实验动物与比较医学 ›› 2025, Vol. 45 ›› Issue (4): 457-465.DOI: 10.12300/j.issn.1674-5817.2025.035

• 动物实验技术与方法 • 上一篇    下一篇

探头式激光共聚焦成像技术用于小鼠消化道组织形态特征分析

刘月琴1,2, 薛卫国1, 王淑友2, 申耀华1, 贾术永2, 王广军2, 宋晓晶2()()   

  1. 1.北京中医药大学针灸推拿学院, 北京 100029
    2.中国中医科学院针灸研究所, 北京 100700
  • 收稿日期:2025-03-04 修回日期:2025-04-13 出版日期:2025-08-25 发布日期:2025-09-01
  • 通讯作者: 宋晓晶(1984—),女,博士,副研究员,硕士生导师,研究方向:针灸效应机制和经穴理化特性的影像学表征。E-mail:xts2010@163.com。ORCID:0000-0002-9382-285X
  • 作者简介:刘月琴(1999—),女,硕士研究生,研究方向:针刺治疗脑病的机制研究。E-mail:15614330337@163.com
  • 基金资助:
    国家自然科学基金面上项目“电针调控‘肠-肝轴’保护酒精性肝损伤的效应和机制研究”(82274650)

Observation of Digestive Tract Tissue Morphology in Mice Using Probe-Based Confocal Laser Endomicroscopy

LIU Yueqin1,2, XUE Weiguo1, WANG Shuyou2, SHEN Yaohua1, JIA Shuyong2, WANG Guangjun2, SONG Xiaojing2()()   

  1. 1.School of Acupuncture-Moxibustion and Tuina, Beijing University of Chinese Medicine, Beijing 100029, China
    2.Institute of Acupuncture & Moxibustion, China Academy of Chinese Medical Sciences, Beijing 100700, China
  • Received:2025-03-04 Revised:2025-04-13 Published:2025-08-25 Online:2025-09-01
  • Contact: SONG Xiaojing (ORCID: 0000-0002-9382-285X), E-mail: xts2010@163.com

摘要:

目的 探讨活体探头式激光共聚焦成像(probe-based confocal laser endomicroscopy,pCLE)技术在快速检测与评估小鼠消化道组织形态特征中的应用价值。 方法 将12只6周龄的SPF级雄性昆明小鼠随机分为两组,其中6只使用乙醇体积分数为52%的红星牌二锅头白酒进行灌胃造模,另外6只使用生理盐水灌胃作为对照。连续灌胃28 d后,每组随机选取3只,经3%异氟烷吸入深度麻醉后颈椎脱臼处死,即刻剖取胃、十二指肠、空肠和直肠组织,浸入1%荧光素钠溶液染色,采用pCLE技术观察各组织黏膜表面的微观结构。剩余模型组和对照组小鼠经3%异氟烷吸入深度麻醉后,先后使用生理盐水、4%多聚甲醛溶液进行心脏灌流,剪取胃、十二指肠、空肠及直肠各组织进行脱水、切片和苏木精-伊红(hematoxylin-eosin,HE)染色,于光学显微镜下观察各节段组织的形态学变化。 结果 pCLE观察显示,对照组小鼠消化道黏膜组织表面荧光染色均匀,胃小凹、肠绒毛及肠隐窝形状完整,排列紧密,边缘结构清晰;而模型组小鼠消化道黏膜发生肿胀和变形,并伴有荧光染色不均匀和荧光素渗漏的现象。此外,部分组织出现缺损或细胞脱落,相邻特征结构(如胃小凹、肠隐窝)之间的边界模糊。HE染色观察显示,对照组小鼠消化道组织结构正常,细胞排列整齐、无缺损,黏膜下腺体大小一致且无增生性改变,无明显炎性细胞浸润;而模型组小鼠部分消化道组织结构缺损,细胞排列紊乱稀疏,黏膜下腺体萎缩,并伴有明显的炎性细胞浸润。pCLE观察结果与HE染色的组织学特征一致。 结论 pCLE能够实现对消化道黏膜结构特征的快速、实时、大范围、高分辨率显微成像,并且能更真实、全面地展示其生理及微观结构特征,该技术在小动物消化系统损伤的组织学评估研究中显示出良好的应用前景和实际应用价值。

关键词: 慢性酒精暴露损伤, 探头式激光共聚焦显微镜, 消化道, 小鼠

Abstract:

Objective To explore the application value of probe-based confocal laser endomicroscopy (pCLE) in rapidly detecting and evaluating the morphological characteristics of digestive tract tissues in mice. Methods Twelve male SPF Kunming mice aged 6 weeks were randomly divided into two groups. Six mice were subjected to gastric gavage with 52% Red Star Erguotou to establish the model, and six were given saline by gastric gavage as a control. After 28 days of modeling, 3 mice were randomly selected from each group. After deep anesthesia induced by inhalation of 3% isoflurane, the mice were sacrificed by cervical dislocation. The stomach, duodenum, jejunum, and rectum tissues were excised and immersed in 1% fluorescein sodium solution for staining. The microstructure of the mucosal surface of each tissue was observed using pCLE. The remaining mice in the model group and the control group were deeply anesthetized by inhaling 3% isoflurane, then cardiac perfusion was performed successively with saline and 4% paraformaldehyde. The stomach, duodenum, jejunum, and rectum tissues were excised for dehydration, section and hematoxylin-eosin (HE) staining, and the morphological changes of the tissues were observed under a microscope. Results Under pCLE imaging, fluorescence staining on the surface of the gastrointestinal mucosa was uniform in the control group; the morphology of gastric pits, intestinal villi, and intestinal crypts was intact, arranged compactly, and had distinct boundaries. In the model group, the gastrointestinal mucosa exhibited mucosal swelling and deformation, with uneven fluorescence staining and fluorescein leakage. Furthermore, some tissues showed defects or cell shedding, and the boundaries between adjacent characteristic structures (e.g., gastric pits, intestinal crypts) were blurred. HE staining showed that the gastrointestinal tissue structure of the control group mice was normal and well-organized, with no structural defects. Moreover, submucosal glands were uniform in size, with no hyperplasia observed, and no obvious inflammatory cell infiltration. In the model group, some gastrointestinal mucosal structures were defective and sparsely arranged; submucosal glands showed atrophy, accompanied by obvious inflammatory cell infiltration. The histological characteristics detected by pCLE were consistent with those of HE staining. Conclusion pCLE can be used to obtain rapid, real-time, large-scale, and high-resolution microscopic imaging of the gastrointestinal mucosa, realistically and comprehensively displaying its physiological and microstructural characteristics. It shows promising prospects and practical utility in the histological evaluation of digestive system injuries in small animals.

Key words: Chronic alcohol exposure injury, Probe-based confocal laser endomicroscopy, Digestive tracts, Mice

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