Hypoglycosylated hFSH Has Greater Bioactivity Than Fully Glycosylated Recombinant hFSH in Human Granulosa Cells

J Clin Endocrinol Metab. 2015 Jun;100(6):E852-60. doi: 10.1210/jc.2015-1317. Epub 2015 Apr 27.

Abstract

Context: Previous studies suggest that aging in women is associated with a reduction in hypoglycosylated forms of FSH.

Objective: Experiments were performed to determine whether glycosylation of the FSHβ subunit modulates the biological activity of FSH in human granulosa cells.

Design and setting: Recombinant human FSH (hFSH) derived from GH3 pituitary cells was purified into fractions containing hypoglycosylated hFSH(21/18) and fully glycosylated hFSH(24). The response to FSH glycoforms was evaluated using the well-characterized, FSH-responsive human granulosa cell line, KGN at an academic medical center.

Interventions: Granulosa cells were treated with increasing concentrations of fully- or hypoglycosylated FSH glycoforms for periods up to 48 hours.

Main outcome measure(s): The main outcomes were indices of cAMP-dependent cell signaling and estrogen and progesterone synthesis.

Results: We observed that hypoglycosylated FSH(21/18) was significantly more effective than fully glycosylated FSH(24) at stimulating cAMP accumulation, protein kinase A (PKA) activity, and cAMP response element binding protein (CREB) (S133) phosphorylation. FSH(21/18) was also much more effective than hFSH(24) on the stimulation CREB-response element-mediated transcription, expression of aromatase and STAR proteins, and synthesis of estrogen and progesterone. Adenoviral-mediated expression of the endogenous inhibitor of PKA, inhibited FSH(21/18)- and FSH(24)-stimulated CREB phosphorylation, and steroidogenesis.

Conclusions: Hypoglycosylated FSH(21/18) has greater bioactivity than fully glycosylated hFSH(24), suggesting that age-dependent decreases in hypoglycosylated hFSH contribute to reduced ovarian responsiveness. Hypoglycosylated FSH may be useful in follicle stimulation protocols for older patients using assisted reproduction technologies.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Carbohydrate Sequence
  • Cells, Cultured
  • Cyclic AMP / metabolism
  • Cyclic AMP Response Element-Binding Protein / metabolism
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Enzyme Activation / drug effects
  • Female
  • Follicle Stimulating Hormone, Human / metabolism*
  • Follicle Stimulating Hormone, Human / pharmacology*
  • Glycosylation
  • Granulosa Cells / drug effects*
  • Granulosa Cells / metabolism
  • Humans
  • Phosphorylation
  • Protein Isoforms
  • Recombinant Proteins / metabolism*
  • Recombinant Proteins / pharmacology*

Substances

  • Cyclic AMP Response Element-Binding Protein
  • Follicle Stimulating Hormone, Human
  • Protein Isoforms
  • Recombinant Proteins
  • Cyclic AMP
  • Cyclic AMP-Dependent Protein Kinases