Fig 1.
(A) Southern blot analysis of genomic DNA. SacI and BamHI digest of C3(1)Tag mouse genomic DNA (Tag-2724 and Tag 2725) and REAR mouse genomic DNA (REAR-2539 and REAR 2548) probed with the 7.5 kb C3(1)Tag transgene [16] as depicted in (B). Note that the 7.5 kb band representing the entire transgene is present in the C3(1)/Tag mice but absent in the REAR mice. However, an approximately 12 kb band representing the transgene with genomic flanking region is found in both the C3(1)/Tag and REAR mice. (C) FISH analysis of a metaphase spread of splenocytes from the original C3(1)/Tag mouse line using the C3(1)/Tag construct (B) as probe demonstrating transgene on chromosome 6 as previously reported. (D) FISH analyses of chromosome 6 from spleens of normal FVB/N mice, C3(1)/Tag mice, and REAR mice, probed with the same probe as in (C).
Fig 2.
C3(1)/Tag-REAR mammary glands.
(A) Minimal to mild ductal hyperplastic changes were seen (10X, Bar = 100 μm), consisting of crowding and piling of epithelial cells within glandular lumens, often with enlarged nuclei and increased mitoses. (B) higher magnification of highlighted hyperplastic lesion of A (arrowhead, 60X, Bar = 20 μm). (C) These hyperplasias were negative for the expression of SV40 T-antigen by IHC staining (20X, Bar = 100 μm). (D) higher magnification of a lesion in C (60X, Bar = 20 μm). Inset in D is a C3(1)/Tag mammary tumor as positive control for Tag staining).
Table 1.
Incidental histology findings in REAR mice.
Table 2.
Xenograft growth of various cell lines in C3(1)/Tag-Rear Mice.
Fig 3.
REAR mice support the growth of SV40 T-Antigen TRAMP prostate tumors.
(A) Tumor incidence of TRAMP tumors subcutaneously transplanted into the flank region of male REAR and wt male FVB/N mice. 7/8 (87.5%) male REAR mice developed TRAMP tumors by 7 weeks following implantation. However, 4/10 FVB/N mice implanted with TRAMP tumors developed tumors at a much slower rate taking up to 7 months for tumor development. (B) Subcutaneous xenografts of TRAMP tumors were composed of solid sheets of small polygonal to ovoid neoplastic cells with scant cytoplasm and dense nuclei. (C) Tumors often exhibited neuroendocrine features including angular, carrot-shaped nuclei or (D) rosettes radiating from a centralized core of eosinophilic material (arrows). (E) Often numerous lobules of neoplastic cells exhibited central necrosis in a comedo fashion (asterisk), leaving a rim of viable tumor cells along the periphery (arrowheads, 20X, Bar = 50 μm).
Fig 4.
REAR mice support the growth of M6 cell line mammary tumors.
(A) Growth curves of tumors arising from 1 X 106 M6 cells implanted into the #2 mammary fat pad of C3(1)-REAR mice (N = 6). (B) Survival curves of REAR (N = 9) and FVB/N (N = 10) mice tail vein injected with 1 X 106 M6 cells. All mice died by 12 weeks post-injection. Note that no tumors grew when M6 cells were implanted into wild-type FVB/N mice (N = 10). (C) Representative primary mammary tumor arising following the implantation of 1 X 106 M6 cells into the #2 mammary fat pad (4X, Bar = 200 μm) were characterized as expansile and locally infiltrative proliferations of solid lobules and clusters of poorly differentiated epithelial cells exhibiting marked nuclear pleomorphism and atypia, and a high mitotic rate. (D) Higher magnification of C (40X, Bar = 50 μm).
Fig 5.
Metastatic tumors arising in REAR mice from M6 cells.
(A) M6-derived mammary fat pad xenografts metastasized to the lung (4X). (B) Higher magnification of A (20X) and (C) liver (4X). (D) Higher magnification of C (10X). Tail vein M6-derived xenografts were associated predominantly with embolic growth in the lung, liver, and E) brain (10X). F) Higher magnification of brain metastasis (40X). A, C, E, Bar = 50 μm; B, D, F, Bar = 20 μm.
Table 3.
Lymphoid cell analyses in REAR and FVB/N mice expressed as percentages of cells.
Fig 6.
Vaccination of mice with irradiated M6 cells or irradiated M6 cells expressing GM-SCF results in reduced tumor volume and increased immunotoxicity of splenocytes.
(A) Mice were given four injections of irradiated cells prior to mammary gland implantation of 106 live M6 cells. Tumor volume was monitored over a 50 day period. (B) At the end point of experiment spleens of 2 mice from each group were used for splenocyte cytotoxicity measured by ELISPOT assay for different ratio of effector:target cells.