A humanized version of Foxp2 affects cortico-basal ganglia circuits in mice.

It has been proposed that two amino acid substitutions in the transcription factor FOXP2 have been positively selected during human evolution due to effects on aspects of speech and language. Here, we introduce these substitutions into the endogenous Foxp2 gene of mice. Although these mice are generally healthy, they have qualitatively different ultrasonic vocalizations, decreased exploratory behavior and decreased dopamine concentrations in the brain suggesting that the humanized Foxp2 allele affects basal ganglia. In the striatum, a part of the basal ganglia affected in humans with a speech deficit due to a nonfunctional FOXP2 allele, we find that medium spiny neurons have increased dendrite lengths and increased synaptic plasticity. Since mice carrying one nonfunctional Foxp2 allele show opposite effects, this suggests that alterations in cortico-basal ganglia circuits might have been important for the evolution of speech and language in humans.

Epigenetic alterations in longevity regulators, reduced life span, and exacerbated aging-related pathology in old father offspring mice.

Advanced age is not only a major risk factor for a range of disorders within an aging individual but may also enhance susceptibility for disease in the next generation. In humans, advanced paternal age has been associated with increased risk for a number of diseases. Experiments in rodent models have provided initial evidence that paternal age can influence behavioral traits in offspring animals, but the overall scope and extent of paternal age effects on health and disease across the life span remain underexplored. Here, we report that old father offspring mice showed a reduced life span and an exacerbated development of aging traits compared with young father offspring mice. Genome-wide epigenetic analyses of sperm from aging males and old father offspring tissue identified differentially methylated promoters, enriched for genes involved in the regulation of evolutionarily conserved longevity pathways. Gene expression analyses, biochemical experiments, and functional studies revealed evidence for an overactive mTORC1 signaling pathway in old father offspring mice. Pharmacological mTOR inhibition during the course of normal aging ameliorated many of the aging traits that were exacerbated in old father offspring mice. These findings raise the possibility that inherited alterations in longevity pathways contribute to intergenerational effects of aging in old father offspring mice.

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.

Low catalytic activity is insufficient to induce disease pathology in triosephosphate isomerase deficiency.

Triosephosphate isomerase (TPI) deficiency is a fatal genetic disorder characterized by hemolytic anemia and neurological dysfunction. Although the enzyme defect in TPI was discovered in the 1960s, the exact etiology of the disease is still debated. Some aspects indicate the disease could be caused by insufficient enzyme activity, whereas other observations indicate it could be a protein misfolding disease with tissue-specific differences in TPI activity. We generated a mouse model in which exchange of a conserved catalytic amino acid residue (isoleucine to valine, Ile170Val) reduces TPI specific activity without affecting the stability of the protein dimer. TPIIle170Val/Ile170Val mice exhibit an approximately 85% reduction in TPI activity consistently across all examined tissues, which is a stronger average, but more consistent, activity decline than observed in patients or symptomatic mouse models that carry structural defect mutant alleles. While monitoring protein expression levels revealed no evidence for protein instability, metabolite quantification indicated that glycolysis is affected by the active site mutation. TPIIle170Val/Ile170Val mice develop normally and show none of the disease symptoms associated with TPI deficiency. Therefore, without the stability defect that affects TPI activity in a tissue-specific manner, a strong decline in TPI catalytic activity is not sufficient to explain the pathological onset of TPI deficiency.

Chronic schistosomiasis during pregnancy epigenetically reprograms T-cell differentiation in offspring of infected mothers.

Schistosomiasis is a nontransplacental helminth infection. Chronic infection during pregnancy suppresses allergic airway responses in offspring. We addressed the question whether in utero exposure to chronic schistosome infection (Reg phase) in mice affects B-cell and T-cell development. Therefore, we focused our analyses on T-cell differentiation capacity induced by epigenetic changes in promoter regions of signature cytokines in offspring. Here, we show that naïve T cells from offspring of schistosome infected female mice had a strong capacity to differentiate into TH 1 cells, whereas TH 2 differentiation was impaired. In accordance, reduced levels of histone acetylation of the IL-4 promoter regions were observed in naïve T cells. To conclude, our mouse model revealed distinct epigenetic changes within the naïve T-cell compartment affecting TH 2 and TH 1 cell differentiation in offspring of mothers with chronic helminth infection. These findings could eventually help understand how helminths alter T-cell driven immune responses induced by allergens, bacterial or viral infections, as well as vaccines.

Functional compensation among HMGN variants modulates the DNase I hypersensitive sites at enhancers.

DNase I hypersensitive sites (DHSs) are a hallmark of chromatin regions containing regulatory DNA such as enhancers and promoters; however, the factors affecting the establishment and maintenance of these sites are not fully understood. We now show that HMGN1 and HMGN2, nucleosome-binding proteins that are ubiquitously expressed in vertebrate cells, maintain the DHS landscape of mouse embryonic fibroblasts (MEFs) synergistically. Loss of one of these HMGN variants led to a compensatory increase of binding of the remaining variant. Genome-wide mapping of the DHSs in Hmgn1(-/-), Hmgn2(-/-), and Hmgn1(-/-)n2(-/-) MEFs reveals that loss of both, but not a single HMGN variant, leads to significant remodeling of the DHS landscape, especially at enhancer regions marked by H3K4me1 and H3K27ac. Loss of HMGN variants affects the induced expression of stress-responsive genes in MEFs, the transcription profiles of several mouse tissues, and leads to altered phenotypes that are not seen in mice lacking only one variant. We conclude that the compensatory binding of HMGN variants to chromatin maintains the DHS landscape, and the transcription fidelity and is necessary to retain wild-type phenotypes. Our study provides insight into mechanisms that maintain regulatory sites in chromatin and into functional compensation among nucleosome binding architectural proteins.

Lymphotoxin ß receptor signalling executes Helicobacter pylori-driven gastric inflammation in a T4SS-dependent manner.

OBJECTIVE: Lymphotoxin ß receptor (LTßR) signalling has been implicated in inflammation-associated tumour development in different tissues. We have analysed the role of LTßR and alternative NF-κB signalling in Helicobacter pylori-mediated gastric inflammation and pathology. DESIGN: We analysed several ligands and receptors of the alternative NF-κB pathway, RelB, p52 nuclear translocation and target genes in tissue samples of H. pylori-infected patients with different degrees of gastritis or early gastric tumours by in situ hybridisation, immunohistochemistry, Western blot and real-time PCR analyses. Molecular mechanisms involved in LTßR activation by H. pylori were assessed in vitro using human gastric cancer cell lines and distinct H. pylori isolates. The effects of blocking or agonistically activating LTßR on gastric pathology during challenge with a human pathogenic H. pylori strain were studied in a mouse model. RESULTS: Among the tested candidates, LT was significantly increased and activated alternative NF-κB signalling was observed in the gastric mucosa of H. pylori-infected patients. H. pyloriinduced LTßR-ligand expression in a type IV secretion system-dependent but CagA-independent manner, resulting in activation of the alternative NF-κB pathway, which was further enhanced by blocking canonical NF-κB during infection. Blocking LTßR signalling in vivo suppressed H. pylori-driven gastritis, whereas LTßR activation in gastric epithelial cells of infected mice induced a broadened pro-inflammatory chemokine milieu, resulting in exacerbated pathology. CONCLUSIONS: LTßR-triggered activation of alternative NF-κB signalling in gastric epithelial cells executes H. pylori-induced chronic gastritis, representing a novel target to restrict gastric inflammation and pathology elicited by H. pylori, while exclusively targeting canonical NF-κB may aggravate pathology by enhancing the alternative pathway.

Early pulmonary response is critical for extra-pulmonary carbon nanoparticle mediated effects: comparison of inhalation versus intra-arterial infusion exposures in mice.

BACKGROUND: The death toll associated with inhaled ambient particulate matter (PM) is attributed mainly to cardio-vascular rather than pulmonary effects. However, it is unclear whether the key event for cardiovascular impairment is particle translocation from lung to circulation (direct effect) or indirect effects due to pulmonary particle-cell interactions. In this work, we addressed this issue by exposing healthy mice via inhalation and intra-arterial infusion (IAI) to carbon nanoparticles (CNP) as surrogate for soot, a major constituent of (ultrafine) urban PM. METHODS: Equivalent surface area CNP doses in the blood (30mm2 per animal) were applied by IAI or inhalation (lung-deposited dose 10,000mm2; accounting for 0.3% of lung-to-blood CNP translocation). Mice were analyzed for changes in hematology and molecular markers of endothelial/epithelial dysfunction, pro-inflammatory reactions, oxidative stress, and coagulation in lungs and extra-pulmonary organs after CNP inhalation (4 h and 24 h) and CNP infusion (4 h). For methodological reasons, we used two different CNP types (spark-discharge and Printex90), with very similar physicochemical properties [≥98 and ≥95% elemental carbon; 10 and 14 nm primary particle diameter; and 800 and 300 m2/g specific surface area] for inhalation and IAI respectively. RESULTS: Mild pulmonary inflammatory responses and significant systemic effects were observed following 4 h and 24 h CNP inhalation. Increased retention of activated leukocytes, secondary thrombocytosis, and pro-inflammatory responses in secondary organs were detected following 4 h and 24 h of CNP inhalation only. Interestingly, among the investigated extra-pulmonary tissues (i.e. aorta, heart, and liver); aorta revealed as the most susceptible extra-pulmonary target following inhalation exposure. Bypassing the lungs by IAI however did not induce any extra-pulmonary effects at 4 h as compared to inhalation. CONCLUSIONS: Our findings indicate that extra-pulmonary effects due to CNP inhalation are dominated by indirect effects (particle-cell interactions in the lung) rather than direct effects (translocated CNPs) within the first hours after exposure. Hence, CNP translocation may not be the key event inducing early cardiovascular impairment following air pollution episodes. The considerable response detected in the aorta after CNP inhalation warrants more emphasis on this tissue in future studies.

Loss of mitochondrial peptidase Clpp leads to infertility, hearing loss plus growth retardation via accumulation of CLPX, mtDNA and inflammatory factors.

The caseinolytic peptidase P (CLPP) is conserved from bacteria to humans. In the mitochondrial matrix, it multimerizes and forms a macromolecular proteasome-like cylinder together with the chaperone CLPX. In spite of a known relevance for the mitochondrial unfolded protein response, its substrates and tissue-specific roles are unclear in mammals. Recessive CLPP mutations were recently observed in the human Perrault variant of ovarian failure and sensorineural hearing loss. Here, a first characterization of CLPP null mice demonstrated complete female and male infertility and auditory deficits. Disrupted spermatogenesis already at the spermatid stage and ovarian follicular differentiation failure were evident. Reduced pre-/post-natal survival and marked ubiquitous growth retardation contrasted with only light impairment of movement and respiratory activities. Interestingly, the mice showed resistance to ulcerative dermatitis. Systematic expression studies detected up-regulation of other mitochondrial chaperones, accumulation of CLPX and mtDNA as well as inflammatory factors throughout tissues. T-lymphocytes in the spleen were activated. Thus, murine Clpp deletion represents a faithful Perrault model. The disease mechanism probably involves deficient clearance of mitochondrial components and inflammatory tissue destruction.

High mobility group N proteins modulate the fidelity of the cellular transcriptional profile in a tissue- and variant-specific manner.

The nuclei of most vertebrate cells contain members of the high mobility group N (HMGN) protein family, which bind specifically to nucleosome core particles and affect chromatin structure and function, including transcription. Here, we study the biological role of this protein family by systematic analysis of phenotypes and tissue transcription profiles in mice lacking functional HMGN variants. Phenotypic analysis of Hmgn1(tm1/tm1), Hmgn3(tm1/tm1), and Hmgn5(tm1/tm1) mice and their wild type littermates with a battery of standardized tests uncovered variant-specific abnormalities. Gene expression analysis of four different tissues in each of the Hmgn(tm1/tm1) lines reveals very little overlap between genes affected by specific variants in different tissues. Pathway analysis reveals that loss of an HMGN variant subtly affects expression of numerous genes in specific biological processes. We conclude that within the biological framework of an entire organism, HMGNs modulate the fidelity of the cellular transcriptional profile in a tissue- and HMGN variant-specific manner.

An ENU mutagenesis-derived mouse model with a dominant Jak1 mutation resembling phenotypes of systemic autoimmune disease.

Within the Munich, Germany, N-ethyl-N-nitrosourea mouse mutagenesis program, we isolated a dominant Jak1 mouse model resembling phenotypic characteristics related to autoimmune disease. Chromosomal sequencing revealed a new Jak1 (p.Ser645Pro) point mutation at the conserved serine of the pseudokinase domain, corresponding to a somatic human mutation (p.Ser646Phe) inducing a constitutive activation of the Janus kinase (JAK)/STAT pathway. Morphologically, all Jak1(S645P+/-) mice showed a progressive structural deterioration of ears starting at the age of 4 months, with mononuclear cell infiltration into the dermis. Female mutant mice, in particular, developed severe skin lesions in the neck from 7 months of age. The IHC analysis of these lesions showed an activation of Stat3 downstream to Jak1(S645P) and elevated tissue levels of IL-6. Histopathological analysis of liver revealed a nodular regenerative hyperplasia. In the spleen, the number of Russell bodies was doubled, correlating with significant increased levels of all immunoglobulin isotypes and anti-DNA antibodies in serum. Older mutant mice developed thrombocytopenia and altered microcytic red blood cell counts. Jak1(S645P+/-) mice showed phenotypes related to impaired bone metabolism as increased carboxy-terminal collagen cross-link-1 levels and alkaline phosphatase activities in plasma, hypophosphatemia, and strongly decreased bone morphometric values. Taken together, Jak1(S645P+/-) mice showed an increased activation of the IL-6-JAK-STAT pathway leading to a systemic lupus erythematosus-like phenotype and offering a new valuable tool to study the role of the JAK/STAT pathway in disease development.

Immune modulation by Fas ligand reverse signaling: lymphocyte proliferation is attenuated by the intracellular Fas ligand domain.

Fas ligand (FasL) not only induces apoptosis in Fas receptor-bearing target cells, it is also able to transmit signals into the FasL-expressing cell via its intracellular domain (ICD). Recently, we described a Notch-like proteolytic processing of FasL that leads to the release of the FasL ICD into the cytoplasm and subsequent translocation into the nucleus where it may influence gene transcription. To study the molecular mechanism underlying such reverse FasL signaling in detail and to analyze its physiological importance in vivo, we established a knockout/knockin mouse model, in which wild-type FasL was replaced with a deletion mutant lacking the ICD. Our results demonstrate that FasL ICD signaling impairs activation-induced proliferation in B and T cells by diminishing phosphorylation of phospholipase C γ, protein kinase C, and extracellular signal-regulated kinase 1/2. We also demonstrate that the FasL ICD interacts with the transcription factor lymphoid-enhancer binding factor-1 and inhibits lymphoid-enhancer binding factor-1-dependent transcription. In vivo, plasma cell numbers, generation of germinal center B cells, and, consequently, production of antigen-specific immunoglobulin M antibodies in response to immunization with T cell-dependent or T cell-independent antigen are negatively affected in presence of the FasL ICD, suggesting that FasL reverse signaling participates in negative fine-tuning of certain immune responses.

Cytochrome c oxidase subunit 4 isoform 2-knockout mice show reduced enzyme activity, airway hyporeactivity, and lung pathology.

Cytochrome c oxidase (COX) is the terminal enzyme of the mitochondrial electron transport chain. The purpose of this study was to analyze the function of lung-specific cytochrome c oxidase subunit 4 isoform 2 (COX4i2) in vitro and in COX4i2-knockout mice in vivo. COX was isolated from cow lung and liver as control and functionally analyzed. COX4i2-knockout mice were generated and the effect of the gene knockout was determined, including COX activity, tissue energy levels, noninvasive and invasive lung function, and lung pathology. These studies were complemented by a comprehensive functional screen performed at the German Mouse Clinic (Neuherberg, Germany). We show that isolated cow lung COX containing COX4i2 is about twice as active (88 and 102% increased activity in the presence of allosteric activator ADP and inhibitor ATP, respectively) as liver COX, which lacks COX4i2. In COX4i2-knockout mice, lung COX activity and cellular ATP levels were significantly reduced (-50 and -29%, respectively). Knockout mice showed decreased airway responsiveness (60% reduced P(enh) and 58% reduced airway resistance upon challenge with 25 and 100 mg methacholine, respectively), and they developed a lung pathology deteriorating with age that included the appearance of Charcot-Leyden crystals. In addition, there was an interesting sex-specific phenotype, in which the knockout females showed reduced lean mass (-12%), reduced total oxygen consumption rate (-8%), improved glucose tolerance, and reduced grip force (-14%) compared to wild-type females. Our data suggest that high activity lung COX is a central determinant of airway function and is required for maximal airway responsiveness and healthy lung function. Since airway constriction requires energy, we propose a model in which reduced tissue ATP levels explain protection from airway hyperresponsiveness, i.e., absence of COX4i2 leads to reduced lung COX activity and ATP levels, which results in impaired airway constriction and thus reduced airway responsiveness; long-term lung pathology develops in the knockout mice due to impairment of energy-costly lung maintenance processes; and therefore, we propose mitochondrial oxidative phosphorylation as a novel target for the treatment of respiratory diseases, such as asthma.

Requirement of the RNA-editing enzyme ADAR2 for normal physiology in mice.

ADAR2, an RNA editing enzyme that converts specific adenosines to inosines in certain pre-mRNAs, often leading to amino acid substitutions in the encoded proteins, is mainly expressed in brain. Of all ADAR2-mediated edits, a single one in the pre-mRNA of the AMPA receptor subunit GluA2 is essential for survival. Hence, early postnatal death of mice lacking ADAR2 is averted when the critical edit is engineered into both GluA2 encoding Gria2 alleles. Adar2(-/-)/Gria2(R/R) mice display normal appearance and life span, but the general phenotypic effects of global lack of ADAR2 have remained unexplored. Here we have employed the Adar2(-/-)/Gria2(R/R) mouse line, and Gria2(R/R) mice as controls, to study the phenotypic consequences of loss of all ADAR2-mediated edits except the critical one in GluA2. Our extended phenotypic analysis covering ∼320 parameters identified significant changes related to absence of ADAR2 in behavior, hearing ability, allergy parameters and transcript profiles of brain.

Innovations in phenotyping of mouse models in the German Mouse Clinic.

Under the label of the German Mouse Clinic (GMC), a concept has been developed and implemented that allows the better understanding of human diseases on the pathophysiological and molecular level. This includes better understanding of the crosstalk between different organs, pleiotropy of genes, and the systemic impact of envirotypes and drugs. In the GMC, experts from various fields of mouse genetics and physiology, in close collaboration with clinicians, work side by side under one roof. The GMC is an open-access platform for the scientific community by providing phenotypic analysis in bilateral collaborations ("bottom-up projects") and as a partner and driver in international large-scale biology projects ("top-down projects"). Furthermore, technology development is a major topic in the GMC. Innovative techniques for primary and secondary screens are developed and implemented into the phenotyping pipelines (e.g., detection of volatile organic compounds, VOCs).

A novel N-ethyl-N-nitrosourea-induced mutation in phospholipase Cγ2 causes inflammatory arthritis, metabolic defects, and male infertility in vitro in a murine model.

OBJECTIVE: It is difficult to identify a single causative factor for inflammatory arthritis because of the multifactorial nature of the disease. This study was undertaken to dissect the molecular complexity of systemic inflammatory disease, utilizing a combined approach of mutagenesis and systematic phenotype screening in a murine model. METHODS: In a large-scale N-ethyl-N-nitrosourea mutagenesis project, the Ali14 mutant mouse strain was established because of dominant inheritance of spontaneous swelling and inflammation of the hind paws. Genetic mapping and subsequent candidate gene sequencing were conducted to find the causative gene, and systematic phenotyping of Ali14/+ mice was performed in the German Mouse Clinic. RESULTS: A novel missense mutation in the phospholipase Cγ2 gene (Plcg2) was identified in Ali14/+ mice. Because of the hyperreactive external entry of calcium observed in cultured B cells and other in vitro experiments, the Ali14 mutation is thought to be a novel gain-of-function allele of Plcg2. Findings from systematic screening of Ali14/+ mice demonstrated various phenotypic changes: an abnormally high T cell:B cell ratio, up-regulation of Ig, alterations in body composition, and a reduction in cholesterol and triglyceride levels in peripheral blood. In addition, spermatozoa from Ali14/+ mice failed to fertilize eggs in vitro, despite the normal fertility of the Ali14/+ male mice in vivo. CONCLUSION: These results suggest that the Plcg2-mediated pathways play a crucial role in various metabolic and sperm functions, in addition to initiating and maintaining the immune system. These findings may indicate the importance of the Ali14/+ mouse strain as a model for systemic inflammatory diseases and inflammation-related metabolic changes in humans.

Mouse phenotyping.

Model organisms like the mouse are important tools to learn more about gene function in man. Within the last 20 years many mutant mouse lines have been generated by different methods such as ENU mutagenesis, constitutive and conditional knock-out approaches, knock-down, introduction of human genes, and knock-in techniques, thus creating models which mimic human conditions. Due to pleiotropic effects, one gene may have different functions in different organ systems or time points during development. Therefore mutant mouse lines have to be phenotyped comprehensively in a highly standardized manner to enable the detection of phenotypes which might otherwise remain hidden. The German Mouse Clinic (GMC) has been established at the Helmholtz Zentrum München as a phenotyping platform with open access to the scientific community (www.mousclinic.de; [1]). The GMC is a member of the EUMODIC consortium which created the European standard workflow EMPReSSslim for the systemic phenotyping of mouse models (http://www.eumodic.org/[2]).

New C3H KitN824K/WT cancer mouse model develops late-onset malignant mammary tumors with high penetrance.

Gastro-intestinal stromal tumors and acute myeloid leukemia induced by activating stem cell factor receptor tyrosine kinase (KIT) mutations are highly malignant. Less clear is the role of KIT mutations in the context of breast cancer. Treatment success of KIT-induced cancers is still unsatisfactory because of primary or secondary resistance to therapy. Mouse models offer essential platforms for studies on molecular disease mechanisms in basic cancer research. In the course of the Munich N-ethyl-N-nitrosourea (ENU) mutagenesis program a mouse line with inherited polycythemia was established. It carries a base-pair exchange in the Kit gene leading to an amino acid exchange at position 824 in the activation loop of KIT. This KIT variant corresponds to the N822K mutation found in human cancers, which is associated with imatinib-resistance. C3H KitN824K/WT mice develop hyperplasia of interstitial cells of Cajal and retention of ingesta in the cecum. In contrast to previous Kit-mutant models, we observe a benign course of gastrointestinal pathology associated with prolonged survival. Female mutants develop mammary carcinomas at late onset and subsequent lung metastasis. The disease model complements existing oncology research platforms. It allows for addressing the role of KIT mutations in breast cancer and identifying genetic and environmental modifiers of disease progression.

Microphthalmia, parkinsonism, and enhanced nociception in Pitx3 ( 416insG ) mice.

A new spontaneous mouse mutant was characterized by closed eyelids at weaning and without apparent eyes (provisional gene name, eyeless; provisional gene symbol, eyl). The mutation follows a recessive pattern of inheritance and was mapped to the region of chromosome 19 containing Pitx3. Genetic complementation tests using Pitx3 ( ak/+ ) mice confirmed eyl as a new allele of Pitx3 (Pitx3 ( eyl )). Sequencing of the Pitx3 gene in eyl mutants identified an inserted G after cDNA position 416 (416insG; exon 4). The shifted open reading frame is predicted to result in a hybrid protein still containing the Pitx3 homeobox, but followed by 121 new amino acids. The novel Pitx3 ( eyl/eyl ) mutants expressed ophthalmological and brain defects similar to Pitx3 ( ak/ak ) mice: microphthalmia or anophthalmia and loss of dopamine neurons of the substantia nigra. In addition, we observed in the homozygous eyeless mutants increased extramedullary hematopoiesis in the spleen, frequently liver steatosis, and reduced body weight. There were also several behavioral changes in the homozygous mutants, including reduced forelimb grip strength and increased nociception. In addition to these alterations in both sexes, we observed in female Pitx3 ( eyl/eyl ) mice increased anxiety-related behavior, reduced locomotor activity, reduced object exploration, and increased social contacts; however, we observed decreased anxiety-related behavior and increased arousal in males. Most of these defects identified in the new Pitx3 mutation are observed in Parkinson patients, making the Pitx3 ( eyl ) mutant a valuable new model. It is the first mouse mutant carrying a point mutation within the coding region of Pitx3.

Increased estrogen to androgen ratio enhances immunoglobulin levels and impairs B cell function in male mice.

Sex steroids, such as estrogens and androgens, are important regulators of the humoral immune response. Studies in female mice have demonstrated that alteration of circulating estrogen concentration regulates antibody-mediated immunity. As males have normally little endogenous estrogen, we hypothesized that in males high estrogens and low androgens affect the immune system and enhance the allergic inflammatory response. Here, we studied transgenic male mice expressing human aromatase (AROM+). These animals have a high circulating estrogen to androgen ratio (E/A), causing female traits such as gynecomastia. We found that AROM+ male mice had significantly higher plasma immunoglobulin levels, particularly IgE. Flow cytometry analyses of splenocytes revealed changes in mature/immature B cell ratio together with a transcriptional upregulation of the Igh locus. Furthermore, higher proliferation rate and increased IgE synthesis after IgE class-switching was found. Subsequently, we utilized an ovalbumin airway challenge model to test the allergic response in AROM+ male mice. In line with above observations, an increase in IgE levels was measured, albeit no impact on immune cell infiltration into the lungs was detected. Together, our findings suggest that high circulating E/A in males significantly alters B cell function without any significant enhancement in allergic inflammation.