A study of the relationship between C-type natriuretic peptide and pregnancy in sheep and red deer

Abstract

C-type natriuretic peptide (CNP) is a signaling molecule with important roles in mammalian growth and vascular function. Unexpectedly high plasma CNP concentrations have been reported in late gestation ewes, suggesting a possible role during pregnancy. However, the temporal pattern of CNP in maternal circulation, site/s of production and function during pregnancy have not been identified. Consequently, the aim of this PhD project was to address these unexplored aspects of CNP biology in order to improve our understanding of CNP’s role in pregnancy.

To characterise the temporal pattern of maternal circulating CNP during pregnancy, a longitudinal survey was conducted in two representative species of ruminant: sheep and red deer. Plasma concentrations of both CNP and a presumed bio-inactive fragment of its precursor (NTproCNP) increased in twin-bearing ewes at 40-50 days of gestation with peak values attained (CNP 31 ± 5 pmol l⁻¹, NTproCNP 270 ± 16 pmol l-1) at about day 120. A similar temporal pattern was evident in red deer hinds. In pregnant ewes studied at day 120, there was a positive relationship between maternal CNP forms and fetal number (P < 0.01). Collectively, these results strongly implicated the uteroplacental unit as the major source of CNP during pregnancy. Therefore, studies were conducted to determine the relative contribution of uterine and placental tissues to circulating CNP concentration. The concentration of both CNP forms in the placenta exceeded that in intercaruncular uterine tissue throughout pregnancy (P < 0.05), with the highest concentrations measured in the fetal cotyledon. Immunohistochemistry revealed staining of CNP and NTproCNP around placental blood vessels and in trophoblast binucleate cells (BNC), identifying these cells as a probable source of maternal circulating CNP during pregnancy.

Having identified the source of CNP during pregnancy, further studies were undertaken to test the hypothesis that CNP production by the ovine uteroplacental unit is homeostatically regulated by nutritional status, using models of nutrient restriction (3-day fast) or nutrient abundance (caloric loading or mid-pregnancy shearing). Maternal circulating NTproCNP concentration, expressed as a percentage of pre-fasting concentration, was significantly elevated in ewes fasted in late pregnancy compared with controls (117 ± 5.1 % vs 107 ± 3.5 %, P < 0.05). In contrast, there was no effect of nutrient abundance on the concentration of either CNP form, suggesting that CNP production is selectively upregulated during situations of fetal restriction.

CNP’s localisation to the BNC and high circulating concentrations in the pregnant ruminant suggest a novel endocrine role for the peptide during pregnancy, in addition to likely paracrine actions within the placental vasculature. The strong relationship between maternal circulating CNP and placental peptide production identified in this thesis makes the pregnant ewe a unique in vivo model for studying the regulation of CNP during pregnancy, and may reveal potential applications for this peptide as a marker of maternal and/or fetal health and wellbeing in both human medicine and animal production.