Genomic Instability and Telomere Fusion of Canine Osteosarcoma Cells

Abstract

Canine osteosarcoma (OSA) is known to present with highly variable and chaotic karyotypes, including hypodiploidy, hyperdiploidy, and increased numbers of metacentric chromosomes. The spectrum of genomic instabilities in canine OSA has significantly augmented the difficulty in clearly defining the biological and clinical significance of the observed cytogenetic abnormalities. In this study, eight canine OSA cell lines were used to investigate telomere fusions by fluorescence in situ hybridization (FISH) using a peptide nucleotide acid probe. We characterized each cell line by classical cytogenetic studies and cellular phenotypes including telomere associated factors and then evaluated correlations from this data. All eight canine OSA cell lines displayed increased abnormal metacentric chromosomes and exhibited numerous telomere fusions and interstitial telomeric signals. Also, as evidence of unstable telomeres, colocalization of γ-H2AX and telomere signals in interphase cells was observed. Each cell line was characterized by a combination of data representing cellular doubling time, DNA content, chromosome number, metacentric chromosome frequency, telomere signal level, cellular radiosensitivity, and DNA-PKcs protein expression level. We have also studied primary cultures from 10 spontaneous canine OSAs. Based on the observation of telomere aberrations in those primary cell cultures, we are reasonably certain that our observations in cell lines are not an artifact of prolonged culture. A correlation between telomere fusions and the other characteristics analyzed in our study could not be identified. However, it is important to note that all of the canine OSA samples exhibiting telomere fusion utilized in our study were telomerase positive. Pending further research regarding telomerase negative canine OSA cell lines, our findings may suggest telomere fusions can potentially serve as a novel marker for canine OSA.

Relative biological effectiveness in canine osteosarcoma cells irradiated with accelerated charged particles

“…The present study used canine OSA cell lines with various radiation sensitivities as reported previously (22). Abrams, D17 and Moresco were fairly resistant to gamma-rays (D 10 =7.12-9.33 Gy), while Grey was relatively radiosensitive (D 10 =2.72 Gy).…”

“…Cells were grown in minimal essential media (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% (v/v) fetal bovine serum (FBS; Sigma-Aldrich, St. Louis, MO, USA) and 1% (v/v) Pen/Strep and fungizone solution (Invitrogen; Thermo Fisher Scientific, Inc.), and they were maintained in a tissue culture incubator at 37˚C in a 100% humidified atmosphere of 5% CO 2 . The cell doubling times of these cells were 19 h for Abrams, 22 h for D17, 18 h for Grey and 22 h for Moresco (22).…”

“…The canine OSA cell lines Abrams, D17, Grey and Moresco were a gift from the Animal Cancer Center of Colorado State University (Fort Collins, CO, USA) (22,24). Cells were grown in minimal essential media (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% (v/v) fetal bovine serum (FBS; Sigma-Aldrich, St. Louis, MO, USA) and 1% (v/v) Pen/Strep and fungizone solution (Invitrogen; Thermo Fisher Scientific, Inc.), and they were maintained in a tissue culture incubator at 37˚C in a 100% humidified atmosphere of 5% CO 2 .…”

“…RBE values of 1.96-2.50 in canine squamous cell carcinoma, fibrosarcoma and hemangiopericytoma cell lines for carbon ions (LET=108 keV/µm) have been previously reported (21). Our previous study described canine OSA cell lines that were either radioresistant or highly radiosensitive to photon radiation, along with their basic cellular characteristics (22). In the present study, an in vitro comparison of the cell-killing effects of photons, protons and heavy ions on canine OSA cells was performed.…”

Genomische Aberrationen in caninen Mammakarzinomen und deren Nachweis in zellfreier plasmatischer DNA

“…copy-number Imbalanzen (CNI), oder Kopienzahl neutraler chromosomaler Umlagerungen (Negrini et al, 2010;Tang et al, 2010;Angstadt et al, 2011;Maeda et al, 2012). Bei humanen Tumoren sind außerdem tumorspezifische chromosomale Veränderungen beschrieben worden (Beroukhim et al, 2010).…”