Table_1_The increased cfRNA of TNFSF4 in peripheral blood at late gestation and preterm labor: its implication as a noninvasive biomarker for premature delivery.csv

IntroductionGiven the important roles of immune tolerance and inflammation in both preterm and term labor, some inflammation-related genes could be related to the initiation of labor, even preterm labor. Inspection of cell-free RNA (cfRNA) engaged in inflammation in maternal blood may represent the varied gestational age and may have significant implications for the development of noninvasive diagnostics for preterm birth.
MethodsTo identify potential biomarkers of preterm birth, we investigated the cfRNA and exosomal miRNA in the peripheral blood of pregnant women at different gestational ages that undergo term labor or preterm labor. 17 inflammatory initiation-related cfRNAs were screened by overlapping with the targets of decreasing miRNAs during gestation and highly expressed cfRNAs at late gestation in maternal blood. To reveal the origins and mechanisms of these screened cfRNAs, the datasets of single-cell RNA sequencing from peripheral blood mononuclear cells of pregnant women, the fetal lung, and the placenta across different gestational ages were analyzed.
ResultsDuring late gestation, TNFSF4 expression increased exclusively in pro-inflammatory macrophages of maternal blood, whereas its receptor, TNFRSF4, increased expression in T cells from the decidua, which suggested the potential cell-cell communication of maternally-originated pro-inflammatory macrophages with the decidual T cells and contributed to the initiation of labor. Additionally, the cfRNA of TNFSF4 was also increased in preterm labor compared to term labor in the validation cohorts. The EIF2AK2 and TLR4 transcripts were increased in pro-inflammatory macrophages from both fetal lung and placenta but not in those from maternal mononuclear cells at late gestation, suggesting these cfRNAs are possibly derived from fetal tissues exclusively. Moreover, EIF2AK2 and TLR4 transcripts were found highly expressed in the pro-inflammatory macrophages from decidua as well, which suggested these specific fetal-origin macrophages may function at the maternal-fetal interface to stimulate uterine contractions, which have been implicated as the trigger of parturition and preterm labor.
DiscussionTaken together, our findings not only revealed the potential of peripheral TNFSF4 as a novel cfRNA biomarker for noninvasive testing of preterm labor but further illustrated how maternal and fetal signals coordinately modulate the inflammatory process at the maternal-fetal interface, causing the initiation of term or preterm labor.

引言：鉴于免疫耐受与炎症在早产临产及足月临产中均发挥关键作用，部分炎症相关基因可能与临产启动（乃至早产临产）相关。对母体外周血中参与炎症过程的无细胞RNA（cell-free RNA, cfRNA）进行检测，可反映个体的孕周差异，同时对开发早产无创诊断技术具有重要指导意义。
方法：为筛选早产潜在生物标志物，本研究对不同孕周、分别经历足月临产或早产临产的孕妇外周血中的无细胞RNA（cfRNA）与外泌体微小RNA（exosomal miRNA）进行分析。通过与妊娠期表达下调的微小RNA靶点取交集，并结合母体外周血晚孕期高表达的cfRNA，最终筛选出17个与炎症启动相关的cfRNA。为明确上述筛选得到的cfRNA的来源与作用机制，本研究对不同孕周孕妇外周血单个核细胞、胎儿肺组织及胎盘组织的单细胞RNA测序数据集进行了分析。
结果：晚孕期时，母体外周血促炎巨噬细胞中TNFSF4的表达仅显著升高，而其受体TNFRSF4在蜕膜T细胞中的表达则出现上调，这提示母源促炎巨噬细胞与蜕膜T细胞间存在潜在的细胞通讯机制，该机制可能参与临产启动。此外，在验证队列中，相较于足月临产组，早产临产组外周血中TNFSF4的cfRNA水平同样升高。EIF2AK2与TLR4转录本在胎儿肺及胎盘来源的促炎巨噬细胞中均表达上调，但晚孕期孕妇外周血单个核细胞来源的促炎巨噬细胞中并无此现象，提示此类cfRNA可能仅来源于胎儿组织。进一步分析发现，蜕膜来源的促炎巨噬细胞中同样高表达EIF2AK2与TLR4转录本，这表明这类特定的胎儿源性巨噬细胞可在母胎界面发挥作用，刺激子宫收缩——而子宫收缩已被证实是临产及早产临产的触发因素。
讨论：综上，本研究不仅证实了外周血TNFSF4作为新型cfRNA生物标志物用于早产临产无创检测的潜力，还阐明了母源与胎儿信号如何协同调控母胎界面的炎症过程，进而介导足月临产或早产临产的启动。