天然雌激素和环境雌激素对输卵管细胞中环腺苷酸水平的调节。

Modulation of Cyclic AMP Levels in Fallopian Tube Cells by Natural and Environmental Estrogens.

机构信息

Department of Reproductive Endocrinology, University Hospital Zurich, Areal Wagi Schlieren, Wagistrasse 14, CH 8952 Schlieren, Switzerland.

R&D Scientific Affairs, IBSA Institut Biochimique SA, Via del Piano 29, P.O. Box 266, CH 6915 Pambio-Noranco, Switzerland.

出版信息

Cells. 2021 May 19;10(5):1250. doi: 10.3390/cells10051250.

DOI:10.3390/cells10051250

PMID:34069403

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8158772/

Abstract

Autocrine/paracrine factors generated in response to 17β-estradiol (E2) within the fallopian tube (FT) facilitate fertilization and early embryo development for implantation. Since cyclic AMP (cAMP) plays a key role in reproduction, regulation of its synthesis by E2 may be of biological/pathophysiological relevance. Herein, we investigated whether cAMP production in FT cells (FTCs) is regulated by E2 and environmental estrogens (EE's; xenoestrogens and phytoestrogens). Under basal conditions, low levels of extracellular cAMP were detectable in bovine FTCs (epithelial cells and fibroblasts; 1:1 ratio). Treatment of FTCs with forskolin (AC; adenylyl cyclase activator), isoproterenol (β-adrenoceptor agonist) and IBMX (phosphodiesterase (PDE) inhibitor) dramatically (>10 fold) increased cAMP; whereas LRE1 (sAC; soluble AC inhibitor) and 2',5'-dideoxyadenosine (DDA; transmembrane AC (tmAC)) inhibitor decreased cAMP. Comparable changes in basal and stimulated intracellular cAMP were also observed. Ro-20-1724 (PDE-IV inhibitor), but not milrinone (PDE-III inhibitor) nor mmIBMX (PDE-I inhibitor), augmented forskolin-stimulated cAMP levels, suggesting that PDE-IV dominates in FTCs. E2 increased cAMP levels and CREB phosphorylation in FTCs, and these effects were mimicked by EE's (genistein, 4-hydroxy-2',4',6'-trichlorobiphenyl, 4-hydroxy-2',4',6'-dichlorobiphenyl). Moreover, the effects of E2 and EE were blocked by the tmAC inhibitor DDA, but not by the ERα/β antagonist ICI182780. Moreover, BAPTA-AM (intracellular-Ca chelator) abrogated the effects of E2, but not genistein, on cAMP suggesting differential involvement of Ca. Treatment with non-permeable E2-BSA induced cAMP levels and CREB-phosphorylation; moreover, the stimulatory effects of E2 and EEs on cAMP were blocked by G15, a G protein-coupled estrogen receptor (GPER) antagonist. E2 and IBMX induced cAMP formation was inhibited by LRE1 and DDA suggesting involvement of both tmAC and sAC. Our results provide the first evidence that in FTCs, E2 and EE's stimulate cAMP synthesis via GPER. Exposure of the FT to EE's and PDE inhibitors may result in abnormal non-cyclic induction of cAMP levels which may induce deleterious effects on reproduction.

摘要

自分泌/旁分泌因子对 17β-雌二醇（E2）在输卵管（FT）内的反应有助于受精和早期胚胎发育植入。由于环磷酸腺苷（cAMP）在生殖中起着关键作用，E2 对其合成的调节可能具有生物学/病理生理学意义。在此，我们研究了 E2 和环境雌激素（EE；xenoestrogens 和 phytoestrogens）是否调节 FT 细胞（FTCs）中的 cAMP 产生。在基础条件下，可检测到牛 FTCs（上皮细胞和成纤维细胞；1:1 比例）中外源 cAMP 的低水平。用 forskolin（AC；腺苷酸环化酶激活剂）、异丙肾上腺素（β-肾上腺素能受体激动剂）和 IBMX（磷酸二酯酶（PDE）抑制剂）处理 FTCs 可显著（>10 倍）增加 cAMP；而 LRE1（sAC；可溶性 AC 抑制剂）和 2'，5'-二脱氧腺苷（DDA；跨膜 AC（tmAC）抑制剂）降低了 cAMP。在基础和刺激的细胞内 cAMP 中也观察到类似的变化。Ro-20-1724（PDE-IV 抑制剂）而非米力农（PDE-III 抑制剂）或 mmIBMX（PDE-I 抑制剂）增加了 forskolin 刺激的 cAMP 水平，表明 PDE-IV 在 FTCs 中占主导地位。E2 增加了 FTCs 中的 cAMP 水平和 CREB 磷酸化，EE（染料木黄酮、4-羟基-2'，4'，6'-三氯联苯、4-羟基-2'，4'，6'-二氯联苯）模拟了这些作用。此外，E2 和 EE 的作用被 tmAC 抑制剂 DDA 阻断，但被 ERα/β拮抗剂 ICI182780 阻断。此外，BAPTA-AM（细胞内-Ca 螯合剂）消除了 E2 但不是染料木黄酮对 cAMP 的作用，表明 Ca 的差异参与。用非渗透性 E2-BSA 处理可诱导 cAMP 水平和 CREB 磷酸化；此外，E2 和 EE 对 cAMP 的刺激作用被 GPER 拮抗剂 G15 阻断。LRE1 和 DDA 抑制 E2 和 IBMX 诱导的 cAMP 形成，表明 tmAC 和 sAC 的参与。我们的结果首次证明，在 FTCs 中，E2 和 EE 通过 GPER 刺激 cAMP 合成。FT 暴露于 EE 和 PDE 抑制剂可能导致异常的非循环诱导的 cAMP 水平，这可能对生殖产生有害影响。