Institut National de la Santéc et de la Recherche Médicale, Lipides Nutrition Cancer, Dijon, France.
Université de Bourgogne Franche-Comté, Lipides Nutrition Cancer, Dijon, France.
Biol Reprod. 2020 May 26;102(6):1326-1339. doi: 10.1093/biolre/ioaa032.
At labor, the myometrium is infiltrated by a massive influx of macrophages that secrete high levels of pro-inflammatory cytokines inducing the expression of specific labor-associated markers. However, the interactions between myocytes and macrophages and the role of macrophages in the myometrium at labor remain to be elucidated. In this work, we studied the role of myometrium-infiltrated macrophages and their interaction with myocytes in lipopolysaccharide-induced preterm labor. A co-culture model of human primary myometrial cells and macrophages was developed and validated. Collagen lattices were used to evaluate myocyte contraction. Differentiation steps were assessed by (i) phalloidin and vinculin staining for cytoskeleton reorganization, (ii) gap junction protein alpha 1 expression and scrape loading/dye transfer with Lucifer Yellow for gap junction intercellular communication, and (iii) calcium imaging for cell excitability. We demonstrated that macrophages favored lipopolysaccharide-induced contraction and early differentiation of myometrial cells. Transwell assays showed that previous activation of macrophages by lipopolysaccharide was essential for this differentiation and that macrophage/myocyte interactions involved macrophage release of reactive oxygen species (ROS). The effects of macrophage-released ROS in myometrial cell transactivation were mimicked by H2O2, suggesting that superoxide anion is a major intermediate messenger in macrophage/myocyte crosstalk during labor. These novel findings provide the foundation for innovative approaches to managing preterm labor, specifically the use of antioxidants to inhibit the initial stages of labor before the contractile phenotype has been acquired. In addition, the co-culture model developed by our team could be used in future research to decipher pathophysiological signaling pathways or screen/develop new tocolytics.
在分娩过程中，大量的巨噬细胞浸润到子宫肌层，分泌高水平的促炎细胞因子，诱导特定的分娩相关标志物的表达。然而，肌细胞和巨噬细胞之间的相互作用以及巨噬细胞在分娩时在子宫肌层中的作用仍有待阐明。在这项工作中，我们研究了浸润到子宫肌层的巨噬细胞及其与肌细胞在脂多糖诱导的早产中的相互作用。建立并验证了人原代子宫平滑肌细胞和巨噬细胞的共培养模型。使用胶原格子评估肌细胞收缩。通过（i）鬼笔环肽和微丝相关蛋白 vimentin 染色评估细胞骨架重排，（ii）缝隙连接蛋白 alpha 1 表达和荧光黄划痕加载/染料转移评估缝隙连接细胞间通讯，（iii）钙成像评估细胞兴奋性来评估分化步骤。我们证明了巨噬细胞有利于脂多糖诱导的子宫平滑肌细胞收缩和早期分化。Transwell 实验表明，巨噬细胞预先被脂多糖激活对于这种分化是必需的，并且巨噬细胞/肌细胞的相互作用涉及巨噬细胞释放活性氧物质（ROS）。巨噬细胞释放的 ROS 在子宫平滑肌细胞转激活中的作用可以被 H2O2 模拟，这表明超氧阴离子是分娩时巨噬细胞/肌细胞串扰中的主要中间信使。这些新发现为管理早产的创新方法提供了基础，特别是使用抗氧化剂在获得收缩表型之前抑制分娩的初始阶段。此外，我们团队开发的共培养模型可用于未来的研究，以解析生理病理信号通路或筛选/开发新的保胎药。
