哺乳动物卵泡早期发育调控的研究进展

哺乳动物卵泡发育是一个被众多内分泌、旁分泌和自分泌因素精确调控的生理过程 ,本文结合本试验的研究工作 ,综述了卵泡早期发育调控的研究进展     

Ovarian function in ruminants

The purpose of this overview is to highlight important steps of ovarian regulation during follicle development, ovulation and the life span of corpus luteum (CL) in ruminants. The ovarian cycle is central to reproductive function. It is characterized by repeating patterns of cellular proliferation, differentiation and transformation that encompass follicular development and ovulation as well as the formation, function and regression of the CL. In the first part, the importance and regulation of final follicle growth and especially of angiogenesis and blood flow during folliculogenesis, dominant follicle development and CL formation are described. Our results underline the importance of growth factors especially of insulin-like growth factor (IGF), vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) for development and completion of a dense network of capillaries (angiogenesis) during follicle growth and CL formation. In the second part, the regulation of CL function by endocrine/paracrine and autocrine acting regulators is discussed. There is evidence that besides the main endocrine hormones luteinizing hormone (LH) and growth hormone (GH) local regulators as growth factors, peptides, steroids and prostaglandins are important modulators of luteal function. During early CL development until midluteal stage oxytocin (OT), prostaglandins and progesterone (P) itself stimulate luteal cell proliferation and function supported by the luteotropic action of a number of growth factors. The still high mRNA expression, protein concentration and localization of VEGF, FGF and IGF family members in the cytoplasm of luteal cells during midluteal stage suggest that they play pivotal role in the maintenance (survival) of this endocrine tissue. The major function of the CL is to secrete P. Progesterone itself regulates the length of the estrous cycle via influencing the timing of the luteolytic PGF2alpha signal from the endometrium. At the end of a nonfertile cycle, the regression of CL commences, steroidogenic capacity is lost (functional luteolysis), cell death is initiated, and tissue involution as well as resorption occurs within a few days (structural luteolysis). The cascade of mediators during luteolysis is very complex and still awaits elucidation. Evidence is given for participation of blood flow, inflammatory cytokines, vasoactive peptides (angiotensin II and endothelin-1), and decrease of the classical luteotropic mediators.     

Monitoring follicular development in cattle by real-time ultrasonography: a review.

The application of real-time ultrasonography to monitoring ovarian function in mammals has advanced the understanding of follicular dynamics and its regulation. Follicular development is a wave-like sequence of organised events. The waves consist of the synchronous growth of small (4 to 5 mm) antral follicles, followed by the selection and growth of one dominant follicle which achieves the largest diameter and suppresses the growth of the subordinate follicles. In the absence of luteal regression, the dominant follicle eventually regresses (becomes atretic) and a new follicular wave begins. The dominant follicle regulates the growth of the subordinate follicles, because the appearance of the next wave is accelerated if the dominant follicle is ablated, and delayed if the lifespan of the dominant follicle is prolonged. During bovine oestrous cycles, two or three successive waves emerge, on average, on the day of ovulation (day 0) and day 10 for two-wave cycles, and on days 0, 9 and 16 for three-wave cycles. During the oestrous cycle there are thus two or three successive dominant follicles, and the last of these ovulates. Ovarian folliculogenesis is a complex process involving interactions between pituitary gonadotrophins, ovarian steroids and non-steroidal factors. Subtle changes in the hormonal milieu regulate folliculogenesis and the emergence of a follicular wave is preceded by a small increase in the concentration of plasma follicle-stimulating hormone. The mechanisms that promote the selection of a dominant follicle have not been elucidated, but considerable progress has been made in understanding follicular development and its regulation. Most treatments designed to control the development of follicular waves have been based on the physical or hormonal removal of the suppressive effect of the dominant follicle, and the consequent controlled induction of the emergence of a new follicular wave. The studies reviewed here describe current methods for regulating the bovine ovarian cycle, interesting models for future studies, and information that may be used for improving reproductive efficiency.     

原始卵泡形成和发育调节机制研究进展

原始卵泡的形成及其向初级卵泡的转化是卵泡发育过程中的一个重要阶段,是由多种内分泌、旁分泌和自分泌因素调控的生理过程。高水平的类固醇激素会抑制卵泡的形成,局部分泌的kit配基、碱性成纤维细胞生长因子、白血病抑制因子、角质形成细胞生长因子、骨形态发生蛋白等可促进原始卵泡向初级卵泡的转化,而抗缪勒氏激素则对这一过程有抑制作用。     

Cellular and extracellular vesicular origins of miRNAs within the bovine ovarian follicle

The ovarian follicle components must provide an ideal environment to ensure the success of reproductive processes, and communication between follicular cells is crucial to support proper oocyte growth. Recently, it has been demonstrated that the presence of extracellular vesicles (EVs) carrying microRNAs (miRNAs) in follicular fluid represents an important autocrine and paracrine communication mechanism inside the ovarian follicle. In this study, we tested the hypothesis that the miRNA content of EVs isolated from ovarian follicular (granulosa and cumulus–oocyte complexes) cell‐conditioned culture media is dependent upon cell type. We initially screened bovine granulosa cells (GCs) and cumulus–oocyte complexes (COCs), as well as their derived EVs for 348 miRNAs using real‐time PCR, and detected 326 miRNAs in GCs and COCs cells and 62 miRNAs in GCs and COCs EVs. A bioinformatics analysis of the identified cell‐specific and differentially expressed miRNAs predicted that they likely modulate important cellular processes, including signalling pathways such as the PI3K‐Akt, MAPK and Wnt pathways. By investigating the origins of miRNAs within the follicular fluid, the results of this study provide novel insights into follicular miRNA content and intercellular communication that may be of invaluable use in the context of reproductive technologies, diagnostic of ovarian‐related diseases and/or the identification of biomarkers for oocyte and embryo quality.     

Endocrine mechanisms responsible for different follicular development during the estrous cycle in Hatano high- and low-avoidance rats

Hatano high- and low-avoidance rats (HAA and LAA strains, respectively) were selected and bred according to the avoidance rate in a shuttle-box task. Although they have clear strain differences in ovarian function, their endocrine mechanisms still remain to be clarified. Differences in female reproductive endocrinology between the strains were investigated by means of measuring the plasma concentration of reproductive hormones during the estrous cycle. LAA rats showed approximately threefold lower basal and surge levels of LH, a more than fourfold lower level of FSH surges and higher levels of inhibin A and inhibin B during the estrous cycle compared with the levels seen in HAA rats. The concentration of estradiol-17β in the proestrous stage was significantly lower in LAA rats than in HAA rats. Additionally, LH and FSH secretions from primary cultured anterior pituitary cells with or without in vitro GnRH stimulation were lower in the cells derived from LAA rats and, in terms of FSH secretion, were unresponsive to GnRH in contrast to cells derived from HAA rats. Although an increased number of preantral follicles in diestrus were observed in LAA rats, number of hCG-induced ovulation was lower in LAA rats. LAA rats may have much more follicle growth during the early stage of folliculogenesis, but most follicles might not grow into mature follicles. These results strongly suggest that the strain difference in ovarian function of these two Hatano rats is due to the difference in the regulation of hypothalamo-hypophyseal system for gonadotropins secretion.     

反刍家畜卵泡发育机理的研究进展

反刍动物的卵泡发育是一个复杂且高度协调的过程，最新的研究肯定了卵泡发育以波的形式存在，但在影响卵泡发育的因素中，除了传统的下丘脑—垂体—性腺轴外，一些非促性腺激素内源性因子及卵巢内部因子也起到非常重要的作用，即便如此，要想对反刍动物卵泡发育进行更精确的控制，仍需作更进一步的研究     

Altered Expression of Transforming Growth Factor‐Beta Isoforms in Bovine Cystic Ovarian Disease

Cystic ovarian disease (COD) is one of the main causes of infertility in dairy cattle. It has been shown that intra‐ovarian factors may contribute to follicular persistence. Transforming growth factor‐beta (TGFB) isoforms are important paracrine and autocrine signalling molecules that regulate ovarian follicle growth and physiology. Considering the importance of these factors in the ovarian physiology, in this study, we examined the expression of TGFB isoforms (TGFB1, TGFB2 and TGFB3) in the ovary of healthy cows and animals with spontaneous and adrenocorticotrophic hormone (ACTH)‐induced COD. In the oestrous‐synchronized control group, the expression of TGFB1 in granulosa and theca cells was higher in spontaneous cysts than in atretic or tertiary follicles. When we compared TGFB2 expression in granulosa cells from atretic or tertiary follicles from the oestrous‐synchronized control group with that in ACTH‐induced or spontaneous follicular cysts, we found a higher expression in the latter. The expression of the TGFB isoforms studied was also altered during folliculogenesis in both the spontaneous and ACTH‐induced COD groups. As it has been previously shown that TGFB influences steroidogenesis, ovarian follicular proliferation and apoptosis, an alteration in its expression may contribute to the pathogenesis of this disease.     

Effect of Oxytocin,IBMX and dbcAMP on Rabbit Ovarian Follicles

Oxytocin (OT) and protein kinase A (PKA), a possible intracellular mediator of hormone action in the ovary, can be potent activators of ovarian functions and fertility. Nevertheless, action of OT on ovarian follicle atresia has not been studied yet. Only single administration of PKA activators [3‐isobutyl‐1‐methyl‐xanthine (IBMX) and dibutyryl cyclic adenosine monophosphate (dbcAMP)] on ovarian follicle atresia was studied previously. The aim of this study was to examine the effect of OT (single treatment per one reproductive cycle, multiple treatments for three cycles), IBMX and dbcAMP (multiple treatments) on folliculogenesis and follicular atresia in rabbit. The ovarian cycle in control females was induced only by gonadotropins. Experimental females received co‐administration of gonadotropins with OT, IBMX or dbcAMP (at 50 μg/female). All females were artificially inseminated. Single‐treated females were euthanized after 18–19 h. Multiple‐treated females were euthanized after the third reproductive cycle. Histological sections of the ovaries were prepared and evaluated by a light microscopy. The follicles were divided into four classes according to the structure of granulosa and theca cells as follows: none or small atresia, cystic atresia, obliterative atresia and atresia associated with luteinization. The ovaries from the control and experimental females, treated during one reproductive cycle or three cycles, were compared. Single OT co‐administration increased proportion of follicles with atresia associated with luteinization, but not other types of atresia. No influence of multiple OT co‐administration on follicular atresia was recorded. Multiple IBMX and dbcAMP co‐administration decreased the proportion of atretic follicles and increased the proportion of healthy follicles without atresia.     

Regulation of primordial follicle formation,dormancy, and activation in mice

In female reproduction, the oocyte number is limited after birth. To achieve a continuous ovulatory cycle, oocytes are stored in primordial follicles. Therefore, the regulation of primordial follicle dormancy and activation is important for reproductive sustainability, and its collapse leads to premature ovarian insufficiency. In this review, we summarize primordial follicle development and the molecular mechanisms underlying primordial follicle maintenance and activation in mice. We also overview the mechanisms discovered through in vitro culture of functional oocytes, including the establishment of primordial follicle induction by environmental factors, which revealed the importance of hypoxia and compression by the extra cellular matrix (ECM) for primordial follicle maintenance in vivo.     

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.     

马属动物卵巢发育研究进展

随着国内经济发展,马、驴产业经济价值越来越高,产业需求量也日益增长,但马属动物是繁殖周期较长的季节性单胎动物,其独特的繁殖生理结构特性成为马属动物繁殖效益和相关繁殖技术开发应用的主要因素之一.卵巢作为雌性动物生殖系统中的核心器官,在神经系统和内分泌系统的作用和调节下,具有周期性产生卵子和分泌生殖激素的功能.然而由于排卵...     

Control of reproductive processes by growth hormone: extra- and intracellular mechanisms

Recent data on the association between growth hormone (GH) and male and female reproductive processes, as well as the effects of GH on these processes and on some reproductive and non-reproductive disorders, and possible extra- and intracellular mediators of its action are reviewed. The available data suggest that GH is an important endocrine and autocrine/paracrine regulator of reproduction. It controls proliferation, apoptosis, growth and differentiation and the secretory and generative activities of different reproductive organs. It also regulates their response to gonadotrophin-releasing hormone (GnRH) and gonadotropins. Despite the effects of GH on the IGF/IGFBP (insulin-like growth factor binding protein) system, oxytocin, steroids, activin, gonadotropin and gonadotropin receptors, the majority of GH's actions on the reproductive processes are probably mediated not by these substances but by specific GH receptors acting through cAMP/protein kinase A, protein kinase G, tyrosine kinase-, MAP kinase and CDC2 kinase-dependent intracellular mechanisms. Although GH treatments can increase the risk of some reproductive and non-reproductive disorders, they may be useful in improving gonadal function, inducing superovulation and in embryo production.     

Insulin-like growth factor-I as a possible hormonal mediator of nutritional regulation of reproduction in cattle

The current review aims to establish insulin-like growth factor-1 (IGF-I) as the factor that signals nutritional status to the reproductive axis, and show that assessment of IGF-I in blood early postpartum during the negative energy balance (NEB) period could be used to predict both nutritional and reproductive status in dairy cattle. The review also explores the effect of nutritional status on circulating IGF-I concentrations and the endocrine role of IGF-I on the reproductive axis. IGF-I plays an important role in gonadotropin-induced folliculogenesis, ovarian steroidogenesis and corpus luteum (CL) function. It also modulates pituitary and hypothalamus function. IGF-I clearly has an endocrine role on the reproductive axis. Severe under nutrition significantly reduces plasma IGF-I concentrations. During the critical period of NEB in high yielding dairy cattle early postpartum, IGF-I concentrations are low in blood and its levels are positively correlated to energy status and reproductive function during this period. Changes in circulating IGF-I immediately postpartum may help predict both nutritional and reproductive status in dairy cattle. IGF-I is therefore one of the long sought factors that signal nutritional status to the reproductive axis.     

绵羊卵泡的发育及其调控研究

卵泡是绵羊卵巢上的基本发育单位,卵子发生是在卵泡内进行的,即卵泡为卵子发生提供了一个最佳的微环境。卵泡的发生受内分泌、旁分泌和自分泌调控,本文就对绵羊卵泡的发育及其调控机理进行了概述。     

Promotion of ovarian follicular development by injecting vascular endothelial growth factor (VEGF) and growth differentiation factor 9 (GDF-9) genes

Ovarian follicular development in mammals is the complex process including endocrine, paracrine and autocrine. There is the development of four basic stages of ovarian follicles, i.e. the primordial, primary, secondary and tertiary or Graafian follicles. There are few blood vessels in the cortical area where primordial and primary follicles are assembled. The development of these follicles is stimulated by oocytes derived factor including growth differentiation factor 9 (GDF-9) or bone morphogenetic protein 15 (BMP-15). Porcine GDF-9 complementary DNA (cDNA) cloned, and then injected its gene into the ovary in gilts. The injection of porcine GDF-9 gene resulted in an increase in the number of primary, secondary and tertiary follicles, concomitant with a decrease in the number of primordial follicles, indicating that exogenous GDF-9 can promote early folliculogenesis in the porcine ovary. On the other hand, the development of antral follicles is associated with increased density of blood vessels within the theca cell layers surrounding the follicles. A recent study reported that vascular endothelial growth factor (VEGF) play an important role in the process of thecal angiogenesis during follicular development. To investigate whether additional induction of thecal angiogenesis would support subsequent follicular development, miniature gilts were directly injected VEGF gene into the ovary. Injection of VEGF gene increased the levels of mRNA expression of VEGF 120 and VEGF 164 isoforms in the granulosa cells and VEGF protein contents in the follicular fluid. The number of preovulatory follicles and the capillary density in the theca interna increased significantly in the ovaries injected with VEGF gene compared with those treated with eCG alone, indicating that the regulation of thecal angiogenesis during follicular development is a very important factor in the development of ovulatory follicles. This technique may be an innovative technique for enhanced induction of follicular development in the ovary through gene and hormonal treatment, which may lead to prevention of infertility caused by ovarian dysfunction.     

Gonadotropin releasing hormone receptor gene and protein expression and immunohistochemical localization in bovine uterus and oviducts

Recently GnRH, GnRH-R systems has been demonstrated in various extrahypothalamic and extrapituitary reproductive tissues in different mammalian species, where GnRH acts in an autocrine and or paracrine manner and modulates different biological processes. GnRH-R mRNA has also been demonstrated in bovine ovaries (follicle and corpus luteum) and normal and carcinogenic human endometrium/endometrial cells. This is the first study elucidating presence of GnRH-R mRNA and GnRH-R protein in bovine uterus and oviducts in follicular and luteal phases of the estrous cycle and further localizing the receptors to endometrial and oviductal epithelial cells. To our knowledge this is the first report demonstrating GnRH-R mRNA and protein in mammalian oviducts. We used gene-specific primers and monoclonal GnRH-R antibody to test GnRH-R mRNA and GnRH-R protein through RT-PCR and immunobloting. Immunohistochemistry was employed to localize these receptors to endometrial and oviductal epithelial cells. GnRH-R mRNA and receptor protein were expressed at expected molecular weights of 920bp and 60kD, respectively. Densitometry analysis revealed that expression levels for GnRH-R protein in uterus and oviducts were similar to bovine pituitary. The presence of GnRH receptors in bovine uterus and oviducts is intriguing and it would be imperative to examine the functional role of this system in the regulation of reproductive processes.     

Role of oocyte‐derived paracrine factors in follicular development

Mammalian oocytes secrete transforming growth factor β (TGF‐β) superfamily proteins, such as growth differentiation factor 9 (GDF9), bone morphogenetic protein 6 (BMP6) and BMP15, and fibroblast growth factors (FGFs). These oocyte‐derived paracrine factors (ODPFs) play essential roles in regulating the differentiation and function of somatic granulosa cells as well as the development of ovarian follicles. In addition to the importance of individual ODPFs, emerging evidence suggests that the interaction of ODPF signals with other intra‐follicular signals, such as estrogen, is critical for folliculogenesis. In this review, we will discuss the current understanding of the role of ODPFs in follicular development with an emphasis on their interaction with estrogen signaling in regulation of the differentiation and function of granulosa cells.     

Fertility of a High‐Altitude Sheep Model is Compromised by Deficiencies in Both Preovulatory Follicle Development and Plasma LH Availability

At high altitude, hypoxia and/or oxidative stress may compromise fertility. This study tested the relative effect of short‐ or long‐term exposure to high‐altitude hypobaric hypoxia and oxidative stress in sheep on preovulatory follicle dynamics and gonadotrophin secretion. Thus, growth dynamics, stereidogenic function and competence to ovulate of preovulatory follicles, as well as FSH and LH availability throughout the entire oestrous cycle, were compared among sheep native from low and high altitude, and sheep newcomers to high altitude. The results indicates that short‐term exposure in sheep newcomers to high altitude has a deleterious effect on both the ovarian function (affecting preovulatory follicular development) and the pituitary function (diminishing plasma LH availability). On the other hand, there were no detected differences in the preovulatory follicular development in sheep adapted to high altitude for generations and, conversely, LH secretion was increased, which suggests an adaptive mechanism. The treatment with antioxidant agents during a relative short period for the time of folliculogenesis (approximately 1 month and a half) changed substantially the development of preovulatory follicles in short‐term exposed sheep to similar patterns than in sheep native and living to both high and low altitude. These results highlight the role of oxidative stress in the detriment of the reproductive function in individuals recently exposed to high‐altitude hypoxic environment.