitative Ca2+ influx through the plasma membrane. Consequently, mitochondrial Ca2+ uptake plays a substantial role in shaping [Ca2+]c signals in many cell types. (iii) Impaired mitochondrial Ca2+ uptake alters the spatiotemporal characteristics of cellular [Ca2+]c signalling and downregulates mitochondrial metabolism. (iv) Under pathological conditions of cellular [Ca2+]c overload, particularly in association with oxidative stress, mitochondrial Ca2+ uptake may trigger pathological states that lead to cell death. In the model of glutamate excitotoxicity, microdomains of [Ca2+]c are apparently central, as the pathway to cell death seems to require the local activation of neuronal nitric oxide synthase (nNOS), itself held by scaffolding proteins in close association with the NMDA receptor. Mitochondrial Ca2+ uptake in combination with NO production triggers the collapse of mitochondrial membrane potential, culminating in delayed cell death.
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              Antioxidative protection by melatonin: multiplicity of mechanisms from radical detoxification to radical avoidance.

              Melatonin has been shown to protect against oxidative stress in various, highly divergent experimental systems. There are many reasons for its remarkable protective potential. Signaling effects comprise the upregulation of antioxidant enzymes, such as superoxide dismutases, peroxidases, and enzymes of glutathione supply, down-regulation of prooxidant enzymes, such as nitric oxide synthases and lipoxygenases, and presumably also the control of quinone reductase 2. Other mechanisms are based on direct interactions with several reactive oxygen and nitrogen species. Among these reactions, the capacity of easily undergoing single-electron transfer reactions is of particular importance. Electron donation by melatonin is not only an aspect of direct radical scavenging, but additionally represents the basis for formation of the protective metabolites AFMK (N1-ace-tyl-N2-formyl-5-methoxykynuramine) and AMK (N1-acetyl-5-methoxykynuramine). Recent investigations on mitochondrial metabolism indicate that melatonin as well as AMK are capable of supporting the electron flux through the respiratory chain, of preventing the breakdown of the mitochondrial membrane potential, and of decreasing electron leakage, thereby reducing the formation of superoxide anions. Radical avoidance is a new line of investigation, which exceeds mitochondrial actions and also comprises antiexcitatory effects and contributions to the maintenance of internal circadian phase relationships.
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                Author and article information

                Journal
                Animals (Basel)
                Animals (Basel)
                animals
                Animals : an Open Access Journal from MDPI
                MDPI
                2076-2615
                27 January 2020
                February 2020
                : 10
                : 2
                : 209
                Affiliations
                [1 ]College of Life Sciences and Food Engineering, Hebei University of Engineering, Handan 056021, China; yangling@ 123456hebu.edu.cn (L.Y.); wangqingkai0814@ 123456yeah.net (Q.W.); zhaozm0320@ 123456126.com (Z.Z.)
                [2 ]Animal Husbandry and Veterinary Research Institute of Tianjin, Tianjin 300412, China; tjlqj@ 123456sina.com (Q.L.); mushq@ 123456163.com (S.M.)
                Author notes
                [* ]Correspondence: tjxmcui2014@ 123456126.com
                [†]

                These authors contributed equally to this work as co-first author.

                Author information
                https://orcid.org/0000-0003-4385-0024
                Article
                animals-10-00209
                10.3390/ani10020209
                7070577
                32012669
                cdb14e32-9c8e-4ab3-9515-899a11a2b593
                © 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/).

                History
                : 26 September 2019
                : 24 January 2020
                Categories
                Article

                melatonin,embryo,oocyte,pig
                melatonin, embryo, oocyte, pig

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