Isolation, establishment, and characterization of ex vivo equine melanoma cell cultures.
Gray horses spontaneously develop metastatic melanomas that resemble human disease, and this is often accompanied with metastasis to other organs. Unlike in other species, the establishment of primary equine melanoma cultures that could be used to develop new therapeutic approaches has remained a major challenge. The purpose of the study was to develop a protocol for routine isolation and cultivation of primary equine melanocytes. Melanoma tissues were excised from 13 horses under local anesthesia, mainly from the perianal area. The melanoma cells were isolated from the melanoma tissue by serial enzymatic digestion using dispase and collagenase. Out of the 13 excised melanomas, cell cultures from eight melanomas were established, which corresponded to a success rate 62%. These cells showed different degrees of melanin pigmentation. Characterization of these cells using confocal microscopy, FACs analysis and western blotting showed that they expressed melanoma-associated antigens; Melan-A, MAGE-1, and MAGE-3, and PCNA expression was higher in fast-proliferating isolates. The protocol we developed and established proved successful for routine isolation and cultivation of primary equine melanoma cells. This method provided a large number of primary equine melanoma cells that could be used to study new therapeutic approaches for treatment of equine melanomas.
Chapman
Sarah W Kamau
SW
Institute of Veterinary Biochemistry and Molecular Biology, Vetsuisse Faculty, University of Zurich, Winterthurerstr. 190, CH-8057, Zurich, Switzerland.
Metzger
Nadine
N
Grest
Paula
P
Feige
Karsten
K
von Rechenberg
Brigitte
B
Auer
Jörg A
JA
Hottiger
Michael O
MO
D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism; D015153:Blotting, Western; D002469:Cell Separation / Q000379:methods*; D005434:Flow Cytometry; D006736:Horses; D008545:Melanoma / Q000473:pathology*; D018613:Microscopy, Confocal; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D013194:Staining and Labeling; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D018809:Proliferating Cell Nuclear Antigen