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Effect of aflatoxin B1 on ultrastructural changes of biliary epithelial cells in mice experimentally infected with Clonorchis sinensis
Hong Ki Min
Department of Parasitology, College of Medicine, Ewha Womans University, Seoul 120, Korea.
Abstract
The present study was carried out to examine the effect of a carcinogen, aflatoxin B1 on the ultrastructural changes of biliary epithelial cells in mice infected with Clonorchis sinensis. A total of 93 male albino mice(BALB/c strain) was divided into 3 groups; group I, treated with 1.0 ppm aflatoxin B1 for 12 weeks; group II, given 50 C. sinensis metacercariae, and group III, given 50 metacercariae and treated with 1.0 ppm aflatoxin B1 for 12 weeks. Three mice served for untreated-uninfected controls. From 4 weeks after the treatment and/or infection, three mice from each group were sacrificed at 4 week intervals up to the 40th week, and their hepatobiliary tissues were prepared for transmission electron microscopy. The most prominent ultrastructural changes in group I were remarkable enlargement of nuclear size, separation of nucleolus, dispersed chromatin granules in nuclei and increased dense granules along the inner membrane of nuclei. In the cytoplasm there was slight proliferation of mitochondria and endoplasmic reticulum (ER) at earlier stage. At the 12th week separation of fibrillar and granular components of the nucleolus was a characteristic finding. As the time elapsed, epithelial cells showed flattened-cuboidal form and a tendency of atrophy. Most of the nuclei were elongated and polygonal in shape. In group II the appearance of elaborate interwoven folds of lateral cytoplasm forming a labyrinth of interconnected intercellular space and variety in nuclear shape were the prominent findings at earlier stage. The cytoplasm showed slight proliferation and dilatation of mitochondria and ER, and a small number of mucin droplets. In the basement membrane scanty fibrous cells were seen. With time, variety in nuclear shape, marked proliferation and dilatation of rough ER and some collagen fibrils were demonstrated. Other features of intracellular organelles and mucin droplets persisted. In group III cuboidal epithelial cells showed their remarkably enlarged and irregular nuclei, increased chromatin granules in the nuclei, separated nucleoli, proliferated and dilated rough ER. With time, sequestered mitochondria showed bleb-like evaginations which lacked cristae and dense matrix, and were limited by a single membrane. Since the 20th week, microvilli were relatively scanty and poorly developed. Organelles and inclusions in the cytoplasm of metaplastic cells were poor. Nuclei were variable in shape. The most prominent changes at later stage were separation of nuclei from the cytoplasm, and appearance of numerous and irregularly angled electron dense granules in the nuclei.
Figures
Figs. 1-2 Fig. 1. Electron microscopic findings of normal epithelial cells of a bile ductule; The cells are solidly attached to one another, partly by interdigitations and partly by desmosomes(D). Microvilli are varying in size and length. The cells have an ovoidal nucleus(N) of low density and nucleoli are generally small in size. Scanty mitochondria, rough and smooth endoplasmic reticulum, Golgi complex and lysosome-like dense bodies are visible in the cytoplasm. (Lead stain, ×10,000)
Fig. 2. Electron microscopic findings of epithelial cells in group I at the 4th week; Enlargement of nucleus (N) together with dispersion of chromatin granules and increase of dense granules along the inner membrane of nucleus and nucleolar margination is characteristic. There are slight proliferations of mitochondria(Mit) and rough endoplasmic reticulum(RER). (Lead stain, ×10,000)
Figs. 3-5 Fig. 3. Electron microscopic findings of epithelial cells in group I at the 28th week; Flattened-cuboidal epithelial cells have numerous microvilli and an enlarged nucleus(N). (Lead stain, ×10,000)
Fig. 4. Electron microscopic findings of epithelial cells in group II at the 4th week; Columnar epithelial cells show scanty microvilli and occasionally cytoplasmic projection in the lumen. The presence of elaborate interwoven folds(IF) of lateral cytoplasm, forming a labyrinth of interconnected intercellular space is the most characteristic change. The nucleus(N) appeared to be elongated and irregular in shape. Slightly enlarged nucleoli are seen in evenly distributed nucleoplasm. (Lead stain, ×10,000)
Fig. 5. Electron microscopic findings of epithelial cells in group II at the 12th week; Variety in nuclear shape with electron dense granules along the inner membrane, marked proliferation and swelling of rough endoplasmic reticulum (RER), scanty mucin droplets and some collagen fibrils are seen. Separation of nucleus from cytoplasmic component is demonstrated. (Lead stain, ×7,000)
Figs. 6-8 Fig. 6. Electron microscopic findings of epithelial cells in group III at the 4th week; Enlarged nucleus(N) with increased chromatin granules, proliferated and slightly dilated endoplasmic reticulum, and some mucin droplets are seen. (Lead stain, ×6,000)
Fig. 7. Electron microscopic findings of epithelial cells in group III at the 12th week; Columnar epithelial cells show a greatly enlarged and elongated nucleus(N) with or without nucleolus. In most cells partially sequestered mitochondria(Mit) show bleb-like evaginations which lacked cristae and dense matrix, and are limited by a single membrane. (Lead stain, ×8,000)
Fig. 8. Electron microscopic findings of epithelial cells in group III at the 20th week; Flattened-cuboidal epithelial cells show stratification. The nuclei(N) are elongated and numerous electron dense and irregular granules are seen. Well developed collagen fibrils(CF) are noted in the junction structure and the basement membrane. (Lead stain, ×8,000)
Figs. 9-10 Fig. 9. Electron microscopic findings of epithelial cells in group III at the 32nd week; Microvilli are relatively scanty and poorly developed. The nuclei are varying in shape with numerous, irregularly angled and highly electron dense granules. Separation of the nuclei from cytoplasmic component(↑) is evident. The cytoplasmic organelles are poor. (Lead stain, ×5,000)
Fig. 10. Electron microscopic findings of epithelial cells in group III at the 32nd week; Similar ultrastructural findings with Figure 9 are noted. Some cells with irregular nucleus invades the surrounding tissue. (Lead stain, ×4,000)
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