In vivo bioluminescence imaging for leptomeningeal dissemination of medulloblastoma in mouse models

Abstract

BackgroundThe primary cause of treatment failure in medulloblastomas (MB) is the development of leptomeningeal dissemination (seeding). For translational research on MB seeding, one of the major challenges is the development of reliable experimental models that simulate the seeding and growth characteristics of MBs. To overcome this obstacle, we improved an experimental mouse model by intracisternal inoculation of human MB cells and monitoring with in vivo live images.MethodsHuman MB cells (UW426, D283 and MED8A) were transfected with a firefly luciferase gene and a Thy1.1 (CD90.1) marker linked with IRES under the control of the CMV promoter in a retroviral DNA backbone (effLuc). The MB-effLuc cells were injected into the cisterna magna using an intrathecal catheter, and bioluminescence images were captured. We performed histopathological analysis to confirm the extent of tumor seeding.ResultsThe luciferase activity of MB-effLuc cells displayed a gradually increasing pattern, which correlated with a quantitative luminometric assay. Live imaging showed that the MB-effLuc cells were diffusely distributed in the cervical spinal cord and the lumbosacral area. All mice injected with UW426-effLuc, D283-effLuc and MED8A-effLuc died within 51 days. The median survival was 22, 41 and 12 days after injection of 1.2 × 106 UW426-effLuc, D283-effLuc and MED8A-effLuc cells, respectively. The histopathological studies revealed that the MB-effLuc cells spread extensively and diffusely along the leptomeninges of the brain and spinal cord, forming tumor cell-coated layers. The tumor cells in the subarachnoid space expressed a human nuclei marker and Ki-67. Compared with the intracerebellar injection method in which the subfrontal area and distal spinal cord were spared by tumor cell seeding in some mice, the intracisternal injection model more closely resembled the widespread leptomeningeal seeding observed in MB patients.ConclusionThe results and described method are valuable resources for further translational research to overcome MB seeding.

Tumour necrosis factor alpha and interleukin‐5 inhibit olfactory regeneration via apoptosis of olfactory sphere cells in mice models of allergic rhinitis

“…Anesthetized normal and allergic rhinitis mice were transcardially perfused with PBS and 4% PFA. Immunofluorescence analysis was performed as described previously . Olfactory tissues from normal and allergic mice were dissected and harvested in a tissue culture hood.…”

Panobinostat, a histone deacetylase inhibitor, suppresses leptomeningeal seeding in a medulloblastoma animal model

“…injection of 20 mg/kg Zoletil (Virbac, Carros, France) and 10 mg/kg Rompun (Bayer, Leverkusen, Germany). After the cisterna magna was exposed, the UW426-effLuc (1.2 × 10 6 cells) cells were slowly injected into the subarachnoid space of the cisterna magna using a 30-gauge needle as previously described [ 32 ].…”

“…For the in vivo BLI analysis, the mice were sedated with 2% isoflurane in 100% O 2 through a nose cone. After D-Luciferin (150 mg/kg, Caliper Life Sciences) was administered, bioluminescence images were taken by an In-Vivo Imaging System 100 (Xenogen Corp.) on days 0, 1, 4, 7, 11, 14, 18 and 21, and the images were analyzed as described previously [ 31 , 32 ]. For the survival analysis, the mice were followed until they died (control vs. treatment, N=8 per group).…”

The molecular biology of medulloblastoma metastasis

“…Tumors of the WNT subgroup arise from the lower rhombic lip and are driven by Wingless-related integration site pathway activation (26) through frequent somatic mutations of beta-catenin (CTNBB1) and monosomy 6. These usually show classic histology, tend to affect older children (ages [4][5][6][7][8][9][10][11][12][13][14][15] and are associated with an excellent prognosis, low rates of recurrence, and very low rates of metastatic dissemination (16). Grp 3 and Grp 4 tumors occupy the fourth ventricle, and are less clearly defined at the molecular level than SHH or WNT tumors.…”

“…Resection of primary tumor increases the survival period of the mouse model sufficiently so as to achieve dissemination and leptomeningeal/distant implantation (22). A modified xenograft mouse model generated through tumor cell injection into the cisterna magna has also shown consistent leptomeningeal dissemination along the brain and spinal cord, and provides another in vivo model of metastasis (13). Transgenic models of metastatic MB have proven difficult to generate.…”