Outbred Mouse Model Research Articles

Advanced paternal age directly impacts mouse embryonic placental imprinting.

The placental epigenome plays a critical role in regulating mammalian growth and development. Alterations to placental methylation, often observed at imprinted genes, can lead to adverse pregnancy complications such as intrauterine growth restriction and preterm birth. Similar associations have been observed in offspring derived from advanced paternal age fathers. As parental age at time of conception continues to rise, the impact of advanced paternal age on these reproductive outcomes is a growing concern, but limited information is available on the molecular mechanisms affected in utero. This longitudinal murine research study thus investigated the impact of paternal aging on genomic imprinting in viable F1 embryonic portions of the placentas derived from the same paternal males when they were young (4–6 months) and when they aged (11–15 months). The use of a controlled outbred mouse model enabled analysis of offspring throughout the natural lifetime of the same paternal males and excluded confounding factors like female age or infertility. Firstly, paternal age significantly impacted embryonic placental weight, fetal weight and length. Targeted bisulfite sequencing was utilized to examine imprinted methylation at the Kcnq1ot1 imprinting control region, with significant hypermethylation observed upon natural paternal aging. Quantitative real-time PCR assessed imprinted gene expression levels at various imprinting clusters, resulting in transcript level alterations attributable to advanced paternal age. In summary, our results demonstrate a paternal age effect with dysregulation at numerous imprinted loci, providing a mechanism for future adverse placental and offspring health conditions.

The Galleria mellonella Infection Model Does Not Accurately Differentiate between Hypervirulent and Classical Klebsiella pneumoniae.

Hypervirulent Klebsiella pneumoniae (hvKp) is an emerging pathogen of increasing concern due to its ability to cause serious organ and life-threatening infections in healthy individuals and its increasing acquisition of antimicrobial resistance determinants. Identification of hvKp is critical for patient care and epidemiologic and research studies. Five genotypic markers on the hvKp-specific virulence plasmid can accurately differentiate hvKp from the less virulent classical K. pneumoniae (cKp) strain, but it is unclear whether the possession of fewer markers accurately predicts the hvKp pathotype. Likewise, the effect, if any, of various antimicrobial resistance factors on the pathogenic potential of hvKp has been incompletely explored. The Galleria mellonella infection model is often used to assess virulence, but this tool has not been validated. Therefore, levels of lethality of defined hvKp and cKp strain cohorts were compared in Galleria and outbred mouse models. The murine model, but not the G. mellonella model, accurately differentiated hvKp from cKp strains. Therefore, isolates in which the pathogenic potential is ambiguous due to an incomplete hvKp biomarker profile, an incomplete pLVPK-like hvKp-specific virulence plasmid, antimicrobial resistance that could decrease biofitness, and/or the lack of a characteristic clinical presentation should be validated in an outbred murine model. These data will assist in determining the minimal genomic content needed for full expression of the hypervirulence phenotype. This information, in turn, is critical for the development of the pragmatic point-of-care testing requisite for patient care and for the performance of epidemiologic and research studies going forward.IMPORTANCE Hypervirulent Klebsiella pneumoniae (hvKp) is of increasing concern because it can infect individuals in community and health care settings and because such infections are becoming difficult to treat. Identification of hvKp is important for patient care and to track its global spread. The genetic definition of hvKp, which can be used for its identification and the development of diagnostic tests, has not been optimized. Determination of possession of 4 of 5 genes that are present on the hvKp-specific virulence plasmid is highly accurate for identifying hvKp. However, an ongoing issue is whether strains that possess only some of these markers are still hypervirulent. The Galleria mellonella model and, less commonly, the murine infection model have been used to assess the virulence of these ambiguously identifiable strains. This report demonstrates that the murine model but not the G. mellonella model accurately identifies suspected hvKp strains. This information is critical for the development of diagnostics for patient care and for future research studies.

Establishing a time course for spontaneous opioid withdrawal in male and female outbred mice using an abbreviated dependence paradigm

Opioid withdrawal is often a limiting factor in recovery from opioid dependence due to negative reinforcement caused by continued use of opioids in order to avoid the distressing withdrawal symptoms. In animal models of dependence, opioid withdrawal is frequently precipitated using opioid antagonists and the severity of withdrawal is related to the level of dependence. However, this model lacks face validity when investigating new treatment possibilities for opioid withdrawal because most human addicts are not exposed to an antagonist in order to precipitate withdrawal. Furthermore, results from animal experiments suggest that spontaneous and precipitated withdrawal involve different mechanisms and may favor different symptoms of withdrawal. These experiments were conducted to investigate the timeline of spontaneous withdrawal in an outbred mouse model using an abbreviated dependence paradigm. Male and female NIH Swiss mice were injected with escalating doses of morphine twice daily (50–125 mg/kg morphine) over a period of four days. On the fifth day, mice were given a final dose of morphine and withdrawal symptoms were assessed at 5, 10, 24, 34, 48, 58, 72, 82, 96, and 106 hrs after the final injection. Withdrawal symptoms were assessed by global scores for an overall picture and by counting the number of tremors, wet dog shakes, and jumps. Results showed a robust withdrawal time course that peaked at 24 hrs in both sexes as assessed by global score. When broken down, wet dog shakes and tremors proved to be the most reliable indicator of severity of withdrawal and females showed more tremors than males for the 1st 24 hours of the time course. These results establish a spontaneous withdrawal time course in an outbred strain of mouse that shows that dependence can be reliably established over a four‐day period using fairly high escalating doses of morphine. Furthermore, these results suggest that individual symptoms of withdrawal should be assessed over time as well as global score.Support or Funding InformationSupported in part by the State of Washington Initiative Measure No.171.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Evaluation of an outbred mouse model for Francisella tularensis vaccine development and testing.

Francisella tularensis (Ft) is a biothreat agent for which there is no FDA-approved human vaccine. Currently, there are substantial efforts underway to develop both vaccines and the tools to assess these vaccines. Tularemia laboratory research has historically relied primarily upon a small number of inbred mouse strains, but the utility of such findings to outbred animals may be limited. Specifically, C57BL/6 mice are more susceptible than BALB/c mice to Ft infection and less easily protected against challenge with highly virulent type A Ft. Thus, depending on the inbred mouse strain used, one could be misled as to which immunogen(s)/vaccine will ultimately be effective in an outbred human population. Accordingly, we evaluated an outbred Swiss Webster (SW) mouse model in direct comparison to a well-established, inbred C57BL/6 mouse model. Mucosal vaccination with the live, attenuated Ft LVS superoxide dismutase (sodB) mutant demonstrated significantly higher protection in outbred SW mice compared to inbred C57BL/6 mice against Ft SchuS4 respiratory challenge. The protection observed in vaccinated outbred mice correlated with lower bacterial density, reduced tissue inflammation, and reduced levels of pro-inflammatory cytokine production. This protection was CD4+ and CD8+ T cell-dependent and characterized by lower titers of serum antibody (Ab) that qualitatively differed from vaccinated inbred mice. Enhanced protection of vaccinated outbred mice correlated with early and robust production of IFN-γ and IL-17A. Neutralizing Ab administered at the time of challenge revealed that IFN-γ was central to this protection, while IL-17A neutralization did not alter bacterial burden or survival. The present study demonstrates the utility of the outbred mouse as an alternative vaccination model for testing tularemia vaccines. Given the limited MHC repertoire in inbred mice, this outbred model is more analogous to the human in terms of immunological diversity.

Timing of gestational exposure to Zika virus is associated with postnatal growth restriction in a murine model

Vertical transmission of Zika virus leads to infection of neuroprogenitor cells and destruction of brain parenchyma. Recent evidence suggests that the timing of infection as well as host factors may affect vertical transmission. As a result, congenital Zika virus infection may only become clinically apparent in the postnatal period. We sought to develop an outbred mouse model of Zika virus vertical transmission to determine if the timing of gestational Zika virus exposure yields phenotypic differences at birth and through adolescence. We hypothesized that later gestational inoculations would only become apparent in adolescence. To better recapitulate human exposures, timed pregnant Swiss-Webster dams (n= 15) were subcutaneously inoculated with 1× 104 plaque-forming units of first passage contemporary Zika virus HN16 strain or a mock injection on embryonic day 4, 8, or 12 with bioactive antiinterferon alpha receptor antibody administered in days preceding and proceeding inoculation. The antibody was given to prevent the robust type I interferon signaling cascade that make mice inherently resistant to Zika virus infection. At birth and adolescence (6 weeks of age) offspring were assessed for growth, brain weight, and biparietal head diameters, and Zika virus viral levels by reverse transcription-polymerase chain reaction or in situ hybridization. Pups of Zika virus-infected dams infected at embryonic days 4 and 8 but not 12 were growth restricted (P < .003). Brain weights were significantly smaller at birth (P= .01) for embryonic day 8 Zika virus-exposed offspring. At 6 weeks of age, biparietal diameters were smaller for all Zika virus-exposed males and females (P < .05), with embryonic day 8-exposed males smallest by biparietal diameter and growth-restriction measurements (weight >2 SD, P= .0007). All pups and adolescent mice were assessed for Zika virus infection by reverse transcription-polymerase chain reaction. Analysis of all underweight pups reveled 1 to be positive for neuronal Zika virus infection by in situ hybridization, while a second moribund animal was diffusely positive at 8 days of age by Zika virus infectivity throughout the brain, kidneys, and intestine. These findings demonstrate that postnatal effects of infection occurring at single time points continue to be detrimental to offspring in the postnatal period in a subset of littermates and subject to a window of gestational susceptibility coinciding with placentation. This model recapitulates frequently encountered clinical scenarios in nonendemic regions, including the majority of the United States, where travel-related exposure occurs in short and well-defined windows of gestation. Our low rate of infection and relatively rare evidence of congenital Zika syndrome parallels human population-based data.

Mast cell accumulation drives allergy-driven chronic pain responses in outbred mice

Abstract We are building novel outbred mouse models of a poorly understood yet prevalent vulvar pain condition – localized, provoked vulvodynia – that affects a substantial proportion of 18–60 year old women. Epidemiologic studies have shown a surprising and increased risk for developing this condition in women with a history of skin and airway allergies. We show that repeated exposures to low doses of the common contact irritant haptens oxazolone or dinitrofluorobenzene on the genital skin of mice produce sustained painful sensitivity to touch as well as a non-normative accumulation of mast cells and an overgrowth of cutaneous nerves in the affected tissue – changes that are also seen in vulvar biopsies taken from women diagnosed with vulvodynia providing the first evidence of biological plausibility for the immunological underpinnings of this chronic pain condition. In addition to laboratory haptens, we show that the hapten methylisothiazolinone (a common cosmetic preservative found in soaps, shampoos, and feminine hygiene products) can induce these responses suggesting that this ubiquitous environmental exposure might be a possible immunological trigger of hitherto unexplained genital pain conditions that affect many women. We have further characterized infiltrating T cells at the allergic site to show that FoxP3+ regulatory CD4 T cells and CD103+ resident memory CD8 T accumulate at these sites as well. Our mouse models provide new tools to both dissect allergy driven pain conditions as well as test novel therapies targeting immune cells and mechanisms for the management and treatment of these debilitating disorders.

Muscle‐intensive and High‐Impact Exercises Differentially Influence Whole Bone 3D Morphology in Young Outbred Male Mice

Skeletal morphology is predominantly dictated by the genetic background and the habitual loading environment of individual bones. Mechanical loads placed on bones are derived from intracorporeal contractions of muscles and from extracorporeal ground reaction forces generated by bearing weight and by substrate reaction forces during locomotion. We investigated the relative effects of these two sources of load on the growing skeleton using an outbred mouse model system. The 3D shape of the femur was compared across groups of mice following discrete exercise regimens designed to emphasize specific loading environments. Hsd:ICR (CD‐1) mice (N=80; 20 per group) were randomly assigned to one of four treatment groups at four weeks old: swimming which emphasizes muscle forces, dropping which emphasizes impact forces, wheel running which results in a combination of muscle and impact forces, and non‐exercised controls which were free to move about their cages. Durations of swimming, running, and impact loading were matched so that mice received approximately the same intensity. After 20 days mice were euthanized and right hind limbs were micro‐CT scanned (~20 μm3 voxel size). Femoral shape was defined by digitizing 23 landmarks on each scan. To adjust for size, each individual femur was scaled to the geometric mean of all possible linear distances among landmarks. Group differences were analyzed using Euclidean Distance Matrix Analysis (EDMA), and differences &gt;5% were considered substantial. In all three activity groups, we find decreased robusticity of the femoral diaphysis and increased distal condylar breadth compared to non‐exercised controls. Additionally, each group displays unique femoral morphologies when compared to controls. The swimming treatment group shows increased length of the femoral neck. The wheel running treatment group shows a narrower femoral neck with a larger femoral head. The impact loading group shows a lengthened and reoriented distal diaphysis. Overall, the running and swimming groups show similar patterns of morphology that diverge from the non‐exercised mice, suggesting muscle forces differentially affect the loading environment during running in mice compared to impact forces. Further, these results indicate the regional sensitivity of femoral morphology to muscle‐ and impact‐derived loads, and emphasize how activity patterns during adolescence can lead to distinct morphologies in the limb skeleton.Support or Funding InformationResearch support: University of Missouri School of MedicineThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Reproductive performance primarily depends on the female genotype in a two-factorial breeding experiment using high-fertility mouse lines.

Mouse models showing an improved fertility phenotype are barely described in the literature. In the present study, we further characterized two outbred mouse models that have been selected for the phenotype 'high fertility' for more than 177 generations (fertility lines (FL) 1 and 2). In order to delineate the impact of males and females on fertility parameters, we performed a two-factorial breeding experiment by mating males and females of the three different genotypes (FL1, FL2, unselected control (Ctrl)) in all 9 possible combinations. Reproductive performance, such as number of offspring per litter or total birth weight of the entire pup, mainly depends on the female genotype. Although the reproductive performance of FL1 and FL2 is very similar, their phenotypes differ. FL2 animals of both genders are larger compared to FL1 and control animals. Females of the control line delivered offspring earlier compared to FL1 and FL2 dams. Males of FL1 are the lightest and the only ones who gained weight during the two weeks mating period. To address whether this effect is correlated with differing serum androgen levels, we measured the concentrations of testosterone, dehydroepiandrosterone, 4-androstenedione, androstanediol and dihydrotestosterone in males of all three lines by GC-MS. We measured serum testosterone between 5.0 and 6.4 ng/mL, whereas the concentrations of the other androgens were at least one order of magnitude lower, with no significant differences between the lines. Our data indicate that reproductive outcome largely depends on the genotype of the female in a two-factorial breeding experiment and supports previous findings that the phenotype 'high fertility' is warranted by using different physiological strategies.

Defining the consequences of genetic variation on a proteome-wide scale.

Genetic variation modulates protein expression through both transcriptional and post-transcriptional mechanisms. To characterize the consequences of natural genetic diversity on the proteome, here we combine a multiplexed, mass spectrometry-based method for protein quantification with an emerging outbred mouse model containing extensive genetic variation from eight inbred founder strains. By measuring genome-wide transcript and protein expression in livers from 192 Diversity outbred mice, we identify 2,866 protein quantitative trait loci (pQTL) with twice as many local as distant genetic variants. These data support distinct transcriptional and post-transcriptional models underlying the observed pQTL effects. Using a sensitive approach to mediation analysis, we often identified a second protein or transcript as the causal mediator of distant pQTL. Our analysis reveals an extensive network of direct protein-protein interactions. Finally, we show that local genotype can provide accurate predictions of protein abundance in an independent cohort of collaborative cross mice.

Intravenous transplantation of mouse embryonic stem cells attenuates demyelination in an ICR outbred mouse model of demyelinating diseases.

Induction of demyelination in the central nervous system (CNS) of experimental mice using cuprizone is widely used as an animal model for studying the pathogenesis and treatment of demyelination. However, different mouse strains used result in different pathological outcomes. Moreover, because current medicinal treatments are not always effective in multiple sclerosis patients, so the study of exogenous cell transplantation in an animal model is of great importance. The aims of the present study were to establish an alternative ICR outbred mouse model for studying demyelination and to evaluate the effects of intravenous cell transplantation in the present developed mouse model. Two sets of experiments were conducted. Firstly, ICR outbred and BALB/c inbred mice were fed with 0.2% cuprizone for 6 consecutive weeks; then demyelinating scores determined by luxol fast blue stain or immunolabeling with CNPase were evaluated. Secondly, attenuation of demyelination in ICR mice by intravenous injection of mES cells was studied. Scores for demyelination in the brains of ICR mice receiving cell injection (mES cells-injected group) and vehicle (sham-inoculated group) were assessed and compared. The results showed that cuprizone significantly induced demyelination in the cerebral cortex and corpus callosum of both ICR and BALB/c mice. Additionally, intravenous transplantation of mES cells potentially attenuated demyelination in ICR mice compared with sham-inoculated groups. The present study is among the earliest reports to describe the cuprizone-induced demyelination in ICR outbred mice. Although it remains unclear whether mES cells or trophic effects from mES cells are the cause of enhanced remyelination, the results of the present study may shed some light on exogenous cell therapy in central nervous system demyelinating diseases.

Sexual diergism in antibody response to whole virus trivalent inactivated influenza vaccine in outbred mice

An outbred mouse model was used to determine if antibody response to immunization with whole-virus trivalent inactivated influenza vaccine (TIV) differs between the sexes. The antibody response was examined one (serum titer of IgM antibodies), and three and six weeks post-immunization (serum titer of neutralizing and total IgG antibodies and IgG subclass profile). Compared with male in female mice was found (i) the more robust IgM response against all influenza strains included in TIV and (ii) more vigorous neutralizing antibody and total IgG responses against H1N1 influenza virus at both the examined time points post-immunization. The total IgG antibody response against H3N2 and B influenza viruses was comparable between female and male mice three weeks post-immunization, but significantly greater in female mice six weeks post-immunization. The neutralizing antibody response against H3N2 and B influenza viruses did not significantly differ between sexes at both the examined points post-immunization. Finally, three weeks post-immunization subclass profile of IgG specific to the influenza strains included in TIV differed between female and male mice, reflecting the lower titer of IgG1 antibodies in female ones, so that IgG2a (contributing mainly to the total IgG) to IgG1 ratio in mice of this sex was shifted toward the former. In agreement with this shift, compared with male mice, Th1/Th2 balance in female mice was shifted toward Th1, as shown by ELISPOT. Collectively, the results showed influenza virus strain-dependent sexual dimorphism in the magnitude, dynamics and characteristics of antibody response in outbred mice immunized with TIV.

Comment on "Diversity Outbred: A New Generation of Mouse Model".

A news article by Charles Schmidt propagates the Diversity Outbred mouse model to facilitate the extrapolation of toxicology findings to humans. In this regard, we would like to highlight two sources of animal diversity that could be relevant: 1) the differential—and genetically codetermined—circadian propensity for activity and rest with which the mice are born, and 2) the differential stability of circadian rhythms in later life stages of mice, to which perinatal photoperiods may contribute. Regarding 1), humans come in different chronotypes corresponding to variations in how physiology, endocrinology, metabolism, and behavior are organized and timed over the individual’s biological day and night. Inasmuch as the extent of genetic variability among model mice should be similar to genetic variations in humans, how do new-generation mouse models capture differential chronobiological propensity? Laboratory mice, like humans, come in various chronotypes, and different strains come in different ones (Wicht et al. 2014); therefore, different time windows of biological nights and days should significantly impact when and what we observe or measure, be it parameters of toxicology, behavior, physiology, or anatomy. With evidence that responses to DNA damage are regulated by the circadian clock in mice (Kang et al. 2010), chronotype-dependent lows and highs of DNA repair within a 24-hour period must be considered when extrapolating mouse-based toxicological data to humans. Regarding 2), although inbred mice may be genetically identical, perinatal photoperiods may nevertheless—by imprinting the circadian clock—lead to a differential stability of circadian rhythms. Perinatal exposures to summer versus winter light conditions (i.e., with a light:dark ratio of 16:8 versus 8:16) can determine the susceptibility of a mouse’s circadian rhythm to dysfunction or disruption throughout the animal’s life (Ciarleglio et al. 2011). Furthermore, the integrity of circadian clocks and rest–activity circadian rhythms plays a major role for tumor suppression by controlling cell proliferation and other cellular functions (Fu and Lee 2003). Taken together, if the perinatal photoperiod may codetermine the very robustness of mice to fight off severe circadian dysfunction (Filipski et al. 2002) and the development of tumors, then a fortiori we would have to understand, and possibly control, circadian diversity. Overall, the biological activity of circadian clocks must be taken into account when experimenting with mice. Taking note of 1) and 2), we should consider possible chronotypes in new-generation mice and standards for the light:dark conditions under which laboratory mice are bred and raised. Moreover, the efficacy, toxicity, and carcinogenicity of chemicals or drugs should be tested at different times within a 24-hour period. Given evidence for links between aging clocks and progressive declines of the circadian control of crucial biological processes (Belancio et al. 2014), age should also be factored in when using mice for testing.

High-fat diet-induced adiposity, adipose inflammation, hepatic steatosis and hyperinsulinemia in outbred CD-1 mice.

High-fat diet (HFD) has been applied to a variety of inbred mouse strains to induce obesity and obesity related metabolic complications. In this study, we determined HFD induced development of metabolic disorders on outbred female CD-1 mice in a time dependent manner. Compared to mice on regular chow, HFD-fed CD-1 mice gradually gained more fat mass and consequently exhibited accelerated body weight gain, which was associated with adipocyte hypertrophy and up-regulated expression of adipose inflammatory chemokines and cytokines such as Mcp-1 and Tnf-α. Increased fat accumulation in white adipose tissue subsequently led to ectopic fat deposition in brown adipose tissue, giving rise to whitening of brown adipose tissue without altering plasma level of triglyceride. Ectopic fat deposition was also observed in the liver, which was associated with elevated expression of key genes involved in hepatic lipid sequestration, including Ppar-γ2, Cd36 and Mgat1. Notably, adipose chronic inflammation and ectopic lipid deposition in the liver and brown fat were accompanied by glucose intolerance and insulin resistance, which was correlated with hyperinsulinemia and pancreatic islet hypertrophy. Collectively, these results demonstrate sequentially the events that HFD induces physiological changes leading to metabolic disorders in an outbred mouse model more closely resembling heterogeneity of the human population.

Generation of outbred Ace2 knockout mice by RNA transfection of TALENs displaying colitis reminiscent pathophysiology and inflammation.

The angiotensin I converting enzyme 2 (ACE2) is a key factor in the maintenance of intestinal homeostasis. Dysregulation of homeostasis can lead to inflammation of the colon (colitis), which can cause life-threatening enfeeblement or even cancer. Animal models are valuable surrogates in deciphering the pathology behind such human conditions and for screening of putative therapeutic targets or treatment paradigms. However, development of disease models can be time-consuming and technical demanding, which might hamper their application-value. In this study, we genetically disrupted the mouse Ace2 gene by direct injection of in vitro transcribed mRNA coding for transcription activator-like effector nucleases (TALENs) into the cytoplasm of outbred Kunming mouse zygotes. Consequently, somatic mutations were induced with an efficiency of 57 %, of which 39 % were frameshift mutations. Moreover, all modifications were stably transferred during germline transmission. In Ace2-knockout male mice (Ace2 −/y), we observed severe chemical induced colitis, characterized by considerable weight loss, diarrhea and a shortened colon length. Histologically, Ace2 mutations resulted in the infiltration of leukocytes and the overt damage of the intestinal mucosal barrier. In addition, we detected an increased expression of inflammatory cytokines in the colon tissue of Ace2 −/y mice. Collectively, the data indicate that high targeting efficiency and heritability can be achieved in an outbred mouse model by zygote injection of TALEN mRNA. Furthermore, the generated Ace2 −/y mice display phenotypic traits reminiscent of colitis and we anticipate that such mice can be of value in studies of the intestinal microbiome or fecal transplantation.Electronic supplementary materialThe online version of this article (doi:10.1007/s11248-014-9855-3) contains supplementary material, which is available to authorized users.

Different roads to fertility: distinct molecular and physiological strategies warrant the phenotype 'high fertility' in two outbred mouse models

Different roads to fertility: distinct molecular and physiological strategies warrant the phenotype 'high fertility' in two outbred mouse models

High Fertility: Two outbred mouse models exhibit different molecular and physiological strategies warranting improved fertility

Animal models are valuable tools in fertility research. Worldwide > 400 transgenic or knockout mouse models are available showing a reproductive phenotype; almost all of them exhibit an infertile or at least subfertile phenotype. In contrast, animal models revealing an improved fertility phenotype are barely described. Here we show data on two outbred mouse models showing a 'high fertility' phenotype. These mouse lines have been generated via selection over a time period of > 40 years and 161 generations. During this selection period the number of offspring per litter and the total birth weight of the entire litter nearly doubled. Concomitantly with the increased fertility phenotype several endocrine parameters (e.g. serum testosterone concentrations in male animals), physiological parameters (e.g. body weight, accelerated puberty, life expectancy) and behavioural parameters (e.g. behaviour in an open field, endurance fitness on a treadmill) have been altered. We elucidate that the two independently bred high fertility mouse lines warranted their improved fertility phenotype using different molecular and physiological strategies. The fertility lines display male- as well as female-specific characteristics. These genetically heterogeneous mouse models provide new insights into molecular and cellular mechanisms that enhance fertility. In view of decreasing fertility in men it will therefore be a precious information source for human reproductive medicine.

A Novel Outbred Mouse Model of 2009 Pandemic Influenza and Bacterial Co-Infection Severity

Influenza viruses pose a significant health risk and annually impose a great cost to patients and the health care system. The molecular determinants of influenza severity, often exacerbated by secondary bacterial infection, are largely unclear. We generated a novel outbred mouse model of influenza virus, Staphylococcus aureus, and co-infection utilizing influenza A/CA/07/2009 virus and S. aureus (USA300). Outbred mice displayed a wide range of pathologic phenotypes following influenza virus or co-infection ranging broadly in severity. Influenza viral burden positively correlated with weight loss although lung histopathology did not. Inflammatory cytokines including IL-6, TNF-α, G-CSF, and CXCL10 positively correlated with both weight loss and viral burden. In S. aureus infection, IL-1β, G-CSF, TNF-α, and IL-6 positively correlated with weight loss and bacterial burden. In co-infection, IL-1β production correlated with decreased weight loss suggesting a protective role. The data demonstrate an approach to identify biomarkers of severe disease and to understand pathogenic mechanisms in pneumonia.

High-fertility phenotypes: two outbred mouse models exhibit substantially different molecular and physiological strategies warranting improved fertility

Animal models are valuable tools in fertility research. Worldwide, there are more than 400 transgenic or knockout mouse models available showing a reproductive phenotype; almost all of them exhibit an infertile or at least subfertile phenotype. By contrast, animal models revealing an improved fertility phenotype are barely described. This article summarizes data on two outbred mouse models exhibiting a 'high-fertility' phenotype. These mouse lines were generated via selection over a time period of more than 40 years and 161 generations. During this selection period, the number of offspring per litter and the total birth weight of the entire litter nearly doubled. Concomitantly with the increased fertility phenotype, several endocrine parameters (e.g. serum testosterone concentrations in male animals), physiological parameters (e.g. body weight, accelerated puberty, and life expectancy), and behavioral parameters (e.g. behavior in an open field and endurance fitness on a treadmill) were altered. We demonstrate that the two independently bred high-fertility mouse lines warranted their improved fertility phenotype using different molecular and physiological strategies. The fertility lines display female- as well as male-specific characteristics. These genetically heterogeneous mouse models provide new insights into molecular and cellular mechanisms that enhance fertility. In view of decreasing fertility in men, these models will therefore be a precious information source for human reproductive medicine. Translated abstract A German translation of abstract is freely available at http://www.reproduction-online.org/content/147/4/427/suppl/DC1.

Reduced Th17 type inflammation associated with enhanced Th1, Th2 and Treg responses in a model of reactivation of congenital ocular toxoplasmosis

AbstractPurpose Ocular toxoplasmosis (OT) is a major cause of blindness in the world. Ocular involvement is frequently seen following congenital infection. Many of these infections are quiescent but pose a life‐time risk of reactivation. We previously developed a Swiss‐Webster outbred mouse model for congenital toxoplasmosis by neonatal injection of Toxoplasma gondii cysts. We also used a mouse model of direct intraocular infection to show a deleterious local Th17 type response upon primary infection. However, little is known about the physiopathology of reactivation.Methods In the present study, we combined our two models to study reinfection into neonatally infected mice, in comparison with a primary ocular infection. Intraocular immunological determinants were studied using both BioPlex proteomic assays in aqueous humor and RT‐PCR for crucial transcription factors.Results We observed diminished Th17 type reaction in reinfection, compared to primary infection. In contrast, Th2 and T regulatory responses were enhanced. Interestingly, this was also true for Th1 responses, which was paralleled by a better parasite control. We observed a similar protective immune reaction pattern in the eye upon reinfection with the virulent RH strain, with the notable exception of IFN‐γ.Conclusion In summary, our results show a less pathogenic but more effective anti‐parasite pattern during reinfection.

The local immune response to intraocular Toxoplasma re-challenge: Less pathology and better parasite control through Treg/Th1/Th2 induction

Ocular toxoplasmosis is a major cause of blindness world-wide. Ocular involvement is frequently seen following congenital infection. Many of these infections are quiescent but pose a life-time risk of reactivation. However, the physiopathology of ocular toxoplasmosis reactivation is largely unexplored. We previously developed a Swiss-Webster outbred mouse model for congenital toxoplasmosis by neonatal injection of Toxoplasma gondii cysts. We also used a mouse model of direct intraocular infection to show a deleterious local T helper 17 type response upon primary infection. In the present study, our two models were combined to study intravitreal re-challenge of neonatally infected mice, as an approximate model of reactivation, in comparison with a primary ocular infection. Using BioPlex proteomic assays in aqueous humour and reverse transcription-PCR for T helper cell transcription factors, we observed diminished T helper 17 type reaction in reinfection, compared with primary infection. In contrast, T helper 2 and T regulatory responses were enhanced. Interestingly, this was also true for T helper 1 markers such as IFN-γ, which was paralleled by better parasite control. Secretion of IL-27, a central cytokine for shifting the immune response from T helper 17 to T helper 1, was also greatly enhanced. We observed a similar protective immune reaction pattern in the eye upon reinfection with the virulent RH strain, with the notable exception of IFN-γ. In summary, our results show that the balance is shifted from T helper 17 to a less pathogenic but more effective anti-parasite Treg/T helper 1/T helper 2 pattern in a reactivation setting.