In the present study the concentration of relaxin in peripheral blood plasma was assessed during canine pregnancy for its suitability as a pregnancy indicator, using a newly developed relaxin enzyme immunoassay. A significant relaxin increase was found in pregnancy at day 24 after ovulation. However, this relaxin increase did not correlate either with litter size or with body weight of the bitch. Induction of abortion with prostaglandin F2 alpha resulted in reduced peripheral relaxin levels, suggesting a damage of the placenta due to this medical intervention. Thus, the results confirm that relaxin, which is produced by the placenta, is a useful marker for early pregnancy diagnosis in the bitch. Relaxin measurement is recommended for detection of pregnancy either alone, or as supplement of ultrasonographic findings.
Relaxin was identified in 1926 in the serum of pregnant guinea pigs as an entity that caused relaxation of the pubic ligament. Relaxin is now known to have several important effects during pregnancy and outside of pregnancy however the activity noted with its discovery, softening of the pubic ligament to allow for delivery, is not found in humans and non-human primates. Relaxin is part of a family of peptide hormones that consists of seven members including relaxin-1, relaxin-2, relaxin-3, insulin-like peptide (INSL) 3, INSL4, INSL5 and INSL6. The seven member family is part of the larger insulin superfamily which also includes insulin and insulin-like growth factors 1 and 2. Relaxin family members function in reproduction, the cardiovascular system, metabolism, as a neuropeptide, and as an incretin.
Relaxin family peptides, similar to insulin, are synthesized as preprohormones composed of a signal sequence, a B-chain, C-chain, and carboxy terminal A-chain domains. Posttranslational proteolytic cleavage removes first the signal peptide followed by the C domain peptide. The mature hormone is a heterodimeric protein consisting of the A and B chains linked by two disulfide bonds and one intra-A-chain disulfide bond. Relaxin family peptides share little overall sequence homology but are grouped together based on a conserved RxxxRxxI/V motif in the B-chain, the conserved cysteines that form the three disulfide bonds across all the family members and a common similar three dimensional structure. The relaxin family peptides bind receptors belonging to the G-protein coupled receptor family (GPCR) known as relaxin/insulin family peptide (RXFP) receptors. This is in contrast to insulin which utilizes receptor tyrosine kinases.
Relaxin has a well-characterized dipsogenic effect in the brain after iv or icv administration via direct actions on the circumventricular organs (subfornical organ/OVLT), which lie along the rostral border of the third ventricle outside the blood-brain barrier and have neural axonal projections to other regions (MPO, SON, and PVN) that subserve vasopressin secretion and water drinking. Relaxin is thought to activate these brain pathways via angiotensin II signaling. In the pregnant rat, injection of a relaxin monoclonal antibody into the brain negates the normal increase in drinking observed during the second half of pregnancy. Exogenous relaxin effects in the rat also vary with time of injection during the light–dark cycle, whereby dose-dependent water drinking caused by relaxin is maximal at night compared with that during the day. This effect, while perhaps predictable in nocturnal animals, may suggest an involvement of relaxin in this and other circadian-based functions.
Blood relaxin concentrations are undetectable until 85 days of gestation but remain above basal thereafter. A first peak (>20 ng/mL) is detected at 3.5 months of gestation. Relaxin concentrations then decrease between 5.5 and 7.5 months, followed by a steady increase until parturition.8 Relaxin is a pregnancy-specific hormone. Its source in South American camelids (SACs) is unknown, but large luteal cells of the mature CL and uterine luminal epithelial cells were identified as the source of relaxin in pregnant dromedary camels.15 Pregnancy diagnosis based on blood relaxin concentration seems possible and accurate 85 days after mating.8 However, no commercial tests are available, and measurement of relaxin remains experimental.
Relaxin levels are low to undetectable in the circulation of nonpregnant animals. In pregnancy, relaxin circulates in the blood of all mammals, but the pattern of relaxin secretion differs markedly across species. For example, in pregnant pigs, relaxin is detectable at low levels in the circulation and marked by a large prepartum surge (over 100 ng/mL) just prior to parturition. In rodents, serum relaxin levels are relatively constant until the end of pregnancy, when there is a marked surge in relaxin secretion just before parturition. In the mare, serum relaxin rises in the first trimester of pregnancy and remains elevated until foaling. Following delivery of the placenta, relaxin levels are undetectable in the circulation of the postpartum mare suggesting that the placenta is the main source of circulating relaxin in the horse. In contrast, in humans and other primates, serum relaxin is highest in early pregnancy and peaks (1 ng/mL) near the end of the first trimester, after which relaxin levels decline and remain low for the duration of pregnancy. In pregnant women, luteal relaxin is the source of systemic relaxin as illustrated in ovum-donation pregnancies in which circulating relaxin is not detectable in the absence of functioning ovaries. In contrast, human placental/decidual relaxin is thought to act locally in a paracrine manner, playing an important role in controlling fetal membrane remodeling at the time of parturition.
