Establishment of a New Human Osteosarcoma Cell Line, UTOS-1: Cytogenetic Characterization by Array Comparative Genomic Hybridization
T Yasuda, M Kanamori, T Hori, K Suzuki, S Nogami, T Kimura. University of Toyama, Toyama, Japan; Takaoka City Hospital, Takaoka, Toyama, Japan
Background: Osteosarcoma (OS) is the most common malignant bone tumor characterized by proliferation of tumor cells which produce osteoid or immature bone matrix. There have been several reports describing xenotransplantation models of human OS, but characterization of human OS at the molecular cytogenetic level has been limited. We describe the establishment and characterization of a new human OS cell line, designated as UTOS-1, derived from a conventional osteoblastic OS. In addition, we analyze chromosomal aberrations and DNA copy number changes in UTOS-1 by array comparative genomic hybridization (aCGH).
Design: Tumor cells, UTOS-1, from a typical osteoblastic OS of an 18-year-old man were cultured in RPMI 1640. To determine the tumorigenicity of the UTOS-1 cell line in vivo, tumor cells were injected subcutaneously into the leg of SCID mice. To determine the doubling time in vitro, we used MTT assay. To assess osteoblastic differentiation, we used 3 monoclonal antibodies: anti-osteopontin (OP), anti-osteocalcin (OC), and alkaline phosphatase (ALP). Expression of osteoblastic differentiation markers was also assessed by RT-PCR. For cytogenetic analysis, preparations of metaphase chromosomes from UTOS-1 cells at passage 15 were banded with Giemsa-trypsin. Array CGH was performed using the GenoSensor Array 300 system.
Results: Cultured UTOS-1 cells are spindle-shaped, and have been maintained in vitro for over 50 passages. The population-doubling time of the cells was 40 hours. UTOS-1 also exhibit morphological and immunohistochemical characteristics typical of osteoblastic OS. In RT-PCR, UTOS-1 cells expressed OP, OC and ALP. Chromosomal analysis by G-band showed 73-85 chromosomes with complicated translocations. Array CGH show frequent gains at locus DAB2 at chromosome 5q13, CCND2 at 12p13, MDM2 at 12q14.3-q15, FLI and TOP3A at 17p11.2-p12 and OCRL1 at Xq25, and show frequent losses at HTR1B at 6q13, D6S268 at 6q16.3-q21, SHGC17327 at 18ptel, and STK6 at 20q13.2-q13.3.
Conclusions: We have isolated and characterized a new permanent human cell line, UTOS-1, established from an osteoblastic OS. This cell line retains the morphology, osteoblastic activities and cytogenetic characteristics of the original tumor in vitro. The UTOS-1 cell line is useful for biologic and molecular pathogenetic studies of human OS.
Category: Bone & Soft Tissue
Tuesday, March 23, 2010 9:30 AM
Poster Session III # 13, Tuesday Morning