The dynamic steroid landscape of equine pregnancy mapped by mass spectrometry

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) allowed comprehensive analysis of various steroids detectable in plasma throughout equine gestation. Mares (n=9) were bled serially until they foaled. Certain steroids dominated the profile at different stages of gestation, clearly defining key physiological and developmental transitions. The period (weeks 6-20) coincident with equine chorionic gonadotropic (eCG) stimulation of primary corpora lutea and subsequent formation of secondary luteal structures was defined by increased progesterone, 17OH-progesterone and androstenedione, all Δ4 steroids. The 5α-reduced metabolite of progesterone, dihydroprogesterone (DHP) paralleled progesterone secretion at less than half the concentration until week 12 of gestation when progesterone began to decline but DHP concentrations continued to increase. DHP exceeded progesterone concentrations by week 16, clearly defining the luteo-placental shift in pregnane synthesis from primarily ovarian to primarily placental. The period corresponding to the growth of fetal gonads was defined by increasing dehydroepiandrosterone and pregnenolone (Δ5 steroids) concentrations from week 14, peaking at week 34 and declining to term. Metabolites of DHP (including allopregnanolone) dominated the steroid profile in late gestation, some exceeding DHP by weeks 13 or 14 and near term by almost tenfold. Thus Δ4 steroids dominated during ovarian stimulation by eCG, inversion of the ratio of progesterone: DHP (increasing 5α-pregnanes) marked the luteo-placental shift, Δ5 steroids defined fetal gonadal growth and 5α-reduced metabolites of DHP dominated the steroid profile in mid- to late-gestation. Comprehensive LC-MS/MS steroid analysis provides opportunities to better monitor the physiology and the progress of equine pregnancies, including fetal development.