Morphological and Immunohistochemical Characterization of Canine Osteosarcoma Spheroid Cell Cultures.
Spheroid cell culture emerges as powerful in vitro tool for experimental tumour research. In this study, we established a scaffold-free three-dimensional spheroid system built from canine osteosarcoma (OS) cells (D17). Spheroids (7, 14 and 19 days of cultivation) and monolayer cultures (2 and 7 days of cultivation) were evaluated and compared on light and electron microscopy. Monolayer and spheroid cultures were tested for vimentin, cytokeratin, alkaline phosphatase, osteocalcin and collagen I by means of immunohistochemistry. The spheroid cell culture exhibited a distinct network of collagen I in particular after 19-day cultivation, whereas in monolayer cultures, collagen I was arranged as a lamellar basal structure. Necrotic centres of large spheroids, as observed in 14- and 19-day cultures, were characterized by significant amounts of osteocalcin. Proliferative activity as determined by Ki-67 immunoreactivity showed an even distribution in two-dimensional cultures. In spheroids, proliferation was predominating in the peripheral areas. Metastasis-associated markers ezrin and S100A4 were shown to be continuously expressed in monolayer and spheroid cultures. We conclude that the scaffold-free spheroid system from canine OS cells has the ability to mimic the architecture of the in vivo tumour, in particular cell-cell and cell-matrix interactions.
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine, Vienna, Austria.
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Gebhard
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine, Vienna, Austria.
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Gabriel
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine, Vienna, Austria.|Vienna VetCore Facility for Research, University of Veterinary Medicine, Vienna, Austria.
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Walter
D000469:Alkaline Phosphatase / Q000032:analysis; D000818:Animals; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D024042:Collagen Type I / Q000032:analysis; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004285:Dogs; D007150:Immunohistochemistry / Q000662:veterinary*; D007633:Keratins / Q000032:analysis; D046529:Microscopy, Electron, Transmission / Q000662:veterinary; D015675:Osteocalcin / Q000032:analysis; D012516:Osteosarcoma / Q000473:pathology*; D000071999:S100 Calcium-Binding Protein A4 / Q000378:metabolism*; D018874:Spheroids, Cellular / Q000166:cytology* / Q000648:ultrastructure*; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000032:analysis; D000099149:Ezrin
D000469:Alkaline Phosphatase; D024042:Collagen Type I; D003598:Cytoskeletal Proteins; D007633:Keratins; D015675:Osteocalcin; D000071999:S100 Calcium-Binding Protein A4; D014746:Vimentin; D000099149:Ezrin