Protein arginine methyltransferase 2 controls inflammatory signaling in acute myeloid leukemia.

Arginine methylation is catalyzed by protein arginine methyltransferases (PRMTs) and is involved in various cellular processes, including cancer development. PRMT2 expression is increased in several cancer types although its role in acute myeloid leukemia (AML) remains unknown. Here, we investigate the role of PRMT2 in a cohort of patients with AML, PRMT2 knockout AML cell lines as well as a Prmt2 knockout mouse model. In patients, low PRMT2 expressors are enriched for inflammatory signatures, including the NF-κB pathway, and show inferior survival. In keeping with a role for PRMT2 in control of inflammatory signaling, bone marrow-derived macrophages from Prmt2 KO mice display increased pro-inflammatory cytokine signaling upon LPS treatment. In PRMT2-depleted AML cell lines, aberrant inflammatory signaling has been linked to overproduction of IL6, resulting from a deregulation of the NF-κB signaling pathway, therefore leading to hyperactivation of STAT3. Together, these findings identify PRMT2 as a key regulator of inflammation in AML.

The Rogdi knockout mouse is a model for Kohlschütter-Tönz syndrome.

Kohlschütter-Tönz syndrome (KTS) is a rare autosomal recessive disorder characterized by severe intellectual disability, early-onset epileptic seizures, and amelogenesis imperfecta. Here, we present a novel Rogdi mutant mouse deleting exons 6-11- a mutation found in KTS patients disabling ROGDI function. This Rogdi-/- mutant model recapitulates most KTS symptoms. Mutants displayed pentylenetetrazol-induced seizures, confirming epilepsy susceptibility. Spontaneous locomotion and circadian activity tests demonstrate Rogdi mutant hyperactivity mirroring patient spasticity. Object recognition impairment indicates memory deficits. Rogdi-/- mutant enamel was markedly less mature. Scanning electron microscopy confirmed its hypomineralized/hypomature crystallization, as well as its low mineral content. Transcriptomic RNA sequencing of postnatal day 5 lower incisors showed downregulated enamel matrix proteins Enam, Amelx, and Ambn. Enamel crystallization appears highly pH-dependent, cycling between an acidic and neutral pH during enamel maturation. Rogdi-/- teeth exhibit no signs of cyclic dental acidification. Additionally, expression changes in Wdr72, Slc9a3r2, and Atp6v0c were identified as potential contributors to these tooth acidification abnormalities. These proteins interact through the acidifying V-ATPase complex. Here, we present the Rogdi-/- mutant as a novel model to partially decipher KTS pathophysiology. Rogdi-/- mutant defects in acidification might explain the unusual combination of enamel and rare neurological disease symptoms.

Stefin B Inhibits NLRP3 Inflammasome Activation via AMPK/mTOR Signalling.

Stefin B (cystatin B) is an inhibitor of lysosomal and nuclear cysteine cathepsins. The gene for stefin B is located on human chromosome 21 and its expression is upregulated in the brains of individuals with Down syndrome. Biallelic loss-of-function mutations in the stefin B gene lead to Unverricht-Lundborg disease-progressive myoclonus epilepsy type 1 (EPM1) in humans. In our past study, we demonstrated that mice lacking stefin B were significantly more sensitive to sepsis induced by lipopolysaccharide (LPS) and secreted higher levels of interleukin 1-ß (IL-1ß) due to increased inflammasome activation in bone marrow-derived macrophages. Here, we report lower interleukin 1-ß processing and caspase-11 expression in bone marrow-derived macrophages prepared from mice that have an additional copy of the stefin B gene. Increased expression of stefin B downregulated mitochondrial reactive oxygen species (ROS) generation and lowered the NLR family pyrin domain containing 3 (NLRP3) inflammasome activation in macrophages. We determined higher AMP-activated kinase phosphorylation and downregulation of mTOR activity in stefin B trisomic macrophages-macrophages with increased stefin B expression. Our study showed that increased stefin B expression downregulated mitochondrial ROS generation and increased autophagy. The present work contributes to a better understanding of the role of stefin B in regulation of autophagy and inflammasome activation in macrophages and could help to develop new treatments.

Development of HPV16 mouse and dog models for more accurate prediction of human vaccine efficacy.

BACKGROUND: Animal models are essential to understand the physiopathology of human diseases but also to evaluate new therapies. However, for several diseases there is no appropriate animal model, which complicates the development of effective therapies. HPV infections, responsible for carcinoma cancers, are among these. So far, the lack of relevant animal models has hampered the development of therapeutic vaccines. In this study, we used a candidate therapeutic vaccine named C216, similar to the ProCervix candidate therapeutic vaccine, to validate new mouse and dog HPV preclinical models. ProCervix has shown promising results with classical subcutaneous murine TC-1 cell tumor isografts but has failed in a phase II study. RESULTS: We first generated E7/HPV16 syngeneic transgenic mice in which the expression of the E7 antigen could be switched on through the use of Cre-lox recombination. Non-integrative LentiFlash® viral particles were used to locally deliver Cre mRNA, resulting in E7/HPV16 expression and GFP reporter fluorescence. The expression of E7/HPV16 was monitored by in vivo fluorescence using Cellvizio imaging and by local mRNA expression quantification. In the experimental conditions used, we observed no differences in E7 expression between C216 vaccinated and control groups. To mimic the MHC diversity of humans, E7/HPV16 transgenes were locally delivered by injection of lentiviral particles in the muscle of dogs. Vaccination with C216, tested with two different adjuvants, induced a strong immune response in dogs. However, we detected no relationship between the level of cellular response against E7/HPV16 and the elimination of E7-expressing cells, either by fluorescence or by RT-ddPCR analysis. CONCLUSIONS: In this study, we have developed two animal models, with a genetic design that is easily transposable to different antigens, to validate the efficacy of candidate vaccines. Our results indicate that, despite being immunogenic, the C216 candidate vaccine did not induce a sufficiently strong immune response to eliminate infected cells. Our results are in line with the failure of the ProCervix vaccine that was observed at the end of the phase II clinical trial, reinforcing the relevance of appropriate animal models.

HRAS germline mutations impair LKB1/AMPK signaling and mitochondrial homeostasis in Costello syndrome models.

Germline mutations that activate genes in the canonical RAS/MAPK signaling pathway are responsible for rare human developmental disorders known as RASopathies. Here, we analyzed the molecular determinants of Costello syndrome (CS) using a mouse model expressing HRAS p.G12S, patient skin fibroblasts, hiPSC-derived human cardiomyocytes, a HRAS p.G12V zebrafish model, and human fibroblasts expressing lentiviral constructs carrying HRAS p.G12S or HRAS p.G12A mutations. The findings revealed alteration of mitochondrial proteostasis and defective oxidative phosphorylation in the heart and skeletal muscle of CS mice that were also found in the cell models of the disease. The underpinning mechanisms involved the inhibition of the AMPK signaling pathway by mutant forms of HRAS, leading to alteration of mitochondrial proteostasis and bioenergetics. Pharmacological activation of mitochondrial bioenergetics and quality control restored organelle function in HRAS p.G12A and p.G12S cell models, reduced left ventricle hypertrophy in CS mice, and diminished the occurrence of developmental defects in the CS zebrafish model. Collectively, these findings highlight the importance of mitochondrial proteostasis and bioenergetics in the pathophysiology of RASopathies and suggest that patients with CS may benefit from treatment with mitochondrial modulators.

Overproduction of hydrogen sulfide, generated by cystathionine ß-synthase, disrupts brain wave patterns and contributes to neurobehavioral dysfunction in a rat model of down syndrome.

Using a novel rat model of Down syndrome (DS), the functional role of the cystathionine-ß-synthase (CBS)/hydrogen sulfide (H2S) pathway was investigated on the pathogenesis of brain wave pattern alterations and neurobehavioral dysfunction. Increased expression of CBS and subsequent overproduction of H2S was observed in the brain of DS rats, with CBS primarily localizing to astrocytes and the vasculature. DS rats exhibited neurobehavioral defects, accompanied by a loss of gamma brain wave activity and a suppression of the expression of multiple pre- and postsynaptic proteins. Aminooxyacetate, a prototypical pharmacological inhibitor of CBS, increased the ability of the DS brain tissue to generate ATP in vitro and reversed the electrophysiological and neurobehavioral alterations in vivo. Thus, the CBS/H2S pathway contributes to the pathogenesis of neurological dysfunction in DS, most likely through dysregulation of cellular bioenergetics and gene expression.

Structure-Activity Relationship in the Leucettine Family of Kinase Inhibitors.

The protein kinase DYRK1A is involved in Alzheimer's disease, Down syndrome, diabetes, viral infections, and leukemia. Leucettines, a family of 2-aminoimidazolin-4-ones derived from the marine sponge alkaloid Leucettamine B, have been developed as pharmacological inhibitors of DYRKs (dual specificity, tyrosine phosphorylation regulated kinases) and CLKs (cdc2-like kinases). We report here on the synthesis and structure-activity relationship (SAR) of 68 Leucettines. Leucettines were tested on 11 purified kinases and in 5 cellular assays: (1) CLK1 pre-mRNA splicing, (2) Threonine-212-Tau phosphorylation, (3) glutamate-induced cell death, (4) autophagy and (5) antagonism of ligand-activated cannabinoid receptor CB1. The Leucettine SAR observed for DYRK1A is essentially identical for CLK1, CLK4, DYRK1B, and DYRK2. DYRK3 and CLK3 are less sensitive to Leucettines. In contrast, the cellular SAR highlights correlations between inhibition of specific kinase targets and some but not all cellular effects. Leucettines deserve further development as potential therapeutics against various diseases on the basis of their molecular targets and cellular effects.

Building the Future Therapies for Down Syndrome: The Third International Conference of the T21 Research Society.

Research focused on Down syndrome has increased in the last several years to advance understanding of the consequences of trisomy 21 (T21) on molecular and cellular processes and, ultimately, on individuals with Down syndrome. The Trisomy 21 Research Society (T21RS) is the premier scientific organization for researchers and clinicians studying Down syndrome. The Third International Conference of T21RS, held June 6-9, 2019, in Barcelona, Spain, brought together 429 scientists, families, and industry representatives to share the latest discoveries on underlying cellular and molecular mechanisms of T21, define cognitive and behavioral challenges and better understand comorbidities associated with Down syndrome, including Alzheimer's disease and leukemia. Presentation of cutting-edge results in neuroscience, neurology, model systems, psychology, cancer, biomarkers and molecular and phar-ma-cological therapeutic approaches demonstrate the compelling interest and continuing advancement in all aspects of understanding and ameliorating conditions associated with T21.

Immune Dysregulation and the Increased Risk of Complications and Mortality Following Respiratory Tract Infections in Adults With Down Syndrome.

The risk of severe outcomes following respiratory tract infections is significantly increased in individuals over 60 years, especially in those with chronic medical conditions, i.e., hypertension, diabetes, cardiovascular disease, dementia, chronic respiratory disease, and cancer. Down Syndrome (DS), the most prevalent intellectual disability, is caused by trisomy-21 in ~1:750 live births worldwide. Over the past few decades, a substantial body of evidence has accumulated, pointing at the occurrence of alterations, impairments, and subsequently dysfunction of the various components of the immune system in individuals with DS. This associates with increased vulnerability to respiratory tract infections in this population, such as the influenza virus, respiratory syncytial virus, SARS-CoV-2 (COVID-19), and bacterial pneumonias. To emphasize this link, here we comprehensively review the immunobiology of DS and its contribution to higher susceptibility to severe illness and mortality from respiratory tract infections.

The effects of Cstb duplication on APP/amyloid-ß pathology and cathepsin B activity in a mouse model.

People with Down syndrome (DS), caused by trisomy of chromosome 21 have a greatly increased risk of developing Alzheimer's disease (AD). This is in part because of triplication of a chromosome 21 gene, APP. This gene encodes amyloid precursor protein, which is cleaved to form amyloid-ß that accumulates in the brains of people who have AD. Recent experimental results demonstrate that a gene or genes on chromosome 21, other than APP, when triplicated significantly accelerate amyloid-ß pathology in a transgenic mouse model of amyloid-ß deposition. Multiple lines of evidence indicate that cysteine cathepsin activity influences APP cleavage and amyloid-ß accumulation. Located on human chromosome 21 (Hsa21) is an endogenous inhibitor of cathepsin proteases, CYSTATIN B (CSTB) which is proposed to regulate cysteine cathepsin activity in vivo. Here we determined if three copies of the mouse gene Cstb is sufficient to modulate amyloid-ß accumulation and cathepsin activity in a transgenic APP mouse model. Duplication of Cstb resulted in an increase in transcriptional and translational levels of Cstb in the mouse cortex but had no effect on the deposition of insoluble amyloid-ß plaques or the levels of soluble or insoluble amyloid-ß42, amyloid-ß40, or amyloid-ß38 in 6-month old mice. In addition, the increased CSTB did not alter the activity of cathepsin B enzyme in the cortex of 3-month or 6-month old mice. These results indicate that the single-gene duplication of Cstb is insufficient to elicit a disease-modifying phenotype in the dupCstb x tgAPP mice, underscoring the complexity of the genetic basis of AD-DS and the importance of multiple gene interactions in disease.

Mechanism of cystathionine-ß-synthase inhibition by disulfiram: The role of bis(N,N-diethyldithiocarbamate)-copper(II).

BACKGROUND: Hydrogen sulfide (H2S) is an endogenous mammalian gasotransmitter. Cystathionine ß-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3-MST) are the principal enzymes responsible for its biogenesis. A recent yeast screen suggested that disulfiram (a well-known inhibitor of aldehyde dehydrogenase and a clinically used drug in the treatment of alcoholism) may inhibit CBS in a cell-based environment. However, prior studies have not observed any direct inhibition of CBS by disulfiram. We investigated the potential role of bioconversion of disulfiram to bis(N,N-diethyldithiocarbamate)-copper(II) complex (CuDDC) in the inhibitory effect of disulfiram on H2S production and assessed its effect in two human cell types with high CBS expression: HCT116 colon cancer cells and Down syndrome (DS) fibroblasts. METHODS: H2S production from recombinant human CBS, CSE and 3-MST was measured using the fluorescent H2S probe AzMC. Mouse liver homogenate (a rich source of CBS) was also employed to measure H2S biosynthesis. The interaction of copper with accessible protein cysteine residues was evaluated using the DTNB method. Cell proliferation and viability were measured using the BrdU and MTT methods. Cellular bioenergetics was evaluated by Extracellular Flux Analysis. RESULTS: While disulfiram did not exert any significant direct inhibitory effect on any of the H2S-producing enzymes, its metabolite, CuDDC was a potent inhibitor of CBS and CSE. The mode of its action is likely related to the complexed copper molecule. In cell-based systems, the effects of disulfiram were variable. In colon cancer cells, no significant effect of disulfiram was observed on H2S production or proliferation or viability. In contrast, in DS fibroblasts, disulfiram inhibited H2S production and improved proliferation and viability. Copper, on its own, failed to have any effects on either cell type, likely due to its low cell penetration. CuDDC inhibited H2S production in both cell types studied and exerted the functional effects that would be expected from a CBS inhibitor: inhibition of cell proliferation of cancer cells and a bell-shaped effect (stimulation of proliferation at low concentration and inhibition of these responses at higher concentration) in DS cells. Control experiments using a chemical H2S donor showed that, in addition to inhibiting CBS and CSE, part of the biological effects of CuDDC relates to a direct reaction with H2S, which occurs through its complexed copper. CONCLUSIONS: Disulfiram, via its metabolite CuDDC acts as an inhibitor of CBS and a scavenger of H2S, which, in turn, potently suppresses H2S levels in various cell types. Inhibition of H2S biosynthesis may explain some of the previously reported actions of disulfiram and CuDDC in vitro and in vivo. Disulfiram or CuDDC may be considered as potential agents for the experimental therapy of various pathophysiological conditions associated with H2S overproduction.

The Human-Specific BOLA2 Duplication Modifies Iron Homeostasis and Anemia Predisposition in Chromosome 16p11.2 Autism Individuals.

Human-specific duplications at chromosome 16p11.2 mediate recurrent pathogenic 600 kbp BP4-BP5 copy-number variations, which are among the most common genetic causes of autism. These copy-number polymorphic duplications are under positive selection and include three to eight copies of BOLA2, a gene involved in the maturation of cytosolic iron-sulfur proteins. To investigate the potential advantage provided by the rapid expansion of BOLA2, we assessed hematological traits and anemia prevalence in 379,385 controls and individuals who have lost or gained copies of BOLA2: 89 chromosome 16p11.2 BP4-BP5 deletion carriers and 56 reciprocal duplication carriers in the UK Biobank. We found that the 16p11.2 deletion is associated with anemia (18/89 carriers, 20%, p = 4e-7, OR = 5), particularly iron-deficiency anemia. We observed similar enrichments in two clinical 16p11.2 deletion cohorts, which included 6/63 (10%) and 7/20 (35%) unrelated individuals with anemia, microcytosis, low serum iron, or low blood hemoglobin. Upon stratification by BOLA2 copy number, our data showed an association between low BOLA2 dosage and the above phenotypes (8/15 individuals with three copies, 53%, p = 1e-4). In parallel, we analyzed hematological traits in mice carrying the 16p11.2 orthologous deletion or duplication, as well as Bola2+/- and Bola2-/- animals. The Bola2-deficient mice and the mice carrying the deletion showed early evidence of iron deficiency, including a mild decrease in hemoglobin, lower plasma iron, microcytosis, and an increased red blood cell zinc-protoporphyrin-to-heme ratio. Our results indicate that BOLA2 participates in iron homeostasis in vivo, and its expansion has a potential adaptive role in protecting against iron deficiency.

BIN1 recovers tauopathy-induced long-term memory deficits in mice and interacts with Tau through Thr348 phosphorylation.

The bridging integrator 1 gene (BIN1) is a major genetic risk factor for Alzheimer's disease (AD). In this report, we investigated how BIN1-dependent pathophysiological processes might be associated with Tau. We first generated a cohort of control and transgenic mice either overexpressing human MAPT (TgMAPT) or both human MAPT and BIN1 (TgMAPT;TgBIN1), which we followed-up from 3 to 15 months. In TgMAPT;TgBIN1 mice short-term memory deficits appeared earlier than in TgMAPT mice; however-unlike TgMAPT mice-TgMAPT;TgBIN1 mice did not exhibit any long-term or spatial memory deficits for at least 15 months. After killing the cohort at 18 months, immunohistochemistry revealed that BIN1 overexpression prevents both Tau mislocalization and somatic inclusion in the hippocampus, where an increase in BIN1-Tau interaction was also observed. We then sought mechanisms controlling the BIN1-Tau interaction. We developed a high-content screening approach to characterize modulators of the BIN1-Tau interaction in an agnostic way (1,126 compounds targeting multiple pathways), and we identified-among others-an inhibitor of calcineurin, a Ser/Thr phosphatase. We determined that calcineurin dephosphorylates BIN1 on a cyclin-dependent kinase phosphorylation site at T348, promoting the open conformation of the neuronal BIN1 isoform. Phosphorylation of this site increases the availability of the BIN1 SH3 domain for Tau interaction, as demonstrated by nuclear magnetic resonance experiments and in primary neurons. Finally, we observed that although the levels of the neuronal BIN1 isoform were unchanged in AD brains, phospho-BIN1(T348):BIN1 ratio was increased, suggesting a compensatory mechanism. In conclusion, our data support the idea that BIN1 modulates the AD risk through an intricate regulation of its interaction with Tau. Alteration in BIN1 expression or activity may disrupt this regulatory balance with Tau and have direct effects on learning and memory.

TUBG1 missense variants underlying cortical malformations disrupt neuronal locomotion and microtubule dynamics but not neurogenesis.

De novo heterozygous missense variants in the γ-tubulin gene TUBG1 have been linked to human malformations of cortical development associated with intellectual disability and epilepsy. Here, we investigated through in-utero electroporation and in-vivo studies, how four of these variants affect cortical development. We show that TUBG1 mutants affect neuronal positioning, disrupting the locomotion of new-born neurons but without affecting progenitors' proliferation. We further demonstrate that pathogenic TUBG1 variants are linked to reduced microtubule dynamics but without major structural nor functional centrosome defects in subject-derived fibroblasts. Additionally, we developed a knock-in Tubg1Y92C/+ mouse model and assessed consequences of the mutation. Although centrosomal positioning in bipolar neurons is correct, they fail to initiate locomotion. Furthermore, Tubg1Y92C/+ animals show neuroanatomical and behavioral defects and increased epileptic cortical activity. We show that Tubg1Y92C/+ mice partially mimic the human phenotype and therefore represent a relevant model for further investigations of the physiopathology of cortical malformations.

Ultrasound-Guided Approaches to Improve Orthotopic Mouse Xenograft Models for Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer death worldwide. While curative approaches for early stage HCC exist, effective treatment options for advanced HCC are lacking. Furthermore, there are no efficient chemopreventive strategies to limit HCC development once cirrhosis is established. One challenge for drug development is unsatisfactory animal models. In this article, we describe an orthotopic xenograft mouse model of human liver cancer cell lines through image-guided injection into the liver. This technique provides a less invasive yet highly efficient approach to engraft human HCC into mouse liver. Similarly, image-guided injections are used to deliver chemotherapeutics locally, enabling reduction in potential systemic adverse effects, while reducing the required dose for a therapeutic effect. In summary, this image-guided strategy provides a novel and convenient approach to improve current HCC mouse models. © 2019 The Authors. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.

Amphiphysin 2 modulation rescues myotubular myopathy and prevents focal adhesion defects in mice.

Centronuclear myopathies (CNMs) are severe diseases characterized by muscle weakness and myofiber atrophy. Currently, there are no approved treatments for these disorders. Mutations in the phosphoinositide 3-phosphatase myotubularin (MTM1) are responsible for X-linked CNM (XLCNM), also called myotubular myopathy, whereas mutations in the membrane remodeling Bin/amphiphysin/Rvs protein amphiphysin 2 [bridging integrator 1 (BIN1)] are responsible for an autosomal form of the disease. Here, we investigated the functional relationship between MTM1 and BIN1 in healthy skeletal muscle and in the physiopathology of CNM. Genetic overexpression of human BIN1 efficiently rescued the muscle weakness and life span in a mouse model of XLCNM. Exogenous human BIN1 expression with adeno-associated virus after birth also prevented the progression of the disease, suggesting that human BIN1 overexpression can compensate for the lack of MTM1 expression in this mouse model. Our results showed that MTM1 controls cell adhesion and integrin localization in mammalian muscle. Alterations in this pathway in Mtm1 -/y mice were associated with defects in myofiber shape and size. BIN1 expression rescued integrin and laminin alterations and restored myofiber integrity, supporting the idea that MTM1 and BIN1 are functionally linked and necessary for focal adhesions in skeletal muscle. The results suggest that BIN1 modulation might be an effective strategy for treating XLCNM.

Cbs overdosage is necessary and sufficient to induce cognitive phenotypes in mouse models of Down syndrome and interacts genetically with Dyrk1a.

Identifying dosage-sensitive genes is a key to understand the mechanisms underlying intellectual disability in Down syndrome (DS). The Dp(17Abcg1-Cbs)1Yah DS mouse model (Dp1Yah) shows cognitive phenotypes that need to be investigated to identify the main genetic driver. Here, we report that three copies of the cystathionine-beta-synthase gene (Cbs) in the Dp1Yah mice are necessary to observe a deficit in the novel object recognition (NOR) paradigm. Moreover, the overexpression of Cbs alone is sufficient to induce deficits in the NOR test. Accordingly, overexpressing human CBS specifically in Camk2a-expressing neurons leads to impaired objects discrimination. Altogether, this shows that Cbs overdosage is involved in DS learning and memory phenotypes. To go further, we identified compounds that interfere with the phenotypical consequence of CBS overdosage in yeast. Pharmacological intervention in Tg(CBS) mice with one selected compound restored memory in the NOR test. In addition, using a genetic approach, we demonstrated an epistatic interaction between Cbs and Dyrk1a, another human chromosome 21-located gene (which encodes the dual-specificity tyrosine phosphorylation-regulated kinase 1a) and an already known target for DS therapeutic intervention. Further analysis using proteomic approaches highlighted several molecular pathways, including synaptic transmission, cell projection morphogenesis and actin cytoskeleton, that are affected by DYRK1A and CBS overexpression. Overall, we demonstrated that CBS overdosage underpins the DS-related recognition memory deficit and that both CBS and DYRK1A interact to control accurate memory processes in DS. In addition, our study establishes CBS as an intervention point for treating intellectual deficiencies linked to DS.

Hepatocyte SHP deficiency protects mice from acetaminophen-evoked liver injury in a JNK-signaling regulation and GADD45ß-dependent manner.

Acetaminophen (APAP) overdose is a leading cause of drug-induced acute liver failure. Prolonged c-Jun N-terminal kinase (JNK) activation plays a central role in APAP-induced liver injury; however, growth arrest and DNA damage-inducible 45 beta (GADD45ß) is known to inhibit JNK phosphorylation. The orphan nuclear receptor small heterodimer partner (SHP, NR0B2) acts as a transcriptional co-repressor of various genes. The aim of the present study was to investigate the role of SHP in APAP-evoked hepatotoxicity. We used lethal (750 mg/kg) or sublethal (300 mg/kg) doses of APAP-treated wild-type (WT), Shp knockout (Shp-/-), hepatocyte-specific Shp knockout (Shphep-/-), and Shp and Gadd45ß double knockout (Shp-/-Gadd45ß-/-) mice for in vivo studies. Primary mouse hepatocytes were used for a comparative in vitro study. SHP deficiency protected against APAP toxicity with an increased survival rate, decreased liver damage, and inhibition of prolonged hepatic JNK phosphorylation in mice, which was independent of APAP metabolism regulation. Furthermore, Shphep-/- mice showed diminished APAP hepatotoxicity compared with WT mice. SHP-deficient primary mouse hepatocytes also showed decreased cell death and inhibition of sustained JNK phosphorylation following toxic APAP treatment. While SHP expression declined, GADD45ß expression increased after APAP treatment in WT mice. In Shp-/- mice, APAP-evoked GADD45ß induction was significantly enhanced. Notably, the ameliorative effects of SHP deficiency on APAP-induced liver injury were abolished in Shp-/-Gadd45ß-/- mice. The current study is the first to demonstrate that hepatocyte-specific SHP deficiency protects against APAP overdose-evoked hepatotoxicity in a JNK signaling regulation and GADD45ß dependent manner. SHP is suggested to be a novel therapeutic target for APAP overdose treatment.

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

Purpose: The 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. Methods: A 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. Results: Common 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. Conclusions: Ocular 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.

A new mouse model of ARX dup24 recapitulates the patients' behavioral and fine motor alterations.

The aristaless-related homeobox (ARX) transcription factor is involved in the development of GABAergic and cholinergic neurons in the forebrain. ARX mutations have been associated with a wide spectrum of neurodevelopmental disorders in humans, among which the most frequent, a 24 bp duplication in the polyalanine tract 2 (c.428_451dup24), gives rise to intellectual disability, fine motor defects with or without epilepsy. To understand the functional consequences of this mutation, we generated a partially humanized mouse model carrying the c.428_451dup24 duplication (Arxdup24/0) that we characterized at the behavior, neurological and molecular level. Arxdup24/0 males presented with hyperactivity, enhanced stereotypies and altered contextual fear memory. In addition, Arxdup24/0 males had fine motor defects with alteration of reaching and grasping abilities. Transcriptome analysis of Arxdup24/0 forebrains at E15.5 showed a down-regulation of genes specific to interneurons and an up-regulation of genes normally not expressed in this cell type, suggesting abnormal interneuron development. Accordingly, interneuron migration was altered in the cortex and striatum between E15.5 and P0 with consequences in adults, illustrated by the defect in the inhibitory/excitatory balance in Arxdup24/0 basolateral amygdala. Altogether, we showed that the c.428_451dup24 mutation disrupts Arx function with a direct consequence on interneuron development, leading to hyperactivity and defects in precise motor movement control and associative memory. Interestingly, we highlighted striking similarities between the mouse phenotype and a cohort of 33 male patients with ARX c.428_451dup24, suggesting that this new mutant mouse line is a good model for understanding the pathophysiology and evaluation of treatment.