研究牙周病常见的条件致病菌中间普氏菌（Pi）致蜕膜炎症引发C57BL/6孕鼠死胎的动物模型的建立方法，初步分析蜕膜炎症部位炎症相关基因表达水平。

经尾静脉注射5 × 10⁶ CFU Pi（ATCC25561）至孕15~ 16日雌鼠体内，分别在12、24和48 h后处死小鼠后解剖，剥离胎鼠胎盘组织，其中部分胎盘组织分离为蜕膜组织和非蜕膜胎盘组织，用于后续实时荧光定量聚合酶链反应（PCR）检测10种细胞因子和7种信号通路关键分子的mRNA表达水平。另外，两种对照组分别是尾静脉注射大肠埃希氏菌（E.coli）JM109（E.coli组）和磷酸盐缓冲液（PBS组），尾静脉注射Pi为实验组（Pi组），注射后48 h后处死孕鼠，计数胎鼠数量和死亡胎鼠数量，比较胎鼠死亡率。采用独立样本t检验分析细胞因子、信号分子的mRNA表达水平差异。卡方检验分析Pi组和E.coli组、PBS组之间孕鼠体内胎鼠死亡率的差异。

静脉注射Pi（5 × 10⁶ CFU）、E.coli（5 × 10⁶ CFU）和PBS 48 h后导致孕鼠的胎鼠死亡率分别为39.2%、3.0%和0%，实验组死亡率均显著高于对照组死亡率（Pi组vs. E.coli组：χ²= 17.54，P<0.001；Pi组vs. PBS组：χ²= 21.49，P<0.001）。实时荧光定量PCR检测结果显示，与PBS组相比，Pi组胎盘组织内细胞因子白细胞介素（IL）-1α、IL-1β、肿瘤坏死因子α（TNF-α）、IL-6、粒细胞集落刺激因子（GCSF）、人IL-8同系物（KC）和信号分子环氧化酶2（COX2）、核因子κB （NF-κB）、髓样细胞分化因子88（MyD88）、Toll样受体4（TLR4）至少在注射24 h后mRNA表达水平显著升高（均P<0.05）。而干扰素γ（IFN-γ）、IL-12、IL-18、IL-10、前列腺素E合成酶（PGES）、COX1、诱导白细胞介素B的TIR相关区域接受蛋白（Trif）差异无统计学意义。进一步比较胎盘内蜕膜和非蜕膜组织IL-1α、IL-1β、TNF-α、COX2的表达水平证实炎症发生主要位于胎盘内蜕膜组织。

牙周病常见的条件致病菌Pi静脉注射可导致孕鼠出现死胎，潜在机制与蜕膜部分的急性炎症关系密切。

To construct the model of circulating Prevotella intermedia (Pi) inducing stillbirth in C57BL/6 pregnant mice via decidual inflammation and preliminarily analyze the relevant expression levels of inflammation-related genes in deciduas.

C57BL/6 Pregnant mice for 15 or 16 d of gestation were infected with 5 × 10⁶ CFU Pi (ATCC25561) by I.V. injection method. Then, the mice were sacrificed and dissected at 12, 24 and 48 h post infection separately. The placentas (some placentas were separated as decidual tissues and non-decidual tissues) were harvested from each pregnant mouse, weighed, and homogenized in sterile PBS, and subsequently used as the templates of quantitative Real-time PCR assay for detection of expression levels of 10 kinds of cytokines and 7 kinds of signal molecules. In addition, birth outcome of pregnant mice in response to I.V. infection of 5 × 10⁶ CFU Pi (Pi group) was evaluated in comparison to 5 × 10⁶ CFU E.coli JM109 (E.coli group) and PBS (PBS group) as well. Total pups and dead pups were counted for each mouse. The fetal death rate was expressed as the percentage of dead fetuses from the total number of pups born to each mother. Differences of mRNA expression of cytokines and signal molecules between two groups were compared using the Independent Samples t test. Chi-square test was used to evaluate the differences of mortality rates of pups among Pi, E.coli and PBS groups.

The fetal death rate were calculated as 39.2%, 3.0% and 0% for pregnant mice infected with Pi, E.coli and PBS individually. The rate of Pi group were significantly higher than E.coli group (χ²= 17.54, P<0.001) and PBS group (χ²= 21.49, P<0.001) . compared to PBS group, the mRNA expression of cytokines (IL-1α, IL-1β, TNF-α, IL-6, GCSF and KC) and signal molecules (COX2, NF-κB, MyD88 and TLR4) were significantly enhanced at 24 hours post infection (all P<0.05) . However, the mRNA expression of IFN-γ, IL-12, IL-18, IL-10, PGES, COX1 and Trif didn′t presented significant differences between two groups. Furthermore, the location of placental inflammation was demonstrated to be deciduas after comparison of mRNA expression of IL-1α, IL-1β, TNF-α and COX2 between decidual tissues and non-decudual tissues.

As one of the common oral conditional anaerobe frequently found in periodontal diseases, the I.V. injection of Pi hold the capability to induce the stillbirth in pregnant mice. The potential mechanism was associated with acute inflammatory response occurred in decidual tissues of placentas.