Ultrastructural and histochemical insights into neotenic and metamorphic axolotl lungs with clues to pulmonary regeneration

The axolotl, one of the preferred experimental animal models in embryological development and regeneration studies, shows strong regenerative capacity. This study aims to provide data for research aimed at enhancing the limited regenerative capacity of human lungs by investigating the cellular, interstitial, and ultrastructural changes in axolotl lungs during their adaptation to terrestrial life. Neotenic and metamorphic axolotls (n = 3, each group) were sacrificed under anaesthesia and lung samples were harvested. Lungs were examined with light microscopic (hematoxylin-eosin, Sudan IV, safranin-mallory, immunohistochemistry) and transmission electron microscopic (TEM) studies. Hematoxylin-eosin and safranin-mallory staining examinations revealed that in the neotenic group, alveolar walls were thick, air spaces were narrow, and collagen deposition was prominent. The metamorphic group exhibited expanded air spaces and dense elastic fibres in interstitial regions. Interstitial lipofibroblast-like (ILF-like) cells were shown in both groups with Sudan IV staining. The alveoli of both neotenic and metamorphic axolotls were lined with pneumocytes, characterised by microvilli, cytoplasm containing lamellar bodies, and round nuclei. Additionally, the blood-air barrier and lamellar bodies were found to exhibit mature structure in metamorphic axolotls. In the neotenic group, lamellar bodies are also present in ciliated columnar cells, whereas in the metamorphic group, these cells do not contain lamellar bodies. Club cells were detected in the metamorphic group. As a consequence of externally administered thyroid hormone-inducing metamorphosis in neotenic axolotls, there is a restructuring of histological features in the lungs to facilitate adaptation to terrestrial life. Nevertheless, metamorphic axolotl lung histology differs from the characteristic lung histology in mammals. By documenting the anatomical remodelling of the axolotl lung during metamorphosis, this study provides fundamental insights into vertebrate lung adaptation and tissue plasticity, offering a morphological framework relevant to pulmonary regeneration.