Recent studies of mice with hair defects have resulted in major contributions to the understanding of hair disorders. To use mouse models as a tool to study nail diseases, a basic understanding of the similarities and differences between the human and mouse nail unit is required. In this study we compare the human and mouse nail unit at the macroscopic and microscopic level and use immunohistochemistry to determine the keratin expression patterns in the mouse nail unit. Both species have a proximal nail fold, cuticle, nail matrix, nail bed, nail plate, and hyponychium. Distinguishing features are the shape of the nail and the presence of an extended hyponychium in the mouse. Expression patterns of most keratins are similar. These findings indicate that the mouse nail unit shares major characteristics with the human nail unit and overall represents a very similar structure, useful for the investigation of nail diseases and nail biology.
animal model; microscopic; claw; nail
A number of single gene mutations in laboratory mice produce hair follicle defects resulting in deformed hair shafts. The radiation induced (SB/LeJ-Foxq1sa) satin mutant mice have a satin-like sheen to their hair and dilute coloration. This sheen is due to failure of the hair shafts to develop normal medullas, while the pigment dilution is due to the unrelated beige (lysosomal trafficking regulator, Lystbg) mutation. A new allelic mutation, Foxq1sa-J, arose spontaneously on the albino (tyrosinase, Tyrc) MRL/MpJ-Faslpr background. The Foxq1sa-J allele has a C to T transition at position 490. By contrast, the Foxq1sa mutant allele was confirmed to be a 67 base pair deletion followed by two base changes (GA to AT). Morphologic changes were similar to those seen in Hoxc13 transgenic and targeted mutant mice. This new allelic mutation provides yet another tool to investigate formation of the interior structures of hair shafts.
Foxq1; satin; hair fiber; medulla; gene networks; mouse model
Epidermolysis Bullosa (EB) encompasses a spectrum of mechanobullous disorders caused by rare mutations that result in structural weakening of the skin and mucous membranes. While gene mutated and types of mutations present are broadly predictive of the range of disease to be expected, a remarkable amount of phenotypic variability remains unaccounted for in all but the most deleterious cases. This unexplained variance raises the possibility of genetic modifier effects. We tested this hypothesis using a mouse model that recapitulates a non-Herlitz form of junctional EB (JEB) owing to the hypomorphic jeb allele of laminin gamma 2 (Lamc2). By varying normally asymptomatic background genetics, we document the potent impact of genetic modifiers on the strength of dermal-epidermal adhesion and on the clinical severity of JEB in the context of the Lamc2jeb mutation. Through an unbiased genetic approach involving a combination of QTL mapping and positional cloning, we demonstrate that Col17a1 is a strong genetic modifier of the non-Herlitz JEB that develops in Lamc2jeb mice. This modifier is defined by variations in 1–3 neighboring amino acids in the non-collagenous 4 domain of the collagen XVII protein. These allelic variants alter the strength of dermal-epidermal adhesion in the context of the Lamc2jeb mutation and, consequentially, broadly impact the clinical severity of JEB. Overall the results provide an explanation for how normally innocuous allelic variants can act epistatically with a disease causing mutation to impact the severity of a rare, heritable mechanobullous disorder.
Epidermolysis bullosa (EB) is a group of rare genetic Mendelian disorders that result in mechanical fragility of the skin and mucosal membranes. Junctional EB is a subset caused by mutations that result in cleavage of the dermal-epidermal junction. All forms of EB demonstrate substantial variability in their clinical phenotype that is not readily explained. The possibility of genetic modifiers as the cause of this variability has been difficult to address in humans. We apply a mouse model carrying a hypomorphic allele of the laminin gamma 2 (Lamc2) gene to address the possibility of genetic modifiers of JEB. We document the potent impact of differing genetic backgrounds on multiple facets of the JEB syndrome expressed in these mice and show that three neighboring amino acid changes within the non-collagenous domain 4 of the collagen XVII protein strongly modify their disease. The study provides a molecular explanation of how a primary mutation that weakens one component of the cutaneous basement membrane is influenced by normally innocuous allelic variants of another component to affect strength of dermal-epidermal adhesion and consequently, the severity of JEB. This approach may guide the genetic prognosis and diagnosis of human EB disorders.
databases; genetically engineered mice; phenotyping; record keeping; mouse pathology; mouse pathology informatics
Sharpin-deficient mice display a multiorgan chronic inflammatory phenotype suggestive of altered leukocyte migration. We therefore studied the role of SHARPIN in lymphocyte adhesion, polarization and migration. We found that SHARPIN localizes to the trailing edges (uropods) of both mouse and human chemokine-activated lymphocytes migrating on ICAM-1, which is one of the major endothelial ligands for migrating leukocytes. SHARPIN-deficient cells adhere better to ICAM-1 and show highly elongated tails when migrating. The increased tail lifetime in SHARPIN-deficient lymphocytes decreases the migration velocity. The adhesion, migration and uropod defects in SHARPIN deficient lymphocytes were rescued by reintroducing SHARPIN into the cells. Mechanistically we show that SHARPIN interacts directly with LFA-1, a leukocyte counter-receptor for ICAM-1, and inhibits the expression of intermediate and high-affinity forms of LFA-1. Thus SHARPIN controls lymphocyte migration by endogenously maintaining LFA-1 inactive to allow adjustable detachment of the uropods in polarized cells.
Mice with spontaneous mutations in the Sharpin gene develop chronic proliferative dermatitis that is characterized by eosinophilic inflammation of the skin and other organs with increased expression of type 2 cytokines and dysregulated development of lymphoid tissues. The mutant mice share phenotypic features with human hypereosinophilic syndromes. The biological function of SHARPIN and how its absence leads to such a complex inflammatory phenotype in mice are poorly understood. However, recent studies identified SHARPIN as a novel modulator of immune and inflammatory responses. The emerging mechanistic model suggests that SHARPIN functions as an important adaptor component of the linear ubiquitin chain assembly complex that modulates activation of NF-κB signaling pathway, thereby regulating cell survival and apoptosis, cytokine production and development of lymphoid tissues. In this review, we will summarize the current understanding of the ubiquitin-dependent regulatory mechanisms involved in NF-κB signaling, and incorporate the recently obtained molecular insights of SHARPIN into this pathway. Recent studies identified SHARPIN as an inhibitor of β1-integrin activation and signaling, and this may be another mechanism by which SHARPIN regulates inflammation. Furthermore, the disrupted lymphoid organogenesis in SHARPIN-deficient mice suggests that SHARPIN-mediated NF-κB regulation is important for de novo development of lymphoid tissues.
Sharpin; immune system; inflammation; chronic dermatitis; NF-κB; ubiquitination; lymphoid organogenesis; eosinophilic dermatitis; scaly skin disease
The capture and use of disease-related anatomic pathology data for both model organism phenotyping and human clinical practice requires a relatively simple nomenclature and coding system that can be integrated into data collection platforms (such as computerized medical record-keeping systems) to enable the pathologist to rapidly screen and accurately record observations. The MPATH ontology was originally constructed in 2,000 by a committee of pathologists for the annotation of rodent histopathology images, but is now widely used for coding and analysis of disease and phenotype data for rodents, humans and zebrafish.
Construction and content
MPATH is divided into two main branches describing pathological processes and structures based on traditional histopathological principles. It does not aim to include definitive diagnoses, which would generally be regarded as disease concepts. It contains 888 core pathology terms in an almost exclusively is_a hierarchy nine layers deep. Currently, 86% of the terms have textual definitions and contain relationships as well as logical axioms to other ontologies such the Gene Ontology.
Application and utility
MPATH was originally devised for the annotation of histopathological images from mice but is now being used much more widely in the recording of diagnostic and phenotypic data from both mice and humans, and in the construction of logical definitions for phenotype and disease ontologies. We discuss the use of MPATH to generate cross-products with qualifiers derived from a subset of the Phenotype and Trait Ontology (PATO) and its application to large-scale high-throughput phenotyping studies. MPATH provides a largely species-agnostic ontology for the descriptions of anatomic pathology, which can be applied to most amniotes and is now finding extensive use in species other than mice. It enables investigators to interrogate large datasets at a variety of depths, use semantic analysis to identify the relations between diseases in different species and integrate pathology data with other data types, such as pharmacogenomics.
Pathology; Ontology; Disease; Mouse; Phenotype
Generalized arterial calcification of infancy (GACI), an autosomal recessive disorder, is characterized by early mineralization of blood vessels, often diagnosed by prenatal ultrasound and usually resulting in demise during the first year of life. It is caused in most cases by mutations in the ENPP1 gene, encoding an enzyme that hydrolyzes ATP to AMP and inorganic pyrophosphate, the latter being a powerful anti-mineralization factor. Recently, a novel mouse phenotype was recognized as a result of ENU mutagenesis – those mice developed stiffening of the joints, hence the mutant mouse was named ‘ages with stiffened joints’ (asj). These mice harbor a missense mutation, p.V246D, in the Enpp1 gene. Here we demonstrate that the mutant ENPP1 protein is largely absent in the liver of asj mice, and the lack of enzymatic activity results in reduced inorganic pyrophosphate (PPi) levels in the plasma, accompanied by extensive mineralization of a number of tissues, including arterial blood vessels. The progress of mineralization is highly dependent on the mineral composition of the diet, with significant shortening of the lifespan on a diet enriched in phosphorus and low in magnesium. These results suggest that the asj mouse can serve as an animal model for GACI.
C57BL/6 mice develop dermatitis and scarring alopecia resembling human cicatricial alopecias (CA), particularly the central centrifugal cicatricial alopecia (CCCA) type. To evaluate the role of retinoids in CA, expression of retinoid metabolism components were examined in these mice with mild, moderate, or severe CA compared to hair cycle matched mice with no disease. Two feeding studies were performed with dams fed either NIH 31 diet (study 1) or AIN93G diet (study 2). Adult mice were fed AIN93M diet with 4 (recommended), 28, or 56 IU vitamin A/g diet. Feeding the AIN93M diet to adults increased CA frequency over NIH 31 fed mice. Increased follicular dystrophy was seen in study 1 and increased dermal scars in study 2 in mice fed the 28 IU diet. These results indicate that retinoid metabolism is altered in CA in C57BL/6J mice that require precise levels of dietary vitamin A. Human patients with CCCA, pseudopelade (end stage scarring), and controls with no alopecia were also studied. Many retinoid metabolism proteins were increased in mild CCCA, but were undetectable in pseudopelade. Studies to determine if these dietary alterations in retinoid metabolism seen in C57BL/6J mice are also involved in different types of human CA are needed.
Alopecia areata (AA) is an autoimmune disease that attacks anagen hair follicles. Gene array in graft-induced C3H/HeJ mice revealed that genes involved in retinoic acid (RA) synthesis were increased, while RA degradation genes were decreased in AA compared to sham controls. This was confirmed by immunohistochemistry in biopsies from patients with AA and both mouse and rat AA models. RA levels were also increased in C3H/HeJ mice with AA. C3H/HeJ mice were fed a purified diet containing one of four levels of dietary vitamin A or an unpurified diet two weeks before grafting and disease progression followed. High vitamin A accelerated AA, while mice fed no vitamin A had more severe disease by the end of the study. More hair follicles were in anagen in mice fed high vitamin A. Both the number and localization of granzyme B positive cells were altered by vitamin A. IFNG was also lowest and IL13 highest in mice fed high vitamin A. Other cytokines were reduced and chemokines increased as the disease progressed, but no additional effects of vitamin A were seen. Combined, these results suggest that vitamin A regulates both the hair cycle and immune response to alter the progression of AA.
Normophosphatemic familial tumoral calcinosis, characterized by ectopic mineralization of skin, is caused by mutations in the SAMD9 gene located on human chromosome 7q21, next to a paralogous gene, SAMD9-like (SAMD9L). Mouse does not have a SAMD9 orthologue, Samd9, because it has been deleted during evolution due to genomic rearrangements. It has been suggested that the mouse Samd9l gene serves as a functional paralogue of human SAMD9. In this study we examined Samd9l knockout mice with respect to ectopic mineralization. We also crossed these mice with Abcc6tm1JfK mice, a model system to study pseudoxanthoma elasticum, to see if the absence of the Samd9l gene modifies the mineralization process. Necropsy analysis of Samd9ltm1Homy mice revealed no evidence of ectopic mineralization, and deletion of the Samd9l gene in mice failed to modify the mineralization process on the Abcc6tm1JfK background. Collectively, the results suggest that mouse Samd9l is not a functional paralogue of human SAMD9.
Ectopic connective tissue mineralization; mouse models of skin diseases; familial tumoral calcinosis; pseudoxanthoma elasticum
The systematic investigation of the phenotypes associated with genotypes in model organisms holds the promise of revealing genotype–phenotype relations directly and without additional, intermediate inferences. Large-scale projects are now underway to catalog the complete phenome of a species, notably the mouse. With the increasing amount of phenotype information becoming available, a major challenge that biology faces today is the systematic analysis of this information and the translation of research results across species and into an improved understanding of human disease. The challenge is to integrate and combine phenotype descriptions within a species and to systematically relate them to phenotype descriptions in other species, in order to form a comprehensive understanding of the relations between those phenotypes and the genotypes involved in human disease. We distinguish between two major approaches for comparative phenotype analyses: the first relies on evolutionary relations to bridge the species gap, while the other approach compares phenotypes directly. In particular, the direct comparison of phenotypes relies heavily on the quality and coherence of phenotype and disease databases. We discuss major achievements and future challenges for these databases in light of their potential to contribute to the understanding of the molecular mechanisms underlying human disease. In particular, we discuss how the use of ontologies and automated reasoning can significantly contribute to the analysis of phenotypes and demonstrate their potential for enabling translational research.
phenotype; animal model; disease; database; comparative phenomics; ontology
Epidermolysis bullosa (EB) describes a spectrum of rare, incurable, inherited mechanobullous disorders unified by the fact that they are caused by structural defects in the basement membrane zone which disrupt adhesion between the epidermis and dermis. Mouse models provide valuable tools to define the molecular basis of these diseases and to test novel therapeutic approaches. There is need for rapid, quantitative tests that measure the integrity of dermal-epidermal adhesions in such models. To address this need, we describe a novel quantitative method to determine the mechanical strength of the adhesion between tail skin epidermis and dermis. We show that this test reliably measures the force required to cause dermal-epidermal separation in tails of mice that are genetically predisposed to a form of non-Herlitz Junctional EB which develops as the result of a hypomorphic mutation in the laminin gamma 2 gene (Lamc2jeb). This simple, quantitative method of directly measuring the tensile strength of dermal-epidermal adhesion provides a novel dimension to the pathophysiological screening, evaluation, and therapeutic treatment of mice that may develop progressive forms of EB and potentially other disorders that compromise cutaneous integrity.
junctional epidermolysis bullosa; laminin gamma 2; tail tension test; dermis; epidermis; push-pull force gauge
B6.Cg-Tg(Thy1-YFP)16Jrs/J transgenic mice were created to express the yellow fluorescent protein gene driven by a mouse Thy1 promoter which labeled motor and sensory neurons such that individual nerves could be followed. We used these mice to identify nerves in the skin that innervate the erector pili and panniculus carnosus muscle. Whole mounts demonstrated yellow fluorescent protein (YFP) expression in nerves of the skin which was confirmed by labeling the neuromuscular junction with fluorescinated α-bungarotoxin. Frozen and paraffin embedded skin sections revealed innervation of the panniculus carnosus muscle. Paraffin sections labeled with an anti-GFP antibody revealed innervation of the panniculus carnosus as well as the erector pili muscle and around the hair follicle bulge, the last two of which were not seen by direct fluorescence.
Eccrine sweat glands in the mouse are found only on the footpads and when mature, resemble human eccrine glands. Eccrine gland anlagen were first apparent at 16.5 days post-conception (DPC) in mouse embryos as small accumulations of cells in the mesenchymal tissue beneath the developing epidermis resembling hair follicle placodes. These cells extended into the dermis where significant cell organization, duct development, and evidence of the acrosyringium were observed in 6-7 postpartum day (PPD) mice. Mouse specific keratin 1 (K1) and 10 (K10) expression was confined to the strata spinosum and granulosum. In 16.5 and 18.5 DPC embryos, K14 and K17 were both expressed in the stratum basale and diffusely in the gland anlagen. K5 expression closely mimicked K17 throughout gland development. K6 expression was not observed in the developing glands of the embryo but was apparent in the luminal cell layer of the duct by 6-7 PPD. By 21 PPD the gland apertures appeared as depressions in the surface surrounded by cornified squames and the footpad surface lacked the organized ridge and crease system seen in human fingers. These data serve as a valuable reference for investigators who utilize genetically engineered mice for skin research.
embryology; development; immunohistochemistry; and scanning electron microscopy
The NFKB pathway performs pivotal roles in diverse physiological processes, such as immunity, inflammation, proliferation, and apoptosis. NFKB is kept inactive in the cytoplasm through association with inhibitors (IKB), and translocates to the nucleus to activate its target genes after the IKBs are phosphorylated and degraded. Here we demonstrate that loss of function of SHARPIN leads to activation of NFKB signaling in skin resulting in the development of an idiopathic hypereosinophilic syndrome (IHES) with eosinophilic dermatitis in C57BL/KaLawRij-Sharpincpdm/RijSunJ mice with clonal expansion of B-1 B cells and CD3+CD4−CD8− T cells. Transcription profiling in skin revealed constitutive activation of classical NFKB pathways predominantly by overexpressed members of IL1 family. Compound null mutants for both the IL1 receptor accessory protein (Il1raptm1Roml) and SHARPIN (Sharpincpdm) resulted in mice having decreased skin disease severity. Inhibition of IKBA degradation by the proteasome inhibitor, bortezomib, alleviated the dermatitis in Sharpincpdm mice. These results indicate that absence of SHARPIN causes IHES with eosinopholic dermatitis by NFKB activation and bortezomib may be an effective treatment for skin problem of IHES.
Sharpin; bortezomib; NFKB; HES; mice
The use of standard nomenclatures for describing the strains, genes, and proteins of species is vital for the interpretation, archiving, analysis, and recovery of experimental data on the laboratory mouse. At a time when sharing of data and meta- analysis of experimental results is becoming a dominant mode of scientific investigation, failure to respect formal nomenclatures can cause confusion, errors, and in some cases contribute to poor science. Here we present the basic nomenclature rules for laboratory mice and explain how these rules should be applied to complex genetic manipulations and crosses.
Peyer’s patches (PP) are an important component in the immune response against intestinal pathogens. Two independent, spontaneous mutations in the mouse Sharpin gene (Sharpincpdm and Sharpincpdm-Dem) result in the absence of PP and disrupted splenic white pulp in adult mice, although a full complement of lymph nodes is present. Here we report that rudimentary PP begin to develop in Sharpincpdm mice during embryogenesis, but lack the organizational patterns that are typical of this tissue. In the present study, small intestines examined at weekly intervals from birth to maturity showed spontaneous regression of PP in mutant mice with concurrent infiltration of granulocytes. At 5 to 6 weeks of age, only indistinct remnants of granulocytic accumulations remain. Transplantation of normal bone marrow into Sharpincpdm mice at 7 days of age did not prevent regression of PP in bone marrow chimeras examined at 7 to 8 weeks of age. These findings indicate that SHARPIN expression is required for the normal development and maintenance, but not initiation, of PP.
Pseudoxanthoma elasticum (PXE), a heritable ectopic mineralization disorder, is caused by mutations in the ABCC6 gene. Null mice (Abcc6−/−) recapitulate the genetic, histopathologic and ultrastructural features of PXE, and they demonstrate early and progressive mineralization of vibrissae dermal sheath, which serves as a biomarker of the overall mineralization process. Recently, as part of a mouse aging study at The Jackson Laboratory, 31 inbred mouse strains were necropsied, and two of them, KK/HlJ and 129S1/SvImJ, were noted to have vibrissae dermal mineralization similar to Abcc6−/− mice. These two strains were shown to harbor a single nucleotide polymorphism (rs32756904) in the Abcc6 gene, which resulted in out-of-frame splicing and marked reduction in ABCC6 protein expression in the liver of these mice. The same polymorphism is present in two additional mouse strains, DBA/2J and C3H/HeJ, with similar reduction in Abcc6 protein levels, yet these mice did not demonstrate tissue mineralization when kept on standard rodent diet. However, all four mouse strains, when placed on experimental diet enriched in phosphate and low in magnesium, developed extensive ectopic mineralization. These results indicate that the genetic background of mice and the mineral composition of their diet can profoundly modulate the ectopic mineralization process predicated on mutations in the Abcc6 gene. These mice provide novel model systems to study the pathomechanisms and the reasons for strain background on phenotypic variability of PXE.
Alopecia areata is among the most prevalent autoimmune diseases, yet compared with other autoimmune conditions is not well studied. This in part results from limitations in the C3H/HeJ mouse and DEBR rat model systems most commonly used to study the disease, which display a low frequency and late onset. We describe a novel high incidence model for spontaneous alopecia areata. The 1MOG244 T cell expresses dual TCRA chains, one of which, when combined with the single TCRB present, promotes the development of CD8+ T cells with specificity for hair follicles. Retroviral transgenic mice expressing this TCR develop spontaneous alopecia areata at nearly 100% incidence. Disease initially follows a reticular pattern, with regionally cyclic episodes of hair loss and regrowth, and ultimately progresses to alopecia universalis. Alopecia development is associated with CD8+ T cell activation, migration into the intrafollicular region, and hair follicle destruction. The disease may be adoptively transferred with T lymphocytes, and is class I and not class II MHC-dependent. Pathologic T cells primarily express IFNG and IL17 early in disease, with dramatic increases in cytokine production and recruitment of IL4 and IL10 production with disease progression. Inhibition of individual cytokines did not significantly alter disease incidence, potentially indicating redundancy in cytokine responses. These results therefore characterize a new high incidence model for alopecia areata in C57BL/6J mice, the first to apply a monoclonal TCR, and indicate that class I MHC-restricted CD8+ T lymphocytes can independently mediate the pathologic response.