The proteolytic activity of pregnancy-associated plasma protein-A is potentially regulated by stanniocalcin-1 and -2 during human ovarian follicle development

Abstract

Study question: Is the proteolytic activity of pregnancy-associated plasma protein-A (PAPP-A) regulated by the stanniocalcins (STC1 and STC2) during human follicle maturation?

Summary answer: The STCs and PAPP-A show similar expression by immunohistochemistry in developing follicles, and regulation of PAPP-A proteolytic activity is suggested by the identification of inhibited protein complexes between PAPP-A and STC1 or STC2 in human follicular fluid (FF).

What is known already: The insulin-like growth factor (IGF)-regulating proteinase PAPP-A is secreted by the granulosa cells of estrogen-dominant follicles and is involved in follicle growth. STC1 and STC2 have recently been identified as novel PAPP-A inhibitors, and their expression in non-human mammalian ovaries has previously been observed.

Study design, size, duration: The proteolytic activity of PAPP-A in human follicular fluid was assessed, and the interaction between PAPP-A and the STCs in human ovarian tissues and follicular fluid was analyzed using immunoassays. From 21 women, matched pairs of follicular fluid were obtained from one follicle just prior to final maturation of follicles with human chorionic gonadotrophin (hCG), and from another follicle in connection with oocyte aspiration after hCG treatment. Ovarian tissues were obtained from women having one ovary removed for fertility preservation by cryopreservation prior to gonadotoxic treatment.

Participants/materials, setting, methods: The concentration and activity of PAPP-A were determined in all samples of follicular fluid. Furthermore, to investigate PAPP-A regulation during follicle development, immunohistochemical staining of PAPP-A, STC1, and STC2 was performed on pre-antral and antral human follicles. To attempt the demonstration of native complexes between PAPP-A and the STCs, immunoprecipitation from a pool of human follicular fluid was performed.

Main results and the role of chance: The concentration of PAPP-A antigen in follicular fluid increased upon stimulation of ovulation with hCG (P < 0.02), but at the same time, PAPP-A activity was decreased. PAPP-A, STC1, and STC2 were localized together in primordial, late primary, and antral follicles, indicating that complex formation is possible in ovarian tissue. Covalent PAPP-A:STC2 and non-covalent PAPP-A:STC1 complexes were immunoprecipitated from follicular fluid, documenting for the first time native inhibited complexes between PAPP-A and the STCs.

Limitations, reasons for caution: We have demonstrated the presence of native complexes between PAPP-A and the STCs in the human ovary, indicating STC-mediated PAPP-A proteolytic inhibition. Further investigation is required to extend this principle to other tissues.

Wider implications of the findings: Our data suggest that the STCs contribute to PAPP-A regulation during folliculogenesis and support a general model in which STC1 and STC2 are regulators of mammalian IGF activity through inhibition of PAPP-A. We suggest that future functional studies take both PAPP-A and the STCs into consideration.

Study funding/competing interests: This work was supported by grants from the Novo Nordisk Foundation, and the Danish Council for Independent Research. No competing interests declared.

Keywords: follicle development; human follicles; insulin-like growth factor system; pregnancy-associated plasma protein-A; stanniocalcin.