Di T, Sullivan J A, Rupnow H L, Magness R R, Bird I M
Department of Obstetrics and Gynecology, University of Wisconsin-Madison.
J Soc Gynecol Investig. 1999 Nov-Dec;6(6):301-6. doi: 10.1016/s1071-5576(99)00038-6.
To determine whether pregnancy increases cytosolic phospholipase A2 (cPLA2) expression in uterine artery (UA) endothelial cells and vascular smooth-muscle (VSM) cells, and whether ovarian steroids mediate this effect.
Uterine arteries and omental arteries (systemic control) were isolated from pregnant (120-130 days' gestation) ewes and from nonpregnant ewes synchronized to the corresponding phases of the ovarian cycle (follicular or luteal) or ovariectomized. In addition, ovariectomized ewes were treated with vehicle, estradiol-17 beta (E2 beta), progesterone (P4), or combined E2 beta/P4 for 10 days, and UAs were collected. Arteries from all studies were separated into mechanically isolated endothelial and VSM fractions. Proteins were then solubilized and separated on 7.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western immunoblotting using an affinity-purified mouse monoclonal antibody for cPLA2.
Both UA endothelium and VSM fractions were seen to express cPLA2, detected as a single band with a molecular mass similar to that observed in myometrium (about 100 kD). Distribution of cPLA2, when expressed per microgram of protein, was observed as 40% in UA endothelium compared with 60% in VSM fractions. Uterine artery endothelial cPLA2 expression was specifically increased 1.9-fold in pregnancy (P < .05), whereas there was no significant change from VSM. Furthermore, in ovariectomized sheep versus intact luteal or follicular phase animals, there was no significant change in cPLA2 expression in endothelium or VSM. Administration of E2 beta, P4, or their combination in ovariectomized sheep also failed to reproduce the pregnancy-induced increase in cPLA2 expression in UA endothelium. Omental artery endothelial and VSM cPLA2 expression was observed at similar magnitudes as UA expression, but levels were consistently unchanged by pregnancy, the ovarian cycle, or ovariectomy.
cPLA2 was expressed throughout the endothelium and VSM of both uterine and omental arteries, but only in UA endothelium was pregnancy associated with elevated cPLA2 expression. The lack of change in cPLA2 expression with the ovarian cycle or in ovariectomized animals, even after prolonged treatment with E2 beta, P4, or E2 beta/P4 combined suggests this pregnancy-induced increase in cPLA2 may not be estrogen and/or progesterone dependent.
确定妊娠是否会增加子宫动脉（UA）内皮细胞和血管平滑肌（VSM）细胞中胞质型磷脂酶A2（cPLA2）的表达，以及卵巢类固醇激素是否介导这种作用。
从怀孕（妊娠120 - 130天）的母羊以及与卵巢周期相应阶段（卵泡期或黄体期）同步的未怀孕母羊或去卵巢母羊中分离子宫动脉和网膜动脉（全身对照）。此外，对去卵巢母羊用赋形剂、17β-雌二醇（E2β）、孕酮（P4）或E2β/P4联合处理10天，然后收集子宫动脉。所有研究中的动脉均被分离为通过机械方法分离的内皮部分和VSM部分。然后将蛋白质溶解，并在7.5%十二烷基硫酸钠-聚丙烯酰胺凝胶电泳上进行分离，随后使用针对cPLA2的亲和纯化小鼠单克隆抗体进行Western免疫印迹分析。
UA内皮和VSM部分均可见cPLA2表达，检测为一条单一的条带，其分子量与在子宫肌层中观察到的相似（约100 kD）。以每微克蛋白质计算，cPLA2在UA内皮中的分布为40%，而在VSM部分中为60%。妊娠时UA内皮cPLA2表达特异性增加1.9倍（P <.05），而VSM部分无显著变化。此外，与完整的黄体期或卵泡期动物相比，去卵巢绵羊的内皮或VSM中cPLA2表达无显著变化。在去卵巢绵羊中给予E2β、P4或它们的组合也未能重现妊娠诱导的UA内皮中cPLA2表达的增加。网膜动脉内皮和VSM的cPLA2表达与UA表达水平相似，但妊娠、卵巢周期或去卵巢对其水平均无持续影响。
cPLA2在子宫动脉和网膜动脉的内皮及VSM中均有表达，但仅在UA内皮中，妊娠与cPLA2表达升高相关。即使在用E2β、P4或E2β/P4联合进行长时间处理后，cPLA2表达在卵巢周期或去卵巢动物中仍无变化，这表明这种妊娠诱导的cPLA2增加可能不依赖于雌激素和/或孕酮。
