Endothelial vasodilator production by uterine and systemic arteries. VI. Ovarian and pregnancy effects on eNOS and NO_x

Abstract

Normal pregnancy and the follicular phase of the ovarian cycle are both estrogen-dominated physiological states that are characterized by elevations in uterine blood flow and endothelial nitric oxide synthase (eNOS) protein expression in the uterine artery (UA) endothelium. It is unknown if elevations in mRNA level account for the changes in protein or eNOS activity. We tested the hypothesis that pregnancy and the follicular phase are associated with increases in eNOS mRNA and the consequent elevated expression of eNOS protein results in increased circulating nitric oxide (NO) levels. UA were obtained from pregnant (PREG; n = 8; 110-130 days gestation; term = 145 +/- 3 days), nonpregnant luteal (LUT; n = 6), nonpregnant follicular (FOL; n = 6), and nonpregnant ovariectomized (OVEX; n = 6) sheep. Circulating NO levels were analyzed as total NO(2)-NO(3) (NO(x)). Western analysis performed on UA endothelial-isolated proteins demonstrated that eNOS protein levels were OVEX = LUT < or = FOL < PREG (P < 0.05), whereas eNOS mRNA expression (RT-PCR) in UA endothelial cells obtained by limited collagenase digestion was OVEX < LUT < FOL < PREG (P < 0.05). Pregnancy dramatically elevated eNOS protein (4.1- to 6.9-fold) and mRNA (2.4- to 6.9-fold) over LUT controls (P < 0.01). Circulating NO(x) levels were not altered by ovariectomy or the ovarian cycle but were elevated from 4.4 +/- 1.1 microM in LUT to 12 +/- 4, 22 +/- 3, and 41 +/- 3 microM at 110, 120, and 130 days gestation (P < 0.01). Systemic NO(x) levels in singleton (12.5 +/- 1.6 microM) were less (P < 0.01) than in multiple (twin 27.6 +/- 6.5 microM; triplet = 46 +/- 10 microM) pregnancies. Therefore, the follicular phase and, to a much greater extent, pregnancy are associated with elevations in UA endothelium-derived eNOS expression, although significant increases in systemic NO(x) levels were only observed in the PREG group (multiple > singleton). Thus, although UA endothelial increases in eNOS protein and mRNA levels are associated with high estrogen states, increases in local UA NO production may require additional eNOS protein activation to play its important role in the maintenance of uterine blood flow in pregnancy.

Litter-size-dependent intrauterine growth restriction in sheep

“…Measurements of NOx, defined as total plasma NO 2 and NO 3 in pregnant ewes (Magness et al, 2001), revealed that ewes pregnant with twins and triplets had more NOx than ewes carrying a single foetus.…”

Transcriptional regulation of endothelial nitric oxide synthase expression in uterine artery endothelial cells by c-Jun/AP-1

“…The importance of the NOS3-NO system during pregnancy has recently been reinforced by the findings that uterine artery remodeling is impaired and litter size is reduced in NOS3-null compared to wild-type mice (van der Heijden et al, 2005). In vivo, there is solid evidence showing that uterine artery endothelial NOS3 protein is increased during pregnancy Magness et al, 2001) and in the follicular phase of the estrous cycle or by estrogen replacement therapy in association with elevated circulating estrogen levels (Chen et al, 2006;Rosenfeld et al, 2003;Rupnow et al, 2001;. Available data also suggest that uterine artery endothelial NOS3 expression can be regulated at the level of transcription (Rosenfeld et al, 2003).…”

Simultaneous imaging of [Ca2+]iand intracellular NO production in freshly isolated uterine artery endothelial cells: effects of ovarian cycle and pregnancy

“…During pregnancy, the uterine artery (UA) shows increased vasodilation in response to a number of agonists, including ATP, which in turn contributes to the increase in uterine blood flow (UBF) necessary to meet the needs of the growing fetus (3,(32)(33)(34). The follicular phase of the ovarian cycle is partially characterized by elevation in UBF, and both pregnancy and follicular phase are associated with both increased estrogen levels and eNOS expression in UA endothelium (UAE) (34), as well as estrogen enhancement of flow-induced vasodilation (19,20).…”

“…The follicular phase of the ovarian cycle is partially characterized by elevation in UBF, and both pregnancy and follicular phase are associated with both increased estrogen levels and eNOS expression in UA endothelium (UAE) (34), as well as estrogen enhancement of flow-induced vasodilation (19,20). To date, only indirect measures have implied an increase in UA NO production (33,34,46), and there has been no direct demonstration of NO production in UA or evaluation of whether NO production increases in parallel with increased eNOS expression in these physiological states. This is not only important for the reasons stated above but also because more recent studies in sheep have implied that the pregnancyassociated increase in the endothelium-dependent relaxation of the UA via NO release may not only occur by way of increases in eNOS expression but also by way of reprogramming of kinase signaling (3).…”