Genetic Background Of Mice Research Articles

Mouse Genetic Background influences Endothelial Function along the Mouse Vascular Tree

The endothelium is critical for maintaining vascular integrity and protecting against cardiovascular disease. Endothelial dysfunction is associated with a host of disease states including cardiovascular disease, diabetes, chronic kidney disease, and Alzheimer's disease. However, it is unclear whether endothelial function or dysfunction varies throughout the vasculature. In addition, accumulating evidence suggests that endothelial function, and more generally vasomotor function, is genetically regulated. The purpose of this study was to determine the influence of genetic background on vasomotor function in blood vessels of differing size. Vasomotor function was measured in aorta (thoracic (TA), and abdominal (AA)) and conduit arteries (carotid artery (CA) and femoral artery (FA)) from two inbred strains (NZW/LacJ (NZW) and C57BL/6J (B6)) of mice. Arteries were dissected, cut into 2 mm segments, and mounted in a wire myograph system. Increasing concentrations of phenylephrine (PE, 1×10−9 – 1×10−5M) were used to measure contractile responses, while increasing concentrations of the endothelium‐dependent vasodilator acetylcholine (ACh, 1×10−9 – 1×10−5M) and endothelium‐independent vasodilator sodium nitroprusside (SNP, 1×10−9 – 1×10−5M) were used to assess relaxation responses. Contractile responses to PE were not significantly different between strains across all vessels. Endothelium‐dependent relaxation to ACh was significantly impaired in TA (P = .0001), AA (P = .0019), and FA (P = .0002) from NZW as compared to B6, while CA relaxation responses were not statistically different between groups. Sensitivity to ACh as determined by IC50 was impaired in NZW TA (P = .0006), AA (P = .0005), CA (P = .001), and FA (P = .0003). Relaxation responses to the endothelium‐independent vasodilator SNP were not statistically different in the AA, CA and FA, while TA relaxation to SNP was statistically lower in B6 TA compared to NZW (P = .02). Sensitivity to SNP was not statistically different between groups for TA, CA, and FA but NZW AA showed a greater sensitivity to SNP than did B6 (P = .0304). These findings indicate a strong influence of genetic background on vasomotor function and could be important in understanding the implications that genetics have on vascular response to vasoactive agents.Support or Funding InformationThis work was supported by a Texas A&M Triads for Transformation grant (Michael Massett, Christopher Woodman), a Texas A&M Merit Fellowship (Dylan Holly), and a Texas A&M College of Education Strategic Research Award (Song Yi Shin).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Humanized NOD/SCID/IL2rγnull (hu-NSG) Mouse Model for HIV Replication and Latency Studies.

Ethical regulations and technical challenges for research in human pathology, immunology, and therapeutic development have placed small animal models in high demand. With a close genetic and behavioral resemblance to humans, small animals such as the mouse are good candidates for human disease models, through which human-like symptoms and responses can be recapitulated. Further, the mouse genetic background can be altered to accommodate diverse demands. The NOD/SCID/IL2rγnull (NSG) mouse is one of the most widely used immunocompromised mouse strains; it allows engraftment with human hematopoietic stem cells and/or human tissues and the subsequent development of a functional human immune system. This is a critical milestone in understanding the prognosis and pathophysiology of human-specific diseases such as HIV/AIDS and aiding the search for a cure. Herein, we report a detailed protocol for generating a humanized NSG mouse model (hu-NSG) by hematopoietic stem cell transplantation into a radiation-conditioned neonatal NSG mouse. The hu-NSG mouse model shows multi-lineage development of transplanted human stem cells and susceptibility to HIV-1 viral infection. It also recapitulates key biological characteristics in response to combinatorial antiretroviral therapy (cART).

Silencing of high-affinity insulin-reactive B lymphocytes by anergy and impact of the NOD genetic background in mice.

Previous studies have demonstrated that high-affinity insulin-binding B cells (IBCs) silenced by anergy in healthy humans lose their anergy in islet autoantibody-positive individuals with recent-onset type 1 diabetes, and in autoantibody-negative first-degree relatives carrying certain risk alleles. Here we explore the hypothesis that IBCs are found in the immune periphery of disease-resistant C57BL/6-H2g7 mice, where, as in healthy humans, they are anergic, but that in disease-prone genetic backgrounds (NOD) they become activated and migrate to the pancreas and pancreatic lymph nodes, where they participate in the development of type 1 diabetes. We compared the status of high-affinity IBCs in disease-resistant VH125.C57BL/6-H2g7 and disease-prone VH125.NOD mice. Consistent with findings in healthy humans, high-affinity IBCs reach the periphery in disease-resistant mice and are anergic, as indicated by a reduced expression of membrane IgM, unresponsiveness to antigen and failure to become activated or accumulate in the pancreatic lymph nodes or pancreas. In NOD mice, high-affinity IBCs reach the periphery early in life and increase in number prior to the onset of hyperglycaemia. These cells are not anergic; they become activated, produce autoantibodies and accumulate in the pancreas and pancreatic lymph nodes prior to disease development. These findings are consistent with genetic determination of the escape of high-affinity IBCs from anergy and their early contribution to the development of type 1 diabetes.

Spatial statistical tools for genome-wide mutation cluster detection under a microarray probe sampling system.

Mutation cluster analysis is critical for understanding certain mutational mechanisms relevant to genetic disease, diversity, and evolution. Yet, whole genome sequencing for detection of mutation clusters is prohibitive with high cost for most organisms and population surveys. Single nucleotide polymorphism (SNP) genotyping arrays, like the Mouse Diversity Genotyping Array, offer an alternative low-cost, screening for mutations at hundreds of thousands of loci across the genome using experimental designs that permit capture of de novo mutations in any tissue. Formal statistical tools for genome-wide detection of mutation clusters under a microarray probe sampling system are yet to be established. A challenge in the development of statistical methods is that microarray detection of mutation clusters is constrained to select SNP loci captured by probes on the array. This paper develops a Monte Carlo framework for cluster testing and assesses test statistics for capturing potential deviations from spatial randomness which are motivated by, and incorporate, the array design. While null distributions of the test statistics are established under spatial randomness via the homogeneous Poisson process, power performance of the test statistics is evaluated under postulated types of Neyman-Scott clustering processes through Monte Carlo simulation. A new statistic is developed and recommended as a screening tool for mutation cluster detection. The statistic is demonstrated to be excellent in terms of its robustness and power performance, and useful for cluster analysis in settings of missing data. The test statistic can also be generalized to any one dimensional system where every site is observed, such as DNA sequencing data. The paper illustrates how the informal graphical tools for detecting clusters may be misleading. The statistic is used for finding clusters of putative SNP differences in a mixture of different mouse genetic backgrounds and clusters of de novo SNP differences arising between tissues with development and carcinogenesis.

Antiviral Activity of Fermented Ginseng Extracts against a Broad Range of Influenza Viruses.

Ginseng products used as herb nutritional supplements are orally consumed and fermented to ginsenoside compounds by the intestinal microbes. In this study, we investigated antiviral protective effects of fermented ginseng extracts against different strains of influenza viruses in genetically diverse mouse models. Intranasal coinoculation of mice with fermented ginseng extract and influenza virus improved survival rates and conferred protection against H1N1, H3N2, H5N1, and H7N9 strains, with the efficacy dependent on the dose of ginseng samples. Antiviral protection by fermented ginseng extract was observed in different genetic backgrounds of mice and in the deficient conditions of key adaptive immune components (CD4, CD8, B cell, MHCII). The mice that survived primary virus inoculation with fermented ginseng extract developed immunity against the secondary infection with homologous and heterosubtypic viruses. In vitro cell culture experiments showed moderate virus neutralizing activity by fermented ginseng extract, probably by inhibiting hemagglutination and neuraminidase activity. This study suggests that fermented ginseng extracts might provide a means to treat influenza disease regardless of virus strains.

Recovery from an acute systemic and central LPS-inflammation challenge is affected by mouse sex and genetic background

Genetic and sexual factors influence the prevalence and the pathogenesis of many inflammatory disorders. In this study their relevance on the peripheral and central inflammatory status induced by a peripheral injection of lipopolysaccharide (LPS) was evaluated. BALB/c and CD-1 male and female mice were intraperitoneally injected with LPS. Spleens and brains were collected 2 and 72 hours later to study the levels of IL-6, TNF-α and IL-1β. Percentage of microglia and astrocytes was determined in the cortex and hippocampus. Locomotor activity was registered before and during the 72 hours after LPS-treatment. Two hours after LPS-injection, a peripheral increase of the three cytokines was found. In brains, LPS increased TNF-α only in males with higher levels in CD-1 than BALB/c. IL-1β increased only in CD-1 males. IL-6 increased in both strains with lower levels in BALB/c females. Peripheral and central levels of cytokines decline 72 hrs after LPS-treatment whilst a significantly increase of Iba-1 expression was detected. A dramatic drop of the locomotor activity was observed immediately after LPS injection. Our results show that acute systemic administration of LPS leads to peripheral and central increase of pro-inflammatory cytokines and microglia activation, in a strain and sex dependent manner.

In silico mapping of quantitative trait loci (QTL) regulating the milk ionome in mice identifies a milk iron locus on chromosome 1.

The breast-feeding neonate depends on mother's milk for both macronutrients and micronutrients including minerals. The goals of the present study were to document the effects of genetic background in mice on milk concentrations of select minerals and to use genome-wide association study (GWAS) to identify quantitative trait loci (QTL) regulating milk mineral concentrations. Milk samples from lactating mice in each of 31 different inbred strains of the mouse diversity panel(MDP) were analyzed by inductively coupled plasma-optical emission spectroscopy to determine the concentrations of calcium (Ca), copper (Cu), iron (Fe), potassium (K), magnesium (Mg), sodium (Na), phosphorus (P), sulfur (S), and zinc (Zn). GWAS identified a single pleiotropic milk mineral concentration QTL (Mmcq) on chromosome 3 for Ca, Mg, and P. For the remaining minerals, six QTL were detected for Fe, four for K, three for Zn, and one for S. Intersecting the Mmcq with published chromatin immunoprecipitation sequence data identified 15 out of 4633 high-linkage disequilibrium single-nucleotide polymorphisms that resided in signal transducer and activation of transcription 5 (STAT5) binding regions. A milk Fe-associated locus (Mmcq9) on chromosome 1 contained an SNP that localized to a STAT5 binding region and intersected with a HOMER motif predicted to bind the transcriptional regulator E74-Like ETS transcription factor 5. This locus also contained the genes for solute carrier family (Slc) members Slc9a2, Slc9a4, Slc39a10, and Slc40a1. Expression analysis of these transporters supports the conclusion that Slc9a2 and Slc40a1 within the mammary gland could mediate the effect of Mmcq9 on milk Fe concentration.

A Population Study of Common Ocular Abnormalities in C57BL/6N rd8 Mice.

PurposeThe purpose of this study was to quantify the frequency and severity of ocular abnormalities affecting wild-type C57BL/6N mice, the most common strain used worldwide for the creation of single-gene knockouts.MethodsA total of 2773 animals (5546 eyes) were examined at one colony at UC Davis and in three more colonies at the Institut Clinique de la Souris in Strasbourg, France. Mice were examined at 15 to 16 weeks postnatal age by performing anterior segment biomicroscopy, posterior segment examination by indirect ophthalmoscopy, intraocular pressure measurement, and optical coherence tomography of anterior and posterior segment structures.ResultsCommon ocular findings in the C57BL/6N strain included corneal deposits (3%), increased optical density of the anterior lens capsule (67%), punctate nuclear cataracts (98%), vitreous crystalline deposits (61%), hyaloid vascular remnant (6%), and retinal dysplasia attributed to the rd8 mutation (58%). Interestingly, retinal dysplasia was more common in male mice in all four breeding colonies evaluated in this study. The thickness of ocular tissues and compartments were measured by spectral-domain optical coherence tomography, including the central cornea, anterior chamber, vitreous, and retinal layers. Intraocular pressure was measured by rebound tonometry.ConclusionsOcular abnormalities are common in anterior and posterior segments of the C57BL/6N mouse, the most common background on which single-gene knockout mice have been made. It is important that vision scientists understand the extent and variability of ocular findings associated with this particular genetic background of mice.

Proof‐of‐Concept in a 3D Culture Model of Glioma Invasion: Towards Personalized Therapeutics in Brain Cancer

Glioma is a highly aggressive form of brain cancer, with some subtypes having 5‐year survival rates of less than 5%. Tumor cell invasion into the surrounding parenchyma seems to be the primary driver of these poor outcomes, as most gliomas recur within 2 cm of the original surgically‐resected tumor. Many current approaches to the development of anticancer therapy attempt to target genetic weaknesses in a particular cancer, but may not take into account the microenvironment experienced by a tumor and patient‐specific genetic differences in susceptibility to treatment.The primary aim of our research program is to overcome these limitations by ultimately developing 3D cerebral organoids generated from patient‐derived induced pluripotent stem cells (iPSCs), in which we will screen to identify an optimal treatment regimen by combining the organoids with patient‐derived tumor cells. As a proof‐of‐concept of this approach, we developed a system in which we could model the invasion of human glioma cells into mouse neural progenitor cell‐derived spheroids. Here we demonstrate the utility of such an approach, and find that we can follow invasion of human tumor cells using cell tracking dyes and 3D laser scanning confocal microscopy, both in real time and in fixed samples. These results were validated using conventional cryosectioning. We go on to show broad applicability of this system by using two different tumor cell lines, and two different mouse genetic backgrounds, attempting to mimic differences that might be observed in tumor and patient brain tissue, respectively. Having obtained this proof‐of‐concept, we will next recapitulate these results in a fully humanized system using spheroids and then cerebral organoids from human iPSCs, before moving on to patient‐derived cultures. This stepwise approach could ultimately lead to truly personalized treatments for brain cancer.Support or Funding InformationDvP is supported by the UBC Faculty of Medicine, the UBC Summer Student Research Program, and the Mach‐Gaensslen Foundation. CCN is supported by the Canada Research Chair program and a grant from the Canadian Cancer Society. WCS is supported by the Canadian Cancer Society.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Regulatory Lymphoid and Myeloid Cells Determine the Cardiac Immunopathogenesis of Trypanosoma cruzi Infection.

Chagas disease is a multisystemic disorder caused by the protozoan parasite Trypanosoma cruzi, which affects ~8 million people in Latin America, killing 7,000 people annually. Chagas disease is one of the main causes of death in the endemic area and the leading cause of infectious myocarditis in the world. T. cruzi infection induces two phases, acute and chronic, where the infection is initially asymptomatic and the majority of patients will remain clinically indeterminate for life. However, over a period of 10–30 years, ~30% of infected individuals will develop irreversible, potentially fatal cardiac syndromes (chronic chagasic cardiomyopathy [CCC]), and/or dilatation of the gastro-intestinal tract (megacolon or megaesophagus). Myocarditis is the most serious and frequent manifestation of chronic Chagas heart disease and appears in about 30% of infected individuals several years after infection occurs. Myocarditis is characterized by a mononuclear cell infiltrate that includes different types of myeloid and lymphoid cells and it can occur also in the acute phase. T. cruzi infects and replicates in macrophages and cardiomyocytes as well as in other nucleated cells. The pathogenesis of the chronic phase is thought to be dependent on an immune-inflammatory reaction to a low-grade replicative infection. It is known that cytokines produced by type 1 helper CD4+ T cells are able to control infection. However, the role that infiltrating lymphoid and myeloid cells may play in experimental and natural Chagas disease pathogenesis has not been completely elucidated, and several reports indicate that it depends on the mouse genetic background and parasite strain and/or inoculum. Here, we review the role that T cell CD4+ subsets, myeloid subclasses including myeloid-derived suppressor cells may play in the immunopathogenesis of Chagas disease with special focus on myocarditis, by comparing results obtained with different experimental animal models.

Cross-Protective Efficacy of Influenza Virus M2e Containing Virus-Like Particles Is Superior to Hemagglutinin Vaccines and Variable Depending on the Genetic Backgrounds of Mice.

Influenza virus M2 extracellular domain (M2e) has been a target for developing cross-protective vaccines. However, the efficacy and immune correlates of M2e vaccination are poorly understood in the different host genetic backgrounds in comparison with influenza vaccines. We previously reported the cross-protective efficacy of virus-like particle (M2e5x VLP) vaccines containing heterologous tandem M2e repeats (M2e5x) derived from human, swine, and avian influenza viruses. In this study to gain better understanding of cross-protective influenza vaccines, we compared immunogenicity and efficacy of M2e5x VLP, H5 hemagglutinin VLP (HA VLP), and inactivated H3N2 virus (H3N2i) in wild-type strains of BALB/c and C57BL/6 mice, and CD4 and CD8 knockout (KO) mice. M2e5x VLP was superior to HA VLP in conferring cross-protection whereas H3N2i inactivated virus vaccine provided high efficacy of homologous protection. After M2e5x VLP vaccination and challenge, BALB/c mice induced higher IgG responses, lower lung viral loads, and less body weight loss when compared with those in C57BL/6 mice. M2e5x VLP but not H3N2i immune mice after primary challenges developed strong immunity against a secondary heterosubtypic virus as a model of future pandemics. M2e5x VLP and HA VLP vaccines were able to raise IgG isotypes in CD4 KO mice. T cells were found to contribute to cross-protection by playing a role in reducing lung viral loads. In conclusion, M2e5x VLP vaccination induced better cross-protection than HA VLP, and its efficacy varied depending on the genetic backgrounds of mice, supporting the important roles of T cells.

Long-term maintenance of peripheral blood derived human NK cells in a novel human IL-15- transgenic NOG mouse

We generated a novel mouse strain expressing transgenic human interleukin-15 (IL-15) using the severe immunodeficient NOD/Shi-scid-IL-2Rγnull (NOG) mouse genetic background (NOG-IL-15 Tg). Human natural killer (NK) cells, purified from the peripheral blood (hu-PB-NK) of normal healthy donors, proliferated when transferred into NOG-IL-15 Tg mice. In addition, the cell number increased, and the hu-PB-NK cells persisted for 3 months without signs of xenogeneic graft versus host diseases (xGVHD). These in vivo-expanded hu-PB-NK cells maintained the original expression patterns of various surface antigens, including NK receptors and killer cell immunoglobulin-like receptor (KIR) molecules. They also contained significant amounts of granzyme A and perforin. Inoculation of K562 leukemia cells into hu-PB-NK-transplanted NOG-IL-15 Tg mice resulted in significant suppression of tumor growth compared with non-transplanted mice. Furthermore, NOG-IL-15 Tg mice allowed for engraftment of in vitro-expanded NK cells prepared for clinical cell therapy. These cells exerted antibody-dependent cell-mediated cytotoxicity (ADCC) on Her2-positive gastric cancer cells in the presence of therapeutic anti-Her2 antibody, and subsequently suppressed tumor growth. Our results collectively suggest that the NOG-IL-15 Tg mice are a useful model for studying human NK biology and evaluating human NK cell-mediated in vivo cytotoxicity.

Quantification of Inter-Sample Differences in T-Cell Receptor Repertoires Using Sequence-Based Information.

Inter-sample comparisons of T-cell receptor (TCR) repertoires are crucial for gaining a better understanding of the immunological states determined by different collections of T cells from different donor sites, cell types, and genetic and pathological backgrounds. For quantitative comparison, most previous studies utilized conventional methods in ecology, which focus on TCR sequences that overlap between pairwise samples. Some recent studies attempted another approach that is categorized into Poisson abundance models using the abundance distribution of observed TCR sequences. However, these methods ignore the details of the measured sequences and are consequently unable to identify sub-repertoires that might have important contributions to the observed inter-sample differences. Moreover, the sparsity of sequence data due to the huge diversity of repertoires hampers the performance of these methods, especially when few overlapping sequences exist. In this paper, we propose a new approach for REpertoire COmparison in Low Dimensions (RECOLD) based on TCR sequence information, which can estimate the low-dimensional structure by embedding the pairwise sequence dissimilarities in high-dimensional sequence space. The inter-sample differences between repertoires are then quantified by information-theoretic measures among the distributions of data estimated in the embedded space. Using datasets of mouse and human TCR repertoires, we demonstrate that RECOLD can accurately identify the inter-sample hierarchical structures, which have a good correspondence with our intuitive understanding about sample conditions. Moreover, for the dataset of transgenic mice that have strong restrictions on the diversity of their repertoires, our estimated inter-sample structure was consistent with the structure estimated by previous methods based on abundance or overlapping sequence information. For the dataset of human healthy donors and Sézary syndrome patients, our method also showed robust estimation performance even under the condition of high sparsity in TCR sequences, while previous studies failed to estimate the structure. In addition, we identified the sequences that contribute to the pairwise-sample differences between the repertoires with the different genetic backgrounds of mice. Such identification of the sequences contributing to variation in immune cell repertoires may provide substantial insight for the development of new immunotherapies and vaccines.

Histological study of postnatal development of mouse tongues.

Numerous factors, including trauma, tumors and myophagism, may lead to tongue defects, which are mostly repaired via muscular flaps. However, these methods cannot restore the muscular function and gustation function of the tongue. Intensive research on tongue development may offer useful clues for tongue regeneration based on tissue engineering or stem cell therapy. In the present study, staining results revealed that tongue muscle fibers became larger, mature and stronger, and the foliate and fungiform papillae also became mature from newborn to adult C57BL/6J genetic background mice. Immunofluorescence staining and polymerase chain reaction results revealed that C-kit was dynamically expressed in muscle cells, as well as in foliate and fungiform papilla cells from newborn to adult stages. The expression level decreased from P1 to P15 and increased at P90. The immunofluorescence staining results revealed that Ki-67 was expressed in muscle cells and papilla cells from newborn to adult stages, and high expression was observed at P6 and P15. In addition, the immunofluorescence staining results also demonstrated that msh homeobox 2 (Msx2) was dynamically expressed in postnatal tongue muscle cells; however, almost no expression was detected in papilla cells. There was relative high expression level of Msx2 at P1 and P6 stages, but this gradually decreased from P15, and it was expressed primarily in the muscle cells located in the marginal zone of the tongue at P90. These findings suggest that the amount of c-kit-expressing precursor cells in tongue muscle and papilla cells increases to promote tongue development at the early postnatal stage and to maintain homeostasis and functional adaptation of the tongue in the adult stage. Furthermore, Msx2 may serve an important role in postnatal tongue muscle development. The present study also suggests that C-kit and Msx2 may be used as cell markers for postnatal tongue regeneration and self-repair, and may provide an approach for developing treatment methods for tongue diseases with a postnatal onset.

Genetic driver mutations define the expression signature and microenvironmental composition of high-grade gliomas.

High-grade gliomas (HGG), including glioblastomas, are characterized by invasive growth, resistance to therapy, and high inter- and intra-tumoral heterogeneity. The key histological hallmarks of glioblastoma are pseudopalisading necrosis and microvascular proliferation, which allow pathologists to distinguish glioblastoma from lower-grade gliomas. In addition to being genetically and molecularly heterogeneous, HGG are also heterogeneous with respect to the composition of their microenvironment. The question of whether this microenvironmental heterogeneity is driven by the molecular identity of the tumor remains controversial. However, this question is of utmost importance since microenvironmental, non-neoplastic cells are key components of the most radiotherapy- and chemotherapy-resistant niches of the tumor. Our work demonstrates a versatile, reliable, and reproducible adult HGG mouse model with NF1-silencing as a driver mutation. This model shows significant differences in tumor microenvironment, expression of subtype-specific markers, and response to standard therapy when compared to our established PDGFB-overexpressing HGG mouse model. PDGFB-overexpressing and NF1-silenced murine tumors closely cluster with human proneural and mesenchymal subtypes, as well as PDGFRA-amplified and NF1-deleted/mutant human tumors, respectively, at both the RNA and protein expression levels. These models can be generated in fully immunocompetent mixed or C57BL/6 genetic background mice, and therefore can easily be incorporated into preclinical studies for cancer cell-specific or immune cell-targeting drug discovery studies.

Abstract 2850: A systems genetics approach reveals Rnaseh2c-immune response axis that affects metastasis severity in breast cancer

Abstract Metastatic breast cancer is a devastating disease with a 5-year survival rate of only 26%. This is due to a lack of effective therapies against established metastases and an inability to identify high risk patients who would benefit from specific adjuvant therapies to prevent metastatic progression. We have shown in mouse models that spontaneously arising tumors metastasize with different severity based on the mouse genetic background. Using systems genetics approaches we have identified genes correlated with metastasis and survival in both mice and humans. Rnaseh2c was identified as a novel candidate metastasis susceptibility gene. This gene encodes a scaffolding subunit of the Ribonuclease H2 enzyme which removes ribonucleotides misincorporated into the DNA. Experimentally modulating Rnaseh2c expression in a murine mammary cancer cell line resulted in significant changes in pulmonary metastasis, confirming this gene as a metastasis modifier. Mutations in Rnaseh2c are known to cause Aicardi-Goutieres Syndrome, a neurological autoinflammatory disorder that overlaps clinically with congenital viral infections and the autoimmune disease Systemic Lupus Erythematosus. Given this, we hypothesized that altered expression of Rnaseh2c in breast cancer cells affects metastasis by engaging the immune system. To investigate immune system involvement, we analyzed metastasis in immunocompromised mice. T cell deficiency ablated the effect of reduced Rnaseh2c expression on metastasis, revealing for the first time an Rnaseh2c-immune response axis in metastasis. Gene ontology pathway analysis following mRNA-sequencing of Rnaseh2c knockdown and overexpression tumors revealed that 20% of the genes with altered expression are involved in immune system-related pathways, including T cell signaling and antigen presentation. Furthermore, genes with significant changes included Type I interferons, T cell markers, and immune regulators. These results confirm that Rnaseh2c is a novel metastasis modifier gene and validate our hypothesis that the immune system is mediating the effect of Rnaseh2c on metastasis. This mechanism highlights a potential new target for combination with immune modulatory therapies to combat this devastating disease and adds to a panel of genes we identified that together could determine patients with high risk for metastasis. Citation Format: Sarah Deasy, Kent Hunter. A systems genetics approach reveals Rnaseh2c-immune response axis that affects metastasis severity in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2850. doi:10.1158/1538-7445.AM2017-2850

The relevance of inter- and intrastrain differences in mice and rats and their implications for models of seizures and epilepsy.

It is becoming increasingly clear that the genetic background of mice and rats, even in inbred strains, can have a profound influence on measures of seizure susceptibility and epilepsy. These differences can be capitalized upon through genetic mapping studies to reveal genes important for seizures and epilepsy. However, strain background and particularly mixed genetic backgrounds of transgenic animals need careful consideration in both the selection of strains and in the interpretation of results and conclusions. For instance, mice with targeted deletions of genes involved in epilepsy can have profoundly disparate phenotypes depending on the background strain. In this review, we discuss findings related to how this genetic heterogeneity has and can be utilized in the epilepsy field to reveal novel insights into seizures and epilepsy. Moreover, we discuss how caution is needed in regards to rodent strain or even animal vendor choice, and how this can significantly influence seizure and epilepsy parameters in unexpected ways. This is particularly critical in decisions regarding the strain of choice used in generating mice with targeted deletions of genes. Finally, we discuss the role of environment (at vendor and/or laboratory) and epigenetic factors for inter- and intrastrain differences and how such differences can affect the expression of seizures and the animals' performance in behavioral tests that often accompany acute and chronic seizure testing.

Animal Models of Alcoholic Liver Disease: Pathogenesis and Clinical Relevance.

Alcoholic liver disease (ALD), a leading cause of chronic liver injury worldwide, comprises a range of disorders including simple steatosis, steatohepatitis, cirrhosis, and hepatocellular carcinoma. Over the last five decades, many animal models for the study of ALD pathogenesis have been developed. Recently, a chronic-plus-binge ethanol feeding model was reported. This model induces significant steatosis, hepatic neutrophil infiltration, and liver injury. A clinically relevant model of high-fat diet feeding plus binge ethanol was also developed, which highlights the risk of excessive binge drinking in obese/overweight individuals. All of these models recapitulate some features of the different stages of ALD and have been widely used by many investigators to study the pathogenesis of ALD and to test for therapeutic drugs/components. However, these models are somewhat variable, depending on mouse genetic background, ethanol dose, and animal facility environment. This review focuses on these models and discusses these variations and some methods to improve the feeding protocol. The pathogenesis, clinical relevance, and translational studies of these models are also discussed.

Quantifying three-dimensional morphology and RNA from individual embryos.

Quantitative analysis of morphogenesis aids our understanding of developmental processes by providing a method to link changes in shape with cellular and molecular processes. Over the last decade, many methods have been developed for 3D imaging of embryos using microCT scanning to quantify the shape of embryos during development. These methods generally involve a powerful, cross-linking fixative such as paraformaldehyde to limit shrinkage during the CT scan. However, the extended time frames that these embryos are incubated in such fixatives prevent use of the tissues for molecular analysis after microCT scanning. This is a significant problem because it limits the ability to correlate variation in molecular data with morphology at the level of individual embryos. Here we outline a novel method that allows RNA, DNA, or protein isolation following CT scan while also allowing imaging of different tissue layers within the developing embryo. We show shape differences early in craniofacial development (E11.5) between common mouse genetic backgrounds, and demonstrate that we are able to generate RNA from these embryos after CT scanning that is suitable for downstream real time PCR (RT-PCR) and RNAseq analyses. Developmental Dynamics 246:431-436, 2017. © 2017 Wiley Periodicals, Inc.

Effects of G6pc2 deletion on body weight and cholesterol in mice.

Genome-wide association study (GWAS) data have linked the G6PC2 gene to variations in fasting blood glucose (FBG). G6PC2 encodes an islet-specific glucose-6-phosphatase catalytic subunit that forms a substrate cycle with the beta cell glucose sensor glucokinase. This cycle modulates the glucose sensitivity of insulin secretion and hence FBG. GWAS data have not linked G6PC2 to variations in body weight but we previously reported that female C57BL/6J G6pc2-knockout (KO) mice were lighter than wild-type littermates on both a chow and high-fat diet. The purpose of this study was to compare the effects of G6pc2 deletion on FBG and body weight in both chow-fed and high-fat-fed mice on two other genetic backgrounds. FBG was reduced in G6pc2 KO mice largely independent of gender, genetic background or diet. In contrast, the effect of G6pc2 deletion on body weight was markedly influenced by these variables. Deletion of G6pc2 conferred a marked protection against diet-induced obesity in male mixed genetic background mice, whereas in 129SvEv mice deletion of G6pc2 had no effect on body weight. G6pc2 deletion also reduced plasma cholesterol levels in a manner dependent on gender, genetic background and diet. An association between G6PC2 and plasma cholesterol was also observed in humans through electronic health record-derived phenotype analyses. These observations suggest that the action of G6PC2 on FBG is largely independent of the influences of environment, modifier genes or epigenetic events, whereas the action of G6PC2 on body weight and cholesterol are influenced by unknown variables.