大鼠胎儿期肺组织固定方法的探索

doi:10.12300/j.issn.1674-5817.2025.053

摘要/Abstract

摘要：

目的基于胎鼠独特的肺组织生理结构和现有的大鼠肺组织固定方法，探索胎鼠肺组织病理学研究所需的取材与固定方法。方法对6只妊娠20.5 d的成年SD大鼠实行剖宫产，获得胎鼠。在每只孕鼠所产胎鼠中选取4只体重、生命体征与呼吸状态相近的健康存活胎鼠，按照随机数表法随机分为直接取肺浸润组、气管内灌注后取肺浸润组、胎鼠整体浸润组和气管内灌注后胎鼠整体浸润组。为系统对比分析不同固定方法下肺组织的解剖形态，对4组胎鼠的肺组织样本进行取材、灌注和固定处理后，观察各组肺组织的大体形态；经过石蜡切片及苏木精-伊红（hematoxylin-eosin，HE）染色后，在光学显微镜下观察肺整体、肺泡和肺支气管的组织学形态。结果胎鼠直接取肺浸润组的肺门不清晰、肺组织分叶不明显、形状不规则，肺内空腔小，肺泡与肺支气管皱缩。气管内灌注后取肺浸润组的肺门清晰，肺组织分叶明显、形状规则，肺内空腔大，肺泡与肺支气管饱满。胎鼠整体浸润组与气管内灌注后胎鼠整体浸润组均能看到肺脏、心脏和皮肤等器官，肺组织分叶明显、形状不规则，肺叶边缘有破损。其中胎鼠整体浸润组胎鼠胸廓扁平，肺内空腔小，肺泡与肺支气管皱缩；而气管内灌注后胎鼠整体浸润组胎鼠胸廓饱满，肺内空腔大，肺泡与肺支气管较饱满。结论气管内灌注后取肺浸润法在胎鼠肺组织原始形态还原度及石蜡切片、染色和组织病理学观察与分析等方面皆优于直接取肺浸润法、胎鼠整体浸润法和气管内灌注后胎鼠整体浸润法，且该方法中包埋、切片和染色等操作流程简便、耗材节省。因此，对于大鼠胎儿期的肺组织病理学观察，推荐使用气管内灌注后取肺浸润法进行固定。

关键词: 大鼠, 胎儿期, 肺组织, 固定方法

Abstract:

Objective This study explores the methods of lung tissue extraction and fixation required for pathological studies of fetal rats, based on the unique physiological structure of fetal rat lung tissue and existing lung tissue fixation techniques for adult rats. Methods Six pregnant adult SD rats at 20.5 days of gestation were subjected to cesarean section to obtain fetal rats. Four healthy fetal rats with similar body weight, vital signs, and respiratory status were selected from each pregnant rat, and they were randomly divided into the following groups using a random number table: direct lung infiltration group, lung infiltration group after intratracheal infusion, whole-body infiltration group of fetal rats, and whole-body infiltration group after intratracheal infusion of fetal rats. To systematically compare and analyze the anatomical morphology under different fixation methods, lung tissues from four groups of fetal rats were harvested, perfused, and fixed, and the gross morphology of lung tissues in each group was observed. Paraffin sections were prepared and stained with Hematoxylin-Eosin (H&E). The histological morphology of the whole lung, alveoli, and bronchi was further examined under optical microscopy. Results In the direct lung infiltration group, the hilar structures were unclear, lung lobation was indistinct, the shape was irregular, lung cavities were small, and alveoli and bronchi were shrunken. In the lung infiltration group after intratracheal infusion, the hilar structures were clear, lobation was pronounced, the shape was regular, lung cavities were large, and alveoli and bronchi were full. Both the whole-body infiltration group and whole-body infiltration group after intratracheal infusion of fetal rats exhibited visible lungs, hearts, skins, and other organs. The lung tissues of both groups showed obvious lobulation, irregular shape, and damage at the margins of lung lobes. In the whole-body infiltration group, the thoracic cavities of the fetus were flattened, lung cavities were small, and alveoli and bronchi were shrunken. In the whole-body infiltration group after intratracheal infusion of fetal rats, the fetal thoracic cavities were full, lung cavities were large, and alveoli and bronchi were relatively full. Conclusion The lung infiltration after intratracheal infusion method for fetal rat lung tissue fixation outperforms direct lung infiltration, whole-body infiltration of fetal rats, and whole-body infiltration after intratracheal infusion of fetal rats in terms of preservation of the lung tissue's original morphology, paraffin sectioning, staining, and pathological observation and analysis. The embedding, sectioning, and staining processes are also simple and save consumables. Therefore, intratracheal infusion followed by lung infiltration method is recommended for fixation in histopathological observation of fetal rat lung tissue.

Key words: Rats, Fetal stage, Lung tissue, Fixation methods

中图分类号:

R-332

刘力瑜,嵇波,刘小玄,等. 大鼠胎儿期肺组织固定方法的探索[J]. 实验动物与比较医学, 2025, 45(4): 432-438. DOI: 10.12300/j.issn.1674-5817.2025.053.

LIU Liyu,JI Bo,LIU Xiaoxuan,et al. Exploration of Rat Fetal Lung Tissue Fixation Methods[J]. Laboratory Animal and Comparative Medicine, 2025, 45(4): 432-438. DOI: 10.12300/j.issn.1674-5817.2025.053.

图/表 4

图1 胎鼠肺组织四种固定方法的流程示意图注：A，直接取肺浸润法；B，气管内灌注后取肺浸润法；C，胎鼠整体浸润法；D，气管内灌注后胎鼠整体浸润法。

Figure 1 Flowchart of four fixation methods for fetal rat lung tissuesNote： A， direct lung infiltration method； B， lung infiltration method after intratracheal infusion； C， whole-body infiltration method of fetal rats； D， whole-body infiltration method after intratracheal infusion of fetal rats.

图2 四种固定方法下的胎鼠肺组织形态比较注：A，胎鼠肺组织在四种固定方法下的大体观察结果，左图为冠状面仰卧位，右图为冠状面俯卧位；B~E为胎鼠肺组织在四种固定方法下的HE染色结果，其中B为直接取肺浸润法，C为气管内灌注后取肺浸润法，D为胎鼠整体浸润法，E为气管内灌注后胎鼠整体浸润法。绿色框内为肺门，绿色箭头所示为肺内血管，黑色箭头所示为肋骨、肌肉、心脏和肺脏破损处。

Figure 2 Comparison of fetal rat lung tissue morphology using four fixation methodsNote： A， comparison of fetal rat lung tissue morphology under gross observation using four fixation methods. The left image shows the coronal view in the supine position， while the right image displays the coronal view in the prone position. B-E show the HE staining results of fetal rat lung tissues under four fixation methods. B， direct lung infiltration method； C，lung infiltration method after intratracheal infusion； D， whole-body infiltration method of fetal rats； E， whole-body infiltration method after intratracheal infusion of fetal rats. The green boxes indicate the lung hilum， the green arrows indicate intrapulmonary blood and the black arrows indicate the ribs， muscles， heart， and sites of lung damage.

图3 四种固定方法下的胎鼠肺泡HE染色结果（×200）注：A，直接取肺浸润组；B，气管内灌注后取肺浸润组；C，胎鼠整体浸润组；D，气管内灌注后胎鼠整体浸润组。箭头指示为肺泡，框内为肺泡腔内杂质。

Figure 3 Results of HE staining of fetal rat alveoli using four fixation methods （×200）Note： A， direct lung infiltration group； B， lung infiltration group after intratracheal infusion； C， whole-body infiltration group of fetal rats； D， whole-body infiltration group after intratracheal infusion of fetal rats. Arrows indicate alveoli， and boxes show impurities in the alveolar lumina.

图4 四种固定方法下的胎鼠肺支气管HE染色结果（×200）注：A，直接取肺浸润组；B，气管内灌注后取肺浸润组；C，胎鼠整体浸润组；D，气管内灌注后胎鼠整体浸润组。绿色箭头指示为肺支气管；黑框指示被放大区域内为单层柱状上皮细胞；三角形处为肺组织内的血管。

Figure 4 Results of HE staining of fetal rat lung bronchi usng four fixation methods （×200）Note： A， direct lung infiltration group； B， lung infiltration group after intratracheal infusion； C， whole-body infiltration group of fetal rats； D， whole-body infiltration group after intratracheal infusion of fetal rats. The green arrows indicate bronchi of lung， the boxed areas highlight the monolayer columnar epithelial cells and the triangular regions indicate the blood vessels within the lung tissue.

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