Growth of precultured human glioma biopsy specimens in nude rat brain

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Object. The aim of this study was to develop an improved animal model for brain tumor study. The need for better and more relevant brain tumor models is generally acknowledged. Glioma tissue can be cultured directly from the biopsy specimen as tumor spheroids. Using such precultured tissue, a new in vivo model for studying human gliomas was established.

Methods. Precultured small tumor spheroids (< 300 µm) prepared from cell lines or tumor biopsy fragments were injected into the brains of immunodeficient rats by using a 5-µl Hamilton syringe that had a piston in the needle. Tumors could be established by injecting a single spheroid derived from the U-87MG cell line, whereas inoculation of 10 spheroids resulted in a tumor take comparable to that attained with injection of 106 single cells. Biopsy specimens obtained from six patients who underwent surgery for glioblastoma multiforme were cultured as organotypic spheroids for 11 to 18 days before inoculation into the rats. The animals were killed 3 months after spheroid implantation. Microscopic examination revealed tumor growth in 87.5 to 100% of the animals inoculated with tumor spheroids from all but one of the tumor biopsy specimens. Extensive invasion and cell migration along the nerve tracts of the corpus callosum was found in tumors that originated from four of the six biopsy specimens.

Conclusions. This approach, in which spheroids from precultured biopsy specimens are injected into the brains of immunodeficient animals, provides new means for experimental studies of human malignant brain tumors in a clinically relevant animal model.

Article Information

Address reprint requests to: Olav Engebraaten, M.D., Ph.D., Department of Tumor Biology, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway. email: olave@radium.uio.no.

© AANS, except where prohibited by US copyright law.

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Figures

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    A: Graph showing survival curves of animals inoculated with 106 cells (solid line) 5 × 104 cells (long dashed line), and 10 spheroids (short dashed line) of the U-87MG cell line. B: Graph showing survival curves of animals inoculated with 10 spheroids (solid line), five spheroids (long dashed line), and one spheroid (short dashed line), made from reaggregated U-87MG cells. Cum = cumulative.

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    A: Photomicrograph of a large U-87MG tumor (t) growing in the rat brain (b). Note the sharply defined borderline between the two tissues. Bar = 500 µm. B: Detail from box in A, showing the U-87MG borderline toward normal brain tissue. The two types of tissue may be easily distinguished. Bar = 50 µm.

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    Photomicrographs showing the histopathological appearance of GBM Case II inoculated into the right cerebral hemisphere of a rat (arrow, injection site). The tumor growth caused a midline shift, indenting the left hemisphere (A), and the cells migrated into the corpus callosum (C), and into the opposite cerebral hemisphere (E; magnification of the box in C). Sections A, C, and E are stained with H & E. The Ki-67—positive tumor cells were observed in the tumor at the site of implantation (B), in corpus callosum (D), and in the contralateral hemisphere (F; magnification of the box in A). Sections B, D, and F have not been counterstained; thus only the tumor cells, which are Ki-67 positive, are seen. Bars: A and B = 2 mm, C and D = 500 µm, E and F = 50 µm.

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    Photomicrographs showing the histopathological appearance of the biopsy specimens from the patients (A, C, E, G, and I) compared with the specimens from the corresponding rat brain tumor (B, D, F, H, and J). No tumors grew from the GBM Type V specimen. A and B: GBM Type I. C and D: GBM Type II. E and F: GBM Type III. G and H: GBM Type IV. I and J: GBM Type VI. Bar = 50 µm.

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