Transcription profile of the insulin-like growth factor signaling pathway during human ovarian follicular development
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The IGF signaling cascade exerts important regulatory functions in human ovarian folliculogenesis. The scope of this study was to evaluate the transcription profile of insulin-like growth factor (IGF) genes during human ovarian follicle development and to analyze follicle fluid levels of key IGF proteins.
Gene expression profiling was performed with microarray gene analysis. The analysis was assessed from ovarian follicles and granulosa cells (GCs) obtained from isolated stage-specific human ovarian follicles, including preantral follicles, small antral follicles, and preovulatory follicles. Numerous genes involved in the IGF signaling pathway was evaluated and key genes were validated by qPCR from GCs. Protein levels of various IGF components of human follicular fluid (FF) were measured by ELISA and time-resolved immunofluorometric assays (TRIFMA).
The gene expression levels of PAPPA, IGF2, IGF receptors and intracellular IGF-activated genes increased with increasing follicle size. This was especially prominent in the late preovulatory stage where IGF2 expression peaked. Protein levels of intact IGF binding protein-4 decreased significantly in FF from large preovulatory follicles compared with small antral follicles concomitant with higher protein levels of PAPP-A. The IGF modulators IGF-2 receptor, IGFBPs, stanniocalcins, and IGF-2 mRNA binding proteins were all observed to be expressed in the different follicle stages.
This study confirms and highlights the importance of PAPP-A regulating bioactive IGF levels throughout folliculogenesis and especially for the high rate of granulosa cell proliferation and expression of key ovarian hormones important in the last part of the follicular phase of the menstrual cycle.
KeywordsIGF system IGFBPs PAPP-A Stanniocalcins Human ovarian follicles
We acknowledge the Core Facility for helping with fine microarray analysis. Furthermore, we are thankful for the work performed at the fertility clinics in regard to collecting the granulosa cells and follicular fluids from IVF patients. Finally, we thank Pernille Rimmer Noer from the Department of Molecular Biology and Genetics at University of Aarhus for technical assistance.
The financial support from The Novo Nordisk Foundation, the Lundbeck Foundation, and Gangstedfonden is gratefully acknowledged.
Compliance with ethical standards
Conflict of interest
The authors declare no conflict of interest.
- 10.Laursen LS, Overgaard MT, Söe R, Boldt HB, Sottrup-jensen L, Giudice LC, et al. Pregnancy-associated plasma protein-a (PAPP-A) cleaves insulin-like growth factor binding protein (IGFBP)-5 independent of IGF: implications for the mechanism of IGFBP-4 proteolysis by PAPP-A. FEBS Lett. 2001;504:36–40. https://doi.org/10.1016/S0014-5793(01)02760-0.CrossRefGoogle Scholar
- 15.Law NC, White MF, Hunzicker-Dunn ME. G protein-coupled receptors (GPCRs) that signal via protein kinase a (PKA) cross-talk at insulin receptor substrate 1 (IRS1) to activate the phosphatidylinositol 3-kinase (PI3K)/AKT pathway. J Biol Chem. 2016;291:27160–9. https://doi.org/10.1074/jbc.M116.763235.CrossRefGoogle Scholar
- 19.Dessel THJ, Chandrasekher Y, Yap OW, Lee PD, Hintz RL, Faessen GH, et al. Serum and follicular fluid levels of insulin-like growth factor I (IGF-I), IGF-II, and IGF-binding protein-1 and -3 during the normal menstrual cycle. J Clin Endocrinol Metab. 1996;81:1224–31.Google Scholar
- 20.Geisthovel F, Moretti-Rojas I, Asch RH, Rojas FJ. Expression of insulin-like growth factor-II (IGF-II) messenger ribonucleic acid (mRNA), but not IGF-I mRNA, in human preovulatory granulosa cells. Hum Reprod. 1989;4:899–902. https://doi.org/10.1093/oxfordjournals.humrep.a137007.CrossRefGoogle Scholar
- 31.Bøtkjær JA, Jeppesen JV, Wissing ML, Kløverpris S, Oxvig C, Mason JI, et al. Pregnancy-associated plasma protein a in human ovarian follicles and its association with intrafollicular hormone levels. Fertil Steril. 2015;104:1294–301. https://doi.org/10.1016/j.fertnstert.2015.07.1152.CrossRefGoogle Scholar
- 32.Giorgetti C, Vanden Meerschaut F, De Roo C, Saunier O, Quarello E, Hairion D, et al. Multivariate analysis identifies the estradiol level at ovulation triggering as an independent predictor of the first trimester pregnancy-associated plasma protein-A level in IVF/ICSI pregnancies. Hum Reprod. 2013;28:2636–42. https://doi.org/10.1093/humrep/det295.CrossRefGoogle Scholar
- 34.Dugoff L, Hobbins JC, Malone FD, Porter TF, Luthy D, Comstock CH, et al. First-trimester maternal serum PAPP-A and free-beta subunit human chorionic gonadotropin concentrations and nuchal translucency are associated with obstetric complications: a population-based screening study (the FASTER trial). Am J Obstet Gynecol. 2004;191:1446–51. https://doi.org/10.1016/j.ajog.2004.06.052.CrossRefGoogle Scholar
- 38.Jepsen MR, Kløverpris S, Bøtkjær JA, Wissing ML, Andersen CY, Oxvig C. The proteolytic activity of pregnancy-associated plasma protein-A is potentially regulated by stanniocalcin-1 and -2 during human ovarian follicle development. Hum Reprod. 2016;31:866–74. https://doi.org/10.1093/humrep/dew013.CrossRefGoogle Scholar
- 39.Overgaard MT, Haaning J, Boldt HB, Olsen IM, Laursen LS, Christiansen M, et al. Expression of recombinant human pregnancy-associated plasma protein-A and identification of the proform of eosinophil major basic protein as its physiological inhibitor. J Biol Chem. 2000;275:31128–33. https://doi.org/10.1074/jbc.M001384200.CrossRefGoogle Scholar
- 40.Oxvig C, Sand O, Kristensen T, Gleich GJ, Sottrup-Jensen L. Circulating human pregnancy-associated plasma protein-A is disulfide-bridged to the proform of eosinophil major basic protein. J Biol Chem. 1993;268:12243–6.Google Scholar
- 43.Rosendahl M, Andersen CY, Ernst E, Westergaard LG, Rasmussen PE, Loft A, et al. Ovarian function after removal of an entire ovary for cryopreservation of pieces of cortex prior to gonadotoxic treatment: a follow-up study. Hum Reprod. 2008;23:2475–83. https://doi.org/10.1093/humrep/den248.CrossRefGoogle Scholar
- 45.Jeppesen JV, Anderson RA, Kelsey TW, Christiansen SL, Kristensen SG, Jayaprakasan K, et al. Which follicles make the most anti-Mullerian hormone in humans? Evidence for an abrupt decline in AMH production at the time of follicle selection. Mol Hum Reprod. 2013;19:519–27. https://doi.org/10.1093/molehr/gat024.CrossRefGoogle Scholar
- 47.Wissing ML, Kristensen SG, Andersen CY, Mikkelsen AL, Høst T, Borup R, et al. Identification of new ovulation-related genes in humans by comparing the transcriptome of granulosa cells before and after ovulation triggering in the same controlled ovarian stimulation cycle. Hum Reprod. 2014;29:997–1010. https://doi.org/10.1093/humrep/deu008.CrossRefGoogle Scholar
- 48.Borgbo T, Povlsen BB, Andersen CY, Borup R, Humaidan P, Grøndahl ML. Comparison of gene expression profiles in granulosa and cumulus cells after ovulation induction with either human chorionic gonadotropin or a gonadotropin-releasing hormone agonist trigger. Fertil Steril. 2013;100:994–1001. https://doi.org/10.1016/j.fertnstert.2013.05.038.CrossRefGoogle Scholar
- 50.Bøtkjær JA, Borgbo T, Kløverpris S, Noer PR, Oxvig C, Andersen CY. Effect of pregnancy-associated plasma protein-A (PAPP-A) single-nucleotide polymorphisms on the level and activity of PAPP-A and the hormone profile in fluid from normal human small antral follicles. Fertil Steril. 2016;106:1778–1786.e8. https://doi.org/10.1016/j.fertnstert.2016.09.008.CrossRefGoogle Scholar
- 54.Stamatas GN, Wu J, PAPP-As A, Mirmirani P, McCormick TS, Cooper KD, et al. An analysis of gene expression data involving examination of signaling pathways activation reveals new insights into the mechanism of action of minoxidil topical foam in men with androgenetic alopecia. Cell Cycle. 2017;16:1578–84. https://doi.org/10.1080/15384101.2017.1327492.CrossRefGoogle Scholar
- 56.Conover CA, Faessen GF, Ilg KE, Chandrasekher YA, Christiansen M, Overgaard MT, et al. Pregnancy-associated plasma protein-A is the insulin-like growth factor binding protein-4 protease secreted by human ovarian granulosa cells and is a marker of dominant follicle selection and the corpus luteum. Endocrinology. 2001;142:2155. https://doi.org/10.1210/endo.142.5.8286.CrossRefGoogle Scholar
- 62.El-Roeiy A, Chen X, Roberts VJ, Shimasakai S, Ling N, LeRoith D, et al. Expression of the genes encoding the insulin-like growth factors (IGF-I and II), the IGF and insulin receptors, and IGF-binding proteins-1–6 and the localization of their gene products in normal and polycystic ovary syndrome ovaries. J Clin Endocrinol Metab. 1994;78:1488–96. https://doi.org/10.1210/jcem.78.6.7515389.Google Scholar