Publications by authors named "Leah Rae Donahue"

68 Publications

Developmental constraint through negative pleiotropy in the zygomatic arch.

Evodevo 2018 27;9. Epub 2018 Jan 27.

2Alberta Children's Hospital Institute for Child and Maternal Health, University of Calgary, Calgary, AB Canada.

Background: Previous analysis suggested that the relative contribution of individual bones to regional skull lengths differ between inbred mouse strains. If the negative correlation of adjacent bone lengths is associated with genetic variation in a heterogeneous population, it would be an example of negative pleiotropy, which occurs when a genetic factor leads to opposite effects in two phenotypes. Confirming negative pleiotropy and determining its basis may reveal important information about the maintenance of overall skull integration and developmental constraint on skull morphology.

Results: We identified negative correlations between the lengths of the frontal and parietal bones in the midline cranial vault as well as the zygomatic bone and zygomatic process of the maxilla, which contribute to the zygomatic arch. Through gene association mapping of a large heterogeneous population of Diversity Outbred (DO) mice, we identified a quantitative trait locus on chromosome 17 driving the antagonistic contribution of these two zygomatic arch bones to total zygomatic arch length. Candidate genes in this region were identified and real-time PCR of the maxillary processes of DO founder strain embryos indicated differences in the RNA expression levels for two of the candidate genes, and .

Conclusions: A genomic region underlying negative pleiotropy of two zygomatic arch bones was identified, which provides a mechanism for antagonism in component bone lengths while constraining overall zygomatic arch length. This type of mechanism may have led to variation in the contribution of individual bones to the zygomatic arch noted across mammals. Given that similar genetic and developmental mechanisms may underlie negative correlations in other parts of the skull, these results provide an important step toward understanding the developmental basis of evolutionary variation and constraint in skull morphology.
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http://dx.doi.org/10.1186/s13227-018-0092-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5787316PMC
January 2018

Weak Epistasis Generally Stabilizes Phenotypes in a Mouse Intercross.

PLoS Genet 2016 Feb 1;12(2):e1005805. Epub 2016 Feb 1.

The Jackson Laboratory, Bar Harbor, Maine, United States of America.

The extent and strength of epistasis is commonly unresolved in genetic studies, and observed epistasis is often difficult to interpret in terms of biological consequences or overall genetic architecture. We investigated the prevalence and consequences of epistasis by analyzing four body composition phenotypes--body weight, body fat percentage, femoral density, and femoral circumference--in a large F2 intercross of B6-lit/lit and C3.B6-lit/lit mice. We used Combined Analysis of Pleiotropy and Epistasis (CAPE) to examine interactions for the four phenotypes simultaneously, which revealed an extensive directed network of genetic loci interacting with each other, circulating IGF1, and sex to influence these phenotypes. The majority of epistatic interactions had small effects relative to additive effects of individual loci, and tended to stabilize phenotypes towards the mean of the population rather than extremes. Interactive effects of two alleles inherited from one parental strain commonly resulted in phenotypes closer to the population mean than the additive effects from the two loci, and often much closer to the mean than either single-locus model. Alternatively, combinations of alleles inherited from different parent strains contribute to more extreme phenotypes not observed in either parental strain. This class of phenotype-stabilizing interactions has effects that are close to additive and are thus difficult to detect except in very large intercrosses. Nevertheless, we found these interactions to be useful in generating hypotheses for functional relationships between genetic loci. Our findings suggest that while epistasis is often weak and unlikely to account for a large proportion of heritable variance, even small-effect genetic interactions can facilitate hypotheses of underlying biology in well-powered studies.
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http://dx.doi.org/10.1371/journal.pgen.1005805DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4734753PMC
February 2016

Modulation of unloading-induced bone loss in mice with altered ERK signaling.

Mamm Genome 2016 Feb 6;27(1-2):47-61. Epub 2015 Nov 6.

Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY, 11794-5281, USA.

Genetic variations mediate skeletal responsiveness to mechanical unloading, with individual space travelers exhibiting large variations in the extent of bone loss. We previously identified genomic regions harboring several hundred genes that can modulate the magnitude of skeletal adaptation to mechanical unloading. Here, bioinformatic filters aided in shortlisting 30 genes with bone-related and mechanoregulatory roles. The genes CD44, FGF2, NOD2, and Fas, all associated with ERK signaling, were then functionally tested in hindlimb-unloaded (HLU) knockout (KO) mice. Compared to their respective normally ambulating wildtype (WT) controls, all KO strains, except Fas mice, had lower trabecular bone volume, bone volume fraction, and/or trabecular number. For cortical bone and compared to ambulatory WT mice, CD44(-/-) had impaired properties while FGF2(-/-) showed enhanced indices. NOD2(-/-) and Fas(-/-) did not have a cortical phenotype. In all KO and WT groups, HLU resulted in impaired trabecular and cortical indices, primarily due to trabecular tissue loss and mitigation of cortical bone growth. The difference in trabecular separation between HLU and ambulatory controls was significantly greater in CD44(-/-) and NOD2(-/-) mice than in WT mice. In cortical bone, differences in cortical thickness, total pore volume, and cortical porosity between HLU and controls were aggravated in CD44(-/-) mice. In contrast, deletion of NOD2 and Fas genes mitigated the differences in Po.V between HLU and control mice. Together, we narrowed a previous list of QTL-derived candidate genes from over 300 to 30, and showed that CD44, NOD2, and Fas have distinct functions in regulating changes in trabecular and cortical bone indices during unloading.
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http://dx.doi.org/10.1007/s00335-015-9611-xDOI Listing
February 2016

A Cross-Species Analysis of Animal Models for the Investigation of Preterm Birth Mechanisms.

Reprod Sci 2016 Apr 16;23(4):482-91. Epub 2015 Sep 16.

Center for Biomedical Informatics, Warren Alpert Medical School of Brown University, Providence, RI, USA

Background: Spontaneous preterm birth is the leading cause of neonatal morbidity and mortality worldwide. The ability to examine the exact mechanisms underlying this syndrome in humans is limited. Therefore, the study of animal models is critical to unraveling the key physiologic mechanisms that control the timing of birth. The purpose of this review is to facilitate enhanced assimilation of the literature on animal models of preterm birth by a broad range of investigators.

Methods: Using classical systematic and informatics search techniques of the available literature through 2012, a database of intact animal models was generated. Research librarians generated a list of articles using multiple databases. From these articles, a comprehensive list of Medical Subject Headings (MeSH) was created. Using mathematical modeling, significant MeSH descriptors were determined, and a MEDLINE search algorithm was created. The articles were reviewed for mechanism of labor induction categorized by species.

Results: Existing animal models of preterm birth comprise specific interventions to induce preterm birth, as no animal model was identified that exhibits natural spontaneous preterm birth at an incidence comparable to that of the humans. A search algorithm was developed which when used results in a comprehensive list of agents used to induce preterm delivery in a host of animal species. The evolution of 3 specific animal models--sheep, mice, and rats--has demonstrated a clear shift in focus in the literature from endocrine to inflammatory agents of preterm birth induction.

Conclusion: The process of developing a search algorithm to provide efficient access to information on animal models of preterm birth illustrates the need for a more precise organization of the literature to allow the investigator to focus on distinctly maternal versus fetal outcomes.
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http://dx.doi.org/10.1177/1933719115604729DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5933186PMC
April 2016

Discovery and characterization of spontaneous mouse models of craniofacial dysmorphology.

Dev Biol 2016 07 31;415(2):216-227. Epub 2015 Jul 31.

The Jackson Laboratory, 600 Main St., Bar Harbor, ME 04609, USA. Electronic address:

Craniofacial abnormalities are among the most common features of human genetic syndromes and disorders. The etiology of these conditions is often complex, influenced by both genetic context and the environment. Frequently, craniofacial abnormalities present as part of a syndrome with clear comorbid phenotypes, providing additional insight into mechanisms of the causative gene or pathway. The mouse has been a key tool in our understanding of the genetic mechanisms of craniofacial development and disease, and can provide excellent models for human craniofacial abnormalities. While powerful genetic engineering tools in the mouse have contributed significantly our understanding of craniofacial development and dysmorphology, forward genetic approaches provide an unbiased means to identify new genes and pathways. Moreover, spontaneous mutations can occur on any number of genetic backgrounds, potentially revealing critical genes that require a specific genetic context. Here we report discovery and phenotyping of 43 craniofacial mouse models, derived primarily from a screen for spontaneous mutations in production colonies at the Jackson Laboratory. We identify the causative gene for 33 lines, including novel genes in pathways not previously connected to craniofacial development, and novel alleles of known genes that present with unique phenotypes. Together with our detailed characterization, this work provides a valuable gene discovery resource for the craniofacial community, and a rich source of mouse models for further investigation.
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http://dx.doi.org/10.1016/j.ydbio.2015.07.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4733616PMC
July 2016

High-throughput phenotyping and genetic linkage of cortical bone microstructure in the mouse.

BMC Genomics 2015 Jul 3;16:493. Epub 2015 Jul 3.

Institute for Biomedical Engineering, University and ETH Zurich, Gloriastrasse 35, Zurich, 8092, Switzerland.

Background: Understanding cellular structure and organization, which plays an important role in biological systems ranging from mechanosensation to neural organization, is a complicated multifactorial problem depending on genetics, environmental factors, and stochastic processes. Isolating these factors necessitates the measurement and sensitive quantification of many samples in a reliable, high-throughput, unbiased manner. In this manuscript we present a pipelined approach using a fully automated framework based on Synchrotron-based X-ray Tomographic Microscopy (SRXTM) for performing a full 3D characterization of millions of substructures.

Results: We demonstrate the framework on a genetic study on the femur bones of in-bred mice. We measured 1300 femurs from a F2 cross experiment in mice without the growth hormone (which can confound many of the smaller structural differences between strains) and characterized more than 50 million osteocyte lacunae (cell-sized hollows in the bone). The results were then correlated with genetic markers in a process called quantitative trait localization (QTL). Our findings provide a mapping between regions of the genome (all 19 autosomes) and observable phenotypes which could explain between 8-40 % of the variance using between 2-10 loci for each trait. This map shows 4 areas of overlap with previous studies looking at bone strength and 3 areas not previously associated with bone.

Conclusions: The mapping of microstructural phenotypes provides a starting point for both structure-function and genetic studies on murine bone structure and the specific loci can be investigated in more detail to identify single gene candidates which can then be translated to human investigations. The flexible infrastructure offers a full spectrum of shape, distribution, and connectivity metrics for cellular networks and can be adapted to a wide variety of materials ranging from plant roots to lung tissue in studies requiring high sample counts and sensitive metrics such as the drug-gene interactions and high-throughput screening.
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http://dx.doi.org/10.1186/s12864-015-1617-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4490749PMC
July 2015

Studies of OC-STAMP in Osteoclast Fusion: A New Knockout Mouse Model, Rescue of Cell Fusion, and Transmembrane Topology.

PLoS One 2015 4;10(6):e0128275. Epub 2015 Jun 4.

Department of Cell and Developmental Biology, University of Massachusetts Medical School, Worcester, MA, United States of America.

The fusion of monocyte/macrophage lineage cells into fully active, multinucleated, bone resorbing osteoclasts is a complex cell biological phenomenon that utilizes specialized proteins. OC-STAMP, a multi-pass transmembrane protein, has been shown to be required for pre-osteoclast fusion and for optimal bone resorption activity. A previously reported knockout mouse model had only mononuclear osteoclasts with markedly reduced resorption activity in vitro, but with paradoxically normal skeletal micro-CT parameters. To further explore this and related questions, we used mouse ES cells carrying a gene trap allele to generate a second OC-STAMP null mouse strain. Bone histology showed overall normal bone form with large numbers of TRAP-positive, mononuclear osteoclasts. Micro-CT parameters were not significantly different between knockout and wild type mice at 2 or 6 weeks old. At 6 weeks, metaphyseal TRAP-positive areas were lower and mean size of the areas were smaller in knockout femora, but bone turnover markers in serum were normal. Bone marrow mononuclear cells became TRAP-positive when cultured with CSF-1 and RANKL, but they did not fuse. Expression levels of other osteoclast markers, such as cathepsin K, carbonic anhydrase II, and NFATc1, were not significantly different compared to wild type. Actin rings were present, but small, and pit assays showed a 3.5-fold decrease in area resorbed. Restoring OC-STAMP in knockout cells by lentiviral transduction rescued fusion and resorption. N- and C-termini of OC-STAMP were intracellular, and a predicted glycosylation site was shown to be utilized and to lie on an extracellular loop. The site is conserved in all terrestrial vertebrates and appears to be required for protein stability, but not for fusion. Based on this and other results, we present a topological model of OC-STAMP as a 6-transmembrane domain protein. We also contrast the osteoclast-specific roles of OC- and DC-STAMP with more generalized cell fusion mechanisms.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0128275PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4456411PMC
May 2016

Exome sequencing reveals pathogenic mutations in 91 strains of mice with Mendelian disorders.

Genome Res 2015 Jul 27;25(7):948-57. Epub 2015 Apr 27.

The Jackson Laboratory, Bar Harbor, Maine 04609, USA;

Spontaneously arising mouse mutations have served as the foundation for understanding gene function for more than 100 years. We have used exome sequencing in an effort to identify the causative mutations for 172 distinct, spontaneously arising mouse models of Mendelian disorders, including a broad range of clinically relevant phenotypes. To analyze the resulting data, we developed an analytics pipeline that is optimized for mouse exome data and a variation database that allows for reproducible, user-defined data mining as well as nomination of mutation candidates through knowledge-based integration of sample and variant data. Using these new tools, putative pathogenic mutations were identified for 91 (53%) of the strains in our study. Despite the increased power offered by potentially unlimited pedigrees and controlled breeding, about half of our exome cases remained unsolved. Using a combination of manual analyses of exome alignments and whole-genome sequencing, we provide evidence that a large fraction of unsolved exome cases have underlying structural mutations. This result directly informs efforts to investigate the similar proportion of apparently Mendelian human phenotypes that are recalcitrant to exome sequencing.
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http://dx.doi.org/10.1101/gr.186882.114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4484392PMC
July 2015

A novel allele of Alx4 results in reduced Fgf10 expression and failure of eyelid fusion in mice.

Mamm Genome 2015 Apr 12;26(3-4):173-80. Epub 2015 Feb 12.

The Jackson Laboratory, 600 Main St., Bar Harbor, ME, 04609, USA.

Normal fusion of developing eyelids requires coordination of inductive signals from the eyelid mesenchyme with migration of the periderm cell layer and constriction of the eyelids across the eye. Failure of this process results in an eyelids open at birth (EOB) phenotype in mice. We have identified a novel spontaneous allele of Alx4 that displays EOB, in addition to polydactyly and cranial malformations. Alx4 is expressed in the eyelid mesenchyme prior to and during eyelid fusion in a domain overlapping the expression of genes that also play a role in normal eyelid development. We show that Alx4 mutant mice have reduced expression of Fgf10, a key factor expressed in the mesenchyme that is required for initiation of eyelid fusion by the periderm. This is accompanied by a reduced number of periderm cells expressing phosphorylated c-Jun, consistent with the incomplete ablation of Fgf10 expression. Together, these data demonstrate that eyelid fusion in mice requires the expression of Alx4, accompanied by the loss of normal expression of essential components of the eyelid fusion pathway.
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http://dx.doi.org/10.1007/s00335-015-9557-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4482110PMC
April 2015

Systemic injection of CK2.3, a novel peptide acting downstream of bone morphogenetic protein receptor BMPRIa, leads to increased trabecular bone mass.

J Orthop Res 2015 Feb 20;33(2):208-15. Epub 2014 Oct 20.

Department of Biological Sciences, University of Delaware, Newark, Delaware.

Bone Morphogenetic Protein 2 (BMP2) regulates bone integrity by driving both osteogenesis and osteoclastogenesis. However, BMP2 as a therapeutic has significant drawbacks. We have designed a novel peptide CK2.3 that blocks the interaction of Casein Kinase 2 (CK2) with Bone Morphogenetic Protein Receptor type Ia (BMPRIa), thereby activating BMP signaling pathways in the absence of ligand. Here, we show that CK2.3 induced mineralization in primary osteoblast cultures isolated from calvaria and bone marrow stromal cells (BMSCs) of 8 week old mice. Further, systemic tail vein injections of CK2.3 in 8 week old mice resulted in increased bone mineral density (BMD) and mineral apposition rate (MAR). In situ immunohistochemistry of the femur found that CK2.3 injection induced phosphorylation of extracellular signal-related kinase (ERK), but not Smad in osteocytes and osteoblasts, suggesting that CK2.3 signaling occurred through Smad independent pathway. Finally mice injected with CK2.3 exhibited decreased osteoclast differentiation and osteoclast activity. These data indicate that the novel mimetic peptide CK2.3 activated BMPRIa downstream signaling to enhance bone formation without the increase in osteoclast activity that accompanies BMP 2 stimulation.
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http://dx.doi.org/10.1002/jor.22752DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304894PMC
February 2015

Quantitative trait loci that modulate trabecular bone's risk of failure during unloading and reloading.

Bone 2014 Jul 31;64:25-32. Epub 2014 Mar 31.

Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY 11794, USA.

Genetic makeup of an individual is a strong determinant of the morphologic and mechanical properties of bone. Here, in an effort to identify quantitative trait loci (QTLs) for changes in the simulated mechanical parameters of trabecular bone during altered mechanical demand, we subjected 352 second generation female adult (16 weeks old) BALBxC3H mice to 3 weeks of hindlimb unloading followed by 3 weeks of reambulation. Longitudinal in vivo microcomputed tomography (μCT) scans tracked trabecular changes in the distal femur. Tomographies were directly translated into finite element (FE) models and subjected to a uniaxial compression test. Apparent trabecular stiffness and components of the Von Mises (VM) stress distributions were computed for the distal metaphysis and associated with QTLs. At baseline, five QTLs explained 20% of the variation in trabecular peak stresses across the mouse population. During unloading, three QTLs accounted for 14% of the variability in peak stresses. During reambulation, one QTL accounted for 5% of the variability in peak stresses. QTLs were also identified for mechanically induced changes in stiffness, median stress values and skewness of stress distributions. There was little overlap between QTLs identified for baseline and QTLs for longitudinal changes in mechanical properties, suggesting that distinct genes may be responsible for the mechanical response of trabecular bone. Unloading related QTLs were also different from reambulation related QTLs. Further, QTLs identified here for mechanical properties differed from previously identified QTLs for trabecular morphology, perhaps revealing novel gene targets for reducing fracture risk in individuals exposed to unloading and for maximizing the recovery of trabecular bone's mechanical properties during reambulation.
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http://dx.doi.org/10.1016/j.bone.2014.03.042DOI Listing
July 2014

Effects of housing density in five inbred strains of mice.

PLoS One 2014 21;9(3):e90012. Epub 2014 Mar 21.

The Jackson Laboratory, Bar Harbor, Maine, United States of America.

To evaluate the effect of increased mouse density in a cage, mice were housed at the density recommended by the 1996 Guide for the Care and Use of Laboratory Animals and at densities that were approximately 2, 2.6, and 3 times greater. Five strains of mice (129S1/SvImJ, A/J, BALB/cByJ, C57BL/6J, and DBA/2J) were evaluated throughout 3- and 8-month timeframes for health and well-being, including mortality, cardiac measures, plasma cholesterol, body weight, bone mineral density, organ weights, hematology, behavioral observations, and open field and light-dark tests. For 22 of the 27 traits measured, increased housing density had no significant effect. Kidney weight, adrenal weight, and heart rate decreased as mice were housed more densely, and some of the decreases were statistically significant. Reduced kidney weight, adrenal weight, and heart rate are not considered to be negative outcomes and may even indicate reduced stress. However, all measurements of these three traits were within normal physiological ranges. Percent fat increased slightly in strains 129S1/SvImJ, A/J, and DBA/2J, but did not increase in strains BALB/cByJ, and C57BL/6J. These results indicate that mice can be housed at higher densities than those currently recommended.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0090012PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3962340PMC
January 2015

Derivation and characterization of mouse embryonic stem cells from permissive and nonpermissive strains.

Nat Protoc 2014 Mar 6;9(3):559-74. Epub 2014 Feb 6.

Genetic Resource Science, The Jackson Laboratory, Bar Harbor, Maine, USA.

Mouse embryonic stem cells (mESCs) are key tools for genetic engineering, development of stem cell-based therapies and basic research on pluripotency and early lineage commitment. However, successful derivation of germline-competent embryonic stem cell lines has, until recently, been limited to a small number of inbred mouse strains. Recently, there have been considerable advances in the field of embryonic stem cell biology, particularly in the area of pluripotency maintenance in the epiblast from which the mESCs are derived. Here we describe a protocol for efficient derivation of germline-competent mESCs from any mouse strain, including strains previously deemed nonpermissive. We provide a protocol that is generally applicable to most inbred strains, as well as a variant for nonpermissive strains. By using this protocol, mESCs can be derived in 3 weeks and fully characterized after an additional 12 weeks, at efficiencies as high as 90% and in any strain background.
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http://dx.doi.org/10.1038/nprot.2014.030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4112089PMC
March 2014

Genetic loci that control the loss and regain of trabecular bone during unloading and reambulation.

J Bone Miner Res 2013 Jul;28(7):1537-49

Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY, USA.

Changes in trabecular morphology during unloading and reloading are marked by large variations between individuals, implying that there is a strong genetic influence on the magnitude of the response. Here, we subjected more than 350 second-generation (BALBxC3H) 4-month-old adult female mice to 3 weeks of hindlimb unloading followed by 3 weeks of reambulation to identify the quantitative trait loci (QTLs) that define an individual's propensity to either lose trabecular bone when weight bearing is removed or to gain trabecular bone when weight bearing is reintroduced. Longitudinal in vivo micro-computed tomography (µCT) scans demonstrated that individual mice lost between 15% and 71% in trabecular bone volume fraction (BV/TV) in the distal femur during unloading (average: -43%). Changes in trabecular BV/TV during the 3-week reambulation period ranged from a continuation of bone loss (-18%) to large additions (56%) of tissue (average: +10%). During unloading, six QTLs accounted for 21% of the total variability in changes in BV/TV whereas one QTL accounted for 6% of the variability in changes in BV/TV during reambulation. QTLs were also identified for changes in trabecular architecture. Most of the QTLs defining morphologic changes during unloading or reambulation did not overlap with those QTLs identified at baseline, suggesting that these QTLs harbor genes that are specific for sensing changes in the levels of weight bearing. The lack of overlap in QTLs between unloading and reambulation also emphasizes that the genes modulating the trabecular response to unloading are distinct from those regulating tissue recovery during reloading. The identified QTLs contain the regulatory genes underlying the strong genetic regulation of trabecular bone's sensitivity to weight bearing and may help to identify individuals that are most susceptible to unloading-induced bone loss and/or the least capable of recovering.
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http://dx.doi.org/10.1002/jbmr.1883DOI Listing
July 2013

The importance of the intracortical canal network for murine bone mechanics.

Bone 2013 Mar 3;53(1):120-8. Epub 2012 Dec 3.

Institute for Biomechanics, ETH Zurich, Zurich, Switzerland.

As shown by recent data bone strength estimation can greatly be improved by including microarchitectural parameters in the analysis. Our previous results showed that intracortical canals (the living space of the vasculature and/or remodeling units) are a major contributor to cortical tissue porosity, and therefore, can be linked to mechanical bone properties. Consequently, the goal of this study was to investigate the importance of the intracortical canal network for murine bone mechanics. To study intracortical canals within murine femoral bone, we used a mouse model, including two mouse strains, C57BL/6J-Ghrhr(lit)/J (B6-lit/+) and C3.B6-Ghrhr(lit)/J (C3.B6-lit/+) representing low and high bone mass, respectively. The intracortical canal network was assessed by synchrotron radiation-based micro-computed tomography and the mechanical bone properties were derived from three-point bending experiments. Multiple linear regression models were built to explain the variation in ultimate force, work to fracture, and stiffness in terms of the morphometric parameters. The power to explain the variation in bone mechanics was increased significantly for most mechanical measures when including morphometric parameters of intracortical canals in addition to macroscopic morphometric measures. Specifically, we could derive generalized (mouse strain-independent) models for ultimate force, where the incorporation of intracortical canals in addition to macroscopic bone measures improved the explained variation in ultimate force considerably, which was confirmed by an increase in adjusted R(2) of 73% and 8% for B6-lit/+ and C3.B6-lit/+, respectively. Further, we observed that the heterogeneity of the morphometric measures for the individual canal branches play an important role for explaining the variation in ultimate force. Finally, the current study provides strong evidence that work to fracture of murine bone, which is triggered critically by microcracks, is affected by intracortical canals. In summary, the study suggests that the intracortical canal network is important for bone mechanics.
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http://dx.doi.org/10.1016/j.bone.2012.11.024DOI Listing
March 2013

Generating embryonic stem cells from the inbred mouse strain DBA/2J, a model of glaucoma and other complex diseases.

PLoS One 2012 27;7(11):e50081. Epub 2012 Nov 27.

The Jackson Laboratory, Bar Harbor, Maine, United States of America.

Mouse embryonic stem (ES) cells are derived from the inner cell mass of blastocyst stage embryos and are used primarily for the creation of genetically engineered strains through gene targeting. While some inbred strains of mice are permissive to the derivation of embryonic stem cell lines and are therefore easily engineered, others are nonpermissive or recalcitrant. Genetic engineering of recalcitrant strain backgrounds requires gene targeting in a permissive background followed by extensive backcrossing of the engineered allele into the desired strain background. The inbred mouse strain DBA/2J is a recalcitrant strain that is used as a model of many human diseases, including glaucoma, deafness and schizophrenia. Here, we describe the generation of germ-line competent ES cell lines derived from DBA/2J mice. We also demonstrate the utility of DBA/2J ES cells with the creation of conditional knockout allele for Endothelin-2 (Edn2) directly on the DBA/2J strain background.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0050081PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3507949PMC
June 2013

Centralized mouse repositories.

Mamm Genome 2012 Oct 4;23(9-10):559-71. Epub 2012 Sep 4.

Mutant Mouse Regional Resource Center (MMRRC), The Jackson Laboratory, Bar Harbor, ME, USA.

Because the mouse is used so widely for biomedical research and the number of mouse models being generated is increasing rapidly, centralized repositories are essential if the valuable mouse strains and models that have been developed are to be securely preserved and fully exploited. Ensuring the ongoing availability of these mouse strains preserves the investment made in creating and characterizing them and creates a global resource of enormous value. The establishment of centralized mouse repositories around the world for distributing and archiving these resources has provided critical access to and preservation of these strains. This article describes the common and specialized activities provided by major mouse repositories around the world.
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http://dx.doi.org/10.1007/s00335-012-9420-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3709583PMC
October 2012

Transcriptional programs controlling perinatal lung maturation.

PLoS One 2012 20;7(8):e37046. Epub 2012 Aug 20.

The Perinatal Institute and Section of Neonatology, Perinatal and Pulmonary Biology, University of Cincinnati, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.

The timing of lung maturation is controlled precisely by complex genetic and cellular programs. Lung immaturity following preterm birth frequently results in Respiratory Distress Syndrome (RDS) and Broncho-Pulmonary Dysplasia (BPD), which are leading causes of mortality and morbidity in preterm infants. Mechanisms synchronizing gestational length and lung maturation remain to be elucidated. In this study, we designed a genome-wide mRNA expression time-course study from E15.5 to Postnatal Day 0 (PN0) using lung RNAs from C57BL/6J (B6) and A/J mice that differ in gestational length by ∼30 hr (B6
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0037046PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3423373PMC
May 2013

Discovery Genetics - The History and Future of Spontaneous Mutation Research.

Curr Protoc Mouse Biol 2012 Jun;2:103-118

The Jackson Laboratory, Bar Harbor, Maine, U.S.A.

Historically, spontaneous mutations in mice have served as valuable models of heritable human diseases, contributing substantially to our understanding of both disease mechanisms and basic biological pathways. While advances in molecular technologies have improved our ability to create mouse models of human disease through targeted mutagenesis and transgenesis, spontaneous mutations continue to provide valuable research tools for discovery of novel genes and functions. In addition, the genetic defects caused by spontaneous mutations are molecularly similar to mutations in the human genome and, therefore often produce phenotypes that more closely resemble those characteristic of human disease than do genetically engineered mutations. Due to the rarity with which spontaneous mutations arise and the animal intensive nature of their genetic analysis, large-scale spontaneous mutation analysis has traditionally been limited to large mammalian genetics institutes. More recently, ENU mutagenesis and new screening methods have increased the rate of mutant strain discovery, and high-throughput DNA sequencing has enabled rapid identification of the underlying genes and their causative mutations. Here, we discuss the continued value of spontaneous mutations for biomedical research.
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http://dx.doi.org/10.1002/9780470942390.mo110200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4215558PMC
June 2012

The influence of genetic factors on the osteoinductive potential of calcium phosphate ceramics in mice.

Biomaterials 2012 Aug 15;33(23):5696-705. Epub 2012 May 15.

Department of Tissue Regeneration, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Drienerlolaan 5, 7522 NB Enschede, The Netherlands.

The efficacy of calcium phosphate (CaP) ceramics in healing large bone defects is, in general, not as high as that of autologous bone grafting. Recently, we reported that CaP ceramics with osteoinductive properties were as efficient in healing an ilium defect of a sheep as autologous bone graft was, which makes this subclass of CaP ceramics a powerful alternative for bone regeneration. Although osteoinduction by CaP ceramics has been shown in several large animal models it is sporadically reported in mice. Because the lack of a robust mouse model has delayed understanding of the mechanism, we screened mice from 11 different inbred mouse strains for their responsiveness to subcutaneous implantation of osteoinductive tricalcium phosphate (TCP). In only two strains (FVB and 129S2) the ceramic induced bone formation, and in particularly, in FVB mice, bone was found in all the tested mice. We also demonstrated that other CaP ceramics induced bone formation at the same magnitude as that observed in other animal models. Furthermore, VEGF did not significantly increase TCP induced bone formation. The mouse model here described can accelerate research of osteoinductive mechanisms triggered by CaP ceramics and potentially the development of therapies for bone regeneration.
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http://dx.doi.org/10.1016/j.biomaterials.2012.04.021DOI Listing
August 2012

Genetic evidence that thyroid hormone is indispensable for prepubertal insulin-like growth factor-I expression and bone acquisition in mice.

J Bone Miner Res 2012 May;27(5):1067-79

Musculoskeletal Disease Center, Loma Linda VA HealthCare System, Loma Linda, CA 92357, USA.

Understanding how bone growth is regulated by hormonal and mechanical factors during early growth periods is important for optimizing the attainment of peak bone mass to prevent or postpone the occurrence of fragility fractures later in life. Using genetic mouse models that are deficient in thyroid hormone (TH) (Tshr(-/-) and Duox2(-/-)), growth hormone (GH) (Ghrhr(lit/lit)), or both (Tshr(-/-); Ghrhr(lit/lit)), we demonstrate that there is an important period prior to puberty when the effects of GH are surprisingly small and TH plays a critical role in the regulation of skeletal growth. Daily administration of T3/T4 during days 5 to 14, the time when serum levels of T3 increase rapidly in mice, rescued the skeletal deficit in TH-deficient mice but not in mice lacking both TH and GH. However, treatment of double-mutant mice with both GH and T3/T4 rescued the bone density deficit. Increased body fat in the TH-deficient as well as TH/GH double-mutant mice was rescued by T3/T4 treatment during days 5 to 14. In vitro studies in osteoblasts revealed that T3 in the presence of TH receptor (TR) α1 bound to a TH response element in intron 1 of the IGF-I gene to stimulate transcription. In vivo studies using TRα and TRβ knockout mice revealed evidence for differential regulation of insulin-like growth factor (IGF)-I expression by the two receptors. Furthermore, blockade of IGF-I action partially inhibited the biological effects of TH, thus suggesting that both IGF-I-dependent and IGF-I-independent mechanisms contribute to TH effects on prepubertal bone acquisition.
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http://dx.doi.org/10.1002/jbmr.1551DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3399953PMC
May 2012

Maternal synchronization of gestational length and lung maturation.

PLoS One 2011 9;6(11):e26682. Epub 2011 Nov 9.

The Perinatal Institute and Section of Neonatology, Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center, the Department of Pediatrics and The University of Cincinnati College of Medicine, Cincinnati, Ohio, United States of America.

Among all mammals, fetal growth and organ maturation must be precisely synchronized with gestational length to optimize survival at birth. Lack of pulmonary maturation is the major cause of infant mortality in preterm birth. Whether fetal or maternal genotypes influence the close relationship between the length of gestation and lung function at birth is unknown. Structural and biochemical indicators of pulmonary maturity were measured in two mouse strains whose gestational length differed by one day. Shorter gestation in C57BL/6J mice was associated with advanced morphological and biochemical pulmonary development and better perinatal survival when compared to A/J pups born prematurely. After ovarian transplantation, A/J pups were born early in C57BL/6J dams and survived after birth, consistent with maternal control gestational length. Expression of genes critical for perinatal lung function was assessed in A/J pups born after ovarian transfer. A subset of mRNAs important for perinatal respiratory adaptation was selectively induced in the A/J pups born after ovarian transfer. mRNAs precociously induced after ovarian transfer indicated an important role for the transcription factors C/EBPα and CREB in maternally induced lung maturation. We conclude that fetal lung maturation is determined by both fetal and maternal genotypes. Ovarian transfer experiments demonstrated that maternal genotype determines the timing of birth and can influence fetal lung growth and maturation to ensure perinatal survival.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0026682PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3212521PMC
April 2012

Multiple quantitative trait loci for cortical and trabecular bone regulation map to mid-distal mouse chromosome 4 that shares linkage homology to human chromosome 1p36.

J Bone Miner Res 2012 Jan;27(1):47-57

The Jackson Laboratory, Bar Harbor, ME 04609, USA.

The mid-distal region of mouse chromosome 4 (Chr 4) is homologous with human Chr 1p36. Previously, we reported that mouse Chr 4 carries a quantitative trait locus (QTL) with strong regulatory effect on volumetric bone mineral density (vBMD). The intent of this study is to utilize nested congenic strains to decompose the genetic complexity of this gene-rich region. Adult females and males from 18 nested congenic strains carrying discrete C3H sequences were phenotyped for femoral mineral and volume by pQCT and for trabecular bone volume (BV), tissue volume (TV), trabecular number (Trab.no), and trabecular thickness (Trab.thk) by MicroCT 40. Our data show that the mouse Chr 4 region consists of at least 10 regulatory QTL regions that affected either or both pQCT and MicroCT 40 phenotypes. The pQCT phenotypes were typically similar between sexes, whereas the MicroCT 40 phenotypes were divergent. Individual congenic strains contained one to seven QTL regions. These regions conferred large positive or negative effects in some congenic strains, depending on the particular bone phenotype. The QTL regions II to X are syntenic with human 1p36, containing from 1 to 102 known genes. We identified 13 candidate genes that can be linked to bone within these regions. Six of these genes were linked to osteoblasts, three linked to osteoclasts, and two linked to skeletal development. Three of these genes have been identified in Genome Wide Association Studies (GWAS) linked to 1p36. In region III, there is only one gene, Lck, which conferred negative pQCT and MicroCT 40 phenotypes in both sexes. This gene is important to development and functioning of T cells, has been associated with osteoclast activity, and represents a novel bone regulatory gene that merits further experimental evaluation. In summary, congenic strains are powerful tools for identifying regulatory regions that influence bone biology and offer models for testing hypotheses about gene-gene and gene-environment interactions that are not available to experimental work in humans.
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http://dx.doi.org/10.1002/jbmr.515DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3460065PMC
January 2012

Hypothalamic mitochondrial dysfunction associated with anorexia in the anx/anx mouse.

Proc Natl Acad Sci U S A 2011 Nov 24;108(44):18108-13. Epub 2011 Oct 24.

Department of Molecular Medicine and Surgery, Karolinska Institutet, 171 76 Stockholm, Sweden.

The anorectic anx/anx mouse exhibits disturbed feeding behavior and aberrances, including neurodegeneration, in peptidergic neurons in the appetite regulating hypothalamic arcuate nucleus. Poor feeding in infants, as well as neurodegeneration, are common phenotypes in human disorders caused by dysfunction of the mitochondrial oxidative phosphorylation system (OXPHOS). We therefore hypothesized that the anorexia and degenerative phenotypes in the anx/anx mouse could be related to defects in the OXPHOS. In this study, we found reduced efficiency of hypothalamic OXPHOS complex I assembly and activity in the anx/anx mouse. We also recorded signs of increased oxidative stress in anx/anx hypothalamus, possibly as an effect of the decreased hypothalamic levels of fully assembled complex I, that were demonstrated by native Western blots. Furthermore, the Ndufaf1 gene, encoding a complex I assembly factor, was genetically mapped to the anx interval and found to be down-regulated in anx/anx mice. These results suggest that the anorexia and hypothalamic neurodegeneration of the anx/anx mouse are associated with dysfunction of mitochondrial complex I.
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http://dx.doi.org/10.1073/pnas.1114863108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3207677PMC
November 2011

Molecular characterization of the translocation breakpoints in the Down syndrome mouse model Ts65Dn.

Mamm Genome 2011 Dec 28;22(11-12):685-91. Epub 2011 Sep 28.

The Jackson Laboratory, Genetic Resource Science, ME, 04609, USA.

Ts65Dn is a mouse model of Down syndrome: a syndrome that results from chromosome (Chr) 21 trisomy and is associated with congenital defects, cognitive impairment, and ultimately Alzheimer's disease. Ts65Dn mice have segmental trisomy for distal mouse Chr 16, a region sharing conserved synteny with human Chr 21. As a result, this strain harbors three copies of over half of the human Chr 21 orthologs. The trisomic segment of Chr 16 is present as a translocation chromosome (Mmu17(16)), with breakpoints that have not been defined previously. To molecularly characterize the Chrs 16 and 17 breakpoints on the translocation chromosome in Ts65Dn mice, we used a selective enrichment and high-throughput paired-end sequencing approach. Analysis of paired-end reads flanking the Chr 16, Chr 17 junction on Mmu17(16) and de novo assembly of the reads directly spanning the junction provided the precise locations of the Chrs 16 and 17 breakpoints at 84,351,351 and 9,426,822 bp, respectively. These data provide the basis for low-cost, highly efficient genotyping of Ts65Dn mice. More importantly, these data provide, for the first time, complete characterization of gene dosage in Ts65Dn mice.
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http://dx.doi.org/10.1007/s00335-011-9357-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3505986PMC
December 2011

Mouse genomic variation and its effect on phenotypes and gene regulation.

Nature 2011 Sep 14;477(7364):289-94. Epub 2011 Sep 14.

The Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1HH, UK.

We report genome sequences of 17 inbred strains of laboratory mice and identify almost ten times more variants than previously known. We use these genomes to explore the phylogenetic history of the laboratory mouse and to examine the functional consequences of allele-specific variation on transcript abundance, revealing that at least 12% of transcripts show a significant tissue-specific expression bias. By identifying candidate functional variants at 718 quantitative trait loci we show that the molecular nature of functional variants and their position relative to genes vary according to the effect size of the locus. These sequences provide a starting point for a new era in the functional analysis of a key model organism.
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http://dx.doi.org/10.1038/nature10413DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3276836PMC
September 2011

Mutation discovery in mice by whole exome sequencing.

Genome Biol 2011 Sep 14;12(9):R86. Epub 2011 Sep 14.

The Jackson Laboratory, 600 Main St, Bar Harbor, ME 04609, USA.

We report the development and optimization of reagents for in-solution, hybridization-based capture of the mouse exome. By validating this approach in a multiple inbred strains and in novel mutant strains, we show that whole exome sequencing is a robust approach for discovery of putative mutations, irrespective of strain background. We found strong candidate mutations for the majority of mutant exomes sequenced, including new models of orofacial clefting, urogenital dysmorphology, kyphosis and autoimmune hepatitis.
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http://dx.doi.org/10.1186/gb-2011-12-9-r86DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3308049PMC
September 2011

Mitotic defects lead to pervasive aneuploidy and accompany loss of RB1 activity in mouse LmnaDhe dermal fibroblasts.

PLoS One 2011 Mar 25;6(3):e18065. Epub 2011 Mar 25.

The Jackson Laboratory, Bar Harbor, Maine, United States of America.

Background: Lamin A (LMNA) is a component of the nuclear lamina and is mutated in several human diseases, including Emery-Dreifuss muscular dystrophy (EDMD; OMIM ID# 181350) and the premature aging syndrome Hutchinson-Gilford progeria syndrome (HGPS; OMIM ID# 176670). Cells from progeria patients exhibit cell cycle defects in both interphase and mitosis. Mouse models with loss of LMNA function have reduced Retinoblastoma protein (RB1) activity, leading to aberrant cell cycle control in interphase, but how mitosis is affected by LMNA is not well understood.

Results: We examined the cell cycle and structural phenotypes of cells from mice with the Lmna allele, Disheveled hair and ears (Lmna(Dhe)). We found that dermal fibroblasts from heterozygous Lmna(Dhe) (Lmna(Dhe/+)) mice exhibit many phenotypes of human laminopathy cells. These include severe perturbations to the nuclear shape and lamina, increased DNA damage, and slow growth rates due to mitotic delay. Interestingly, Lmna(Dhe/+) fibroblasts also had reduced levels of hypophosphorylated RB1 and the non-SMC condensin II-subunit D3 (NCAP-D3), a mitosis specific centromere condensin subunit that depends on RB1 activity. Mitotic check point control by mitotic arrest deficient-like 1 (MAD2L1) also was perturbed in Lmna(Dhe/+) cells. Lmna(Dhe/+) fibroblasts were consistently aneuploid and had higher levels of micronuclei and anaphase bridges than normal fibroblasts, consistent with chromosome segregation defects.

Conclusions: These data indicate that RB1 may be a key regulator of cellular phenotype in laminopathy-related cells, and suggest that the effects of LMNA on RB1 include both interphase and mitotic cell cycle control.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0018065PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3064591PMC
March 2011

The FaceBase Consortium: a comprehensive program to facilitate craniofacial research.

Dev Biol 2011 Jul 31;355(2):175-82. Epub 2011 Mar 31.

Department of Biomedical Informatics, University of Pittsburgh, Pittsburgh, PA 15232, USA.

The FaceBase Consortium consists of ten interlinked research and technology projects whose goal is to generate craniofacial research data and technology for use by the research community through a central data management and integrated bioinformatics hub. Funded by the National Institute of Dental and Craniofacial Research (NIDCR) and currently focused on studying the development of the middle region of the face, the Consortium will produce comprehensive datasets of global gene expression patterns, regulatory elements and sequencing; will generate anatomical and molecular atlases; will provide human normative facial data and other phenotypes; conduct follow up studies of a completed genome-wide association study; generate independent data on the genetics of craniofacial development, build repositories of animal models and of human samples and data for community access and analysis; and will develop software tools and animal models for analyzing and functionally testing and integrating these data. The FaceBase website (http://www.facebase.org) will serve as a web home for these efforts, providing interactive tools for exploring these datasets, together with discussion forums and other services to support and foster collaboration within the craniofacial research community.
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http://dx.doi.org/10.1016/j.ydbio.2011.02.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3440302PMC
July 2011