Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for analysis of endogenous steroids in the luteal phase and early pregnancy in dogs: a pilot study.

BACKGROUND Blood samples from dogs are often limited in volume, only allowing few steroids to be quantified with immunoassays. In addition, immunoassays may be compromised by interferences such as anti-reagent antibodies. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) can be used for the simultaneous quantitation of several steroids. This has not been described in dogs before. OBJECTIVES The aims were to use LC-MS/MS to study steroid profiles in early pregnancy and luteal phase in dogs, and to determine if differences exist between pregnant (P) and nonpregnant (NP) dogs. METHODS Nine female dogs were included, 4 during a NP luteal phase, 4 during a P luteal phase, and one during one NP and one P luteal phase. Blood samples were collected around the time of the LH surge (Day 0) and on Day 26. Serum was analyzed for 5 classes of steroids, including glucocorticoids, androgens, estrogens, pregnanes, and progestins, using LC-MS/MS methods. RESULTS The concentration of progesterone was significantly higher on Day 26 in P than in NP bitches. Distribution of concentrations of glucocorticoids, androgens, estrogens, or pregnanes in P and NP dogs were not statistically different. The predominating glucocorticoid was cortisol, and dihydroepiandrosterone (DHEA) was the predominating androgen. Concentration of estrone was comparable to oestradiol, whereas concentrations of pregnenolone were higher than those of 17-OH pregnenolone. CONCLUSIONS Only concentration of progesterone differed between P and NP bitches, being significantly higher on Day 26 in P than in NP bitches. LC-MS/MS offers interesting possibilities for studies of canine reproductive endocrinology.

[1]  B. Hoffmann,et al.  Investigations on hormonal changes around parturition in the dog and the occurrence of pregnancy-specific non conjugated oestrogens. , 2009, Experimental and clinical endocrinology.

[2]  P. Concannon.  Reproductive cycles of the domestic bitch. , 2011, Animal reproduction science.

[3]  C. Welt,et al.  Simultaneous Measurement of Thirteen Steroid Hormones in Women with Polycystic Ovary Syndrome and Control Women Using Liquid Chromatography-Tandem Mass Spectrometry , 2014, PloS one.

[4]  F. Stanczyk,et al.  Advantages and challenges of mass spectrometry assays for steroid hormones , 2010, The Journal of Steroid Biochemistry and Molecular Biology.

[5]  A. Rockwood,et al.  Cortisol and cortisone analysis in serum and plasma by atmospheric pressure photoionization tandem mass spectrometry. , 2004, Clinical biochemistry.

[6]  A. Rockwood,et al.  Liquid chromatography-tandem mass spectrometry assay for androstenedione, dehydroepiandrosterone, and testosterone with pediatric and adult reference intervals. , 2010, Clinical chemistry.

[7]  J. Bergquist,et al.  High-sensitivity tandem mass spectrometry assay for serum estrone and estradiol. , 2008, American journal of clinical pathology.

[8]  J. West,et al.  Steroid hormone concentration profiles in healthy intact and neutered dogs before and after cosyntropin administration. , 2003, Domestic animal endocrinology.

[9]  M. Isaksson,et al.  Canine herpesvirus during pregnancy and non-pregnant luteal phase. , 2012, Reproduction in domestic animals = Zuchthygiene.

[10]  P. Concannon,et al.  Changes in LH, progesterone and sexual behavior associated with preovulatory luteinization in the bitch. , 1977, Biology of reproduction.

[11]  P. Chakraborty.  Reproductive hormone concentrations during estrus, pregnancy, and pseudopregnancy in the Labrador bitch☆☆☆ , 1987 .

[12]  J. Kratzsch,et al.  Preclinical challenges in steroid analysis of human samples , 2010, The Journal of Steroid Biochemistry and Molecular Biology.

[13]  G. Hortin,et al.  Effect of blood collection tubes on total triiodothyronine and other laboratory assays. , 2005, Clinical chemistry.

[14]  A. Rockwood,et al.  Development and performance evaluation of a tandem mass spectrometry assay for 4 adrenal steroids. , 2006, Clinical chemistry.

[15]  H. Field.  Tandem mass spectrometry in hormone measurement. , 2013, Methods in molecular biology.

[16]  P. Concannon,et al.  Serum androstenedione and testosterone concentrations during pregnancy and nonpregnant cycles in dogs. , 1985, Biology of reproduction.