MTA1在子痫前期胎盘组织中的表达及其对滋养细胞功能的影响

doi:10.3969/j.issn.1674-8115.2024.11.005

上海交通大学学报（医学版） ›› 2024, Vol. 44 ›› Issue (11): 1383-1390.doi: 10.3969/j.issn.1674-8115.2024.11.005

• 论著 · 基础研究 • 上一篇

MTA1在子痫前期胎盘组织中的表达及其对滋养细胞功能的影响

耿瑶¹^,²(), 张杨², 赵洁², 李伟², 蔡国青³()

^1.西北大学生命科学学院，西安 710069
^2.中国人民解放军空军军医大学基础医学院人体解剖与组织胚胎学教研室，西安 710032
^3.中国人民解放军空军军医大学第一附属（西京）医院妇产科，西安 710032

收稿日期:2024-01-02 接受日期:2024-05-13 出版日期:2024-11-28 发布日期:2024-11-28
通讯作者: 蔡国青 E-mail:ygeng1119@163.com;184634529@qq.com
作者简介:耿　瑶（1999—），女，硕士生；电子信箱：ygeng1119@163.com。
基金资助:
国家自然科学基金(31971070);陕西省重点研发计划(2022SF-241)

Expression of MTA1 in preeclamptic placental tissue and its effects on trophoblast function

GENG Yao¹^,²(), ZHANG Yang², ZHAO Jie², LI Wei², CAI Guoqing³()

^1.College of Life Science, Northwest University, Xi′an 710069, China
^2.Department of Human Anatomy, Histology and Embryology, Basic Medical Science Academy, Air Force Military Medical University, Xi′an 710032, China
^3.Department of Gynaecology and Obstetrics, Xijing Hospital, Air Force Military Medical University, Xi′an 710032, China

Received:2024-01-02 Accepted:2024-05-13 Online:2024-11-28 Published:2024-11-28
Contact: CAI Guoqing E-mail:ygeng1119@163.com;184634529@qq.com
Supported by:
National Natural Science Foundation of China(31971070);General Projects of Shaanxi Provincial Key R & D Plan(2022SF-241)

摘要/Abstract

摘要：

目的·探讨转移相关蛋白1（metastasis-associated protein 1，MTA1）在子痫前期（preeclampsia，PE）胎盘组织中的表达及其对滋养细胞功能的影响。方法·采集PE孕妇胎盘标本（PE组，20例），以健康妊娠孕妇胎盘标本作为对照（对照组，35例）。采用蛋白质印迹法、免疫荧光双重染色分析MTA1的表达变化情况；培养人早孕期胎盘滋养层细胞系 HTR8/SVneo；通过细胞伤口愈合实验检测细胞迁移能力；通过Transwell细胞侵袭实验检测细胞侵袭能力；体外条件下模拟绒毛外滋养层细胞侵入子宫时的低氧环境，通过定量实时聚合酶链式反应检测低氧诱导下蛋白酶MMP-2、MMP-9的mRNA表达情况；构建绒毛外植体模型，通过绒毛外植体培养检测外植体整体绒毛外延能力；通过免疫组织化学方法检测内皮标志物CD31，进行小鼠胎盘血管形成分析；将Mta1^-/-雌性小鼠、野生型C57雌性小鼠各15只，与野生型雄性C57小鼠合笼，进行生育力测试。结果·蛋白质印迹法分析结果显示，与对照组比较，MTA1蛋白在PE组胎盘中的表达异常降低；免疫荧光双重染色结果显示MTA1主要定位于滋养层细胞胞核；细胞伤口愈合实验表明稳定敲低MTA1的人绒毛膜滋养层细胞HTR8/SVneo的细胞迁移能力弱于对照组（P=0.002），Transwell细胞侵袭实验显示细胞的侵袭性被显著抑制，明显低于对照组（P=0.015）；低氧刺激诱导的MMP-2、MMP-9表达水平显著降低（P=0.020，P=0.003）；MTA1敲低后外植体整体绒毛外延能力较对照组下降（P=0.003）；免疫组织化学结果显示Mta1^-/-雌鼠胎盘CD31表达明显低于野生型雌鼠（P=0.004）；Mta1^-/-雌鼠产仔数显著降低（P=0.000）。结论·MTA1表达水平与PE密切相关；内源性MTA1可能参与滋养细胞浸润子宫内膜及绒毛毛细血管重塑过程。

关键词: 子痫前期, 转移相关蛋白1, 绒毛外滋养层细胞

Abstract:

Objective ·To investigate the expression of metastasis-associated protein 1 (MTA1) in placental tissues of preeclampsia (PE) patients and its impact on trophoblast cell function. Methods ·Placental specimens were collected from pregnant women with PE (PE group, 20 cases) patients and healthy pregnant women as controls (control group, 35 cases). Western blotting and immunofluorescent double staining were performed to analyze the expression changes of MTA1. The human first-trimester placental trophoblast cell line HTR8/SVneo was cultured, and the cell migration ability was assessed through wound healing assay. The cell invasion ability was detected using Transwell invasion assay. Under hypoxic conditions simulating the invasion of extravillous trophoblasts into the uterus, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to analyze the mRNA expression of hypoxia-induced matrix metalloproteinases (MMP-2 and MMP-9), thus assessing their secretion levels. An extravillous trophoblast explant model was constructed to assess the overall villus outgrowth capacity of the explants. Immunohistochemistry (IHC) was performed to confirm the presence of the endothelial marker CD31 for placental angiogenesis analysis in mice. Fifteen Mta1^-/- female mice and fifteen wild-type C57 female mice were mated with wild-type male C57 mice for fertility testing. Results ·Western blotting revealed significantly decreased expression of MTA1 protein in placental tissues of the PE group compared to the control group. Immunofluorescent double staining showed that MTA1 was mainly localized in the nuclei of trophoblast cells. The wound healing assay demonstrated that HTR8/SVneo with stable MTA1 knockdown exhibited weaker cell migration ability compared to the control group (P=0.002). The Transwell invasion assay demonstrated a marked decrease in invasiveness in MTA1-knockdown cells, significantly lower than the control group (P=0.015). Hypoxia-induced expression levels of matrix metalloproteinases MMP-2 and MMP-9 were significantly reduced (P=0.020, P=0.003). After MTA1 knockdown, the overall villus outgrowth capacity of the explants was decreased compared to the control group (P=0.003). IHC results showed that CD31 expression in the placenta of Mta1^-/- female mice was significantly lower than that of wild-type female mice (P=0.004). The litter size of Mta1^-/- female mice was significantly reduced (P=0.000). Conclusion ·The expression level of MTA1 is closely related to PE. Endogenous MTA1 may be involved in trophoblast invasion into the endometrium and villous capillary remodeling.

Key words: preeclampsia (PE), metastasis-associated protein 1 (MTA1), extravillous trophoblast (EVT)

中图分类号:

R714.24

耿瑶, 张杨, 赵洁, 李伟, 蔡国青. MTA1在子痫前期胎盘组织中的表达及其对滋养细胞功能的影响[J]. 上海交通大学学报（医学版）, 2024, 44(11): 1383-1390.

GENG Yao, ZHANG Yang, ZHAO Jie, LI Wei, CAI Guoqing. Expression of MTA1 in preeclamptic placental tissue and its effects on trophoblast function[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2024, 44(11): 1383-1390.

图/表 4

图1 人胎盘组织MTA1蛋白的表达Note： A. Western blotting. B. Immunofluorescence assay. C. Expression site of MTA1 protein located by immunofluorescence double staining.

Fig 1 Expression of MTA1 protein in human placental tissues

图2 敲低 MTA1 对HTR8/SVneo细胞功能的影响Note： A. Expression of MTA1 protein in HTR8/SVneo cells with stable MTA1 knockdown by Western blotting. B. Wound healing assay. Representative images were taken at the beginning of wound healing (0 h) and 24 h later. The level of cell migration towards the wound scratches was quantified by measuring the distance of wound closure. C. Invasive capacity of the cells by Transwell in vitro invasion assay and statistical analysis. D. Expression levels of MMP-2 and MMP-9 by qRT-PCR. n=4. ①P=0.002, ②P=0.015, ③P=0.020, ④P=0.003.

Fig 2 Impact of MTA1 knockdown on the functional properties of HTR8/SVneo cells

图3 敲低 MTA1 对体外培养的绒毛外植体外延能力的影响Note： A. Expression of MTA1 protein in villous explants by Western blotting. B. The growth state of villus explants (arrows) under the inverted microscope. C. The average migration distance of villous explants. n=4. ①P=0.003.

Fig 3 Effect of knocking down MTA1 on epitaxial ability of cultured villous explants in vitro

图4 Mta1-/-小鼠的鉴定及其胎盘组织中CD31表达情况Note： A. PCR identification results of mouse tail genomic DNA. B. Detection of MTA1 expression in placental tissues by Western blotting. C. Immunohistochemical staining and quantitative analysis of CD31 protein in placental tissues of mice. n=4. ①P=0.004.

Fig 4 Identification of Mta1-/- mice and expression of CD31 in the placental tissues

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MTA1在子痫前期胎盘组织中的表达及其对滋养细胞功能的影响

Expression of MTA1 in preeclamptic placental tissue and its effects on trophoblast function

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