The regulation of ovarian granulosa cell death by pro- and anti-apoptotic molecules

In the mammalian ovary, follicular development and atresia are closely regulated by cell death and survival-promoting factors, including hormones (gonadotropins) and intraovarian regulators (gonadal steroids, cytokines, and intracellular proteins). Several hundred thousand primordial follicles are present in the mammalian ovary; however, only a limited number of primordial follicles develop to the preovulatory stage and ovulate. The others, more than 99% of follicles, will be eliminated via a degenerative process known as "atresia". The endocrinological regulatory mechanisms involved in follicular development and atresia have been characterized to a large extent, but the precise temporal and molecular mechanisms involved in the regulation of these events have remained largely unknown. Recent studies suggest that the apoptosis of ovarian granulosa cells plays a major role in follicular atresia. In this review, we provide an overview of development and atresia of follicles, and apoptosis of granulosa cells in mammals.     

颗粒细胞凋亡影响家禽卵泡闭锁的研究进展

卵巢是家禽的重要繁殖器官,会产生大量卵泡,而卵泡在生长发育的各个阶段中都可能因为不同因素的调控而发生闭锁,最终导致繁殖性能衰退。颗粒细胞对卵泡的生长发育有重要调控作用,其凋亡会诱导卵泡发生闭锁。诱导颗粒细胞发生凋亡的因素较多,包括激素、细胞因子、氧化应激、线粒体及其他体外因素。颗粒细胞凋亡主要由线粒体途径导致,其涉及到半胱天冬酶（Caspase）家族参与,当线粒体裂解时会释放细胞色素C （Cyt-C）,随后形成凋亡小体激活Caspase-3和Caspase-8,最终激活Caspase-9导致颗粒细胞凋亡;当颗粒细胞发生凋亡,家禽体内卵泡丧失生物功能并且卵泡细胞之间的调控失衡,促使卵泡内卵母细胞和膜细胞凋亡,最终导致卵泡发生闭锁;颗粒细胞在存活状态下所分泌的生长因子、性腺类固醇、细胞因子能减少卵母细胞氧化损伤,防止细胞内活性氧（ROS）水平过高导致的线粒体DNA损伤,从而避免线粒体功能障碍而造成的颗粒细胞凋亡。作者从颗粒细胞凋亡及其影响因素、颗粒细胞凋亡和卵泡闭锁的关系、颗粒细胞凋亡对卵泡闭锁的影响3个方面进行阐述,以期为减少卵泡闭锁、提高家禽繁殖性能提供理论依据。     

生殖激素控制卵泡细胞凋亡的研究进展

研究表明颗粒细胞凋亡是导致卵泡闭锁的重要原因，而颗粒细胞凋亡涉及许多因素，其中生殖激素，如GnRH、FSH、LH、P4、E、A、GH、Mel、inhibin、activin、follistatin等间接地和直接地对卵巢卵泡细胞凋亡发挥重要的综合控制作用，因此正确理解激素对体内、外卵泡及颗粒细胞发育和衰亡的调节网络具有很重要的理论和实践意义。     

家禽卵泡颗粒细胞体外培养方法研究进展

家禽卵巢中颗粒细胞在卵泡发育和闭锁中起着重要作用,包括信号传递、营养供给以及离子平衡等。卵泡颗粒细胞的增殖与分化自身也受到生殖激素和细胞因子的综合调节,其作用机制是当前研究热点。家禽卵泡颗粒细胞的体外培养可作为研究繁殖生理调节的理想细胞模型。本文对家禽卵泡颗粒细胞的体外培养研究现状及颗粒细胞在卵泡生长发育和闭锁中的作用进行简要概述,并总结了原代颗粒细胞体外培养模型的建立方法。     

细胞自噬对动物卵泡发育调控的研究进展

卵泡是雌性哺乳动物发挥其繁殖能力的基础,其发育是一个动态的过程,主要涉及原始卵泡的形成、卵泡的募集、优势卵泡的选择、成熟卵泡的排卵以及排卵后卵泡的黄体化。卵泡发育的整个过程受内分泌系统、细胞自噬、细胞凋亡等的调控。自噬是一种进化上保守的应激反应过程,通过将细胞内物质包裹形成自噬体并传递到溶酶体中进行降解,以帮助细胞维持胞内物质代谢平衡,其在卵泡发育的过程中发挥着重要作用,一方面它能够通过降解或回收受损的蛋白质或有害代谢产物缓解应激造成的卵泡损伤,另一方面它又通过产生大量自噬体导致细胞器过度降解而引起卵泡闭锁。自噬对卵泡发育的调控需要PI3K-Akt-mTOR、MAPK-ULK1、ERK1/2、Sirt1-FOXO1-Atg7等多种经典信号通路的参与,这些信号通路在激素、氧化应激、细胞饥饿等的刺激下,通过独立作用或相互作用促进或抑制自噬调控卵泡细胞的生理活动。目前已知不同的自噬水平对卵泡细胞的存活具有不同作用,但关于决定细胞能否存活的自噬水平的研究还比较少。此外,自噬对卵泡发育调控的研究主要集中在颗粒细胞中,而对卵母细胞的成熟和卵泡膜细胞的作用的报道较少。文章简述了自噬在卵巢储备的形成、生长卵泡的发育、黄体的形成和退化及卵泡闭锁中的作用,并分析了一些常见的化工产品和应激诱导的自噬对卵泡发育的影响,以期为全面了解自噬在卵泡发育中的调控作用提供一定的参考。     

Regulation of ovarian folliculogenesis by IGF and BMP system in domestic animals

Involvement of insulin-like growth factors (IGFs) and IGF binding proteins (IGFBPs) in ovarian folliculogenesis has been extensively studied during the last decade. In all mammalian species, IGF-I stimulates granulosa cell proliferation and steroidogenesis. The concentrations of IGF-I and -II do not vary during terminal follicular growth and atresia. In contrast, the levels of IGFBP-2 and -4, as well as IGFBP-5 in ruminants, dramatically decrease and increase during terminal follicular growth and atresia, respectively. These changes are responsible for an increase and a decrease in IGF bioavailability during follicular growth and atresia, respectively. They are partly explained by changes in ovarian expression. In particular, expression of IGFBP-2 mRNA decreases during follicular growth in ovine, bovine and porcine ovaries, and expression of IGFBP-5 mRNA dramatically increases in granulosa cells of bovine and ovine atretic follicles. Changes in IGFBP-2 and -4 levels are also due to changes in intrafollicular levels of specific proteases. Recently, we have shown that the pregnancy-associated plasma protein-A (PAPP-A) is responsible for the degradation of IGFBP-4 in preovulatory follicles of domestic animals. Expression of PAPP-A mRNA is restricted to the granulosa cell compartment, and is positively correlated to expression of aromatase and LH receptor. From recent evidence, the bone morphogenetic protein (BMP) family would also play a key role in ovarian physiology of domestic animals. In particular, we and others have recently shown that a non-conservative substitution (Q249R) in the bone morphogenetic protein-receptor type IB (BMPR-IB) coding sequence is fully associated with the hyperprolific phenotype of FecB(B)/FecB(B) Booroola ewes. BMP-4 and GDF-5, natural ligands of BMPR-IB, strongly inhibit secretion of progesterone by ovine granulosa cells in vitro, but granulosa cells from FecB(B)/FecB(B) ewes are less responsive than those from FecB(+)/FecB(+) to the action of these peptides. It is suggested that in FecB(B)/FecB(B) ewes, Q249R substitution would impair the function of BMPR-IB, leading to a precocious differentiation of granulosa cells and of follicular maturation. Interestingly, recent findings have described mutations in BMP-15 gene associated with hyperprolific phenotypes in Inverdale and Hanna ewes, suggesting that the BMP pathway plays a crucial role in the control of ovulation rate.     

Role of cell-death ligand-receptor system of granulosa cells in selective follicular atresia in porcine ovary

In the mammalian ovary, more than 99% of follicles degenerate without ovulation and few oocytes ovulate and succeed to the next generation. Granulosa cell apoptosis plays a critical role in this process, follicular atresia. However, the molecular mechanisms responsible for the regulation of granulosa cell apoptosis have not been clarified. Death ligand and receptor systems are major apoptosis-inducing factors. This review describes the granulosa cell apoptosis via death ligand and receptor systems during follicular atresia in the porcine ovary.     

Regulation of granulosa cell apoptosis by death ligand–receptor signaling

Several hundred thousand primordial follicles are present in the mammalian ovary, however, only 1% develop to the preovulatory stage and finally ovulate. The remainder will be eliminated via a degenerative process called ‘atresia’. The endocrinological regulatory mechanisms involved in follicular development and atresia have largely been characterized but the precise temporal and molecular mechanisms involved in the regulation of these events remain unknown. Many recent studies suggest that apoptosis in ovarian granulosa cells plays a crucial role in follicular atresia. Notably, death ligand‐receptor interaction and subsequent intracellular signaling have been demonstrated to be the key mechanisms regulating granulosa cell apoptosis. In this review we provide an overview of granulosa cell apoptosis regulated by death ligand‐receptor signaling. The roles of death ligands and receptors [Fas ligand (FasL)]‐Fas, tumor necrosis factor α (TNFα)‐TNF receptor and TNFα‐related apoptosis‐inducing ligand (TRAIL)‐TRAIL receptor (TRAILR)] and intracellular death‐signal mediating molecules (Fas‐associated death domain protein), TNF receptor 1‐associated death domain protein, caspases, apoptotic protease‐activating factor 1, TNFR‐associated factor 2 and cellular FLICE‐like inhibitory protein in granulosa cells are discussed.     

The Insulin‐like Growth Factor System: a Key Determinant Role in the Growth and Selection of Ovarian Follicles? A Comparative Species Study

The aim of the present paper is to make a comparative study of the expression of the elements of the insulin‐like growth factor (IGF) system in different mammalian species and thus illuminate their potential role in the process of ovarian folliculogenesis in mammals. In most mammalian species, IGFs and IGFBPs (in particular IGFBP‐2 and IGFBP‐4) are considered, respectively, as stimulators and inhibitors of follicular growth and maturation. In mammalian species, IGFs might play a key role in sensitizing ovarian granulosa cells to FSH action during terminal follicular growth. Concentrations of IGFBP‐2 and IGFBP‐4 in follicular fluid strongly decrease and increase during follicular growth and atresia, respectively, leading to an increase and a decrease in IGF bioavailability, respectively. The decrease in these IGFBPs is because of a decrease in mRNA expression (IGFBP‐2) and an increase in proteolytic degradation by PAPP‐A in follicular fluid (IGFBP‐2, IGFBP‐4 and IGFBP‐5), and likely participates in the selection of dominant follicles. In contrast, levels and/or sites of expression of IGF‐I, IGF‐II, IGFBP‐4, IGFBP‐5 and type II receptor in follicular cells strongly differ between mammalian species, suggesting that these phenomena might play species‐specific or secondary roles in ovarian folliculogenesis.     

Expression and localization of growth factors and their receptors in the mammalian testis. Part I: Fibroblast growth factors and insulin-like growth factors

It is now well established that normal development and function of testis are mediated by endocrine and paracrine pathways including hormones, growth factors and cytokines as well as by direct cell-to-cell contacts depending on tight, adhering and gap junctions. In the last two decades, several growth factors were identified in the testis of various mammalian species. Growth factors are shown to promote cell proliferation, regulate tissue differentiation, and modulate organogenesis. Interestingly, most of these peptides are expressed not only in the adult mammalian testis during spermatogenesis but also during testicular morphogenesis in prenatal and postnatal life. Our study was launched to provide an overview of the expression, localization, and putative physiological roles of growth factors and their receptors in the mammalian testis. The growth factors considered in this part of our review are fibroblast growth factors and insulin-like growth factors. These factors are found in testicular cells in prenatal, postnatal, and adult animals and are implicated in the regulation of important testicular activities including testicular cord morphogenesis, modulation of testicular hormone secretion and control of spermatogenesis.     

Regulation mechanism of selective atresia in porcine follicles: regulation of granulosa cell apoptosis during atresia

More than 99% of follicles undergo a degenerative process known as "atresia", in mammalian ovaries, and only a few follicles ovulate during ovarian follicular development. We have investigated the molecular mechanism of selective follicular atresia in mammalian ovaries, and have reported that follicular selection dominantly depends on granulosa cell apoptosis. However, we have little knowledge of the molecular mechanisms that control apoptotic cell death in granulosa cells during follicle selection. To date, at least five cell death ligand-receptor systems [tumor necrosis factor (TNF)alpha and receptors, Fas (also called APO-1/CD95) ligand and receptors, TNF-related apoptosis-inducing ligand (TRAIL; also called APO-2) and receptors, APO-3 ligand and receptors, and PFG-5 ligand and receptors] have been reported in granulosa cells of porcine ovaries. Some cell death ligand-receptor systems have "decoy" receptors, which act as inhibitors of cell death ligand-induced apoptosis in granulosa cells. Moreover, we showed that the porcine granulosa cell is a type II apoptotic cell, which has the mitochondrion-dependent apoptosis-signaling pathway. Briefly, the cell death receptor-mediated apoptosis signaling pathway in granulosa cells has been suggested to be as follows. (1) A cell death ligand binds to the extracellular domain of a cell death receptor, which contains an intracellular death domain (DD). (2) The intracellular DD of the cell death receptor interacts with the DD of the adaptor protein (Fas-associated death domain: FADD) through a homophilic DD interaction. (3) FADD activates an initiator caspase (procaspase-8; also called FLICE), which is a bipartite molecule, containing an N-terminal death effector domain (DED) and a C-terminal DD. (4) Procaspase-8 begins auto-proteolytic cleavage and activation. (5) The auto-activated caspase-8 cleaves Bid protein. (6) The truncated Bid releases cytochrome c from mitochondrion. (7) Cytochrome c and ATP-dependent oligimerization of apoptotic protease-activating factor-1 (Apaf-1) allows recruitment of procaspase-9 into the apoptosome complex. Activation of procaspase-9 is mediated by means of a conformational change. (8) The activated caspase-9 cleaves downstream effector caspases (caspase-3). (9) Finally, apoptosis is induced. Recently, we found two intracellular inhibitor proteins [cellular FLICE-like inhibitory protein short form (cFLIPS) and long form (cFLIPL)], which were strongly expressed in granulosa cells, and they may act as anti-apoptotic/survival factors. Further in vivo and in vitro studies will elucidate the largely unknown molecular mechanisms, e. g. which cell death ligand-receptor system is the dominant factor controlling the granulosa cell apoptosis of selective follicular atresia in mammalian ovaries. If we could elucidate the molecular mechanism of granulosa cell apoptosis (follicular selection), we could accurately diagnose the healthy ovulating follicles and precisely evaluate the oocyte quality. We hope that the mechanism will be clarified and lead to an integrated understanding of the regulation mechanism.     

Nutritional influences on folliculogenesis

Folliculogenesis is an intricate process that involves the proliferation and differentiation of both somatic and germ cells. This process depends on complex interactions between systemic factors such as both pituitary gonadotrophins and metabolic hormones and/or local factors produced by the ovarian somatic and germ cells, such as the IGF system and TGF-β superfamily members. In domestic ruminants, follicular development begins during foetal life with formation of primordial follicles from the association of germ cells and pre-granulosa cells. After follicular formation, folliculogenesis begins with a primordial follicle progressing into more developed stages (i.e. primary, secondary, pre-antral and antral) in a continuous, progressive process to either ovulation or, as in most cases, to atresia. Even early stages of follicular formation and subsequent development are influenced by both internal (e.g. genotype) and/or external environmental (e.g. nutrition and season) factors. Among these external factors, nutrition is one of the most important affecting reproductive function, and this is the focus of this review, because other reviews in this issue discuss other environmental factors. A number of studies have now shown that nutrition can have both positive and negative effects on follicular growth, oestrous activity, oocyte quality, blastocyst development and pregnancy outcome. Therefore, understanding the intricate processes involved during folliculogenesis and the ways in which factors, such as nutrition, affect them is leading to new opportunities to improve pregnancy rates by influencing follicle development and oocyte quality. This review will focus on follicular development from foetal to adult stages and the influences that nutrition has during some of these developmental stages.     

miRNA调节卵泡颗粒细胞凋亡及其机制的研究进展

微小核糖核酸(microRNA,miRNA)是一类内源性非编码RNA,具有广泛的基因表达调控作用,可以在转录后水平通过影响靶基因来调控相应蛋白质的表达,进而调节细胞的生命活动。miRNA在哺乳动物卵泡颗粒细胞中表达,并调控颗粒细胞的凋亡。颗粒细胞作为卵巢卵泡中数量最多的细胞群,在卵泡发育过程中起着至关重要的作用,不仅为卵母细胞提供营养物质,还调控其发育和成熟。颗粒细胞凋亡是导致卵泡闭锁的重要原因,影响卵泡的数量和质量从而影响雌性动物的繁殖性能。颗粒细胞凋亡过程受多种因素的调控。文章简述了miRNA对卵巢颗粒细胞凋亡的调控作用及其机制,其中包括miRNA通过调控激素分泌和细胞凋亡相关因子的表达进而调节颗粒细胞的凋亡,miRNA对颗粒细胞凋亡相关信号通路的影响,miRNA调控颗粒细胞凋亡导致的卵巢相关疾病,并总结了对颗粒细胞凋亡有调控作用的miRNA,以及miRNA在疾病诊断和治疗中的潜在作用,以期为后续相关卵巢疾病的发病机制和治疗方案研究,以及提高雌性哺乳动物生殖性能提供指导和参考。