AMPK activation in pregnant human myometrial arteries from high-altitude and intrauterine growth-restricted pregnancies

Abstract

High-altitude (>2,500 m) residence increases the incidence of intrauterine growth restriction (IUGR) due, in part, to reduced uterine artery blood flow and impaired myometrial artery (MA) vasodilator response. A role for the AMP-activated protein kinase (AMPK) pathway in protecting against hypoxia-associated IUGR is suggested by genomic and transcriptomic studies in humans and functional studies in mice. AMPK is a hypoxia-sensitive metabolic sensor with vasodilatory properties. Here we hypothesized that AMPK-dependent vasodilation was increased in MAs from high versus low-altitude (<1,700 m) Colorado women with appropriate for gestational age (AGA) pregnancies and reduced in IUGR pregnancies regardless of altitude. Vasoreactivity studies showed that, in AGA pregnancies, MAs from high-altitude women were more sensitive to vasodilation by activation of AMPK with A769662 due chiefly to increased endothelial nitric oxide production, whereas MA responses to AMPK activation in the low-altitude women were endothelium independent. MAs from IUGR compared with AGA pregnancies had blunted vasodilator responses to acetylcholine at high altitude. We concluded that 1) blunted vasodilator responses in IUGR pregnancies confirm the importance of MA vasodilation for normal fetal growth and 2) the increased sensitivity to AMPK activation in AGA pregnancies at high altitude suggests that AMPK activation helped maintain MA vasodilation and fetal growth. These results highlight a novel mechanism for vasodilation of MAs under conditions of chronic hypoxia and suggest that AMPK activation could provide a therapy for increasing uteroplacental blood flow and improving fetal growth in IUGR pregnancies.NEW & NOTEWORTHY Intrauterine growth restriction (IUGR) impairs infant well- being and increases susceptibility to later-in-life diseases for mother and child. Our study reveals a novel role for AMPK in vasodilating the myometrial artery (MA) from women residing at high altitude (>2,500 m) with appropriate for gestational age pregnancies but not in IUGR pregnancies at any altitude.

Keywords: AMPK; altitude; hypoxia; intrauterine growth restriction; vasodilation.

Fig. 1.

AMPK is expressed and phosphorylated in myometrial vessels from women at low and high altitude. A: representative microscope pictures of myometrial tissue from appropriate for gestational age (AGA) or intrauterine growth-restricted (IUGR) pregnant women residing at low or high altitude showing staining of AMPK, p-AMPK, and CD31. Yellow arrowheads show blood vessels, orange arrowheads show blood vessels zoomed-in for each inset, and green staining shows localization of endothelial cells. Scale bars = 50 µm (10 µm, insets). B and C: Quantification of AMPK and p-AMPK staining in myometrial arteries (MAs) from low (closed symbols)- and high-altitude (open symbols) subjects, expressed as percentage of stained-positive pixels within the vessels. Symbols are averaged values per subject; bars are means ± SE. Same italicized letters represent no statistical differences among groups with a P < 0.05 by two-way ANOVA.

Fig. 2.

AMPK activation reduced vasoconstriction in both low- and high-altitude myometrial arteries (MAs). Concentration-response curves of phenylephrine (PE) expressed as percentage of maximal K⁺ constriction (K_max) in the presence (+ A769662, gray symbols) or absence (untreated, black symbols) of 30 µM A760662 in MAs from low (A; closed symbols)- and high-altitude (B; open symbols) women. Number in parentheses are number of subjects per group. Symbols are mean values ± SE. Half-maximal effective concentration (EC₅₀) and maximal responses (Max. response) analysis of PE concentration-response curves at low (C and D) and high altitude (E and F). Symbols are averaged vessels per subject; bars are means ± SE. Different italicized letters represent statistical differences among EC₅₀ or maximal responses with a P < 0.05 by Kruskal-Wallis.

Fig. 3.

AMPK-dependent vasodilation was increased by altitude in myometrial arteries (MAs) in an endothelium-dependent manner. A: A769662 concentration-response curves expressed as percentage of 10 µM phenylephrine (PE) in MAs from low (closed circles)- or high-altitude (open symbols) women. C and E: before A769662 curves were performed, MAs from low (C, closed symbols) or high altitude (E, open symbols) were pretreated with 10 µM N^G-nitro-l-arginine methyl ester (l-NAME (squares) or 10 µM l-NAME + 10 µM Indo (diamonds), or their endothelium was removed (−endothelium, triangles). Numbers in parentheses are number of subjects per group. Symbols are means values ± SE. B, D, and F: area under the curve (AUC) analysis of A769662 concentration curves in A, C, and E, respectively. Symbols are averaged vessels per subject; bars are means ± SE. Different italicized letters represent statistical differences among AUC with a P < 0.05 by two-way ANOVA.

Fig. 4.

Myometrial artery (MA) vasodilator response to AMPK activation was similar in intrauterine growth restriction (IUGR) pregnancies at low altitude but not at high altitude. A769662 concentration-response curves expressed as percentage of 10 µM phenylephrine (PE) in MAs from women with appropriate for gestational age (AGA; black symbols) or IUGR (gray symbols) pregnancies residing at low (A; closed symbols) or high altitude (B; open symbols). Numbers in parentheses are number of subjects per group. Symbols are means values ± SE. C: area under the curve (AUC) analysis of A769662 concentration curves at low and high altitude. Symbols are averaged vessels per subject; bars are means ± SE. Different italicized letters represent statistical differences among AUC with a P < 0.05 by two-way ANOVA.

Fig. 5.

Acetylcholine (ACh) but not bradykinin (BK) vasodilator responses were reduced in intrauterine growth restriction (IUGR) pregnancies at both altitudes. ACh (A) and BK (C) concentration-response curves expressed as percentage of 10 µM phenylephrine (PE) in myometrial artery (MA) from IUGR (gray symbols) pregnant women residing at low (closed symbols) or high altitude (open symbols). Numbers in parentheses are number of subjects per group. Symbols are means values ± SE. B and D: area under the curve (AUC) analysis of ACh and BK concentration-curves in A and C, respectively. Symbols are averaged vessels per subject, bars are means ± SE. Same italicized letters represent no statistical differences among groups with a P < 0.05 by two-way ANOVA.