Figure 3. Increased FRMD6 expression inhibits subcutaneous and intracranial GBM growth and progression.

(A) 1 × 10⁶ U251 and 5 × 10⁶ U87MG GBM cells were injected subcutaneously into each immunocompromised Rag-2/II2rg mouse. Subcutaneous GBM growth experiments were terminated when the fastest growing gliomas reach ~1 cm in their longest diameters in accordance with the IACUC regulation and dissected subcutaneous tumors were weighted, recorded, and expressed as the mean weight +/− SD. n = 10 mice/group. **p < 0.01. (B) Representative images of the H&E stained cross sections of mouse brain bearing glioblastomas derived from U87MG-control (left two panels) and U87MG-FRMD6 (right two panels) cells 25 days after i.c. injection of the GBM cells. Bar is 40 μm in upper two panels and 160 μm in bottom two panels. Red circles in two right panels highlight a smaller glioblastoma derived from U87MG-FRMD6 cells. (C) Morphology and in situ anti-Ki67 antibody reactivity that highlights proliferating GBM cells in tumors are shown. The GBM sections were stained with H&E to outline histological morphology of the tumors (upper 4 panels). In vivo proliferating GBM cells were detected using an anti-Ki67 antibody (Fisher Scientific, bottom 4 panels). These GBM sections were derived from U87MG-control (first upper and bottom panels), U87MG-FRMD6 (second upper and bottom panels), U251-control (third upper and bottom panels), and U251-FRMD6 (last upper and bottom panels). Bar, 100 μm. (D) For intracranial GBM growth experiments, U87MG (3 × 10⁵ cells in 10 μl HBSS/Rag2 mouse)/U251 cells (2 × 10⁵cells in 10 μl HBSS/Rag2 mouse) were injected intracranially. n = 10 mice/group. Following the intracranial injections, mice were monitored closely and durations of their survival were recorded. Increased expression of FRMD6 was found to inhibit intracranial growth/progression of the GBM cells and extends survival of the experimental mice. **p < 0.01.