Orthotopic transplantation of v-src–expressing glioma cell lines into immunocompetent mice: establishment of a new transplantable in vivo model for malignant glioma

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Object

The aim of this study was to develop and characterize a new orthotopic, syngeneic, transplantable mouse brain tumor model by using the cell lines Tu-9648 and Tu-2449, which were previously isolated from tumors that arose spontaneously in glial fibrillary acidic protein (GFAP)-v-src transgenic mice.

Methods

Striatal implantation of a 1-μl suspension of 5000 to 10,000 cells from either clone into syngeneic B6C3F1 mice resulted in tumors that were histologically identified as malignant gliomas. Prior subcutaneous inoculations with irradiated autologous cells inhibited the otherwise robust development of a microscopically infiltrating malignant glioma. Untreated mice with implanted tumor cells were killed 12 days later, when the resultant gliomas were several millimeters in diameter. Immunohistochemically, the gliomas displayed both the astroglial marker GFAP and the oncogenic form of signal transducer and activator of transcription–3 (Stat3). This form is called tyrosine-705 phosphorylated Stat3, and is found in many malignant entities, including human gliomas. Phosphorylated Stat3 was particularly prominent, not only in the nucleus but also in the plasma membrane of peripherally infiltrating glioma cells, reflecting persistent overactivation of the Janus kinase/Stat3 signal transduction pathway. The Tu-2449 cells exhibited three non-random structural chromosomal aberrations, including a deletion of the long arm of chromosome 2 and an apparently balanced translocation between chromosomes 1 and 3. The GFAP-v-src transgene was mapped to the pericentromeric region of chromosome 18.

Conclusions

The high rate of engraftment, the similarity to the high-grade malignant glioma of origin, and the rapid, locally invasive growth of these tumors should make this murine model useful in testing novel therapies for human malignant gliomas.

Abbreviations used in this paper:FISH = fluorescence in situ hybridization; GBM = glioblastoma multiforme; GFAP = glial fibrillary acidic protein; IL-6 = interleukin-6; PNST = peripheral nerve sheath tumor; pY-Stat3 = tyrosine-705 phosphorylated signal transducer and activator of transcription–3; SSC = standard saline citrate; TBS = Tris-buffered saline.

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Contributor Notes

Address reprint requests to: Henry M. Smilowitz, Ph.D., Department of Pharmacology, University of Connecticut Health Center, Farmington, Connecticut 06030. email: smilowitz@nso1.uchc.edu.
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