Bioluminescence imaging of angiogenesis in a murine orthotopic pancreatic cancer model

Abstract

Purpose: Angiogenesis is essential for physiological processes as well as for carcinogenesis. New approaches to cancer therapy include targeting angiogenesis. One target is VEGF-A and its receptor VEGFR2. In this study, we sought to investigate pancreatic cancer angiogenesis in a genetically modified VEGFR2-luc-KI mouse.

Procedures: Live in vivo bioluminescence imaging of angiogenesis was performed continuously until sacrifice in subcutaneous tumors as well as in orthotopically transplanted tumors. Tumor tissue was immunostained for CD-31 and VEGFR2.

Results: Peritumoral angiogenesis measured by light emission was detected beginning at week 3 following subcutaneous injection. In the orthotopic model, light emission began at day 4, which likely corresponds to wound healing, and continued throughout the experimental period during tumor growth. Peritumoral CD-31 vessel- and VEGFR2-staining were positive.

Conclusions: The VEGFR2-luc-KI mouse is a valuable tool to demonstrate tumor angiogenesis and seems to be suitable to evaluate anti-angiogenic approaches in pancreatic cancer.

Fig. 1

Imaging of a subcutaneous donor tumor. Male donor VEGFR2-luc-KI mice were injected with 2 × 10⁶ cells subcutaneously. They were imaged as described in “Materials and Methods” section. These mice have a relatively high background of VEGFR2 promoter activity and specific activity in the testicles, the intra-abdominal sexual organs, spleen, and liver. There was a steady increase in light emission at the tumor site over time. The insert shows the increase in tumor area (mm²). Error bars represent the SD of the measurement at each time point.

Fig. 2

Imaging of an orthotopic tumor. Pieces of the donor tumors were transplanted into male recipient VEGFR2-luc-KI mice, which were imaged weekly. Localized light emission was detected. Error bars represent the SD of the measurement at each time point.

Fig. 3

Imaging at sacrifice. After 6 weeks of tumor growth, the animals were imaged in vivo and post-mortem. The animal, tumors, and the skin which covered the tumors were imaged and quantified separately. In the post-mortem imaging, the total light emission is stronger because no light is absorbed through the tissue. Error bars represent the SD of the measurement at each time point

Fig. 4

Immunohistochemistry for CD-31 and VEGFR2. The peritumoral area shows distinct staining for CD-31 and weak staining for VEGFR2.