蜕膜组织NF-κB介导趋化因子与细胞因子调控子宫NK功能

NF-κB mediated chemokines and cytokines regulate of uterine NK cell function in decidual tissue

摘要

目的 探讨人子宫内膜基质细胞(hESC)蜕膜化过程中核因子κB(NF-κB)介导调控趋化因子、细胞因子产生进而影响子宫自然杀伤细胞(uNK)功能。方法 选取2022年10月1日至2023年3月1日在空军军医大学唐都医院妇产科行人工流产术终止妊娠的20名健康孕妇,收集其蜕膜组织并分离纯化出人uNK.将hESC转染siRelA以抑制NF-κB的活性,随后体外诱导蜕膜化,采用实时荧光定量聚合酶链式反应(RT-qPCR)检测相关趋化因子、细胞因子的mRNA水平.再转染siRelA的hESC诱导蜕膜化后与人原代uNK共培养,分别收获uNK和细胞上清液,采用流式细胞术检测uNK细胞因子分泌水平,通过体外成管实验、侵袭和划痕实验评估uNK上清体外促血管形成和促滋养层细胞侵袭迁移的能力。结果 转染siRelA后诱导蜕膜化产生的蜕膜基质细胞(DSC)内干扰素γ(IFN-γ)、基质金属蛋白酶(MMP)9、趋化因子CCL2的mRNA表达水平均显著降低(t值分别为3.290、6.415、5.095,P<0.05).抑制NF-κB活性并诱导蜕膜化产生的DSC与uNK共培养后,抑制了uNK相关因子血管内皮生长因子(VEGF)、胎盘生长因子(PLGF)和IFN-γ的分泌(t值分别为6.641、7.743、8.886,P<0.05).此外,共培养后产生的uNK上清体外促血管形成能力受损(t值分别为4.629、5.522,P<0.05),并且促滋养层细胞侵袭和迁移能力均显著下降(t值分别为8.329、7.190,P<0.05)。结论 NF-κB调控蜕膜基质细胞趋化因子、细胞因子的表达,进而影响子宫自然杀伤细胞功能.

Abstract

Objective To investigate the role of nuclear factor κB (NF-κB) in mediating the regulation of chemokine and cytokine production during human endometrial stromal cell (hESC) decidualization, thus affecting the function of uterine natural killer cell (uNK). Methods 20 healthy pregnant women who underwent artificial abortion to terminate pregnancy at Department of Gynecology and Obstetrics, Tangdu Hospital, Air Force Medical University from October 1, 2022 to March 1, 2023 were selected. Their decidual tissues were collected, and pure human uNK cells were isolated. The hESC were transfected with siRelA to inhibit NF-κB activity, followed by in vitro decidualization induction. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used to detect mRNA levels of relevant chemokines and cytokines. After inducing decidualization of siRelA-transfected hESC, they were co-cultured with primary human uNK cells, and uNK cells and cell supernatants were harvested separately. Flow cytometry was used to detect the cytokine secretion function of uNK. The supernatant of uNK was used for in vitro tube formation assay, invasion assay and scratch assay to investigate the function of adjusting angiogenesis and trophoblast cells invasion and migration ability. Results Transfection with siRelA significantly decreased the mRNA expression levels of interferon-γ (IFN-γ), matrix metalloproteinase (MMP) 9, and chemokine CCL2 in decidual stromal cells (DSC) induced by decidualization (t=3.290, 6.415 and 5.095, respectively, P<0.05). Inhibiting NF-κB activity and inducing decidualization of DSC suppressed the secretion of uNK-related factors vascular endothelial growth factor (VEGF), placentail growth factor (PLGF), and IFN-γ after co-culture with uNK cells (t=6.641, 7.743 and 8.886, respectively, P<0.05). The ability of uNK cell supernatants to promote angiogenesis was impaired after co-culture (t=4.629 and 5.522, P<0.05), and trophoblast invasion and migration were significantly reduced (t=8.329 and 7.190, P<0.05). Conclusion NF-κB regulates the expression of chemokines and cytokines in DSC, thus influencing the function of uNK.

关键词

NF-κB / 蜕膜基质细胞 / 细胞因子 / 趋化因子 / 自然杀伤细胞

Key words

NF-κB / decidual stromal cell / cytokines / chemokines / natural killer cell

参考文献

[1] Munro S K, Balakrishnan B, Lissaman A C, et al. Cytokines and pregnancy: potential regulation by histone deacetylases[J]. Mol Reprod Dev, 2021, 88(5): 321-337.

[2] 朱霞, 李慧珍, 刘丹, 等. 胚胎植入的相关信号通路[J]. 国际生殖健康/计划生育杂志, 2022, 41(5): 409-413.

[3] Yang X, Tian Y, Zheng L, et al. The update immune-regulatory role of pro- and anti-inflammatory cytokines in recurrent pregnancy losses[J]. Int J Mol Sci, 2022, 24(1): 132.

[4] Zhang Q, Lenardo M J, Baltimore D. 30 Years of NF-κB: a blossoming of relevance to human pathobiology[J]. Cell, 2017, 168(1-2): 37-57.

[5] 张丹妮, 刘迪, 王丽. 核因子-κB对妊娠及分娩影响[J]. 临床军医杂志, 2019, 47(11): 1276-1278.

[6] Liu H, Huang X, Mor G, et al. Epigenetic modifications working in the decidualization and endometrial receptivity[J]. Cell Mol Life Sci, 2020, 77(11): 2091-2101.

[7] Ma Q, Beal J R, Bhurke A, et al. Extracellular vesicles secreted by human uterine stromal cells regulate decidualization, angiogenesis, and trophoblast differentiation[J]. Proc Natl Acad Sci USA, 2022, 119(38): e2200252119.

[8] Ramhorst R, Grasso E, Vota D, et al. From decidualization to pregnancy progression: an overview of immune and metabolic effects of VIP[J]. Am J Reprod Immunol, 2022, 88(4): e13601.

[9] Wu H M, Chen L H, Hsu L T, et al. Immune tolerance of embryo implantation and pregnancy: the role of human decidual stromal cell- and embryonic-derived extracellular vesicles[J]. Int J Mol Sci, 2022, 23(21): 13382.

[10] Bao S, Chen Z, Qin D, et al. Single-cell profiling reveals mechanisms of uncontrolled inflammation and glycolysis in decidual stromal cell subtypes in recurrent miscarriage[J]. Hum Reprod, 2023, 38(1): 57-74.

[11] Sojka D K, Yang L, Yokoyama W M. Uterine natural killer cells[J]. Front Immunol, 2019, 10: 960.

[12] 盛秋, 李芬. 子宫自然杀伤细胞的研究进展[J]. 中国妇幼健康研究, 2013, 24(6): 960-962.

[13] Xu X, Zhou Y, Fu B, et al. Uterine NK cell functions at maternal-fetal interface[J]. Biol Reprod, 2022, 107(1): 327-338.

[14] Díaz-Hernández I, Alecsandru D, García-Velasco J A, et al. Uterine natural killer cells: from foe to friend in reproduction[J]. Hum Reprod Update, 2021, 27(4): 720-746.

[15] Gamliel M, Goldman-Wohl D, Isaacson B, et al. Trained memory of human uterine NK cells enhances their function in subsequent pregnancies[J]. Immunity, 2018, 48(5): 951-962.e5.

[16] Strunz B, Bister J, Jönsson H, et al. Continuous human uterine NK cell differentiation in response to endometrial regeneration and pregnancy[J]. Sci Immunol, 2021, 6(56): eabb7800.

[17] Gómez-Chávez F, Correa D, Navarrete-Meneses P, et al. NF-κB and its regulators during pregnancy[J]. Front Immunol, 2021, 12: 679106.

[18] Sakowicz A. The role of NFκB in the three stages of pregnancy-implantation, maintenance, and labour: a review article[J]. BJOG, 2018, 125(11): 1379-1387.

[19] Yan C, Boyd D D. Regulation of matrix metalloproteinase gene expression[J]. J Cell Physiol, 2007, 211(1): 19-26.

[20] 黎俏. 子宫内膜NK细胞在胚胎种植中的研究进展[J]. 海南医学, 2023, 34(15): 2277-2280.

[21] James-Allan L B, Buckley R J, Whitley G S, et al. The phenotype of decidual CD56+ lymphocytes is influenced by secreted factors from decidual stromal cells but not macrophages in the first trimester of pregnancy[J]. J Reprod Immunol, 2020, 138: 103082.

[22] Gordon S M. Interleukin-15 in outcomes of pegnancy[J]. Int J Mol Sci, 2021, 22(20): 11094.

[23] Ashley R L, Runyan C L, Maestas M M, et al. Inhibition of the C-X-C motif chemokine 12 (CXCL12) and its receptor CXCR4 reduces utero-placental expression of the VEGF system and increases utero-placental autophagy[J]. Front Vet Sci, 2021, 8: 650687.

[24] Wang Q, Sun Y, Fan R, et al. Role of inflammatory factors in the etiology and treatment of recurrent implantation failure[J]. Reprod Biol, 2022, 22(4): 100698.

[25] Zheng J, Wang H, Zhou W. Modulatory effects of trophoblast-secreted CXCL12 on the migration and invasion of human first-trimester decidual epithelial cells are mediated by CXCR4 rather than CXCR7[J]. Reprod Biol Endocrinol, 2018, 16(1): 17.

[26] Meyer N, Zenclussen A C. Immune cells in the uterine remodeling: are they the target of endocrine disrupting chemicals? [J]. Front Immunol, 2020, 11: 246.

[27] 曹小宛, 陈黎, 宋珍珍, 等. 白细胞介素33在妊娠相关疾病中的作用[J]. 国际生殖健康/计划生育杂志, 2021, 40(5): 427-430.

基金资助

国家自然科学基金(8227060612)