Toll樣受體對(duì)子宮NK細(xì)胞的調(diào)控特征及其與母胎耐受的關(guān)系
發(fā)布時(shí)間：2018-09-17 14:11
【摘要】： 長(zhǎng)久以來(lái)胎兒被認(rèn)為是一種自體移植物，但令人困惑的是妊娠的建立和維持無(wú)法用經(jīng)典的移植免疫耐受或腫瘤免疫逃逸理論來(lái)解釋。由于母體激素的作用，會(huì)有很多可溶性免疫介質(zhì)和免疫細(xì)胞聚集在哺乳動(dòng)物的生殖道內(nèi)，它們不僅沒(méi)有引起強(qiáng)烈的免疫排斥反應(yīng)，反而可以維持和促進(jìn)正常的月經(jīng)周期、胚胎植入和胎兒發(fā)育。通過(guò)對(duì)人和嚙齒類動(dòng)物的生殖免疫研究，發(fā)現(xiàn)過(guò)去被視為對(duì)腫瘤組織具有殺傷能力的NK細(xì)胞和整個(gè)妊娠過(guò)程有著極為密切的關(guān)系，子宮NK細(xì)胞還可以從不同方面影響母胎耐受。哺乳動(dòng)物胚泡在著床開(kāi)始后，為了進(jìn)一步為入侵子宮內(nèi)膜基質(zhì)做準(zhǔn)備，子宮內(nèi)膜蛻膜化。此時(shí)，大量的子宮NK細(xì)胞在一系列粘附分子、趨化因子和細(xì)胞因子的作用下從外周淋巴組織被招募到母胎界面處發(fā)揮其免疫調(diào)節(jié)功能。 在妊娠這一特殊的生理階段，機(jī)體任何的細(xì)胞免疫反應(yīng)都會(huì)受到雌激素、孕激素等的調(diào)控，子宮NK細(xì)胞或其前體細(xì)胞向蛻膜的遷移也是一樣。我們首先通過(guò)細(xì)胞轉(zhuǎn)輸實(shí)驗(yàn)分析了妊娠早期的C5781／6J小鼠的子宮NK細(xì)胞遷移特征。給懷孕第6.5天的小鼠輸入CFDA-SE標(biāo)記的正常脾淋巴細(xì)胞，24小時(shí)后發(fā)現(xiàn)NK1.1~+細(xì)胞比CD3~+細(xì)胞細(xì)胞更具有遷移優(yōu)勢(shì)，流式檢測(cè)表明：相對(duì)于肝臟(0.96±0.14％)、脾臟(0.15±0.08％)，蛻膜組織有1.13±0.18％的CFDA-SE~+NK1.1~+細(xì)胞駐留。這說(shuō)明至少在蛻膜反應(yīng)階段，母胎界面是NK細(xì)胞歸巢的主要和重要部位。但是Toll樣受體(TLR)信號(hào)的激活可以打破上述NK細(xì)胞的穩(wěn)態(tài)平衡：在進(jìn)行細(xì)胞轉(zhuǎn)輸?shù)耐瑫r(shí)給受體鼠腹腔注射聚肌胞苷酸(polyⅠ：C，TLR3配體)后，發(fā)現(xiàn)轉(zhuǎn)輸?shù)腃FSE-DA~+NK1.1~+細(xì)胞大量的在肝臟聚集，而不是子宮或脾臟。這也表明妊娠期間外周淋巴細(xì)胞向子宮募集是有前提條件的，外界dsRNA刺激可以從時(shí)間和空間角度影響NK細(xì)胞向母胎界面遷移。 本研究還證實(shí)炎性刺激可以導(dǎo)致子宮內(nèi)膜TLR介導(dǎo)的免疫應(yīng)答。TLR3信號(hào)可以調(diào)節(jié)機(jī)體抗病毒反應(yīng)，促進(jìn)細(xì)胞因子分泌。由于許多生殖道病毒包涵或表達(dá)dsRNA，因此我們利用小鼠流產(chǎn)模型進(jìn)行polyⅠ：C干預(yù)，并以子宮NK細(xì)胞為中心，著眼于妊娠早期，分析母胎界面TLR3的表達(dá)特征以及TLR3激動(dòng)劑干預(yù)誘發(fā)的子宮局部免疫反應(yīng)對(duì)妊娠結(jié)果的影響，闡述子宮NK細(xì)胞的調(diào)節(jié)性作用及局部微環(huán)境的天然免疫學(xué)特點(diǎn)。在妊娠第6.5天，對(duì)CBA×DBA／2小鼠模型分別腹腔注射polyⅠ：C和PBS，于第7.5、8.5、10.5、12.5天處死動(dòng)物進(jìn)行相應(yīng)研究。結(jié)果表明在妊娠中期，CBA×DBA／2小鼠胚胎吸收率為11.0±3.0％，polyⅠ：C處理使其顯著增加到40.2±1.7％，并伴隨有明顯的子宮螺旋動(dòng)脈重鑄抑制等病理變化，如管腔變小、管壁變厚，量化分析發(fā)現(xiàn)其血管外、內(nèi)橫截面積比值顯著增加。同時(shí)研究還發(fā)現(xiàn)，盡管polyⅠ：C處理24小時(shí)后(妊娠第7.5天)表觀檢查未能發(fā)現(xiàn)CBA×DBA／2小鼠有胚胎吸收的明顯征兆，與對(duì)照組相比母胎界面IFN-γ含量也沒(méi)有明顯變化，但其蛻膜區(qū)TLR3表達(dá)明顯上調(diào)，且局部組織TNF-α含量增加2.7倍。上述結(jié)果說(shuō)明子宮蛻膜TNF-α分泌是一個(gè)重要的炎性信號(hào)，它可能抑制子宮螺旋動(dòng)脈重鑄并最終影響CBA×DBA／2小鼠的妊娠結(jié)局。 另外，24小時(shí)后檢測(cè)發(fā)現(xiàn)polyⅠ：C處理明顯上調(diào)了蛻膜NK(DX5~+CD3~-)細(xì)胞的胞內(nèi)因子TNF-α和IFN-γ的陽(yáng)性率。出乎意料的是小鼠注射polyⅠ：C后，流式檢測(cè)(DX5~+CD3~+)和細(xì)胞組化(DBA~+)分析結(jié)果都證實(shí)母胎界面子宮NK細(xì)胞數(shù)目顯著減少。此外，局部NKT(DX5~+CD3~-)、T(DX5~+CD3~-)細(xì)胞以及樹(shù)突狀細(xì)胞(CD11c~+)均不受polyⅠ：C的影響。實(shí)際上，母胎界面存在的每一種免疫細(xì)胞都對(duì)妊娠過(guò)程是有益的。由于DX5~+細(xì)胞下調(diào)，使得母胎界面DC／NK比例發(fā)生改變，從而影響了包括細(xì)胞因子在內(nèi)的局部免疫反饋調(diào)節(jié)過(guò)程。由于DC-NK的相互作用不要求細(xì)胞的直接接觸，，因而盡管蛻膜DC和NK細(xì)胞的空間分布沒(méi)有完全重合，二者的相互影響也不容忽視。 綜上所述，本研究證明淋巴細(xì)胞在小鼠妊娠早期(蛻膜化階段)向母胎界面募集時(shí)，外周NK細(xì)胞比T細(xì)胞有更大的遷移優(yōu)勢(shì)。PolyⅠ：C作用不僅可以影響NK細(xì)胞的歸巢和組織特異性分布，還可以激活蛻膜TLR3信號(hào)。子宮NK細(xì)胞的高度活化表現(xiàn)為細(xì)胞因子分泌能力的增強(qiáng)，繼而促進(jìn)母胎界面的炎癥反應(yīng)，加劇在妊娠過(guò)程中的病理性變化如螺旋動(dòng)脈重鑄抑制，導(dǎo)致胚胎吸收的增加。由此可見(jiàn)子宮NK細(xì)胞和局部TLR信號(hào)對(duì)小鼠妊娠具有重要調(diào)節(jié)功能，本動(dòng)物實(shí)驗(yàn)的結(jié)果可以外推到臨床研究和實(shí)踐中，即通過(guò)關(guān)注子宮NK細(xì)胞與其它免疫細(xì)胞的相互作用來(lái)治療生殖免疫疾病、預(yù)測(cè)妊娠結(jié)果。
[Abstract]:The fetus has long been thought of as an autograft, but the establishment and maintenance of pregnancy cannot be explained by classical transplant tolerance or tumor immune escape theories. Through the study of reproductive immunity in humans and rodents, it was found that NK cells, which were previously considered to be cytotoxic to tumor tissues, were closely related to the whole pregnancy process, and uterine NK cells were also found. Mammalian blastocysts are recruited from peripheral lymphoid tissue to the maternal-fetal interface by a series of adhesion molecules, chemokines, and cytokines to prepare for invasion of endometrial matrix. Its immunomodulatory function.
In this particular physiological stage of pregnancy, any cellular immune response is regulated by estrogen, progesterone, and so on. The migration of uterine NK cells or their precursors to decidua is also the same. We first analyzed the migration characteristics of uterine NK cells in C5781/6J mice in early pregnancy by cell transfer experiments. The normal splenic lymphocytes labeled with CFDA-SE were transfused into the spleen of day 1 mice. NK1.1~+ cells had more migration advantages than CD3~+ cells 24 hours later. Flow cytometry showed that 1.13 (+ 0.18%) CFDA-SE ~+ NK1.1~+ cells resided in the decidua tissue than in the liver (0.96 (+ 0.14%) and the spleen (0.15 (+ 0.08%). The maternal-fetal interface is the main and important site of NK cell homing. However, the activation of Toll-like receptor (TLR) signal can break the homeostasis of NK cells mentioned above. After intraperitoneal injection of Polyinosine (poly I: C, TLR3 ligand) into recipient mice, it was found that a large number of C FSE-DA~+NK1.1~+ cells accumulated in the liver, while Toll-like receptor (TLR) signal was transported. This also indicates that peripheral lymphocyte recruitment to the uterus during pregnancy is preconditioned. External dsRNA stimulation can influence the migration of NK cells to the maternal-fetal interface in time and space.
TLR3 signal can regulate the body's antiviral response and promote the secretion of cytokines. Because many reproductive tract viruses contain or express dsRNA, we used the mouse abortion model poly I:C intervention, and the uterine NK cells as the center, focusing on pregnancy. In early pregnancy, the expression of TLR3 at maternal-fetal interface and the effect of TLR3 agonist on the outcome of pregnancy were analyzed. The regulatory effect of uterine NK cells and the natural immunological characteristics of local microenvironment were elaborated. 7.5, 8.5, 10.5 and 12.5 days after the treatment, the animals were sacrificed. The results showed that in the second trimester of pregnancy, the embryo absorption rate of CBA *DBA/2 mice was 11.0 +3.0%. Poly I: C treatment significantly increased the embryo absorption rate to 40.2 +1.7%. It was accompanied by obvious pathological changes, such as the reduction of lumen and the thickening of tube wall. At the same time, the study also found that although the apparent examination of poly I:C treatment 24 hours after pregnancy (7.5 days) did not show obvious signs of embryonic absorption in CBA *DBA/2 mice, and compared with the control group, there was no significant change in IFN-gamma content at the maternal-fetal interface, but the expression of TLR3 in the decidual region was significantly up-regulated, and local. These results suggest that the secretion of TNF-a in decidua is an important inflammatory signal, which may inhibit the recasting of uterine spiral artery and ultimately affect the pregnancy outcome of CBA *DBA/2 mice.
In addition, 24 hours later, poly I:C treatment significantly increased the positive rates of TNF-alpha and IFN-gamma in decidual NK (DX5~+CD3~-) cells. To our surprise, flow cytometry (DX5~+CD3~+) and cytohistochemistry (DBA~+) analysis showed a significant decrease in the number of uterine NK cells at the maternal-fetal interface. NKT (DX5~+CD3~-), T (DX5~+CD3~-) cells and dendritic cells (CD11c~+) were not affected by poly I: C. In fact, every immune cell at the maternal-fetal interface was beneficial to the pregnancy process. Because of the downregulation of DX5~+ cells, the DC/NK ratio at the maternal-fetal interface was changed, which affected local factors including cytokines. Because DC-NK interactions do not require direct cell contact, the spatial distribution of DC and NK cells in decidua does not coincide completely, and the interaction between DC and NK cells should not be neglected.
In conclusion, this study demonstrates that peripheral NK cells have a greater migration advantage than T cells when lymphocytes are recruited at the maternal-fetal interface in early pregnancy (decidualization stage). Poly I:C not only affects the homing and tissue-specific distribution of NK cells, but also activates TLR3 signal in decidua. The highly activated uterine NK cells are manifested by high activation of TLR3 signal. The enhancement of cytokine secretion and the subsequent inflammatory response at the maternal-fetal interface can aggravate the pathological changes during pregnancy, such as the inhibition of spiral artery recasting, leading to the increase of embryo absorption. In bed research and practice, we treat reproductive immune diseases and predict pregnancy outcomes by focusing on the interaction between uterine NK cells and other immune cells.
【學(xué)位授予單位】：中國(guó)科學(xué)技術(shù)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2007
【分類號(hào)】：R392
本文編號(hào)：2246177
[Abstract]:The fetus has long been thought of as an autograft, but the establishment and maintenance of pregnancy cannot be explained by classical transplant tolerance or tumor immune escape theories. Through the study of reproductive immunity in humans and rodents, it was found that NK cells, which were previously considered to be cytotoxic to tumor tissues, were closely related to the whole pregnancy process, and uterine NK cells were also found. Mammalian blastocysts are recruited from peripheral lymphoid tissue to the maternal-fetal interface by a series of adhesion molecules, chemokines, and cytokines to prepare for invasion of endometrial matrix. Its immunomodulatory function.
In this particular physiological stage of pregnancy, any cellular immune response is regulated by estrogen, progesterone, and so on. The migration of uterine NK cells or their precursors to decidua is also the same. We first analyzed the migration characteristics of uterine NK cells in C5781/6J mice in early pregnancy by cell transfer experiments. The normal splenic lymphocytes labeled with CFDA-SE were transfused into the spleen of day 1 mice. NK1.1~+ cells had more migration advantages than CD3~+ cells 24 hours later. Flow cytometry showed that 1.13 (+ 0.18%) CFDA-SE ~+ NK1.1~+ cells resided in the decidua tissue than in the liver (0.96 (+ 0.14%) and the spleen (0.15 (+ 0.08%). The maternal-fetal interface is the main and important site of NK cell homing. However, the activation of Toll-like receptor (TLR) signal can break the homeostasis of NK cells mentioned above. After intraperitoneal injection of Polyinosine (poly I: C, TLR3 ligand) into recipient mice, it was found that a large number of C FSE-DA~+NK1.1~+ cells accumulated in the liver, while Toll-like receptor (TLR) signal was transported. This also indicates that peripheral lymphocyte recruitment to the uterus during pregnancy is preconditioned. External dsRNA stimulation can influence the migration of NK cells to the maternal-fetal interface in time and space.
TLR3 signal can regulate the body's antiviral response and promote the secretion of cytokines. Because many reproductive tract viruses contain or express dsRNA, we used the mouse abortion model poly I:C intervention, and the uterine NK cells as the center, focusing on pregnancy. In early pregnancy, the expression of TLR3 at maternal-fetal interface and the effect of TLR3 agonist on the outcome of pregnancy were analyzed. The regulatory effect of uterine NK cells and the natural immunological characteristics of local microenvironment were elaborated. 7.5, 8.5, 10.5 and 12.5 days after the treatment, the animals were sacrificed. The results showed that in the second trimester of pregnancy, the embryo absorption rate of CBA *DBA/2 mice was 11.0 +3.0%. Poly I: C treatment significantly increased the embryo absorption rate to 40.2 +1.7%. It was accompanied by obvious pathological changes, such as the reduction of lumen and the thickening of tube wall. At the same time, the study also found that although the apparent examination of poly I:C treatment 24 hours after pregnancy (7.5 days) did not show obvious signs of embryonic absorption in CBA *DBA/2 mice, and compared with the control group, there was no significant change in IFN-gamma content at the maternal-fetal interface, but the expression of TLR3 in the decidual region was significantly up-regulated, and local. These results suggest that the secretion of TNF-a in decidua is an important inflammatory signal, which may inhibit the recasting of uterine spiral artery and ultimately affect the pregnancy outcome of CBA *DBA/2 mice.
In addition, 24 hours later, poly I:C treatment significantly increased the positive rates of TNF-alpha and IFN-gamma in decidual NK (DX5~+CD3~-) cells. To our surprise, flow cytometry (DX5~+CD3~+) and cytohistochemistry (DBA~+) analysis showed a significant decrease in the number of uterine NK cells at the maternal-fetal interface. NKT (DX5~+CD3~-), T (DX5~+CD3~-) cells and dendritic cells (CD11c~+) were not affected by poly I: C. In fact, every immune cell at the maternal-fetal interface was beneficial to the pregnancy process. Because of the downregulation of DX5~+ cells, the DC/NK ratio at the maternal-fetal interface was changed, which affected local factors including cytokines. Because DC-NK interactions do not require direct cell contact, the spatial distribution of DC and NK cells in decidua does not coincide completely, and the interaction between DC and NK cells should not be neglected.
In conclusion, this study demonstrates that peripheral NK cells have a greater migration advantage than T cells when lymphocytes are recruited at the maternal-fetal interface in early pregnancy (decidualization stage). Poly I:C not only affects the homing and tissue-specific distribution of NK cells, but also activates TLR3 signal in decidua. The highly activated uterine NK cells are manifested by high activation of TLR3 signal. The enhancement of cytokine secretion and the subsequent inflammatory response at the maternal-fetal interface can aggravate the pathological changes during pregnancy, such as the inhibition of spiral artery recasting, leading to the increase of embryo absorption. In bed research and practice, we treat reproductive immune diseases and predict pregnancy outcomes by focusing on the interaction between uterine NK cells and other immune cells.
【學(xué)位授予單位】：中國(guó)科學(xué)技術(shù)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2007
【分類號(hào)】：R392
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 張建紅;C57Bl/6J小鼠妊娠早-中期子宮自然殺傷細(xì)胞研究[J];生殖醫(yī)學(xué)雜志;2005年01期
本文編號(hào)：2246177
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