Serotonin reduction in post-acute sequelae of viral infection.

Post-acute sequelae of COVID-19 (PASC, "Long COVID") pose a significant global health challenge. The pathophysiology is unknown, and no effective treatments have been found to date. Several hypotheses have been formulated to explain the etiology of PASC, including viral persistence, chronic inflammation, hypercoagulability, and autonomic dysfunction. Here, we propose a mechanism that links all four hypotheses in a single pathway and provides actionable insights for therapeutic interventions. We find that PASC are associated with serotonin reduction. Viral infection and type I interferon-driven inflammation reduce serotonin through three mechanisms: diminished intestinal absorption of the serotonin precursor tryptophan; platelet hyperactivation and thrombocytopenia, which impacts serotonin storage; and enhanced MAO-mediated serotonin turnover. Peripheral serotonin reduction, in turn, impedes the activity of the vagus nerve and thereby impairs hippocampal responses and memory. These findings provide a possible explanation for neurocognitive symptoms associated with viral persistence in Long COVID, which may extend to other post-viral syndromes.

A microbiome-dependent gut-brain pathway regulates motivation for exercise.

Exercise exerts a wide range of beneficial effects for healthy physiology1. However, the mechanisms regulating an individual's motivation to engage in physical activity remain incompletely understood. An important factor stimulating the engagement in both competitive and recreational exercise is the motivating pleasure derived from prolonged physical activity, which is triggered by exercise-induced neurochemical changes in the brain. Here, we report on the discovery of a gut-brain connection in mice that enhances exercise performance by augmenting dopamine signalling during physical activity. We find that microbiome-dependent production of endocannabinoid metabolites in the gut stimulates the activity of TRPV1-expressing sensory neurons and thereby elevates dopamine levels in the ventral striatum during exercise. Stimulation of this pathway improves running performance, whereas microbiome depletion, peripheral endocannabinoid receptor inhibition, ablation of spinal afferent neurons or dopamine blockade abrogate exercise capacity. These findings indicate that the rewarding properties of exercise are influenced by gut-derived interoceptive circuits and provide a microbiome-dependent explanation for interindividual variability in exercise performance. Our study also suggests that interoceptomimetic molecules that stimulate the transmission of gut-derived signals to the brain may enhance the motivation for exercise.

Survey of spiking in the mouse visual system reveals functional hierarchy.

The anatomy of the mammalian visual system, from the retina to the neocortex, is organized hierarchically1. However, direct observation of cellular-level functional interactions across this hierarchy is lacking due to the challenge of simultaneously recording activity across numerous regions. Here we describe a large, open dataset-part of the Allen Brain Observatory2-that surveys spiking from tens of thousands of units in six cortical and two thalamic regions in the brains of mice responding to a battery of visual stimuli. Using cross-correlation analysis, we reveal that the organization of inter-area functional connectivity during visual stimulation mirrors the anatomical hierarchy from the Allen Mouse Brain Connectivity Atlas3. We find that four classical hierarchical measures-response latency, receptive-field size, phase-locking to drifting gratings and response decay timescale-are all correlated with the hierarchy. Moreover, recordings obtained during a visual task reveal that the correlation between neural activity and behavioural choice also increases along the hierarchy. Our study provides a foundation for understanding coding and signal propagation across hierarchically organized cortical and thalamic visual areas.

Liquid and Hydrogel Phases of PrPC Linked to Conformation Shifts and Triggered by Alzheimer's Amyloid-ß Oligomers.

Protein phase separation by low-complexity, intrinsically disordered domains generates membraneless organelles and links to neurodegeneration. Cellular prion protein (PrPC) contains such domains, causes spongiform degeneration, and is a receptor for Alzheimer's amyloid-ß oligomers (Aßo). Here, we show that PrPC separates as a liquid phase, in which α-helical Thr become unfolded. At the cell surface, PrPC Lys residues interact with Aßo to create a hydrogel containing immobile Aßo and relatively mobile PrPC. The Aßo/PrP hydrogel has a well-defined stoichiometry and dissociates with excess Aßo. NMR studies of hydrogel PrPC reveal a distinct α-helical conformation for natively unfolded amino-terminal Gly and Ala residues. Aßo/PrP hydrogel traps signal-transducing mGluR5 on the plasma membrane. Recombinant PrPC extracts endogenous Aßo from human Alzheimer's soluble brain lysates into hydrogel, and a PrPC antagonist releases Aßo from endogenous brain hydrogel. Thus, coupled phase and conformational transitions of PrPC are driven by Aß species from Alzheimer's disease.

Gaucher disease protects against tuberculosis.

Biallelic mutations in the glucocerebrosidase (GBA1) gene cause Gaucher disease, characterized by lysosomal accumulation of glucosylceramide and glucosylsphingosine in macrophages. Gaucher and other lysosomal diseases occur with high frequency in Ashkenazi Jews. It has been proposed that the underlying mutations confer a selective advantage, in particular conferring protection against tuberculosis. Here, using a zebrafish Gaucher disease model, we find that the mutation GBA1 N370S, predominant among Ashkenazi Jews, increases resistance to tuberculosis through the microbicidal activity of glucosylsphingosine in macrophage lysosomes. Consistent with lysosomal accumulation occurring only in homozygotes, heterozygotes remain susceptible to tuberculosis. Thus, our findings reveal a mechanistic basis for protection against tuberculosis by GBA1 N370S and provide biological plausibility for its selection if the relatively mild deleterious effects in homozygotes were offset by significant protection against tuberculosis, a rampant killer of the young in Europe through the Middle Ages into the 19th century.

Upregulation of non-canonical and canonical inflammasome genes associates with pathological features in Krabbe disease and related disorders.

Infantile Krabbe disease is a rapidly progressive and fatal disorder of myelin, caused by inherited deficiency of the lysosomal enzyme ß-galactocerebrosidase. Affected children lose their motor skills and other faculties; uncontrolled seizures are a frequent terminal event. Overexpression of the sphingolipid metabolite psychosine is a pathogenic factor, but does not fully account for the pleiotropic manifestations and there is a clear need to investigate additional pathological mechanisms. We examined innate immunity, caspase-11 and associated inflammatory pathways in twitcher mice, an authentic model of Krabbe disease. Combined use of molecular tools, RNAscope in situ hybridization and immunohistochemical staining established that the expression of pro-inflammatory non-canonical caspase-11, canonical caspase-1, gasdermin D and cognate genes is induced in nervous tissue. Early onset and progressive upregulation of these genes accompany demyelination and gliosis and although the molecules are scant in healthy tissue, abundance of the respective translation products is greatly increased in diseased animals. Caspase-11 is found in reactive microglia/macrophages as well as astrocytes but caspase-1 and gasdermin D are restricted to reactive microglia/macrophages. The inflammasome signature is not unique to Krabbe disease; to varying degrees, this signature is also prominent in other lysosomal diseases, Sandhoff and Niemann-Pick Type-C1, and the lysolecithin toxin model of focal demyelination. Given the potent inflammatory response here identified in Krabbe disease and the other neurodegenerative disorders studied, a broad induction of inflammasomes is likely to be a dominant factor in the pathogenesis, and thus represents a platform for therapeutic exploration.

A dyadic approach to the delineation of diagnostic entities in clinical genomics.

The delineation of disease entities is complex, yet recent advances in the molecular characterization of diseases provide opportunities to designate diseases in a biologically valid manner. Here, we have formalized an approach to the delineation of Mendelian genetic disorders that encompasses two distinct but inter-related concepts: (1) the gene that is mutated and (2) the phenotypic descriptor, preferably a recognizably distinct phenotype. We assert that only by a combinatorial or dyadic approach taking both of these attributes into account can a unitary, distinct genetic disorder be designated. We propose that all Mendelian disorders should be designated as "GENE-related phenotype descriptor" (e.g., "CFTR-related cystic fibrosis"). This approach to delineating and naming disorders reconciles the complexity of gene-to-phenotype relationships in a simple and clear manner yet communicates the complexity and nuance of these relationships.

GATA3 is essential for separating patterning domains during facial morphogenesis.

Neural crest cells (NCCs) within the mandibular and maxillary prominences of the first pharyngeal arch are initially competent to respond to signals from either region. However, mechanisms that are only partially understood establish developmental tissue boundaries to ensure spatially correct patterning. In the 'hinge and caps' model of facial development, signals from both ventral prominences (the caps) pattern the adjacent tissues whereas the intervening region, referred to as the maxillomandibular junction (the hinge), maintains separation of the mandibular and maxillary domains. One cap signal is GATA3, a member of the GATA family of zinc-finger transcription factors with a distinct expression pattern in the ventral-most part of the mandibular and maxillary portions of the first arch. Here, we show that disruption of Gata3 in mouse embryos leads to craniofacial microsomia and syngnathia (bony fusion of the upper and lower jaws) that results from changes in BMP4 and FGF8 gene regulatory networks within NCCs near the maxillomandibular junction. GATA3 is thus a crucial component in establishing the network of factors that functionally separate the upper and lower jaws during development.

Venglustat combined with imiglucerase for neurological disease in adults with Gaucher disease type 3: the LEAP trial.

Gaucher disease type 3 is a chronic neuronopathic disorder with wide-ranging effects, including hepatosplenomegaly, anaemia, thrombocytopenia, skeletal disease and diverse neurological manifestations. Biallelic mutations in GBA1 reduce lysosomal acid ß-glucosidase activity, and its substrates, glucosylceramide and glucosylsphingosine, accumulate. Enzyme replacement therapy and substrate reduction therapy ameliorate systemic features of Gaucher disease, but no therapies are approved for neurological manifestations. Venglustat is an investigational, brain-penetrant, glucosylceramide synthase inhibitor with potential to improve the disease by rebalancing influx of glucosylceramide with impaired lysosomal recycling. The Phase 2, open-label LEAP trial (NCT02843035) evaluated orally administered venglustat 15 mg once-daily in combination with maintenance dose of imiglucerase enzyme replacement therapy during 1 year of treatment in 11 adults with Gaucher disease type 3. Primary endpoints were venglustat safety and tolerability and change in concentration of glucosylceramide and glucosylsphingosine in CSF from baseline to Weeks 26 and 52. Secondary endpoints included change in plasma concentrations of glucosylceramide and glucosylsphingosine, venglustat pharmacokinetics in plasma and CSF, neurologic function, infiltrative lung disease and systemic disease parameters. Exploratory endpoints included changes in brain volume assessed with volumetric MRI using tensor-based morphometry, and resting functional MRI analysis of regional brain activity and connectivity between resting state networks. Mean (SD) plasma venglustat AUC0-24 on Day 1 was 851 (282) ng•h/ml; Cmax of 58.1 (26.4) ng/ml was achieved at a median tmax 2.00 h. After once-daily venglustat, plasma concentrations (4 h post-dose) were higher compared with Day 1, indicating ∼2-fold accumulation. One participant (Patient 9) had low-to-undetectable venglustat exposure at Weeks 26 and 52. Based on mean plasma and CSF venglustat concentrations (excluding Patient 9), steady state appeared to be reached on or before Week 4. Mean (SD) venglustat concentration at Week 52 was 114 (65.8) ng/ml in plasma and 6.14 (3.44) ng/ml in CSF. After 1 year of treatment, median (inter-quartile range) glucosylceramide decreased 78% (72, 84) in plasma and 81% (77, 83) in CSF; median (inter-quartile range) glucosylsphingosine decreased 56% (41, 60) in plasma and 70% (46, 76) in CSF. Ataxia improved slightly in nine patients: mean (SD, range) total modified Scale for Assessment and Rating of Ataxia score decreased from 2.68 [1.54 (0.0 to 5.5)] at baseline to 1.55 [1.88 (0.0 to 5.0)] at Week 52 [mean change: -1.14 (95% CI: -2.06 to -0.21)]. Whole brain volume increased slightly in patients with venglustat exposure and biomarker reduction in CSF (306.7 ± 4253.3 mm3) and declined markedly in Patient 9 (-13894.8 mm3). Functional MRI indicated stronger connectivity at Weeks 26 and 52 relative to baseline between a broadly distributed set of brain regions in patients with venglustat exposure and biomarker reduction but not Patient 9, although neurocognition, assessed by Vineland II, deteriorated in all domains over time, which illustrates disease progression despite the intervention. There were no deaths, serious adverse events or discontinuations. In adults with Gaucher disease type 3 receiving imiglucerase, addition of once-daily venglustat showed acceptable safety and tolerability and preliminary evidence of clinical stability with intriguing but intrinsically inconsistent signals in selected biomarkers, which need to be validated and confirmed in future research.

Hypomyelination, hypodontia and craniofacial abnormalities in a Polr3b mouse model of leukodystrophy.

RNA polymerase III (Pol III)-related hypomyelinating leukodystrophy (POLR3-HLD), also known as 4H leukodystrophy, is a severe neurodegenerative disease characterized by the cardinal features of hypomyelination, hypodontia and hypogonadotropic hypogonadism. POLR3-HLD is caused by biallelic pathogenic variants in genes encoding Pol III subunits. While approximately half of all patients carry mutations in POLR3B encoding the RNA polymerase III subunit B, there is no in vivo model of leukodystrophy based on mutation of this Pol III subunit. Here, we determined the impact of POLR3BΔ10 (Δ10) on Pol III in human cells and developed and characterized an inducible/conditional mouse model of leukodystrophy using the orthologous Δ10 mutation in mice. The molecular mechanism of Pol III dysfunction was determined in human cells by affinity purification-mass spectrometry and western blot. Postnatal induction with tamoxifen induced expression of the orthologous Δ10 hypomorph in triple transgenic Pdgfrα-Cre/ERT; R26-Stopfl-EYFP; Polr3bfl mice. CNS and non-CNS features were characterized using a variety of techniques including microCT, ex vivo MRI, immunofluorescence, immunohistochemistry, spectral confocal reflectance microscopy and western blot. Lineage tracing and time series analysis of oligodendrocyte subpopulation dynamics based on co-labelling with lineage-specific and/or proliferation markers were performed. Proteomics suggested that Δ10 causes a Pol III assembly defect, while western blots demonstrated reduced POLR3BΔ10 expression in the cytoplasm and nucleus in human cells. In mice, postnatal Pdgfrα-dependent expression of the orthologous murine mutant protein resulted in recessive phenotypes including severe hypomyelination leading to ataxia, tremor, seizures and limited survival, as well as hypodontia and craniofacial abnormalities. Hypomyelination was confirmed and characterized using classic methods to quantify myelin components such as myelin basic protein and lipids, results which agreed with those produced using modern methods to quantify myelin based on the physical properties of myelin membranes. Lineage tracing uncovered the underlying mechanism for the hypomyelinating phenotype: defective oligodendrocyte precursor proliferation and differentiation resulted in a failure to produce an adequate number of mature oligodendrocytes during postnatal myelinogenesis. In summary, we characterized the Polr3bΔ10 mutation and developed an animal model that recapitulates features of POLR3-HLD caused by POLR3B mutations, shedding light on disease pathogenesis, and opening the door to the development of therapeutic interventions.

Homozygosity for a Rare Plec Variant Suggests a Contributory Role in Congenital Insensitivity to Pain.

Congenital insensitivity to pain is a rare human condition in which affected individuals do not experience pain throughout their lives. This study aimed to identify the molecular etiology of congenital insensitivity to pain in two Thai patients. Clinical, radiographic, histopathologic, immunohistochemical, and molecular studies were performed. Patients were found to have congenital insensitivity to pain, self-mutilation, acro-osteolysis, cornea scars, reduced temperature sensation, tooth agenesis, root maldevelopment, and underdeveloped maxilla and mandible. The skin biopsies revealed fewer axons, decreased vimentin expression, and absent neurofilament expression, indicating lack of dermal nerves. Whole exome and Sanger sequencing identified a rare homozygous variant c.4039C>T; p.Arg1347Cys in the plakin domain of Plec, a cytolinker protein. This p.Arg1347Cys variant is in the spectrin repeat 9 region of the plakin domain, a region not previously found to harbor pathogenic missense variants in other plectinopathies. The substitution with a cysteine is expected to decrease the stability of the spectrin repeat 9 unit of the plakin domain. Whole mount in situ hybridization and an immunohistochemical study suggested that Plec is important for the development of maxilla and mandible, cornea, and distal phalanges. Additionally, the presence of dental anomalies in these patients further supports the potential involvement of Plec in tooth development. This is the first report showing the association between the Plec variant and congenital insensitivity to pain in humans.

Genetic Variants in KCTD1 Are Associated with Isolated Dental Anomalies.

KCTD1 plays crucial roles in regulating both the SHH and WNT/ß-catenin signaling pathways, which are essential for tooth development. The objective of this study was to investigate if genetic variants in KCTD1 might also be associated with isolated dental anomalies. We clinically and radiographically investigated 362 patients affected with isolated dental anomalies. Whole exome sequencing identified two unrelated families with rare (p.Arg241Gln) or novel (p.Pro243Ser) variants in KCTD1. The variants segregated with the dental anomalies in all nine patients from the two families. Clinical findings of the patients included taurodontism, unseparated roots, long roots, tooth agenesis, a supernumerary tooth, torus palatinus, and torus mandibularis. The role of Kctd1 in root development is supported by our immunohistochemical study showing high expression of Kctd1 in Hertwig epithelial root sheath. The KCTD1 variants in our patients are the first variants found to be located in the C-terminal domain, which might disrupt protein-protein interactions and/or SUMOylation and subsequently result in aberrant WNT-SHH-BMP signaling and isolated dental anomalies. Functional studies on the p.Arg241Gln variant are consistent with an impact on ß-catenin levels and canonical WNT signaling. This is the first report of the association of KCTD1 variants and isolated dental anomalies.

Transcriptomic analysis reveals the role of SIX1 in mouse cranial neural crest patterning and bone development.

BACKGROUND: Genetic variants of the transcription factor SIX1 and its co-factor EYA1 underlie 50% of Branchio-oto-renal syndrome (BOR) cases. BOR is characterized by craniofacial defects, including malformed middle ear ossicles leading to conductive hearing loss. In this work, we expand our knowledge of the Six1 gene regulatory network by using a Six1-null mouse line to assess gene expression profiles of E10.5 mandibular arches, which give rise to the neural crest (NC)-derived middle ear ossicles and lower jaw, via bulk RNA sequencing. RESULTS: Our transcriptomic analysis led to the identification of 808 differentially expressed genes that are related to translation, NC cell differentiation, osteogenesis, and chondrogenesis including components of the WNT signaling pathway. As WNT signaling is a known contributor to bone development, we demonstrated that SIX1 is required for expression of the WNT antagonist Frzb in the mandibular arch, and determined that SIX1 expression results in repression of WNT signaling. CONCLUSION: Our results clarify the mechanisms by which SIX1 regulates the development of NC-derived craniofacial elements that are altered in SIX1-associated disorders. In addition, this work identifies novel genes that could be causative to this birth defect and establishes a link between SIX1 and WNT signaling during patterning of NC cells.

Expression of Ripk1 and DAM genes correlates with severity and progression of Krabbe disease.

Krabbe disease, an inherited leukodystrophy, is a sphingolipidosis caused by deficiency of ß-galactocerebrosidase: it is characterized by myelin loss, and pathological activation of macrophage/microglia and astrocytes. To define driving pathogenic factors, we explored the expression repertoire of candidate neuroinflammatory genes: upregulation of receptor interacting protein kinase 1 (Ripk1) and disease-associated microglia (DAM) genes, including Cst7 and Ch25h, correlated with severity of Krabbe disease genetically modelled in the twitcher mouse. Upregulation of Ripk1 in Iba1/Mac2-positive microglia/macrophage associated with the pathognomic hypertrophic/globoid phenotype of this disease. Widespread accumulation of ubiquitinin1 in white and grey matter co-localised with p62. In Sandhoff disease, another sphingolipid disorder, neuroinflammation, accumulation of p62 and increased Ripk1 expression was observed. The upregulated DAM genes and macrophage/microglia expression of Ripk1 in the authentic model of Krabbe disease strongly resemble those reported in Alzheimer disease associating with disturbed autophagosomal/lysosomal homeostasis. Activation of this shared molecular repertoire, suggests the potential for therapeutic interdiction at a common activation step, irrespective of proximal causation. To clarify the role of Ripk1 in the pathogenesis of Krabbe disease, we first explored the contribution of its kinase function, by intercrossing twitcher and the K45A kinase-dead Ripk1 mouse and breeding to homozygosity. Genetic ablation of Ripk1 kinase activity neither altered the neuropathological features nor the survival of twitcher mice. We conclude that Ripk1 kinase-dependent inflammatory and degenerative capabilities play no instrumental role in Krabbe disease; however, putative kinase-independent functions of Ripk1 remain formally to be explored in its molecular pathogenesis.

Radiographic Cortical Thickness Index Predicts Fragility Fracture in Gaucher Disease.

Background Patients with Gaucher disease (GD) have a high risk of fragility fractures. Routine evaluation of bone involvement in these patients includes radiography and repeated dual-energy x-ray absorptiometry (DXA). However, osteonecrosis and bone fracture may affect the accuracy of DXA. Purpose To assess the utility of DXA and radiographic femoral cortical thickness measurements as predictors of fragility fracture in patients with GD with long-term follow-up (up to 30 years). Materials and Methods Patients with GD age 16 years and older with a detailed medical history, at least one bone image (DXA and/or radiographs), and minimum 2 years follow-up were retrospectively identified using three merged UK-based registries (Gaucherite study, enrollment 2015-2017; Clinical Bone Registry, enrollment 2003-2006; and Mortality Registry, enrollment 1993-2019). Cortical thickness index (CTI) and canal-to-calcar ratio (CCR) were measured by two independent observers, and inter- and intraobserver reliability was calculated. The fracture-predictive value of DXA, CTI, CCR, and cutoff values were calculated using receiver operating characteristic curves. Statistical differences were assessed using univariable and multivariable analysis. Results Bone imaging in 247 patients (123 men, 124 women; baseline median age, 39 years; IQR, 27-50 years) was reviewed. The median follow-up period was 11 years (IQR, 7-19 years; range, 2-30 years). Thirty-five patients had fractures before or at first bone imaging, 23 patients had fractures after first bone imaging, and 189 patients remained fracture-free. Inter- and intraobserver reproducibility for CTI/CCR measurements was substantial (range, 0.96-0.98). In the 212 patients with no baseline fracture, CTI (cutoff, ≤0.50) predicted future fractures with higher sensitivity and specificity (area under the receiver operating characteristic curve [AUC], 0.96; 95% CI: 0.84, 0.99; sensitivity, 92%; specificity, 96%) than DXA T-score at total hip (AUC, 0.78; 95% CI: 0.51, 0.91; sensitivity, 64%; specificity, 93%), femoral neck (AUC, 0.73; 95% CI: 0.50, 0.86; sensitivity, 64%; specificity, 73%), lumbar spine (AUC, 0.69; 95% CI: 0.49, 0.82; sensitivity, 57%; specificity, 63%), and forearm (AUC, 0.78; 95% CI: 0.59, 0.89; sensitivity, 70%; specificity, 70%). Conclusion Radiographic cortical thickness index of 0.50 or less was a reliable independent predictor of fracture risk in Gaucher disease. Clinical trial registration no. NCT03240653 © RSNA, 2022 Supplemental material is available for this article.

The spontaneous mouse mutant low set ears (Lse) is caused by tandem duplication of Fgf3 and Fgf4.

The external ear develops from an organized convergence of ventrally migrating neural crest cells into the first and second branchial arches. Defects in external ear position are often symptomatic of complex syndromes such as Apert, Treacher-Collins, and Crouzon Syndrome. The low set ears (Lse) spontaneous mouse mutant is characterized by the dominant inheritance of a ventrally shifted external ear position and an abnormal external auditory meatus (EAM). We identified the causative mutation as a 148 Kb tandem duplication on Chromosome 7, which includes the entire coding sequences of Fgf3 and Fgf4. Duplications of FGF3 and FGF4 occur in 11q duplication syndrome in humans and are associated with craniofacial anomalies, among other features. Intercrosses of Lse-affected mice revealed perinatal lethality in homozygotes, and Lse/Lse embryos display additional phenotypes including polydactyly, abnormal eye morphology, and cleft secondary palate. The duplication results in increased Fgf3 and Fgf4 expression in the branchial arches and additional discrete domains in the developing embryo. This ectopic overexpression resulted in functional FGF signaling, demonstrated by increased Spry2 and Etv5 expression in overlapping domains of the developing arches. Finally, a genetic interaction between Fgf3/4 overexpression and Twist1, a regulator of skull suture development, resulted in perinatal lethality, cleft palate, and polydactyly in compound heterozygotes. These data indicate a role for Fgf3 and Fgf4 in external ear and palate development and provide a novel mouse model for further interrogation of the biological consequences of human FGF3/4 duplication.

Long-term effects of eliglustat on skeletal manifestations in clinical trials of patients with Gaucher disease type 1.

PURPOSE: Most patients with Gaucher disease have progressive and often disabling skeletal manifestations. We examined the long-term effect of eliglustat treatment on bone outcomes in clinical trials in adults with Gaucher disease type 1. METHODS: Data from 4 completed phase 2 and 3 trials were evaluated in treatment-naïve patients or patients switching to eliglustat from enzyme replacement therapy (ERT). RESULTS: Overall, 319 of 393 (81%) eliglustat-treated patients remained in their trials until completion or commercial eliglustat became available. Mean eliglustat treatment duration ranged from 3.3 to 6.5 years. In treatment-naïve patients and ERT-switch patients, frequency and severity of bone pain decreased during eliglustat treatment. Mean lumbar spine T-scores shifted from abnormal to normal in treatment-naïve patients and remained in the healthy reference range or improved modestly in ERT-switch patients. Mean total bone marrow burden score shifted from marked-to-severe to moderate in treatment-naïve patients and remained moderate in ERT-switch patients. MIP-1ß (marker of active bone disease) was elevated at baseline and decreased to the healthy reference range in treatment-naïve patients and remained in the healthy reference range among ERT-switch patients. CONCLUSION: These findings confirm the long-term efficacy of eliglustat on skeletal complications of Gaucher disease in treatment-naïve and ERT-switch patients.

A mutational hotspot in AMOTL1 defines a new syndrome of orofacial clefting, cardiac anomalies, and tall stature.

AMOTL1 encodes angiomotin-like protein 1, an actin-binding protein that regulates cell polarity, adhesion, and migration. The role of AMOTL1 in human disease is equivocal. We report a large cohort of individuals harboring heterozygous AMOTL1 variants and define a core phenotype of orofacial clefting, congenital heart disease, tall stature, auricular anomalies, and gastrointestinal manifestations in individuals with variants in AMOTL1 affecting amino acids 157-161, a functionally undefined but highly conserved region. Three individuals with AMOTL1 variants outside this region are also described who had variable presentations with orofacial clefting and multi-organ disease. Our case cohort suggests that heterozygous missense variants in AMOTL1, most commonly affecting amino acid residues 157-161, define a new orofacial clefting syndrome, and indicates an important functional role for this undefined region.

Altering osteoclast numbers using CTSK models in utero affects mice offspring craniofacial morphology.

BACKGROUND: Bone remodelling during development and growth is important for craniofacial integrity of offspring. The aim of this study was to investigate the changes in offspring adult skull morphology when the osteoclasts number was altered in utero, using three-dimensional (3D) geometric morphometric analysis (GMA). MATERIALS AND METHODS: We altered osteoclasts number in utero via two approaches. First, we generated heterozygous CtskCre ;DTAfl/+ (diphtheria toxin A) mice. Second, we altered Ctsk expression in vivo by injecting pregnant wild-type dams at embryonic day (E) 12.5 with in vivo siRNA specific for Ctsk. Mice were collected at 6 weeks and analysed using geometric morphometric analysis via computed tomography, histomorphometry and gene expression analysis. RESULTS: Altering osteoclasts number in utero affected the offspring adult skull morphology. Decreased Ctsk and osteoclast numbers were associated with a decrease in cranial vault height and an increase in mandibular body length. Changes in size and shape were observed with an increased number of osteoclasts in CtskCre ;DTAfl/+ mice, including an increase in cranial vault height, as well as a shortening of mandibular body length and ramus height. CONCLUSION: The findings of this study suggest that modulation of osteoclast numbers during pre- and post-natal development may be a previously unknown factor in the aetiology of skeletal malocclusions. An improved understanding of the factors affecting bone homeostasis during development and growth may help in the development of future therapies that would target the early intervention of skeletal malocclusion.

Altering maternal calcium and phosphorus dietary intake induces persistent sex-specific changes in the dentition of the offspring.

BACKGROUND: The maternal diet is essential to offspring development, but the specific effects on tooth morphology are still unknown. The aim of this study was to evaluate the effects of altering maternal calcium (Ca) and phosphorus (P) supplementation during gestation and lactation on offspring dentition. METHODS: Pregnant mice were fed an experimental diet containing a threefold increase in Ca and a threefold decrease in P compared to the standard mouse chow diet at embryonic Day 0.5 (E0.5). Offspring mice were maintained on standard or experimental diets from post-natal Day 0 to weaning, then fed control diets until 6 weeks of age. Six-week-old offspring heads were collected and scanned using micro-computed tomography. Dental morphometrics of offspring maxillary and mandibular first and third molars (n = 5-6 per diet/per sex) were determined. A two-way ANOVA test was employed to verify the existence of any significant differences between groups. The significance level was set at P < .05. RESULTS: A two-way ANOVA revealed a statistically significant interaction between the effects of diet and sex on the upper and lower dentition. Moreover, experimental diet-fed female offspring exhibited smaller molars with shorter mesiodistal width and larger pulp chambers relative to controls, while experimental diet-fed male offspring possessed larger molars with wider mesiodistal width and smaller pulp chambers. CONCLUSION: Our findings reveal that altering the maternal and offspring dietary Ca:P ratio during gestation, lactation and weaning led to significant, sex-specific changes in the offspring dentition. The differences in dentition appeared to be correlated with the sex-specific changes in the craniofacial skeleton.