OBJECTIVE: To elucidate the natural course of circulating insulin-like peptide 3 (INSL3) levels according to puberty as well as its relation to other reproductive hormones.

DESIGN: Population-based cohort study.

SETTING: Not applicable.

PATIENT(S): Healthy peripubertal girls (n = 10) examined every 6 months; total number of examinations was 84; median (range) per girl: 9 (4-10), including staging of pubertal breast development and blood samples.

INTERVENTION(S): None.

MAIN OUTCOME MEASURE(S): Serum levels of INSL3, inhibin B, E2, antimüllerian hormone, LH, and FSH (validated immunoassays), and T and androstenedione (liquid chromatography-tandem mass spectrometry).

RESULT(S): Serum levels of INSL3 varied considerably between girls (range, 0.01-0.27 ng/mL) and within each girl as puberty progressed; intraindividual variation, median (range) 102% (65%-143%). Insulin-like peptide 3 increased in late puberty (B1 to B4+B5); geometric mean 0.03 ng/mL to 0.15 ng/mL. Insulin-like peptide 3 levels reflected markers of large follicles (T, androstendione, inhibin B, and E2) better than markers of small follicles (antimüllerian hormone), and INSL3 staining was localized in theca interna cells of antral follicles.

CONCLUSION(S): Insulin-like peptide 3 increased in late puberty, albeit inter- and intraindividual variations were substantial. Immunohistochemistry and intraindividual variation, as well as relations to other ovarian hormones, reveal that INSL3 in girls is a unique and specific marker of theca cells surrounding antral follicles. The potential clinical use of INSL3 for evaluation of ovarian function in girls remains to be elucidated.