Department of Physiology, Meharry Medical College, 1005 Dr. D.B. Todd Jr. Blvd, Nashville, TN 37208, USA.
Dig Dis Sci. 2013 Jun;58(6):1507-15. doi: 10.1007/s10620-013-2610-4. Epub 2013 Mar 17.
Gastroparesis affects predominantly females; however, the biological basis for this gender bias is completely unknown. Several lines of evidence suggest that nitrergic dependent stomach motility function is reduced in diabetic gastroparesis and that nNOS is estrogen-regulated.
The purpose of this study was to investigate whether reduced levels of estradiol-17β (E2) down-regulates tetrahydrobiopterin (BH4, a cofactor for nNOS dimerization and enzyme activity) biosynthesis and therefore nNOS mediated gastric motility would be impaired in a mouse model of chronic estrogen deficiency, follicle stimulating hormone receptor knock-out female mice (FORKO).
In-bred 12-week-old female FORKO mice were obtained from our FORKO breeding colony. Gastric emptying was measured in overnight fasting mice. Nitrergic relaxation (AUC/mg tissue) was measured at 2 Hz through electric field stimulation using gastric antrum strips prepared from WT and FORKO mice. Protein expression for nNOSα, BH4 biosynthesis enzymes (GCH-1, DHFR) and estrogen receptors (α, β) were measured in gastric antrum by western blotting. Levels of BH4 and oxidized BH2, B biopterin levels were determined by HPLC.
In FORKO, compared to wild type (WT) stomachs we indentified (1) reduced (%) gastric emptying (64 ± 2.5 vs. 77.6 ± 0.88), (2) greater reduction in nitregic relaxation (-0.13 ± 0.012 vs. -0.28 ± 0.012), (3) increased nNOS dimerization (0.48 ± 0.02 vs. 0.34 ± 0.05), (4) decreased NO release whether measured at 24 h (0.6 ± 0.04 vs. 1.7 ± 0.22, p < 0.05) or at 48 h (3.4 ± 0.26 vs. 5.0 ± 0.15, p < 0.05) of incubation, (5) decreased GCH-1 (1.9 ± 0.06 vs. 2.3 ± 0.04), DHFR (1.8 ± 0.14 vs. 2.4 ± 0.07) and ERα (2.7 ± 0.4 vs. 3.9 ± 0.4) and (6) increased oxidized biopterin levels and decreased ratio of BH4 versus BH2 + B.
We conclude that chronic estrogen deficiency negatively modifies the function of both BH4 and nNOS thereby contributing to the development of gastroparesis in a FORKO mouse model.
胃轻瘫主要影响女性；然而，这种性别偏见的生物学基础完全未知。有几条证据表明，在糖尿病性胃轻瘫中，依赖于氮能的胃动力功能降低，并且 nNOS 受雌激素调节。
本研究的目的是探讨雌二醇-17β（E2）水平降低是否会下调四氢生物蝶呤（BH4，nNOS 二聚化和酶活性的辅助因子）的生物合成，因此，在慢性雌激素缺乏的卵泡刺激素受体敲除雌性小鼠（FORKO）模型中，nNOS 介导的胃动力会受损。
从我们的 FORKO 繁殖群体中获得了 12 周龄的近交雌性 FORKO 小鼠。在隔夜禁食的小鼠中测量胃排空。通过使用从 WT 和 FORKO 小鼠制备的胃窦条带，以 2 Hz 的频率通过电刺激测量氮能松弛（AUC/mg 组织）。通过 Western 印迹法测量胃窦中的 nNOSα、BH4 生物合成酶（GCH-1、DHFR）和雌激素受体（α、β）的蛋白表达。通过 HPLC 测定 BH4 和氧化的 BH2、B 生物蝶呤水平。
与野生型（WT）相比，在 FORKO 中，我们发现（1）胃排空减少（64 ± 2.5% vs. 77.6 ± 0.88%），（2）氮能松弛减少更大（-0.13 ± 0.012% vs. -0.28 ± 0.012%），（3）nNOS 二聚化增加（0.48 ± 0.02% vs. 0.34 ± 0.05%），（4）NO 释放减少，无论是在 24 小时（0.6 ± 0.04% vs. 1.7 ± 0.22%，p < 0.05）还是在 48 小时（3.4 ± 0.26% vs. 5.0 ± 0.15%，p < 0.05）孵育时，（5）GCH-1（1.9 ± 0.06% vs. 2.3 ± 0.04%）、DHFR（1.8 ± 0.14% vs. 2.4 ± 0.07%）和 ERα（2.7 ± 0.4% vs. 3.9 ± 0.4%）降低，（6）氧化的生物蝶呤水平增加，BH4 与 BH2+B 的比值降低。
我们得出结论，慢性雌激素缺乏会改变 BH4 和 nNOS 的功能，从而导致 FORKO 小鼠模型中胃轻瘫的发展。
