Exploration of Rat Fetal Lung Tissue Fixation Methods

doi:10.12300/j.issn.1674-5817.2025.053

Abstract

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

CLC Number:

R-332

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.

Figures/Tables 4

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.

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.

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.

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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