Involvement of growth hormone (GH) and insulin-like growth factor (IGF) system in ovarian folliculogenesis

During the last decade, involvement of growth hormone (GH), insulin-like growth factors (IGFs) and IGF binding proteins (IGFBPs) in ovarian folliculogenesis has been extensively studied. This review provides an update on the GH, IGF system and their role in ovarian follicular development. In vitro studies and knockout experiments demonstrated an important role of GH in preantral follicle growth and differentiation through their binding with GH receptors, which are located both in the oocyte and follicular somatic tissues. Furthermore, GH stimulates the development of small antral follicles to gonadotrophin-dependent stages, as well as maturation of oocytes. With regard to the IGF system, IGF-I has no effects on primordial follicle development, but both IGF-I and IGF-II stimulate growth of secondary follicles. Depending on the species studies and method used, these proteins have been detected in oocytes and/or somatic cells. In antral follicles, these IGFs stimulate granulosa cell proliferation and steroidogenesis in most mammals. The bioavailability of IGFs is regulated by a family of intrafollicular expressed IGF binding proteins (IGFBPs). Facilitation of IGF can be increased through the activity of specific IGFBP proteases, which degrade the IGF/IGFBP complex, resulting in the production of IGFBP fragments and release of attached IGF.     

Type I IGF receptors of Leydig cells are upregulated by human chorionic gonadotropin

The effects of human chorionic gonadotropin (hCG) on type I insulin-like growth factor (IGF) receptors of purified Leydig cells were investigated. Sprague-Dawley rats (50 day-old) were treated with a single injection of hCG 10 units intraperitoneally, type I IGF receptors were then determined daily for 4 days. HCG caused a rapid increase in type I IGF receptors within 24 h, which returned to basal by 72 h. There was no significant change in binding affinity. Our present study indicates that type I IGF receptors of Leydig cells are up regulated by hCG, and this may be one mechanism by which hCG and IGF-I interact to enhance Leydig cell steroidogenesis.     

Growth and the initiation of steroidogenesis in porcine follicles are associated with unique patterns of gene expression for individual componentsof the ovarian insulin-like growth factor system

Ovarian follicular growth and steroidogenesis are controlled by the interaction of insulin-like growth factors (IGFs) and gonadotropins. The objective was to determine the temporal and spatial relationships for gonadotropin receptor, steroidogenic enzyme, and IGF system gene expression during the development of preovulatory porcine follicles. Sows (n = 18) were weaned and follicles were monitored by transrectal ultrasonography. Ovaries were collected from sows when the mean diameter of the preovulatory follicular cohort was approximately 2, 4, 6, or 8 mm. mRNA were measured by in situ hybridization for individual follicles within the preovulatory cohort (3 to 5 follicles per sow). Patterns of gene expression detected by in situ hybridization were confirmed by RNase protection analyses of pooled RNA samples. The amount of LH receptor mRNA and steroidogenic enzyme mRNA (17alpha-hydroxylase and aromatase) increased as the mean diameter of the follicular cohort increased from 2 to 6 mm, but then decreased abruptly for 8-mm follicles. Estradiol concentrations in follicular fluid closely followed the expression patterns of steroidogenic enzymes and LH receptor mRNA. FSH receptor mRNA was present in cohorts of 2-mm follicles but declined in 4-mm follicles and was undetectable in 6- and 8-mm follicles. The expression of IGF-I and type I IGF receptor mRNA were similar for follicles of 2, 4, 6, and 8 mm. In contrast, IGF-II mRNA progressively increased in follicles collected from 2-, 4-, and 6-mm cohorts, and then decreased slightly at 8 mm. Type II IGF receptor mRNA was greatest in 8-mm follicles. IGF binding protein-2 (BP-2) mRNA decreased as follicles achieved progressively larger sizes during the preovulatory period (2 to 8 mm), whereas the IGFBP-4 mRNA remained relatively low for follicles in 2- to 6-mm cohorts but then increased markedly in 8-mm follicles. In summary, temporal and spatial patterns of gene expression for gonadotropin receptor, steroidogenic enzyme, and IGF system genes were characterized in preovulatory porcine follicles by using in situ hybridization and RNase protection analyses. The unique patterns of gene expression suggest interdependence among specific genes that may be essential for preovulatory follicular development.     

用酵母双杂交技术研究IGF-1与其结合蛋白3的相互作用

 为了探讨胰岛素样生长因子结合蛋白 3(IGFBP- 3)分子上的胰岛素样生长因子 1 (IGF- 1 )结合位点并找出其关键氨基酸残基组成 ,首先建立了研究 IGF- 1与 IGFBP- 3相互作用的酵母双杂交模型 ,可以定性和定量地分析两个蛋白质之间的相互作用大小 ;同时利用基因体外定点突变的方法 ,对推断出的 IGF- 1结合位点中的氨基酸突变 ,经 DNA序列分析 ,构建了 5种 IGFBP- 3突变体 .然后在酵母双杂交模型中通过对报告基因活性的定量分析 ,检测 IGF- 1与各种 IGFBP- 3突变体之间相互作用的大小 .结果表明 ,IGFBP- 3分子上的 Lys2 2 2 ,Gln2 2 3突变后 ,与 IGF- 1的结合力大大下降 ,而 Arg2 2 5,Pro2 2 6,Ser2 2 7突变后也导致与 IGF- 1结合力的部分下降 .从而初步确定了IGFBP- 3分子中第 2 2 2～ 2 2 7位的氨基酸区域为 IGF- 1的结合位点 ,并且 IGFBP- 3分子上 Lys2 2 2 ,Gln2 2 3在与 IGF- 1结合中起着重要作用 .     

Insulin like growth factor 1 and regulation of ovarian function in mammals

Various growth factors have been proposed to play endocrine and/or paracrine role in mammalian ovarian follicular development. The insulin like growth factor 1 (IGF-1) is one such factor. More and more reports now support the existence of an intra-ovarian IGF system including receptors and binding proteins. The role of IGF-1 in ovary is to amplify gonadotropin hormone action in terms of increased steroidogenesis by ovarian granulosa cell and granulosa cell proliferation. The synthesis and proteolysis of insulin like growth factor binding proteins, under the control of follicle stimulating hormone, regulate the intra-follicular availability of IGF-1, which further determines the sensitivity of granulosa cells to gonadotropins. Besides gonadotropins, IGF-1 has been implicated in somatotropin hormone action in the ovarian function. Exact mechanism of IGF-1 action in the ovarian follicles needs to be worked out to elucidate whether or not IGF-1 is indispensable in addition to know endocrine factors like gonadotropic and ovarian steroid hormones. This will pave the way for better understanding of control(s) which ensure final development of dominant follicle(s) and atresia of other follicles of the cohort.     

Regulation of insulin-like growth factor (IGF) bioactivity by sequential proteolytic cleavage of IGF binding protein-4 and -5

The biological activity of IGF-I and -II is controlled by six binding proteins (IGFBPs), preventing the IGFs from interacting with the IGF receptor. Proteolytic cleavage of IGFBPs is one mechanism by which IGF can be released to bind the receptor. The IGFBPs are usually studied individually, although the presence of more than one of the IGFBPs in most tissues suggests a cooperative function. Thus, the IGFBPs are part of regulatory networks with proteolytic enzymes in one end and the IGF receptor in the other end. We have established a model system that allows analysis of the dynamics between IGF, IGFBP-4 and -5, the IGF receptor, and the proteolytic enzyme PAPP-A, which specifically cleaves both IGFBP-4 and -5. We demonstrate different mechanisms of IGF release from IGFBP-4 and -5: cooperative binding to IGF is observed for the proteolytic fragments of IGFBP-5, but not fragments of IGFBP-4. Furthermore, we find that PAPP-A-mediated IGF-dependent cleavage of IGFBP-4 is inhibited by IGFBP-5, which sequesters IGF from IGFBP-4, and that cleavage of both IGFBP-4 and -5 is required for the release of bioactive IGF. Finally, we show that cell surface-localized proteolysis of IGFBP-4 represents the final regulatory step of efficient IGF delivery to the receptor. Our data define a regulatory system in which molar ratios between the IGFBPs and IGF and between the different IGFBPs, sequential proteolytic cleavage of the IGFBPs, and surface association of the activating proteinase are key elements in the regulation of IGF receptor stimulation.     

Possible autocrine/paracrine actions of insulin-like growth factors during embryonic development: Expression and action of IGFs in undifferentiated P19 cells

The insulin-like growth factors I and II (IGF I and II) and their cell surface receptors are expressed in the mammalian embryo and may function as autocrine or paracrine growth factors during early development. P19 embryonic carcinoma cells, derived from a 7.5 day mouse embryo, were used as a model for a functional study of the IGF system in post-implantation embryogenesis. Undifferentiated P19 cells synthesized IGF I and II, the type I and II IGF receptors, and IGF binding proteins (IGF BP2, IGF BP3, and IGF BP4). P19 cells showed an increase in thymidine incorporation of 150% of control with a 4 hour incubation of IGF I (10 ng/ml) or IGF II (100 ng/ml) and an increase in cell viability compared to control cells during 24 hours of serum starvation. In both experiments IGF I was more potent than IGF II. Endogenous concentrations of IGF I and II in conditioned media were low compared to the doses of exogenous IGFs required for biologic effect, but nonetheless contributed significantly to baseline DNA synthesis, as demonstrated by inhibition of IGF actions with specific antibodies. Cell surface associated IGF BPs bound more radiolabeled IGF than IGF receptors, as determined by binding studies and affinity cross-linking. IGF I and IGF II appeared to regulate production of IGF BP2, suggesting that the IGFs may regulate their own actions by altering the abundance of their binding proteins. © 1993Wiley-Liss, Inc.     

Insulin-like growth factor inhibition of growth hormone secretion

Somatomedins-insulin-like growth factors (SM/IGF) are growth hormone (GH) dependent serum growth factors. There is some evidence that IGF inhibit GH release (negative feedback) in 3- to 24-h incubations of cultured rat adenohypophysial cells. We have used acutely dispersed noncultured rat adenohypophysial cells to study the dynamics of IGF on GH secretion. In this system both IGF-I and IGF-II (100 ng/mL) slightly, but significantly, decrease the cumulative GH released by human pancreas growth hormone releasing factor 1-40 (GRF) and the phosphodiesterase inhibitor 3-isobutyl-1-methyl xanthine. The inhibition is small (16%) and usually not statistically significant until 2 h of incubation. The inhibition with IGF is additive to that produced with low concentrations of somatostatin. The IGF also significantly decrease the rate of GH release in all time periods tested (0-1, 1-2, 2-3 h). In addition, the IGF decrease the quantity of [14C]leucine protein eluted at the position of labelled rat GH on Sephadex G75, which would include newly synthesized GH extracted from the cells. Thus we conclude that the decreased GH released may be due to an effect of IGF on both rate of release and on GH synthesis.     

胰岛素样生长因子IGF系统与鱼类性腺的研究进展

IGFs系统包含3个配体(IGF-1、IGF-2、IGF-3)、2个受体(IGF-1R、IGF-2R)和6个IGF结合蛋白(IGFBP).生殖和生长是生物体最基本的特征,两者既密切相关又相互区别,胰岛素样生长因子(IGFs)是生长轴和生殖轴相交联的关键因子.最近研究表明:鱼类性腺的发育及成熟伴随着细胞分化和组织生长,传统的生长因子IGF-1、IGF-2和最近发现的IGF-3,对鱼类性腺发挥着重要作用.本文重点介绍鱼类特有的配体IGF-3的结构,鱼类IGFs系统的信号通路及其与鱼类性腺的相关性研究进展.     

Insulin-like growth factors I and II are produced in the metanephros and are required for growth and development in vitro

The role(s) of one family of polypeptide growth factors in a developing organ system was examined. Renal anlagen (metanephroi) were surgically removed from 13-d-old rat embryos and grown in organ culture for up to 6 d. Over this period of time when placed in serum-free defined media, the metanephroi increased in size and morphologic complexity. Messenger RNAs for both insulin-like growth factors (IGFs), IGF I and IGF II, were present in the metanephroi. Immunoreactive IGF I and IGF II were produced by the renal anlagen and released into culture media. Levels were relatively constant during the 6 d in culture and averaged 3.5 X 10(-9) M IGF I and 8.3 X 10(-9) M IGF II in media removed from metanephroi after contact for 24 h. IGF binding protein activity was not detected in culture media. Growth and development of metanephroi in vitro was prevented by the addition of anti-IGF I or anti-IGF II antibodies to organ cultures. IGF II produced by metanephroi was active in an IGF II biological assay system and addition of anti-IGF II receptor antibodies to organ cultures prevented growth and development, consistent with the action of IGF II in metanephroi being mediated via the IGF II receptor. The data demonstrate production of both IGF I and IGF II by developing rat metanephroi in organ culture. Each of these peptides is necessary for growth and development of the renal anlage to take place in vitro. Our findings suggest that both IGF I and IGF II are produced within the developing metanephros in vivo and promote renal organogenesis.     

Localization of the insulin-like growth factor system in a rat model of heart failure induced by myocardial infarction.

Although cardiac effects of growth hormone (GH) and insulin-like growth factor (IGF)-I have been reported in experimental models of heart failure and in human dilated cardiomyopathy, the IGF system has not been comprehensively assessed in the failing heart. We therefore localized the IGF system in the left ventricle during congestive heart failure after myocardial infarction (MI) in the rat. The left anterior descending coronary artery was ligated in adult female Sprague-Dawley rats and hearts were examined after 6 months when congestive heart failure had developed. In situ hybridization histochemistry was used to localize mRNA for the components of the IGF system in the left ventricle of sham and congestive heart failure animals. We were able to detect changes in the spatial distribution of mRNA for IGF-I and IGF binding proteins 3, 4, 5, and 6 in the left ventricle during congestive heart failure after MI. IGF-I and the binding proteins were predominantly increased in the infarct/peri-infarct area of the left ventricle. Other components of the IGF system were indistinguishable from the low to undetectable levels in sham-operated rats. These results demonstrate that the IGF system is altered in the failing heart and suggest that the IGF system plays an important role in the response of the heart to MI and consequent failure.     

The IGF axis and programmed cell death.

Insulin-like growth factors (IGF) are mitogenic peptides that have been implicated as positive regulators of cellular proliferation. In recent years, several studies have suggested an additional role for the IGF axis in the regulation of apoptosis. Signalling through the IGF receptor has been shown to have a potent survival function and protect cells from a variety of apoptotic stimuli. The actions of IGF are regulated by a family of high-affinity IGF binding proteins (IGFBP), which sequester the IGF from the IGF receptor. However, there is some evidence that one of these binding proteins, IGFBP-3, may have its own pro-apoptotic effects that are independent of its ability to modulate IGF bioavailability. In addition, it has been suggested that the tumour suppressor p53, a crucial mediator of apoptosis in response to cellular stress, may elicit several of its apoptotic effects through manipulation of components of the IGF axis. This review summarizes what is currently known about the role of the IGF system in the regulation of apoptosis, highlighting its implications in the context of tumorigenesis.     

Pulse sensitivity of the luteinizing hormone beta promoter is determined by a negative feedback loop Involving early growth response-1 and Ngfi-A binding protein 1 and 2

The hypothalamic-pituitary-gonadal endocrine axis regulates reproduction through estrous phase-dependent release of the heterodimeric gonadotropic glycoprotein hormones, LH and FSH, from the gonadotropes of the anterior pituitary. Gonadotropin synthesis and release is dependent upon pulsatile stimulation by the hypothalamic neuropeptide GnRH. Alterations in pulse frequency and amplitude alter the relative levels of gonadotropin synthesis and release. The mechanism of interpretation of GnRH pulse frequency and amplitude by gonadotropes is not understood. We have examined gene expression in LbetaT2 gonadotropes under various pulse regimes in a cell perifusion system by microarray and identified 1127 genes activated by tonic or pulsatile GnRH. Distinct patterns of expression are associated with each pulse frequency, but the greatest changes occur at a 60-min or less interpulse interval. The immediate early gene mRNAs encoding early growth response (Egr)1 and Egr2, which activate the gonadotropin LH beta-subunit gene promoter, are stably induced at high pulse frequency. In contrast, mRNAs for the Egr corepressor genes Ngfi-A binding protein Nab1 and Nab2 are stably induced at low pulse frequency. We show that Ngfi-A binding protein members inhibit Egr-mediated frequency-dependent induction of the LH beta-subunit promoter. This pattern of expression suggests a model of pulse frequency detection that acts by suppressing activation by Egr family members at low frequency and allowing activation at sustained high-frequency pulses.     

Mannose 6-phosphate/insulin-like growth factor II receptor: distinct binding sites for mannose 6-phosphate and insulin-like growth factor II

Pentamannosyl phosphate substituted bovine serum albumin (PMP-BSA) and insulin like growth factor II (IGF II) bind specifically to immobilized mannose 6-phosphate/insulin like growth factor II receptor. An excess of IGF II inhibited binding of PMP-BSA by less than or equal to 20%, and an excess of PMP-BSA inhibited binding of IGF II by less than or equal to 10%. Polyclonal antibodies against the receptor purified from human liver inhibited preferentially the binding of PMP-BSA, and a monocloncal antibody 2C2 inhibited only the binding of IGF II to the receptor. Similar results were obtained for binding of PMP-BSA and IGF II to human skin fibroblasts. These results suggest that the binding sites for mannose 6-phosphate and IGF II reside in different portions of the receptor.     

Characteristics of binding of insulin-like growth factor (IGF)-I and IGF-II analogues to the type 1 IGF receptor determined by BIAcore analysis.

Insulin-like growth factor (IGF) binding to the type 1 IGF receptor (IGF1R) elicits mitogenic effects, promotion of differentiation and protection from apoptosis. This study has systematically measured IGF1R binding affinities of IGF-I, IGF-II and 14 IGF analogues to a recombinant high-affinity form of the IGF1R using BIAcore technology. The analogues assessed could be divided into two groups: (a) those designed to investigate binding of IGF-binding protein, which exhibited IGF1R-binding affinities similar to those of IGF-I or IGF-II; (b) those generated to probe IGF1R interactions with greatly reduced IGF1R-binding affinities. The relative binding affinities of IGF-I analogues and IGF-I for the IGF1R determined by BIAcore analysis agreed closely with existing data from receptor-binding assays using cells or tissue membranes, demonstrating that BIAcore technology is a powerful tool for measuring affinities of IGFs for IGF1R. In parallel studies, IGF1R-binding affinities were related to ability to protect against serum withdrawal-induced apoptosis in three different assays including Hoechst 33258 staining, cell survival, and DNA fragmentation assays using the rat pheochromocytoma cell line, PC12. In this model system, IGF-I and IGF-II at low nanomolar concentrations are able to prevent apoptosis completely. We conclude that ability to protect against apoptosis is directly related to ability to bind the IGF1R.     

Characterization of the protein which binds insulin-like growth factor in human serum

The binding of the 125I-labelled insulin-like growth factors I and II (125I-IGF I and 125I-IGF II) to the high-molecular-mass binding protein of human serum was characterized. With diluted human serum both growth factors showed optimal specific binding at 4 degrees C and pH 5-6. When 0.1% Triton X-100 was present in the incubation buffer an increase in the affinity of the IGF-binding protein was induced, which produced an enhanced binding of IGF I and IGF II. Competition experiments with various peptide hormones revealed that the native IGF-binding protein complex binds both the IGF I and IGF II with high specificity. Analysis of binding data according to the method of Scatchard resulted in linear plots for IGF I and IGF II respectively, indicating that in human serum only a single class of non-interacting binding sites is present. At optimal binding conditions the dissociation constants were determined to be 0.28 x 10(-9) M for IGF I binding and 0.66 x 10(-9) M for IGF II. Human serum was gel-filtered on Sepharose CL-6B at neutral pH and the eluate was assayed for binding activity with both IGF I and IGF II. One peak with an apparent molecular mass of 175 kDa and a Stokes radius of 4.8 nm was determined for both growth factors. Thus, our data suggest that human serum contains one class of high-molecular-mass binding protein with comparable binding characteristics for IGF I and IGF II.     

胰岛素样生长因子系统与肝癌

肝癌是最常见的人类恶性肿瘤之一.肝癌的发生是一个多因素交互作用逐级发生的过程.包括乙肝、丙肝病毒感染、酗酒、肝硬化等都可能导致肝癌.研究表明,胰岛素样生长因子 (IGF)系统由IGF1、IGF2及其受体（IGF1R、IGF2R）和胰岛素样生长因子结合蛋白（IGFBP）组成,在正常以及恶性肝细胞的增殖、分化和转移过程中起重要作用.本综述回顾近年来胰岛素样生长因子系统与肝癌发生发展的机理研究,及其在肝癌治疗中可能的临床应用前景的研究进展,并对目前该研究领域存在的主要问题和未来的研究展望提出了看法.