Origin of spontaneous and transplacentally induced mouse lung tumors from alveolar type II cells

Mouse lung tumors were induced transplacentally in offspring by treating C3H/HeNCrMTV- and Swiss Webster [Tac:(SW)fBR] mice during different periods of gestation with a single i.p. injection of N-nitrosoethylurea (ENU) at 0.5 mmol or 0.74 mmol/kg. Quantitative and qualitative evaluation of the lung tumors in the offspring at ages ranging from 1 week to 52 weeks was carried out by light microscopic study of hematoxylin and eosin-stained (H&E) serial and step sections. By nitroblue tetrazolium enzyme histochemistry, 3-hydroxybutyrate dehydrogenase (seen predominantly in Clara cells) was localized in frozen tissue sections. By avidin-biotin peroxidase complex immunohistochemistry, various specific cellular and nuclear markers were investigated on paraffin sections (antisera against surfactant apoprotein, Clara cell antigen, lysozyme, and 5-bromo-2' deoxyuridine). Normal lung and lung tumors were also studied by electron microscopy. A histological method was developed to assess all lesions present in the entire lung. It was shown that solid and papillary tumor types arose individually and that mixed solid/papillary forms represented a progression of the benign solid adenoma to the malignant papillary carcinoma. Immunocytochemical localization of DNA synthesis with 5-bromo-2' deoxyuridine gave the highest labeling indices at early stages of tumor growth. As the size of the papillary tumors increased, fewer nuclei were labeled/mm2 of tumor section. Lack of both specific Clara cell antigen and 3-hydroxybutyrate dehydrogenase and the absence of typical nonosmiophilic Clara cell granules indicated a cell of origin other than Clara cells. Evidence for alveolar type II cell origin of both solid and papillary neoplasms in spontaneous and induced tumors was found in the expression of surfactant apoprotein, the presence of mature lamellar bodies (solid tumors) or small lamellar bodies, and immature stages of lamellar bodies (papillary tumors). Lysozyme was present in mature alveolar type II cells and solid tumors but absent in fetal lung and papillary neoplasms. Tumors induced on gestation day 14 or day 16 had all developed by 2 weeks of age and generally did not increase in multiplicity with age, whereas those induced on day 18 showed a protracted development with regard to frequency, growth (size), and progression. The multiplicity of mouse lung tumors induced at different stages of fetal development paralleled the number of alveolar type II precursor cells (i.e., followed a bell-shaped pattern peaking on day 16 of gestation).