Quantifying intra‐osseous growth of osteosarcoma in a murine model with radiographic analysis

Abstract

ABSTRACT:The orthotopic murine osteosarcoma model is an excellent representation of the human condition as mice develop rapid growth of 'primary' tumor with subsequent lung metastasis. Currently, monitoring tumor growth relies on measuring pulmonary metastases occurring four weeks post injection. Studies show that amputation of the tumor-bearing limb is required before pulmonary metastases are detectable due to rapid growth causing morbidity. Thus, a method measuring 'primary' tumor growth independent of metastasis is required. We hypothesized that serial radiography would allow for longitudinal quantification of 'primary' osteosarcoma growth and explored this idea by utilizing the tibial orthotopic model. Tumor growth was monitored weekly by radiography and calipers, and results were compared with mCT and histology. We found that radiographs demonstrate extra and intra-osseous tumor growth by displaying lytic and blastic lesions and the surrounding radio-opaque area enlarged significantly (p < 0.0001) allowing for quantification. Additionally, radiographs proved more precise than indirect caliper measurements (intra-observer error AE6.64%: interobserver error AE15.84%). Therefore, we determined that radiography provides accurate, longitudinal quantification of 'primary' osteosarcoma tumor that can be performed serially in the same mouse, does not require introduction of bioluminescence to the host or cell, and is more precise than the current caliper method. ß

Thrombin induces osteosarcoma growth, a function inhibited by low molecular weight heparin in vitro and in vivo

“…In a previous study, 22 x-ray analysis of the soft tissue masses (at 4 weeks) averaged an intraobserver error (n ¼ 3) of AE6.64% and interobserver error of AE15.84% (n ¼ 3), with a normal distribution and standard deviation of 23%. Five mice per group was determined to detect a 25% reduction in tumor size, with a statistical power (b) of .8 and a ¼ .05 (PS Power and Sample Size Calculations software 24 ).…”

“…Assessment for enlargement of the soft tissues surrounding the injected tibia was conducted by analysis of radiographs using Image J (version 1.44u, National Institutes of Health, Bethesda, Md) as previously described. 22 Tumor growth was calculated weekly as the percentage increase in size of the injected tibia compared with the paired contralateral control. In addition, magnetic resonance imaging (MRI-9.4T Inova imaging system; Varian, Palo Alto, Calif) 23 and histological examination confirmed tumor size (mice that did not develop histological evidence of tumor were excluded from the study).…”

Micro-Computed Tomography Derived Anisotropy Detects Tumor Provoked Deviations in Bone in an Orthotopic Osteosarcoma Murine Model

“…A circumference was drawn around the soft tissue of the tibia in the following manner: defining the circumference with borders defined by the patella superiorly, the tissue/tibia interface inferiorly, the skin anteriorly and the gastrocnemius muscle definition posteriorly. The area of the resultant circle was analyzed [12] [13].Tumor growth was calculated weekly as the percentage increase in size of the injected tibia compared with the paired contralateral, non-injected tibia.…”

“…All tissue was then processed into paraffin and stained with H&E and Martius scarlet blue as previously described [12].…”

Autocrine VEGF/VEGFR1 Signaling in a Subpopulation of Cells Associates with Aggressive Osteosarcoma

“…Weekly radiographic evaluations were performed using a LX-60 Radiography System (Faxitron X Ray; 8 seconds at 35 kV). Assessment of the soft tissue silhouette surrounding the injected tibia was conducted by analysis of radiographs using Image J (version 1.44 u; NIH) as previously described (21). Tumor growth was calculated weekly as the percentage increase in size of the injected tibia compared with the paired contralateral control.…”