Abstract
To compare and summarize the mechanisms, frequencies of occurrence, and classification schemes of spontaneous, experimental, and genetically engineered mouse skeletal neoplasms, the literature was reviewed, and archived case material at The Jackson Laboratory was examined. The frequency of occurrence of spontaneous bone neoplasms was less than 1% for most strains, with the exceptions of osteomas in CF-1 (5.5% and 10% in two studies) and OF-1 outbred strains (35%), and osteosarcomas in NOD/ShiLtJ (11.5%) and NOD-derived (7.1%) mice. The frequency was 100% for osteochondromas induced by conditional inactivation of exostoses (multiple) 1 (
Keywords
Neoplastic lesions of bone are less common in humans, domestic animals, and laboratory mice, compared to neoplasms of other tissues.2,22,42,55-57,68,86,89,95,96 Malignant neoplasms of bone are generally more common than the benign, and osteosarcoma is the most common skeletal neoplastic lesion in all species, including laboratory mice.2,22,42,55,56,68,69,86,89,95,96 Benign tumors of bones that have been reported to occur spontaneously in mice include chondroma, hemangioma, ossifying fibroma, osteoblastoma, and osteoma.2,22,55–57,59,61,78,79,99 Spontaneous malignant neoplasms of bone reported in mice include chondrosarcoma, hemangiosarcoma, and osteosarcoma.2,22,55–57,60,78,79,94,99
This article addresses the similarities and differences in classification schemes for neoplasms of bone in humans, domestic animals, and laboratory mice. It also presents a comparison and summary of neoplasms of bone—spontaneous, genetically engineered, radiation induced, and chemically induced—in various strains of laboratory mice (as noted in the literature), as well as a retrospective evaluation of the frequency of bone neoplasms in various mouse strains submitted for diagnostic necropsy at The Jackson Laboratory (Bar Harbor, Maine).
Materials and Methods
We reviewed reports of spontaneous, experimentally induced, or genetically engineered primary skeletal neoplasms in inbred, hybrid, genetically engineered, and outbred strains and stocks of mice and summarized the mechanisms, frequencies of occurrence, and other features of these neoplasms. Additional details on the strains of mice, genetic manipulations, and experimental procedures are available in the cited references.
For retrospective evaluation of the frequency of bone neoplasms at The Jackson Laboratory, we reviewed archived necropsy records and glass slides for diagnoses of bone neoplasms in various strains submitted to the diagnostic service for necropsy between January 1987 and January 2010. Criteria for submission were clinical assessment of a “sick mouse” or the presence of a grossly visible tumor. Gross necropsy examination was performed on all mice, and collected tissues were fixed in Tellyesniczky/Fekete fixative (70% ethanol, 100 ml; 37–40% formalin, 5 ml; glacial acetic acid, 5 ml).
88
Appropriate tissues were decalcified with 10% formic acid (Formical-2000, Decal Chemical Corporation, Tallman, New York) or 3% hydrochloric acid (Cal-Ex, Fisher Scientific, Fairlawn, New Jersey) for 24 hours. Tissues were paraffin embedded and stained with hematoxylin and eosin. Diagnoses were reviewed and neoplasms classified according to criteria described in the
For any strain that had fewer than 100 total necropsy submissions, the frequency of occurrence was not considered representative. Details of the age of occurrence and frequency of benign and malignant neoplasms sorted by sex are available in Supplementary Tables 1 and 2 (available at http://vet.sagepub.com/supplemental).
Classification Schemes for Bone Tumors a
a NA, not applicable to the species; NCDE, no corresponding designation or entry in this classification scheme; NCDL, not commonly diagnosed or listed as a primary tumor of bone or joint; NCLM, no commonly listed genetically engineered mouse model with primary involvement of bone or joint; NCLS, not commonly listed as a subtype.
Benign Bone Tumors in Various Strains of Mice as Reported in the Literature
a Age of incidence as indicated in the original reference.
b Number of tumor-bearing mice and mice examined by necropsy not available from the reference.
The Mouse Tumor Biology Database (http://www.informatics.jax.org/, September 2010), 52 Mouse Genome (http://www.informatics.jax.org/, September 2010), 21 and JAX Mice Database (http://www.jaxmice.org/query/, September 2010) at The Jackson Laboratory were used to verify information on tumor types, tumor frequencies, genes, alleles, and mouse strains. Tumor frequencies or comments pertaining to tumor frequencies that reference the Mouse Tumor Biology Database are identified in subscript as (MTB).
Genes with a confirmed involvement in the pathogenesis of bone neoplasms in genetically engineered mouse models and genes known to significantly interact with the aforementioned genes were listed for gene network analysis. For performing the network analysis, each gene was assigned a hypothetical fold change score. If the loss or inactivation of a gene resulted in the development of a bone neoplasm in a mouse model, that gene was considered
Malignant Bone Tumors in Various Strains of Mice as Reported in the Literature
a Age of incidence as indicated in the original reference.
b Number of mice examined by necropsy is not available from the reference.
c Numbers of tumor bearing mice and mice examined by necropsy are not available from the reference.
d Induction time: time to appearance of the neoplasm after inoculation with the virus (at 1 to 2 days of age).
Results
Comparison of Classification Schemes
The
The
The most commonly used scheme for classification of mouse bone tumors is the one detailed in the
Cartilaginous Tumors
Osteochondroma
Osteochondromas are benign, cartilage-capped, partially ossified tumors that are also known as
Chondroma
Chondromas are benign tumors of chondrocytes consisting mostly of mature hyaline cartilage. They are typically circumscribed, encapsulated, slow growing, expansile, and smooth to nodular, and they usually occur in bones of enchondral origin. They can arise within the medullary cavity, where they are known as
Chondromyxoid fibroma
Chondromyxoid fibromas are benign cartilaginous tumors that typically involve the metaphyses of long bones. Histologically, chondromyxoid fibromas are usually lobulated, consisting of nodules of poorly formed hyaline cartilage mixed with variable amounts of myxoid matrix delinated by fibrous septae.42,86,96,99,100 Chondromyxoid fibromas are extremely rare in humans,42,86,96 domestic animals, 89 and mice and were reported in C57BL/10J mice (0.04% frequency in males).99,100
Chondrosarcoma
Chondrosarcomas are malignant cartilaginous neoplasms that arise de novo in bone or undergo malignant transformation within a preexisting benign cartilaginous tumor. They are typically expansile and sometimes lobulated, and they more commonly involve flat bones than long bones in most species. They have also been reported to arise in locations with extraskeletal cartilage, such as the larynx in humans and mice. Chondrosarcomas may arise from the medullary cavity (central chondrosarcoma) or periosteum (peripheral chondrosarcoma), with the former being more common. Histologically, chondrosarcomas are often lobulated, consisting of different proportions of hyaline or myxoid cartilage composed of chondrocytes that vary in cytologic atypia, mitotic activity, and degree of differentiation (Figs. 3, 4). They primarily produce chondroid and fibrillar matrix but rarely osteoid. However, occasional chondrosarcomas may contain reactive or metaplastic bone matrix.22,42,55–57,86,89,95,96 Chondrosarcoma is the third-most common malignancy of bone in humans, after myeloma and osteosarcoma,42,86,96 and it is reported in cats, dogs, and sheep89,95 but is extremely rare in mice.2,22,55–57 Spontaneous chondrosarcomas were reported in B6C3F1 mice (0.13% frequency in females), 2 in C57BL/10J mice (0.03% frequency in females),99,100 and in a ST/Eh mouse (single case) 92 and observed in a 325 day-old male C57BL/6J mouse (Figs. 3, 4) at The Jackson Laboratory.
Osteogenic Tumors
Osteoma
Osteomas are rare benign tumors of osteoblasts composed of mature well-differentiated bone with a predominantly woven arrangement. They are typically solitary, dense, and smooth, and they preferentially arise from the periosteal surface and merge with underlying cortical bone. Osteomas most commonly occur in bones of the skull in most species (Fig. 5). An osteoma may rarely be seen on the surface of a long bone. Histologically, osteomas are composed of dense coalescing spicules and trabeculae of bone lined by osteoblasts and osteoclasts. Fatty or hematopoietic marrow may be present within the trabeculae (Figs. 6, 7). The intertrabecular connective tissue is composed of spindle-shaped cells, and it becomes increasingly sparse as the tumor becomes more compact and sclerotic.22,42,55-57,61,86,89,95,96 Spontaneous osteomas are rare in most strains of mice and seem to be more common in females than in males, with the difference being attributed partly to a female hormonal influence.
Cases of spontaneous osteoma were documented in the following mice strains: AKR, 61 B6C3F1,2,9 C57BL/6NNia(MTB), CBA/H(MTB), CD-1,63,66,72 CF-1,2,10,61 CFW/CarWHanJena(MTB), NMRI2,7 NZO/Bl(MTB), Him:OF1, 36 and OF-1. 108 CF-1, Him:OF1, and OF-1, which are genetically related outbred strains of mice and have the highest documented frequency of osteomas among all mouse strains.2,61 The reported frequency of spontaneous osteomas in CF-1 mice was 5.5% in one study 2 (4.4% in males and 6.5% in females) and 10% in another (8.5% in males and 11.5% in females). 10 In the latter, the combined average age of occurrence for both sexes was 96 weeks. 10 Spontaneous osteomas are quite rare in other strains, with frequencies ranging from 0.02 to 0.20% in various reports. 2 In one study, a frequency of 0.13% was observed in female B6C3F1 mice (1 case in 770 mice examined). 2
Reports exist of higher frequencies of osteomas with concurrent identification and isolation of retroviral particles.
61
In a study on skeletal lesions of sodium fluoride toxicity in Crl:CD-1 (ICR) outbred mice, the reported frequency was 12.5% in sodium fluoride–treated mice (11% in males and 14% in females) and 3.2% in controls (1.1% in males and 5.3% in females) with a suspected retroviral etiology at 55 to 97 weeks of age.
66
In a study on the carcinogenic potential of cyclosporine A in OF-1 mice, a 26.3% frequency was reported in cyclosporine-treated mice (14.7% in males and 38% in females), with 35% in controls (20% in males and 30% in females), at 40 to 78 weeks of age.
108
Type C retroviral particles were identified in tumors from both groups, but an etiologic correlation was not conclusively established. In an aging study, Him:OF-1 mice were reported to have a frequency of 51.8% (50.9% in males and 52.7% in females) at 533 ± 98 days of age, and viruslike particles were seen on ultrastructural observation of tumor cells and necrotic tissues.
36
Osteomas induced by neonatal administration of murine polyomavirus were documented in C3H/BiDa mice.
17
At The Jackson Laboratory, osteomas were diagnosed in mice of strains A/J, C57BL/6J (Fig. 5), DBA/2J, B6C3F1/J, C3.SW-
Osteosarcoma
Osteosarcomas are malignant mesenchymal neoplasms in which tumor cells typically produce osteoid or bony matrix.2,22,25,42,56,57,60,69,75,86,89,95,96,102They are typically invasive, and they frequently metastasize.2,22,42,55-57,60,75,86,89,95,96,102 Osteosarcomas most commonly arise within the medullary cavities of bones—particularly, the metaphyseal regions of long bones—and invade the adjacent cortex (Fig. 8); these are referred to as
Osteosarcomas are the most commonly reported bone tumors in laboratory mice,2,22,55,57,68,69 but the average frequency of spontaneous osteosarcomas in most strains of mice is relatively low(MTB).2,56 There have been single-case reports of osteosarcoma in a 5-month-old AKR/J male mouse,
1
300-day-old AKR/Ms mouse,
70
5-week-old C57BL/6J male mouse,
14
224-day-old C57BL/6J female mouse(MTB),757-day-old C57BL/6J-
Virally induced osteosarcomas were periodically reported, and most notable among them are the ones induced by retroviruses (Finkel Biskis Jenkins murine osteosarcoma virus and Moloney murine sarcoma virus)23,73,77,105 and murine polyomavirus.17,97 Periosteal chondro-osseous tumors resembling human parosteal osteosarcomas were induced in National Institutes of Health Swiss mice by neonatal intraperitoneal inoculation of Finkel Biskis Jenkins virus. 105 Osteosarcomas with and without metastases in CzechII/EiJ and C3H/BiDa mice, respectively, were induced by neonatal intraperitoneal inoculation of the RA strain of murine polyomavirus. 97 Osteosarcomas in cranial bones are a relatively frequent observation in many SV40 Large T antigen transgenic mice regardless of the promoter, suggesting a direct effect (J.P.S., unpublished observation).
Osteosarcomas induced by intraperitoneal administration of bone-seeking radionuclides 224Ra, 223Ra, and 227Th were reported in C3H/HeEl.102, (BALB/c x CBA) F2, (C3Hx101) F1, and NMRI mice 64 and by external irradiation with 90Sr-90Y beta rays in ICR mice. 71 In other studies, administration of radiostrontium (90Sr) was reported to cause osteosarcomas and skeletal hemangiosarcomas in CBA and C3H mice.
Strains of mice varied widely with respect to metastases of osteosarcomas, and metastases were infrequent or nonexistent in some. Metastases of spontaneous osteosarcomas were noted in BALB/c,
27
C3Hf/Sed;CB17-
At The Jackson Laboratory osteosarcomas were diagnosed in mice of strains 129Sv/J, A/J, AKR/J (Figs. 9, 10), B6;129P2-
Vascular Tumors
Hemangioma
Hemangiomas are benign vascular tumors composed of blood-filled, endothelium-lined spaces. They are typically soft tissue tumors but can rarely arise as primary bone tumors, occurring more frequently in the axial skeleton than elsewhere in humans.42,96 Histologically, hemangiomas consist of a partially circumscribed network of vascular spaces, usually lined by a single layer of well-differentiated endothelial cells embedded in a fibrous stroma. Intraosseous hemangiomas are extremely infrequent in laboratory mice2,55 as well as dogs and cats.89,95 They were reported in BALB/c mice (0.95% frequency). 55 An intraosseous hemangioma was observed in an HRS/J +/+ mouse at The Jackson Laboratory.
Hemangiosarcoma
Hemangiosarcomas (hemangioendothelial sarcomas) are malignant vascular tumors composed of endothelium-lined vascular channels supported by various proportions of stroma.30,42,89,95,96 They are typically soft tissue tumors but can occasionally arise as primary bone tumors.30,42,89,95,96 Histologically, hemangiosarcomas consist of malignant endothelial cells that differ in their degree of differentiation and cytologic atypia and form vascular channels and solid sheets embedded in a fibrovascular stroma (Figs. 13, 14).30,42,89,96 Intraosseous hemangiosarcomas are extremely infrequent in laboratory mice and domestic animals.2,55,56,89 The primary differential diagnosis for an intraosseous hemangiosarcoma is a telangiectatic osteosarcoma in which osteoblasts rather than neoplastic endothelial cells line the blood-filled spaces.42,89,95,96 A spontaneous intraosseous hemangiosarcoma (angiosarcoma) was reported in an NZB mouse.
41
Hemangiosarcomas of bone were found in mice at The Jackson Laboratory of strains CD-1, DBA/2J, HRS/J, FVB/NJ-
Miscellaneous Tumors and Lesions
Ossifying fibroma, osteofibroma, and osteofibrous dysplasia
Ossifying fibromas are rare fibro-osseous tumors that usually involve the mandible and maxilla in most species. They are typically solitary, slow growing, expansile, and sharply demarcated, and they distort the normal contour of the affected bone. They are morphologically similar to a fibroma but contain metaplastic bone. Histologically, ossifying fibromas are well demarcated from surrounding tissue by bone, and they consist of spindle-shape fibroblasts. The fibroblasts appear to transform into osteoblasts forming irregular spicules of woven bone. In general, ossifying fibromas are of relatively low cellularity with a higher proportion of fibrous stroma than bone. Lamellar bone is extremely rare in an ossifying fibroma but may be formed where woven bone trabeculae are resorbed and replaced.2,22,57,58,89,95
A few authors consider ossifying fibromas as being synonymous with osteofibroma or fibrous osteoma, 89 while others describe them as being synonymous with cementifying fibroma when teeth are involved.22,57,58 The latter authors describe osteofibromas as being distinct from ossifying fibromas and more similar to osteoblastomas. Such osteofibromas are typified as benign, expansile, noninvasive bone-forming tumors with a predilection for the vertebral column. Histologically, osteofibromas are characterized by mature trabecular bone within a densely cellular spindle cell stroma.22,57,59 Human tumors previously described as ossifying fibroma are currently designated as osteofibrous dysplasia.25,42,96 Osteofibrous dysplasia is characterized by a hypocellular spindle cell proliferation with the production of immature woven bone. 42 Spontaneous ossifying fibromas (incidence not available) were reported in CD-1, CF-1, and NMRI mice.2,57 A single case was diagnosed in a NONcNZO10/LtJ mouse (Figs. 15, 16) at The Jackson Laboratory. The latter lesion had some microscopic features resembling those described in osteofibrous dysplasia of humans.42,96
Experimental and Genetically Engineered Mouse Models
Numerous genetically engineered mouse models have contributed to the elucidation of genetic mechanisms involved in skeletal tumors. 49 Selected models and their characteristics are summarized below.
Exostoses (multiple) 1 and 2 (Ext1, Ext2)
The genes exostoses (multiple) 1 (
GLI-Kruppel family member oncogene (Gli2)
Enchondromas were shown to develop with high frequency in mice with transgenic overexpression of the GLI-Kruppel family member oncogene (
Nuclear factor of activated T cells, cytoplasmic calcineurin dependent-2 gene (Nfatc2)
Mice with targeted mutations in the
Osteosarcoma-associated genes
The molecular mechanisms and genetic pathways implicated in osteosarcomagenesis include aberrations involving transformation-related protein 53 (
Mice with mutations of
Mice that bear the tumor antigen gene from SV40 Tag in their genome were also shown to develop osteosarcomas5,40,51,65,106 and lesions resembling synovial chondromatosis. 98 The lesions of chondromatosis reportedly involved the articular cartilages of the legs and feet in these mice. 98 In SV40 Tag–induced osteosarcomas, sequestration and inactivation of the P53 and RB1 proteins by the SV40 Tag (usually, large T antigen) resulted in tumorigenesis in a promoter-independent manner. Tumors arose predominantly in the axial skeleton with variable involvement of the skull, vertebral column, sternum, ribs, shoulder, and pelvic girdle, with less frequent involvement of the humerus and femur.5,40,51,65,106
Human synovial sarcomas are marked by a signature translocation-mediated fusion of the Synovial Sarcoma Translocation, chromosome 18 (
Gene Network Analysis
A list of candidate genes was created from genes with a confirmed involvement in the pathogenesis of neoplasms of bone and additional genes known to significantly influence the former. Hypothetical fold change scores were assigned to each listed gene, and gene network analysis was performed to deduce network interactions between genes.
Results of the analysis (Supplementary Table 4 and Supplementary Figs. 1–4, available at http://vet.sagepub.com/supplemental) suggested that transformation related protein 53 (
Benign and Malignant Bone Tumors in Strains of Mice From The Jackson Laboratory Diagnostic Necropsy Archives a
a F, frequency of tumors in percentages (number of cases by number of necropsies); A, average age of observation of tumor (average age at necropsy) in days. Tumor frequencies and average ages for males and females are available in Supplementary Tables 1 and 2 (available at http://vet.sagepub.com/supplemental). No bone tumors were found in the diagnostic archives for the inbred and hybrid strains highlighted in gray. The frequencies were considered nonrepresentative for strains with less than 100 total necropsies (highlighted in gray). Columns highlighted in darker gray indicate that there were no documented cases of that tumor type for that strain in the necropsy archives.
Discussion
Spontaneous bone tumors were infrequent in most strains of mice with the exceptions of osteosarcomas in NZBR/BI, NOD, and NOD-derived mice and osteomas in CF-1 and OF-1 mice. The most popular inbred strains of mice currently available through The Jackson Laboratory are 129S1/SvImJ, 129X1/SvJ, A/J, AKR/J, BALB/cByJ, BALB/cJ, C3H/HeJ, C57BL/6J, C57BL/6NJ, C57BL/10J, CBA/J, DBA/1J, DBA/2J, FVB/NJ, NOD/ShiLtJ, and SJL/J (JAX Mice, http://www.jaxmice.org/, September 2010). Among these, 129S1/SvImJ, C57BL/10J, DBA/1J, FVB/NJ, and SJL/J mice had no documented bone neoplasms in the diagnostic necropsy archives. This is noteworthy even though the colony sizes of these strains have varied through the years, many of the submitted mice were 8 months or younger, and focus of the diagnostic necropsy program was evaluation of sick mice rather than a systematic study of cancer with equal numbers of age- and sex-matched mice. Systematic aging studies
109
have been designed to better address such questions. In those mice with documented neoplasms, spontaneous osteogenic neoplasms occurred more frequently than cartilaginous, vascular, and hematopoietic ones, with osteoma being the most common benign bone tumor and with osteosarcoma being the most common malignancy of bone. Spontaneous cartilaginous neoplasms were the least frequent tumors of bone in mice, in direct contrast to chondrogenic neoplasms, the second-most common bone tumor type, after hematolymphoid malignancies in humans. In this context it is worth noting that chondrogenic tumors were produced by targeted mutations of
Another noteworthy aspect is that strains of mice with spontaneous neoplasms of bone varied in their propensity to develop metastases. Metastases were infrequent or nonexistent in many studies. At The Jackson Laboratory, the age at necropsy for mice with spontaneous or genetically engineered malignant neoplasms of bone ranged from 41 to 537 days (
Conspicuous disparities exist between classification systems for human, veterinary, and mouse bone and joint tumors. Unified systems of classification and characterization for human, veterinary, mouse, and genetically engineered bone tumors that reconcile differences in designation for similar entities would enhance comparative studies of tumors of the skeleton across species. The Mouse Pathology Ontology (Pathbase; http://eulep.pdn.cam.ac.uk/Pathology_Ontology/index.php) developed by a consortium of veterinary and physician pathologists has reconciled these differences.
Footnotes
Acknowledgements
We gratefully acknowledge D. Boggess for retrieval of archived necropsy records and materials; J. Miller, E. Taylor, and M. McKluskey for retrieval of archived glass slides; the Department of Histopathology and Microscopy Sciences for preparation of histologic sections from archived paraffin blocks; J. Hammer for assistance with preparation of figure panels; T. Stearns for assistance with the Ingenuity IPA Pathway Analysis Software and preparation of gene networks; N. Buckley for assistance with formatting some references; Dr. C. Bult for advice on interpretation of the gene network analyses; and Drs E. Leiter, L. Shultz, and W. Beamer for critical comments and suggestions.
The authors declared that they received no commercial financial support for research or authorship of this article.
This work was supported in part by grants from the National Institutes of Health (CA34196, JPS), the Ellison Medical Foundation (JPS), and institutional shared services at The Jackson Laboratory (AK, OF).
References
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
