A comprehensive and comparative phenotypic analysis of the collaborative founder strains identifies new and known phenotypes.

The collaborative cross (CC) is a large panel of mouse-inbred lines derived from eight founder strains (NOD/ShiLtJ, NZO/HILtJ, A/J, C57BL/6J, 129S1/SvImJ, CAST/EiJ, PWK/PhJ, and WSB/EiJ). Here, we performed a comprehensive and comparative phenotyping screening to identify phenotypic differences and similarities between the eight founder strains. In total, more than 300 parameters including allergy, behavior, cardiovascular, clinical blood chemistry, dysmorphology, bone and cartilage, energy metabolism, eye and vision, immunology, lung function, neurology, nociception, and pathology were analyzed; in most traits from sixteen females and sixteen males. We identified over 270 parameters that were significantly different between strains. This study highlights the value of the founder and CC strains for phenotype-genotype associations of many genetic traits that are highly relevant to human diseases. All data described here are publicly available from the mouse phenome database for analyses and downloads

A large scale hearing loss screen reveals an extensive unexplored genetic landscape for auditory dysfunction

The developmental and physiological complexity of the auditory system is likely reflected in the underlying set of genes involved in auditory function. In humans, over 150 non-syndromic loci have been identified, and there are more than 400 human genetic syndromes with a hearing loss component. Over 100 non-syndromic hearing loss genes have been identified in mouse and human, but we remain ignorant of the full extent of the genetic landscape involved in auditory dysfunction. As part of the International Mouse Phenotyping Consortium, we undertook a hearing loss screen in a cohort of 3006 mouse knockout strains. In total, we identify 67 candidate hearing loss genes. We detect known hearing loss genes, but the vast majority, 52, of the candidate genes were novel. Our analysis reveals a large and unexplored genetic landscape involved with auditory function

AOX delays the onset of the lethal phenotype in a mouse model of Uqcrh (complex III) disease

The alternative oxidase, AOX, provides a by-pass of the cytochrome segment of the mitochondrial respiratory chain when the chain is unavailable. AOX is absent from mammals, but AOX from Ciona intestinalis is benign when expressed in mice. Although non-protonmotive, so does not contribute directly to ATP production, it has been shown to modify and in some cases rescue phenotypes of respiratory-chain disease models. Here we studied the effect of C. intestinalis AOX on mice engineered to express a disease-equivalent mutant of Uqcrh, encoding the hinge subunit of mitochondrial respiratory complex III, which results in a complex metabolic phenotype beginning at 4–5 weeks, rapidly progressing to lethality within a further 6–7 weeks. AOX expression delayed the onset of this phenotype by several weeks, but provided no long-term benefit. We discuss the significance of this finding in light of the known and hypothesized effects of AOX on metabolism, redox homeostasis, oxidative stress and cell signaling. Although not a panacea, the ability of AOX to mitigate disease onset and progression means it could be useful in treatment.Peer reviewe

Laboratory mouse housing conditions can be improved using common environmental enrichment without compromising data.

Animal welfare requires the adequate housing of animals to ensure health and well-being. The application of environmental enrichment is a way to improve the well-being of laboratory animals. However, it is important to know whether these enrichment items can be incorporated in experimental mouse husbandry without creating a divide between past and future experimental results. Previous small-scale studies have been inconsistent throughout the literature, and it is not yet completely understood whether and how enrichment might endanger comparability of results of scientific experiments. Here, we measured the effect on means and variability of 164 physiological parameters in 3 conditions: with nesting material with or without a shelter, comparing these 2 conditions to a "barren" regime without any enrichments. We studied a total of 360 mice from each of 2 mouse strains (C57BL/6NTac and DBA/2NCrl) and both sexes for each of the 3 conditions. Our study indicates that enrichment affects the mean values of some of the 164 parameters with no consistent effects on variability. However, the influence of enrichment appears negligible compared to the effects of other influencing factors. Therefore, nesting material and shelters may be used to improve animal welfare without impairment of experimental outcome or loss of comparability to previous data collected under barren housing conditions

Extensive identification of genes involved in congenital and structural heart disorders and cardiomyopathy

Clinical presentation of congenital heart disease is heterogeneous, making identification of the disease-causing genes and their genetic pathways and mechanisms of action challenging. By using in vivo electrocardiography, transthoracic echocardiography and microcomputed tomography imaging to screen 3,894 single-gene-null mouse lines for structural and functional cardiac abnormalities, here we identify 705 lines with cardiac arrhythmia, myocardial hypertrophy and/or ventricular dilation. Among these 705 genes, 486 have not been previously associated with cardiac dysfunction in humans, and some of them represent variants of unknown relevance (VUR). Mice with mutations in Casz1, Dnajc18, Pde4dip, Rnf38 or Tmem161b genes show developmental cardiac structural abnormalities, with their human orthologs being categorized as VUR. Using UK Biobank data, we validate the importance of the DNAJC18 gene for cardiac homeostasis by showing that its loss of function is associated with altered left ventricular systolic function. Our results identify hundreds of previously unappreciated genes with potential function in congenital heart disease and suggest causal function of five VUR in congenital heart disease

Influence of main factors (enrichment, sex, cohort) on mean of parameters in 2 mouse strains.

<p>(A) The heatmap shows the results for all metric parameters (rows) and influencing factors and their double interactions (columns). The B6 and D2 data are on the left and right of each column, respectively. The influence of main factors was evaluated using linear models. A variable selection using the BIC was performed. Reference categories are enrichment none, sex female, and the first cohort. The differences between means of parameters are expressed as color-coded estimators β of the respective model in percent of the intercept of the model. The estimator β describes the change induced by the respective factors. The intercept is the mean of the parameter of the reference group and is set as 100%. Grey shade: variable selection using the BIC yielded nonrelevant influencing factor, thus no effect can be assumed; violet shade: factor is relevant for the model, i.e., mean is increased compared to reference group; yellow shade: factor is relevant for the model, i.e., mean is decreased compared to reference group; strength of color reflects size of difference. “X” marks parameters that were not analyzed. (B) Color grading scheme: the color grading illustrates the change in percent of the intercept; the strength of color reflects the size of difference. ABR, auditory brain stem response; ALAT/GPT, Alanine aminotransferase/Glutamat pyruvat transaminase; ALP, Alkaline phosphatase; ASAT/GOT, Aspartate aminotransferase/Glutamat oxalacetat transaminase; ASR, acoustic startle response; AU400, name of clinical chemistry analyzer; AUC, area under the curve; BIC, Bayesian Information Criterion; BN, background noise; CNPG3, substrate of test reaction (2-Chloro-4-Nitrophenyl-α-D-Maltotriosid); DEXA, dual-energy x-ray absorptiometry; HR, heart rate; HRV, heart rate variability; IgE, immunoglobulin E; IpGTT, intraperitoneal glucose tolerance test; LDH, Lactat-dehydrogenase; LVIDd, left ventricular end-diastolic internal diameter; LVIDs, left ventricular end-systolic internal diameter; LVPW, left ventricular posterior wall; P1-P4, startle amplitude following pulse of 67 (P1), 69 (P2), 73 (P3), 81 (P4) dB; PP (67, 69, 73, 81), startle amplitude and PPI following startle pulse (110 dB) with preceding pre-pulse of 67, 69, 73, 81 dB; PPI, pre-pulse inhibition; qNMR, quantitative nuclear magnetic resonance; QTc, corrected QT; R1, region analyzed (whole body excluding the skull); SHIRPA, Smithkline Beecham, MRC Harwell, Imperial College, the Royal London Hospital Phenotype Assessment; ST 110, acoustic startle response at 110 dB.</p

Workflow for the screens of this study.

<p>DEXA, dual-energy x-ray absorptiometry; IpGTT, intraperitoneal glucose tolerance test; NMR, nuclear magnetic resonance; PPI, pre-pulse inhibition; SHIRPA, Smithkline Beecham, MRC Harwell, Imperial College, the Royal London Hospital Phenotype Assessment.</p

Raw data of selected parameters on the background of the biological range for B6.

<p>Raw data of the 4 main parameters of the open field test: (A) “distance traveled total,” (B) “number of rears total,” (C) “percent center distance total,” and (D) “center permanence time” as box- and whisker-plots. The box represents 25th percentile, median, and 75th percentile; the length of whiskers is maximally the 1.5-fold interquartile range but is determined by the last value within this range. All individual values are shown for each experimental group (“con,” nest, double) and every cohort (1, 2, 3) for female and male mice in the upper and lower plot, respectively, for each selected parameter (A–D). The range in the background gives 1 SD (dark shading) and 2 SD (bright shading) of >200 reference B6 female (red) and male (blue) mice. The reference mice were same-aged wild-type control mice from other phenotyping projects of the GMC and were measured within the same timespan as the mice used in this project. “con”, control; GMC, German Mouse Clinic.</p

Animal numbers used for this study.

<p>Animal numbers used for this study.</p

Influence of main factors (enrichment, sex, cohort) on bootstrapped CVs in 2 strains.

<p>The heatmap shows the results for all metric parameters (rows) and influencing factors (columns) for B6 (left column) and D2 (right column). Reference categories are enrichment none, sex female, and the first cohort. Results of the bootstrap method are summarized by computed confidence intervals for estimators (β), and bootstrapped CVs are then classified into the following 3 categories: confidence interval for β includes 0, thus no effect is assumed (grey); confidence interval for β is greater than 0, i.e., bootstrapped CVs are increased compared to reference group (violet); confidence interval for β is below 0, i.e., bootstrapped CVs are decreased compared to reference group (yellow). “X” mark parameters that were not analyzed. ABR, auditory brain stem response; ALAT/GPT, Alanine aminotransferase/Glutamat pyruvat transaminase; ALP, Alkaline phosphatase; ASAT/GOT, Aspartate aminotransferase/Glutamat oxalacetat transaminase; ASR, acoustic startle response; AU400, name of clinical chemistry analyzer; AUC, area under the curve; BIC, Bayesian Information Criterion; BN, background noise; CNPG3, substrate of test reaction (2-Chloro-4-Nitrophenyl-α-D-Maltotriosid); CV, coefficient of variation; DEXA, dual-energy x-ray absorptiometry; HR, heart rate; HRV, heart rate variability; IgE, immunoglobulin E; IpGTT, intraperitoneal glucose tolerance test; LDH, Lactat-dehydrogenase; LVIDd, left ventricular end-diastolic internal diameter; LVIDs, left ventricular end-systolic internal diameter; LVPW, left ventricular posterior wall; P1-P4, startle amplitude following pulse of 67 (P1), 69 (P2), 73 (P3), 81 (P4) dB; PP (67, 69, 73, 81), startle amplitude and PPI following startle pulse (110 dB) with preceding pre-pulse of 67, 69, 73, 81 dB; PPI, pre-pulse inhibition; qNMR, quantitative nuclear magnetic resonance; QTc, corrected QT; R1, region analyzed (whole body excluding the skull); SHIRPA, Smithkline Beecham, MRC Harwell, Imperial College, the Royal London Hospital Phenotype Assessment; ST 110, acoustic startle response at 110 dB.</p