MRC Centre for Reproductive Health and Tommy's Centre for Maternal and Fetal Health, University of Edinburgh, Queen's Medical Research Institute, Edinburgh, UK.
MRC Centre for Inflammation Research, University of Edinburgh, Queen's Medical Research Institute, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Mol Hum Reprod. 2017 Oct 1;23(10):708-724. doi: 10.1093/molehr/gax038.
Is labour, both at term and preterm, associated with alterations in decidual lymphocyte densities and widespread changes to the decidual transcriptome?
The onset of parturition, both at term and preterm, is associated with widespread gene expression changes in the decidua, many of which are related to inflammatory signalling, but is not associated with changes in the number of any of the decidual lymphocyte populations examined.
Given its location, directly at the maternal-foetal interface, the decidua is likely to play a pivotal role in the onset of parturition, however, the molecular events occurring in the decidua in association with the onset of labour, both at term and preterm, remain relatively poorly defined. Using flow cytometry and microarray analysis, the present study aimed to investigate changes to the immune cell milieu of the decidua in association with the onset of parturition and define the decidual gene signature associated with term and preterm labour (PTL).
STUDY DESIGN, SIZE, DURATION: This study used decidual samples collected from 36 women across four clinical groups: term (38-42 weeks of gestation) not in labour, TNL; term in labour, TL; preterm (<35 weeks of gestation)not in labour, PTNL; and preterm in labour, PTL.
PARTICIPANTS/MATERIALS, SETTING, METHODS: Decidual lymphocytes were isolated from fresh decidual tissue collected from women in each of our four patient groups and stained with a panel of antibodies (CD45, CD3, CD19, CD56, CD4, CD8 and TCRVα24-Jα18) to investigate lymphocyte populations present in the decidua (TNL, n = 8; TL, n = 7; PTNL, n = 5; PTL, n = 5). RNA was extracted from decidual tissue and subjected to Illumina HT-12v4.0 BeadChip expression microarrays (TNL, n = 11; TL, n = 8; PTNL, n = 7; PTL, n = 10). Quantitative real-time PCR (qRT-PCR) was used to validate the microarray results.
The relative proportions of decidual lymphocytes (T cells, NK cells, B cells and invariant natural killer (iNKT) cells) were unaffected by either gestation or labour status. However, we found elevated expression of the non-classical MHC-protein, CD1D, in PTL decidua samples (P < 0.05), suggesting the potential for increased activation of decidual invariant NKT (iNKT) cells in PTL. Both term and PTL were associated with widespread gene expression changes, particularly related to inflammatory signalling. Up-regulation of candidate genes in TL (IL-6, PTGS2, ATF3, IER3 and TNFAIP3) and PTL (CXCL8, MARCO, LILRA3 and PLAU) were confirmed by qRT-PCR analysis.
Microarray data are available at www.ebi.ac.uk/arrayexpress under accession number E-MTAB-5353.
Whilst no changes in lymphocyte number were observed across our patient samples, we did not investigate the activation state of any of the immune cell sub-populations examined, therefore, it is possible that the function of these cells may be altered in association with labour onset. Additionally, the results of our transcriptomic analyses are descriptive and at this stage, we cannot prove direct causal link with the up-regulation of any of the genes examined and the onset of either term or PTL.
Our findings demonstrate that the onset of parturition is associated with widespread changes to the decidual transcriptome, and there are distinct gene expression changes associated with term and PTL. We confirmed that an inflammatory signature is present within the decidua, and we also report the up-regulation of several genes involved in regulating the inflammatory response. The identification of genes involved in regulating the inflammatory response may provide novel molecular targets for the development of new, more effective therapies for the prevention of preterm birth (PTB). Such targets are urgently required.
STUDY FUNDING AND COMPETING INTEREST(S): This work was supported by Medical Research Council (grant number MR/L002657/1) and Tommy's, the baby charity. Jane Norman has had research grants from the charity Tommy's and from the National Institute for Health Research on PTB during the lifetime of this project. Jane Norman also sits on a data monitoring committee for GSK for a study on PTB prevention and her institution receives financial recompense for this. The other authors do not have any conflicts of interest to declare.
足月和早产分娩时，蜕膜淋巴细胞密度是否会发生改变，蜕膜转录组是否会发生广泛变化？
足月和早产分娩时，分娩的开始与蜕膜中广泛的基因表达变化相关，其中许多与炎症信号有关，但与所检查的任何蜕膜淋巴细胞群数量的变化无关。
由于其位置直接位于母体-胎儿界面，蜕膜很可能在分娩开始时发挥关键作用，然而，与足月和早产分娩开始相关的蜕膜中发生的分子事件仍相对定义不明确。本研究使用流式细胞术和微阵列分析，旨在研究与分娩开始相关的蜕膜免疫细胞环境的变化，并确定与足月和早产分娩（PTL）相关的蜕膜基因特征。
研究设计、规模、持续时间：本研究使用了来自四个临床组的 36 名妇女的蜕膜样本：足月（38-42 周妊娠）未分娩（TNL）；足月分娩（TL）；早产（<35 周妊娠）未分娩（PTNL）；和早产分娩（PTL）。
参与者/材料、设置、方法：从每个患者组的女性的新鲜蜕膜组织中分离出蜕膜淋巴细胞，并使用一组抗体（CD45、CD3、CD19、CD56、CD4、CD8 和 TCRVα24-Jα18）对蜕膜中的淋巴细胞群进行染色（TNL，n=8；TL，n=7；PTNL，n=5；PTL，n=5）。从蜕膜组织中提取 RNA，并进行 Illumina HT-12v4.0 BeadChip 表达微阵列分析（TNL，n=11；TL，n=8；PTNL，n=7；PTL，n=10）。使用定量实时 PCR（qRT-PCR）验证微阵列结果。
蜕膜淋巴细胞（T 细胞、NK 细胞、B 细胞和不变自然杀伤（iNKT）细胞）的相对比例不受妊娠或分娩状态的影响。然而，我们发现 PTL 蜕膜样本中 MHC 蛋白 CD1D 的表达升高（P<0.05），提示 PTL 中可能增加了蜕膜不变自然杀伤（iNKT）细胞的激活。足月和 PTL 都与广泛的基因表达变化相关，特别是与炎症信号相关。TL（IL-6、PTGS2、ATF3、IER3 和 TNFAIP3）和 PTL（CXCL8、MARCO、LILRA3 和 PLAU）中候选基因的上调通过 qRT-PCR 分析得到证实。
微阵列数据可在 www.ebi.ac.uk/arrayexpress 上以 accession number E-MTAB-5353 获得。
限制/原因：尽管我们在患者样本中未观察到淋巴细胞数量的变化，但我们没有研究任何检查的免疫细胞亚群的激活状态，因此，这些细胞的功能可能与分娩开始时的变化有关。此外，我们的转录组分析结果是描述性的，在现阶段，我们不能证明任何检查基因的上调与足月或 PTL 的开始有直接的因果关系。
我们的研究结果表明，分娩的开始与蜕膜转录组的广泛变化相关，并且与足月和 PTL 相关的基因表达变化有明显的区别。我们证实了炎症特征存在于蜕膜中，并且我们还报告了几个参与调节炎症反应的基因的上调。参与调节炎症反应的基因的鉴定可能为预防早产（PTB）的新的、更有效的治疗方法提供新的分子靶点。目前迫切需要这些靶点。
这项工作得到了医学研究委员会（grant number MR/L002657/1）和 Tommy's，婴儿慈善机构的支持。Jane Norman 在这个项目的整个过程中，都有 Tommy's 的研究资助和关于 PTB 的国家卫生研究院资助。Jane Norman 还担任 GSK 预防 PTB 研究的数据监测委员会成员，她的机构因此获得经济补偿。其他作者没有任何利益冲突需要声明。
