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Abstract

To develop effective therapeutic strategies aimed at treating tumor metastasis, critical steps in this process must be better understood. For this purpose we have established a new model to visualize and quantify early metastasis. Murine CT-26 colon adenocarcinoma cells were stably transfected with green fluorescent protein (GFP). Tumor cells were intraportally delivered to the liver of Balb/c mice and subsequently tracked by intravital fluorescence microscopy. Coinjection of fluorescent beads and in vivo propidium iodide staining allowed examination of initial tumor cell arrest, extravasation, viability and proliferation. Results showed that GFP-transfection compared to conventional labeling procedures (Calcein, cytoplasmic microspheres) did not alter early metastatic properties. However, the long-term development of liver metastases expressing GFP was markedly reduced compared to wild type CT-26 tumor cells. An increase in the size and the number of liver metastases in T- and B-cell-deficient SCID mice suggested an immune response to the GFP transfected cells responsible for the reduced metastatic growth in wild-type mice. Based on our findings, this model can be used to examine the early steps of metastasis in vivo. However, in immunocompetent mice, the use of GFP-labeled tumor cells should be limited to tracking cell arrest and extravasation, whereas evaluations of long-term metastatic growth should be performed in immunodeficient mice.

Reduced Expression of the ROCK Inhibitor Rnd3 Is Associated with Increased Invasiveness and Metastatic Potential in Mesenchymal Tumor Cells

“…We first inoculated mock-transfected control cells and Rnd3 clone16 cells subcutaneously in nude mice, to verify their tumorigenic potential. Cells from both transfected populations developed tumors; however, after about 10 days, when tumors were around 250 mm 3 , the tumors were rejected, probably because of an immune response to EGFP expressing cells [36]. To avoid rejection, we inoculated the cells into SCID (Severe Combined Immuno-Deficiency) mice, which are deficient in T and B cells.…”

Role of Tumor Cell Adhesion and Migration in Organ-Specific Metastasis Formation

“…Mechanical arrest was observed by different groups in experimental studies where human TCs were injected intravenously in different animal models [8,9]. While multi-cellular emboli would arrest in the microcirculation in experimental metastasis models, parental tumors rather release single TCs into the circulation that can easily pass through the microvasculature of different organs [10].…”

Activation of NF-κB by the intracellular expression of NF-κB-inducing kinase acts as a powerful vaccine adjuvant

“…We also examined whether NIK can enhance immune responses against specific antigens by taking advantage of the immunogenicity of GFP (21,22), a jellyfish protein already incorporated into our adenoviral vectors. We generated human GFP-specific T cell lines by culturing peripheral blood mononuclear cells (PBMCs) with recombinant GFP protein and IL-2, as described for tetanusspecific T cell lines (23).…”