Publications by authors named "Sarah E Smith"

79 Publications

Repeated evolution of circadian clock dysregulation in cavefish populations.

PLoS Genet 2021 Jul 12;17(7):e1009642. Epub 2021 Jul 12.

Ecology, Evolution, and Behavior, University of Minnesota, Saint Paul, Minnesota, United States of America.

Circadian rhythms are nearly ubiquitous throughout nature, suggesting they are critical for survival in diverse environments. Organisms inhabiting largely arrhythmic environments, such as caves, offer a unique opportunity to study the evolution of circadian rhythms in response to changing ecological pressures. Populations of the Mexican tetra, Astyanax mexicanus, have repeatedly invaded caves from surface rivers, where individuals must contend with perpetual darkness, reduced food availability, and limited fluctuations in daily environmental cues. To investigate the molecular basis for evolved changes in circadian rhythms, we investigated rhythmic transcription across multiple independently-evolved cavefish populations. Our findings reveal that evolution in a cave environment has led to the repeated disruption of the endogenous biological clock, and its entrainment by light. The circadian transcriptome shows widespread reductions and losses of rhythmic transcription and changes to the timing of the activation/repression of core-transcriptional clock. In addition to dysregulation of the core clock, we find that rhythmic transcription of the melatonin regulator aanat2 and melatonin rhythms are disrupted in cavefish under darkness. Mutants of aanat2 and core clock gene rorca disrupt diurnal regulation of sleep in A. mexicanus, phenocopying circadian modulation of sleep and activity phenotypes of cave populations. Together, these findings reveal multiple independent mechanisms for loss of circadian rhythms in cavefish populations and provide a platform for studying how evolved changes in the biological clock can contribute to variation in sleep and circadian behavior.
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http://dx.doi.org/10.1371/journal.pgen.1009642DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8297936PMC
July 2021

Adults with Cerebral Palsy Require Ongoing Neurologic Care: A Systematic Review.

Ann Neurol 2021 05 26;89(5):860-871. Epub 2021 Feb 26.

Department of Neurology, Division of Pediatric Neurology, Washington University School of Medicine, St Louis, MO, USA.

Cerebral palsy (CP) neurologic care and research efforts typically focus on children. However, most people with CP are adults. Adults with CP are at increased risk of new neurologic conditions, such as stroke and myelopathy, that require ongoing neurologic surveillance to distinguish them from baseline motor impairments. Neurologic factors could also contribute to the motor function decline, chronic pain, and chronic fatigue that are commonly experienced by adults with CP. Based on a systematic literature review, we suggest (1) guidelines for neurologic surveillance and neurologist referral and (2) clinical research questions regarding the evolving neurologic risks for adults with CP. ANN NEUROL 2021;89:860-871.
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http://dx.doi.org/10.1002/ana.26040DOI Listing
May 2021

A computational method for immune repertoire mining that identifies novel binders from different clonotypes, demonstrated by identifying anti-pertussis toxoid antibodies.

MAbs 2021 Jan-Dec;13(1):1869406

Department of Statistics, University of Oxford , Oxford, UK.

Due to their shared genetic history, antibodies from the same clonotype often bind to the same epitope. This knowledge is used in immune repertoire mining, where known binders are used to search bulk sequencing repertoires to identify new binders. However, current computational methods cannot identify epitope convergence between antibodies from different clonotypes, limiting the sequence diversity of antigen-specific antibodies that can be identified. We describe how the antibody binding site, the paratope, can be used to cluster antibodies with common antigen reactivity from different clonotypes. Our method, paratyping, uses the predicted paratope to identify these novel cross clonotype matches. We experimentally validated our predictions on a pertussis toxoid dataset. Our results show that even the simplest abstraction of the antibody binding site, using only the length of the loops involved and predicted binding residues, is sufficient to group antigen-specific antibodies and provide additional information to conventional clonotype analysis. : BCR: B-cell receptor; CDR: complementarity-determining region; PTx: pertussis toxoid.
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http://dx.doi.org/10.1080/19420862.2020.1869406DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7808390PMC
January 2021

Gait features of dystonia in cerebral palsy.

Dev Med Child Neurol 2021 Jun 7;63(6):748-754. Epub 2021 Jan 7.

Division of Pediatric Neurology, Department of Neurology, Washington University School of Medicine and St. Louis Children's Hospital, St. Louis, MO, USA.

Aim: To determine the features cited by motor phenotyping experts when identifying dystonia in people with cerebral palsy (CP).

Method: Dystonia identification in CP, particularly when comorbid with spasticity, can be difficult. The dystonia diagnostic criterion standard remains subjective visual identification by expert consensus. For this qualitative study, we conducted an inductive thematic analysis of consensus-building discussions between three pediatric movement disorder physicians as they identified the presence or absence of dystonia in gait videos of 40 participants with spastic CP and periventricular leukomalacia.

Results: Unanimous consensus about the presence or absence of dystonia was achieved for 34 out of 40 videos. Two main themes were present during consensus-building discussions as videos were evaluated for dystonia: (1) unilateral leg or foot adduction that was variable over time, and (2) difficulty in identifying dystonia. Codes contributing to the first theme were more likely to be cited by a discussant when they felt dystonia was present (as opposed to absent) in a video (χ test, p=0.004).

Discussion: These results describe the gait features cited by experts during consensus-building discussion as they identify dystonia in ambulatory people with CP. Qualitative thematic analysis of these discussions could help codify the subjective process of dystonia diagnosis.
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http://dx.doi.org/10.1111/dmcn.14802DOI Listing
June 2021

Astrocyte deletion of α2-Na/K ATPase triggers episodic motor paralysis in mice via a metabolic pathway.

Nat Commun 2020 12 2;11(1):6164. Epub 2020 Dec 2.

Department of Neuroscience, Washington University School of Medicine, St. Louis, MO, 63110, USA.

Familial hemiplegic migraine is an episodic neurological disorder characterized by transient sensory and motor symptoms and signs. Mutations of the ion pump α2-Na/K ATPase cause familial hemiplegic migraine, but the mechanisms by which α2-Na/K ATPase mutations lead to the migraine phenotype remain incompletely understood. Here, we show that mice in which α2-Na/K ATPase is conditionally deleted in astrocytes display episodic paralysis. Functional neuroimaging reveals that conditional α2-Na/K ATPase knockout triggers spontaneous cortical spreading depression events that are associated with EEG low voltage activity events, which correlate with transient motor impairment in these mice. Transcriptomic and metabolomic analyses show that α2-Na/K ATPase loss alters metabolic gene expression with consequent serine and glycine elevation in the brain. A serine- and glycine-free diet rescues the transient motor impairment in conditional α2-Na/K ATPase knockout mice. Together, our findings define a metabolic mechanism regulated by astrocytic α2-Na/K ATPase that triggers episodic motor paralysis in mice.
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http://dx.doi.org/10.1038/s41467-020-19915-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7710756PMC
December 2020

In Situ Hematopoietic Stem Cell Imaging.

Methods Mol Biol 2021 ;2185:373-382

Institute for Stem Cell and Regenerative Medicine Research, Albert Einstein College of Medicine, Bronx, NY, USA.

Although immunohistochemistry of tissue sections has been the gold standard for analyzing tissue structure and cellular localization, this approach has significant shortcomings when it comes to analyzing complex and heterogeneous tissues such as the bone marrow with rare cells like hematopoietic stem cells (HSCs). Hence, studying rare cells and their relationship with the surrounding heterogenous microenvironment requires visualization of specifically labeled cells within large intact tissues in three dimensions. Here, we describe a whole mount sternal bone marrow imaging method which has enabled detailed quantitative and qualitative analysis of rare HSCs within the sternal tissue. The methodology is broadly applicable for examining the 3D architecture of niche cells in relation to HSCs.
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http://dx.doi.org/10.1007/978-1-0716-0810-4_23DOI Listing
March 2021

A Pesco-Mediterranean Diet With Intermittent Fasting: JACC Review Topic of the Week.

J Am Coll Cardiol 2020 09;76(12):1484-1493

Lipid Clinic, Endocrinology and Nutrition Service, Institut d'Investigacions Biomediques August Pi Sunyer, Hospital Clinic, University of Barcelona, and CIBER Fisiopatología de la Obesidad y Nutrición, Instituto de Salud Carlos III, Madrid, Spain.

As opportunistic omnivores, humans are evolutionarily adapted to obtain calories and nutrients from both plant and animal food sources. Today, many people overconsume animal products, often-processed meats high in saturated fats and chemical additives. Alternatively, strict veganism can cause nutritional deficiencies and predispose to osteopenia, sarcopenia, and anemia. A logical compromise is a plant-rich diet with fish/seafood as principal sources of animal food. This paper reviews cumulative evidence regarding diet and health, incorporating data from landmark clinical trials of the Mediterranean diet and recommendations from recent authoritative guidelines, to support the hypothesis that a Pesco-Mediterranean diet is ideal for optimizing cardiovascular health. The foundation of this diet is vegetables, fruits, nuts, seeds, legumes, whole grains, and extra-virgin olive oil with fish/seafood and fermented dairy products. Beverages of choice are water, coffee, and tea. Time-restricted eating is recommended, whereby intermittent fasting is done for 12 to 16 h each day.
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http://dx.doi.org/10.1016/j.jacc.2020.07.049DOI Listing
September 2020

β-Catenin and Associated Proteins Regulate Lineage Differentiation in Ground State Mouse Embryonic Stem Cells.

Stem Cell Reports 2020 09 20;15(3):662-676. Epub 2020 Aug 20.

Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, MO 64110, USA; University of Kansas Medical Center, Kansas City, KS, USA. Electronic address:

Mouse embryonic stem cells (ESCs) cultured in defined medium resemble the pre-implantation epiblast in the ground state, with full developmental capacity including the germline. β-Catenin is required to maintain ground state pluripotency in mouse ESCs, but its exact role is controversial. Here, we reveal a Tcf3-independent role of β-catenin in restraining germline and somatic lineage differentiation genes. We show that β-catenin binds target genes with E2F6 and forms a complex with E2F6 and HMGA2 or E2F6 and HP1γ. Our data indicate that these complexes help β-catenin restrain and fine-tune germ cell and neural developmental potential. Overall, our data reveal a previously unappreciated role of β-catenin in preserving lineage differentiation integrity in ground state ESCs.
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http://dx.doi.org/10.1016/j.stemcr.2020.07.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7486223PMC
September 2020

A novel therapeutic antibody screening method using bacterial high-content imaging reveals functional antibody binding phenotypes of Escherichia coli ST131.

Sci Rep 2020 07 24;10(1):12414. Epub 2020 Jul 24.

Department of Medicine, Cambridge Institute for Therapeutic Immunology & Infectious Disease, University of Cambridge, Cambridge, UK.

The increase of antimicrobial resistance (AMR), and lack of new classes of licensed antimicrobials, have made alternative treatment options for AMR pathogens increasingly attractive. Recent studies have demonstrated anti-bacterial efficacy of a humanised monoclonal antibody (mAb) targeting the O25b O-antigen of Escherichia coli ST131. To evaluate the phenotypic effects of antibody binding to diverse clinical E. coli ST131 O25b bacterial isolates in high-throughput, we designed a novel mAb screening method using high-content imaging (HCI) and image-based morphological profiling to screen a mAb targeting the O25b O-antigen. Screening the antibody against a panel of 86 clinical E. coli ST131 O25:H4 isolates revealed 4 binding phenotypes: no binding (18.60%), weak binding (4.65%), strong binding (69.77%) and strong agglutinating binding (6.98%). Impaired antibody binding could be explained by the presence of insertion sequences or mutations in O-antigen or lipopolysaccharide core biosynthesis genes, affecting the amount, structure or chain length of the O-antigen. The agglutinating binding phenotype was linked with lower O-antigen density, enhanced antibody-mediated phagocytosis and increased serum susceptibly. This study highlights the need to screen candidate mAbs against large panels of clinically relevant isolates, and that HCI can be used to evaluate mAb binding affinity and potential functional efficacy against AMR bacteria.
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http://dx.doi.org/10.1038/s41598-020-69300-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7382476PMC
July 2020

Changing role of coral reef marine reserves in a warming climate.

Nat Commun 2020 04 24;11(1):2000. Epub 2020 Apr 24.

Department of Biodiversity, Conservation and Attractions, Perth, WA, 6151, Australia.

Coral reef ecosystems are among the first to fundamentally change in structure due to climate change, which leads to questioning of whether decades of knowledge regarding reef management is still applicable. Here we assess ecological responses to no-take marine reserves over two decades, spanning a major climate-driven coral bleaching event. Pre-bleaching reserve responses were consistent with a large literature, with higher coral cover, more species of fish, and greater fish biomass, particularly of upper trophic levels. However, in the 16 years following coral mortality, reserve effects were absent for the reef benthos, and greatly diminished for fish species richness. Positive fish biomass effects persisted, but the groups of fish benefiting from marine reserves profoundly changed, with low trophic level herbivores dominating the responses. These findings highlight that while marine reserves still have important roles on coral reefs in the face of climate change, the species and functional groups they benefit will be substantially altered.
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http://dx.doi.org/10.1038/s41467-020-15863-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181733PMC
April 2020

Pre-treatment loss to follow-up among children with multidrug-resistant tuberculosis in South Africa, 2008-2010.

PLoS One 2020 7;15(4):e0230504. Epub 2020 Apr 7.

U.S. Centers for Disease Control and Prevention, Atlanta, Georgia United States of America.

Multidrug-resistant (MDR) TB is more difficult to diagnose and treat compared with drug-susceptible TB. Young children are at greater risk of severe TB disease and death when treatment is delayed compared to adults. We sought to describe characteristics of children (<13 years) diagnosed with MDR TB between 2008-2010 in three South African provinces and assess factors associated with pre-treatment loss to follow-up. We matched laboratory and medical records at treatment facilities to identify pre-treatment loss and examined demographic and clinical characteristics for association with loss. Categorical variables were examined for association using Pearson's x2 or Fisher's exact test, employing Bonferroni correction for multiple pairwise comparisons. Between 2008-2010, 156 children were diagnosed with laboratory-confirmed MDR TB. Only 44% (n = 69) were documented as having received treatment. Young children (<2 years) (47/59, 80%), children with extrapulmonary (EP) TB (27/34, 79%), and children diagnosed at general hospitals (60/97, 62%) were most likely to be lost before treatment. Children most vulnerable to death from TB are most likely to be lost before treatment, possibly leading to underestimates of disease burden, case notifications, and poor outcomes among this population. Point-of-care diagnosis and robust follow-up may reduce pre-treatment loss in this population.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0230504PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7138297PMC
July 2020

Bat IFITM3 restriction depends on S-palmitoylation and a polymorphic site within the CD225 domain.

Life Sci Alliance 2020 01 11;3(1). Epub 2019 Dec 11.

Medical Research Council Laboratory for Molecular Cell Biology, University College London, London, UK.

Host interferon-induced transmembrane proteins (IFITMs) are broad-spectrum antiviral restriction factors. Of these, IFITM3 potently inhibits viruses that enter cells through acidic endosomes, many of which are zoonotic and emerging viruses with bats (order Chiroptera) as their natural hosts. We previously demonstrated that microbat IFITM3 is antiviral. Here, we show that bat IFITMs are characterized by strong adaptive evolution and identify a highly variable and functionally important site-codon 70-within the conserved CD225 domain of IFITMs. Mutation of this residue in microbat IFITM3 impairs restriction of representatives of four different virus families that enter cells via endosomes. This mutant shows altered subcellular localization and reduced S-palmitoylation, a phenotype copied by mutation of conserved cysteine residues in microbat IFITM3. Furthermore, we show that microbat IFITM3 is S-palmitoylated on cysteine residues C71, C72, and C105, mutation of each cysteine individually impairs virus restriction, and a triple C71A-C72A-C105A mutant loses all restriction activity, concomitant with subcellular re-localization of microbat IFITM3 to Golgi-associated sites. Thus, we propose that S-palmitoylation is critical for Chiropteran IFITM3 function and identify a key molecular determinant of IFITM3 S-palmitoylation.
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http://dx.doi.org/10.26508/lsa.201900542DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6907390PMC
January 2020

Yeast centrosome components form a noncanonical LINC complex at the nuclear envelope insertion site.

J Cell Biol 2019 05 12;218(5):1478-1490. Epub 2019 Mar 12.

Stowers Institute for Medical Research, Kansas City, MO

Bipolar spindle formation in yeast requires insertion of centrosomes (known as spindle pole bodies [SPBs]) into fenestrated regions of the nuclear envelope (NE). Using structured illumination microscopy and bimolecular fluorescence complementation, we map protein distribution at SPB fenestrae and interrogate protein-protein interactions with high spatial resolution. We find that the Sad1-UNC-84 (SUN) protein Mps3 forms a ring-like structure around the SPB, similar to toroids seen for components of the SPB insertion network (SPIN). Mps3 and the SPIN component Mps2 (a Klarsicht-ANC-1-Syne-1 domain [KASH]-like protein) form a novel noncanonical linker of nucleoskeleton and cytoskeleton (LINC) complex that is connected in both luminal and extraluminal domains at the site of SPB insertion. The LINC complex also controls the distribution of a soluble SPIN component Bbp1. Taken together, our work shows that Mps3 is a fifth SPIN component and suggests both direct and indirect roles for the LINC complex in NE remodeling.
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http://dx.doi.org/10.1083/jcb.201809045DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6504903PMC
May 2019

Distribution of Proteins at the Inner Nuclear Membrane Is Regulated by the Asi1 E3 Ligase in .

Genetics 2019 04 1;211(4):1269-1282. Epub 2019 Feb 1.

Stowers Institute for Medical Research, Kansas City, Missouri 64110

Inner nuclear membrane (INM) protein composition regulates nuclear function, affecting processes such as gene expression, chromosome organization, nuclear shape, and stability. Mechanisms that drive changes in the INM proteome are poorly understood, in part because it is difficult to definitively assay INM composition rigorously and systematically. Using a split-GFP complementation system to detect INM access, we examined the distribution of all C-terminally tagged membrane proteins in wild-type cells and in mutants affecting protein quality control pathways, such as INM-associated degradation (INMAD), ER-associated degradation, and vacuolar proteolysis. Deletion of the E3 ligase Asi1 had the most specific effect on the INM compared to mutants in vacuolar or ER-associated degradation pathways, consistent with a role for Asi1 in the INMAD pathway. Our data suggest that Asi1 not only removes mistargeted proteins at the INM, but also controls the levels and distribution of native INM components, such as the membrane nucleoporin Pom33 Interestingly, loss of Asi1 does not affect Pom33 protein levels but instead alters Pom33 distribution in the nuclear envelope through Pom33 ubiquitination, which drives INM redistribution. Taken together, our data demonstrate that the Asi1 E3 ligase has a novel function in INM protein regulation in addition to protein turnover.
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http://dx.doi.org/10.1534/genetics.119.301911DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6456303PMC
April 2019

N-Cadherin-Expressing Bone and Marrow Stromal Progenitor Cells Maintain Reserve Hematopoietic Stem Cells.

Cell Rep 2019 01;26(3):652-669.e6

Stowers Institute for Medical Research, Kansas City, MO 66110, USA; Department of Pathology and Laboratory Medicine, University of Kansas Medical Center, Kansas City, KS 66160, USA. Electronic address:

Regulation of hematopoietic stem cells (HSCs) by bone marrow (BM) niches has been extensively studied; however, whether and how HSC subpopulations are distinctively regulated by BM niches remain unclear. Here, we functionally distinguished reserve HSCs (rHSCs) from primed HSCs (pHSCs) based on their response to chemotherapy and examined how they are dichotomously regulated by BM niches. Both pHSCs and rHSCs supported long-term hematopoiesis in homeostasis; however, pHSCs were sensitive but rHSCs were resistant to chemotherapy. Surviving rHSCs restored the HSC pool and supported hematopoietic regeneration after chemotherapy. The rHSCs were preferentially maintained in the endosteal region that enriches N-cadherin (N-cad) bone-lining cells in homeostasis and post-chemotherapy. N-cad cells were functional bone and marrow stromal progenitor cells (BMSPCs), giving rise to osteoblasts, adipocytes, and chondrocytes in vitro and in vivo. Finally, ablation of N-cad niche cells or deletion of SCF from N-cad niche cells impaired rHSC maintenance during homeostasis and regeneration.
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http://dx.doi.org/10.1016/j.celrep.2018.12.093DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6890378PMC
January 2019

Functional analysis of Salmonella Typhi adaptation to survival in water.

Environ Microbiol 2018 11;20(11):4079-4090

The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, UK.

Contaminated water is a major risk factor associated with the transmission of Salmonella enterica serovar Typhi (S. Typhi), the aetiological agent of human typhoid. However, little is known about how this pathogen adapts to living in the aqueous environment. We used transcriptome analysis (RNA-seq) and transposon mutagenesis (TraDIS) to characterize these adaptive changes and identify multiple genes that contribute to survival. Over half of the genes in the S. Typhi genome altered expression level within the first 24 h following transfer from broth culture to water, although relatively few did so in the first 30 min. Genes linked to central metabolism, stress associated with arrested proton motive force and respiratory chain factors changed expression levels. Additionally, motility and chemotaxis genes increased expression, consistent with a scavenging lifestyle. The viaB-associated gene tviC encoding a glcNAc epimerase that is required for Vi polysaccharide biosynthesis was, along with several other genes, shown to contribute to survival in water. Thus, we define regulatory adaptation operating in S. Typhi that facilitates survival in water.
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http://dx.doi.org/10.1111/1462-2920.14458DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6282856PMC
November 2018

The Transcriptional Regulator SnoN Promotes the Proliferation of Cerebellar Granule Neuron Precursors in the Postnatal Mouse Brain.

J Neurosci 2019 01 13;39(1):44-62. Epub 2018 Nov 13.

Department of Neuroscience, Washington University School of Medicine, St. Louis, Missouri 63110,

Control of neuronal precursor cell proliferation is essential for normal brain development, and deregulation of this fundamental developmental event contributes to brain diseases. Typically, neuronal precursor cell proliferation extends over long periods of time during brain development. However, how neuronal precursor proliferation is regulated in a temporally specific manner remains to be elucidated. Here, we report that conditional KO of the transcriptional regulator SnoN in cerebellar granule neuron precursors robustly inhibits the proliferation of these cells and promotes their cell cycle exit at later stages of cerebellar development in the postnatal male and female mouse brain. In laser capture microdissection followed by RNA-Seq, designed to profile gene expression specifically in the external granule layer of the cerebellum, we find that SnoN promotes the expression of cell proliferation genes and concomitantly represses differentiation genes in granule neuron precursors Remarkably, bioinformatics analyses reveal that SnoN-regulated genes contain binding sites for the transcription factors N-myc and Pax6, which promote the proliferation and differentiation of granule neuron precursors, respectively. Accordingly, we uncover novel physical interactions of SnoN with N-myc and Pax6 in cells. In behavior analyses, conditional KO of SnoN impairs cerebellar-dependent learning in a delayed eye-blink conditioning paradigm, suggesting that SnoN-regulation of granule neuron precursor proliferation bears functional consequences at the organismal level. Our findings define a novel function and mechanism for the major transcriptional regulator SnoN in the control of granule neuron precursor proliferation in the mammalian brain. This study reports the discovery that the transcriptional regulator SnoN plays a crucial role in the proliferation of cerebellar granule neuron precursors in the postnatal mouse brain. Conditional KO of SnoN in granule neuron precursors robustly inhibits the proliferation of these cells and promotes their cycle exit specifically at later stages of cerebellar development, with biological consequences of impaired cerebellar-dependent learning. Genomics and bioinformatics analyses reveal that SnoN promotes the expression of cell proliferation genes and concomitantly represses cell differentiation genes Although SnoN has been implicated in distinct aspects of the development of postmitotic neurons, this study identifies a novel function for SnoN in neuronal precursors in the mammalian brain.
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http://dx.doi.org/10.1523/JNEUROSCI.0688-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6325268PMC
January 2019

Volatile Acid-Solvent Evaporation (VASE): Molecularly Homogeneous Distribution of Acyclovir in a Bioerodable Polymer Matrix for Long-Term Treatment of Herpes Simplex Virus-1 Infections.

J Drug Deliv 2018 26;2018:6161230. Epub 2018 Sep 26.

Towson University Herpes Virus Lab, Department of Biological Sciences, Towson University, Towson, MD 21252, USA.

Treatment for herpes simplex virus-1 and -2 (HSV-1 and -2) patients who suffer from recurrent outbreaks consists of multiple daily doses of the antiviral drugs acyclovir (ACV), penciclovir, or their more orally bioavailable derivatives valacyclovir or famciclovir. Drug troughs caused by missed doses may result in viral replication, which can generate drug-resistant mutants along with clinical sequelae. We developed a molecularly homogeneous mixture of ACV with the bioerodable polymer polycaprolactone. Through scanning electron microscopy, infrared spectroscopy, gel permeation chromatography, 1H NMR, and differential scanning calorimetry, our method of combining drug and polymer, termed Volatile Acid-Solvent Evaporation (VASE), does not compromise the integrity of polymer or drug. Furthermore, VASE creates materials that deliver therapeutic amounts of drug consistently for approximately two months. Devices with high enough drug loads diminish primary infection of HSV-1 in Vero cells to the same level as seen with a single dose of ACV. Our data will lead to further experiments in animal models, demonstrating efficacy in preventing reactivation of these viruses with a single intervention, and with other antiviral drugs amenable to such manipulation. Additionally, this type of treatment would leave no trace after its useful lifetime, as drug is released and polymer matrix is degraded .
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http://dx.doi.org/10.1155/2018/6161230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6178155PMC
September 2018

Treatment correlates of successful outcomes in pulmonary multidrug-resistant tuberculosis: an individual patient data meta-analysis.

Lancet 2018 09;392(10150):821-834

Centre for Lung Infection and Immunity, Department of Medicine & UCT Lung Institute, University of Cape Town, Cape Town, South Africa.

Background: Treatment outcomes for multidrug-resistant tuberculosis remain poor. We aimed to estimate the association of treatment success and death with the use of individual drugs, and the optimal number and duration of treatment with those drugs in patients with multidrug-resistant tuberculosis.

Methods: In this individual patient data meta-analysis, we searched MEDLINE, Embase, and the Cochrane Library to identify potentially eligible observational and experimental studies published between Jan 1, 2009, and April 30, 2016. We also searched reference lists from all systematic reviews of treatment of multidrug-resistant tuberculosis published since 2009. To be eligible, studies had to report original results, with end of treatment outcomes (treatment completion [success], failure, or relapse) in cohorts of at least 25 adults (aged >18 years). We used anonymised individual patient data from eligible studies, provided by study investigators, regarding clinical characteristics, treatment, and outcomes. Using propensity score-matched generalised mixed effects logistic, or linear regression, we calculated adjusted odds ratios and adjusted risk differences for success or death during treatment, for specific drugs currently used to treat multidrug-resistant tuberculosis, as well as the number of drugs used and treatment duration.

Findings: Of 12 030 patients from 25 countries in 50 studies, 7346 (61%) had treatment success, 1017 (8%) had failure or relapse, and 1729 (14%) died. Compared with failure or relapse, treatment success was positively associated with the use of linezolid (adjusted risk difference 0·15, 95% CI 0·11 to 0·18), levofloxacin (0·15, 0·13 to 0·18), carbapenems (0·14, 0·06 to 0·21), moxifloxacin (0·11, 0·08 to 0·14), bedaquiline (0·10, 0·05 to 0·14), and clofazimine (0·06, 0·01 to 0·10). There was a significant association between reduced mortality and use of linezolid (-0·20, -0·23 to -0·16), levofloxacin (-0·06, -0·09 to -0·04), moxifloxacin (-0·07, -0·10 to -0·04), or bedaquiline (-0·14, -0·19 to -0·10). Compared with regimens without any injectable drug, amikacin provided modest benefits, but kanamycin and capreomycin were associated with worse outcomes. The remaining drugs were associated with slight or no improvements in outcomes. Treatment outcomes were significantly worse for most drugs if they were used despite in-vitro resistance. The optimal number of effective drugs seemed to be five in the initial phase, and four in the continuation phase. In these adjusted analyses, heterogeneity, based on a simulated I method, was high for approximately half the estimates for specific drugs, although relatively low for number of drugs and durations analyses.

Interpretation: Although inferences are limited by the observational nature of these data, treatment outcomes were significantly better with use of linezolid, later generation fluoroquinolones, bedaquiline, clofazimine, and carbapenems for treatment of multidrug-resistant tuberculosis. These findings emphasise the need for trials to ascertain the optimal combination and duration of these drugs for treatment of this condition.

Funding: American Thoracic Society, Canadian Institutes of Health Research, US Centers for Disease Control and Prevention, European Respiratory Society, Infectious Diseases Society of America.
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http://dx.doi.org/10.1016/S0140-6736(18)31644-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6463280PMC
September 2018

Sinonasal T-cell expression of cytotoxic mediators granzyme B and perforin is reduced in patients with chronic rhinosinusitis.

Am J Rhinol Allergy 2017 Nov;31(6):352-356

Department of Otolaryngology-Head and Neck Surgery, Medical University of South Carolina, Charleston, South Carolina, USA.

Background: CD8+ T cells and natural killer (NK) cells are cytotoxic cells that use granzyme B (GrB) and perforin. Defective cytotoxic function is known to play a role in dysregulated immune response as seen in chronic sinusitis, also referred to as chronic rhinosinusitis (CRS). However, to our knowledge, in the United States, neither GrB or perforin expression has been reported in patients with CRS.

Objective: The aim of this study was to investigate sinonasal cytotoxic cells, their mediators, and cell-specific distribution of these mediators in patients with CRS with nasal polyp (CRSwNP) and in patients with CRS without nasal polyp (CRSsNP).

Methods: Blood and sinus tissue samples were taken from patients with CRSsNP (n = 8) and CRSwNP (n = 8) at the time of surgery. Control subjects (n = 8) underwent surgery for cerebrospinal fluid leak repair or to remove non-hormone-secreting pituitary tumors. The cells were analyzed via flow cytometry by using CD8 expression to identify cytotoxic T cells and CD56 expression to identify NK cells. Intracellular GrB and perforin expression were analyzed with flow cytometry.

Results: We observed no significant differences in plasma or peripheral blood immune cell numbers or specific levels of GrB or perforin among the groups. In the sinonasal mucosa of the patients with CRSsNP and the patients with CRSwNP, there was a significant decrease in GrB and perforin levels (p < 0.05) despite similar or increased numbers of cytotoxic cells when compared with the controls. The overall decrease in GrB and perforin in the sinonasal mucosa of the patients with CRSsNP and the patients with CRSwNP was due to decreased T cell production. There was no difference in total NK cell count or expression of perforin or GrB among all the groups.

Conclusion: Total levels of sinonasal GrB and perforin were decreased in the sinonasal mucosa of both the patients with CRSwNP and the patients with CRSsNP compared with the controls, whereas sinonasal CD8+ T cells, (but not NK cells,), intracellular stores of GrB and perforin were reduced in the patients with CRSwNP compared with the controls.
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http://dx.doi.org/10.2500/ajra.2017.31.4474DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5691237PMC
November 2017

The role of regulatory T cells in the regulation of upper airway inflammation.

Am J Rhinol Allergy 2017 Nov;31(6):345-351

Department of Otolaryngology-Head and Neck Surgery, Medical University of South Carolina, Charleston, South Carolina, USA.

Allergic rhinitis (AR) and chronic rhinosinusitis with nasal polyps (CRSwNP) are inflammatory diseases of the upper airway, with a similar immunologic profile, characterized by aberrant and persistent type 2 inflammation. One cell population that has been identified as altered in both disease types is regulatory T cell (Treg). Tregs have the capacity to modulate T-effector function and suppress inflammatory cytokine production in a broad range of cell types. Given the ability of Tregs to control inflammation, the role of Tregs in respiratory diseases has attracted much attention. As discussed in this article, alterations in the Treg numbers and function, or both, have been identified in AR and CRSwNP, although much of the data is conflicting. Here, we explored what is known and, in many cases, unknown about the mechanisms by which Tregs differentiate and function, and how these functions can be controlled in the mucosal microenvironment. By gaining a greater understanding of these processes, it may be possible to harness the natural immunosuppressive activity of Tregs to ameliorate the chronic inflammation associated with AR and CRSwNP.
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http://dx.doi.org/10.2500/ajra.2017.31.4472DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5691236PMC
November 2017

Nanoscale architecture of the contractile ring.

Elife 2017 09 15;6. Epub 2017 Sep 15.

Department of Cell and Developmental Biology, Vanderbilt University, Nashville, United States.

The contractile ring is a complex molecular apparatus which physically divides many eukaryotic cells. Despite knowledge of its protein composition, the molecular architecture of the ring is not known. Here we have applied super-resolution microscopy and FRET to determine the nanoscale spatial organization of contractile ring components relative to the plasma membrane. Similar to other membrane-tethered actin structures, we find proteins localize in specific layers relative to the membrane. The most membrane-proximal layer (0-80 nm) is composed of membrane-binding scaffolds, formin, and the tail of the essential myosin-II. An intermediate layer (80-160 nm) consists of a network of cytokinesis accessory proteins as well as multiple signaling components which influence cell division. Farthest from the membrane (160-350 nm) we find F-actin, the motor domains of myosins, and a major F-actin crosslinker. Circumferentially within the ring, multiple proteins proximal to the membrane form clusters of different sizes, while components farther from the membrane are uniformly distributed. This comprehensive organizational map provides a framework for understanding contractile ring function.
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http://dx.doi.org/10.7554/eLife.28865DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5779233PMC
September 2017

Aneuploidy as a cause of impaired chromatin silencing and mating-type specification in budding yeast.

Elife 2017 08 25;6. Epub 2017 Aug 25.

Department of Cell Biology, Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, United States.

Aneuploidy and epigenetic alterations have long been associated with carcinogenesis, but it was unknown whether aneuploidy could disrupt the epigenetic states required for cellular differentiation. In this study, we found that ~3% of random aneuploid karyotypes in yeast disrupt the stable inheritance of silenced chromatin during cell proliferation. Karyotype analysis revealed that this phenotype was significantly correlated with gains of chromosomes III and X. Chromosome X disomy alone was sufficient to disrupt chromatin silencing and yeast mating-type identity as indicated by a lack of growth response to pheromone. The silencing defect was not limited to cryptic mating type loci and was associated with broad changes in histone modifications and chromatin localization of Sir2 histone deacetylase. The chromatin-silencing defect of disome X can be partially recapitulated by an extra copy of several genes on chromosome X. These results suggest that aneuploidy can directly cause epigenetic instability and disrupt cellular differentiation.
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http://dx.doi.org/10.7554/eLife.27991DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5779231PMC
August 2017

Low-dose tamoxifen treatment in juvenile males has long-term adverse effects on the reproductive system: implications for inducible transgenics.

Sci Rep 2017 08 21;7(1):8991. Epub 2017 Aug 21.

MRC Centre for Reproductive Health, The Queen's Medical Research Institute, 47 Little France Crescent, Edinburgh, EH16 4TJ, UK.

The tamoxifen-inducible Cre system is a popular transgenic method for controlling the induction of recombination by Cre at a specific time and in a specific cell type. However, tamoxifen is not an inert inducer of recombination, but an established endocrine disruptor with mixed agonist/antagonist activity acting via endogenous estrogen receptors. Such potentially confounding effects should be controlled for, but >40% of publications that have used tamoxifen to generate conditional knockouts have not reported even the minimum appropriate controls. To highlight the importance of this issue, the present study investigated the long-term impacts of different doses of a single systemic tamoxifen injection on the testis and the wider endocrine system. We found that a single dose of tamoxifen less than 10% of the mean dose used for recombination induction, caused adverse effects to the testis and to the reproductive endocrine system that persisted long-term. These data raise significant concerns about the widespread use of tamoxifen induction of recombination, and highlight the importance of including appropriate controls in all pathophysiological studies using this means of induction.
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http://dx.doi.org/10.1038/s41598-017-09016-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5566418PMC
August 2017

Treatment Outcomes of Children With HIV Infection and Drug-resistant TB in Three Provinces in South Africa, 2005-2008.

Pediatr Infect Dis J 2017 Dec;36(12):e322-e327

From the *Department of Epidemiology, Emory University, Atlanta, Georgia; †U.S. Centers for Disease Control and Prevention, Atlanta, Georgia; ‡National Institute for Communicable Diseases, National Health Laboratory Service, Johannesburg, South Africa; and §South African Medical Research Council, Pretoria, South Africa.

Objective: To describe outcomes of HIV-infected pediatric patients with drug-resistant tuberculosis (DR TB).

Methods: Demographic, clinical and laboratory data from charts of pediatric patients treated for DR TB during 2005-2008 were collected retrospectively from 5 multi-DR TB hospitals in South Africa. Data were summarized, and Pearson χ test or Fisher exact test was used to assess differences in variables of interest by HIV status. A time-to-event analysis was conducted using days from start of treatment to death. Variables of interest were first assessed using the Kaplan-Meier method. Cox proportional hazard models were fit to estimate crude and adjusted hazard ratios.

Results: Of 423 eligible participants, 398 (95%) had culture-confirmed DR TB and 238 (56%) were HIV infected. A total of 54% were underweight, 42% were male and median age was 10.7 years (interquartile range: 5.5-15.3). Of the 423 participants, 245 (58%) were successfully treated, 69 (16%) died, treatment failed in 3 (1%), 36 (9%) were lost to follow-up and 70 (17%) were still on treatment, transferred or had unknown outcomes. Time to death differed by HIV status (P = 0.008), sex (P < 0.001), year of tuberculosis diagnosis (P = 0.05) and weight status (P = 0.002). Over the 2-year risk period, the adjusted rate of death was 2-fold higher among participants with HIV compared with HIV-negative participants (adjusted hazard ratio = 2.28; 95% confidence interval: 1.11-4.68).

Conclusions: Male, underweight and HIV-infected children with DR TB were more likely to experience death when compared with other children with DR TB within this study population.
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http://dx.doi.org/10.1097/INF.0000000000001691DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5797992PMC
December 2017

Table Saw Injuries: Are Our Safety Features Really Keeping Us Safe?

Hand (N Y) 2018 Mar 1;13(2):181-183. Epub 2017 Mar 1.

1 Bassett Medical Center, Cooperstown, NY, USA.

Background: Currently, table saws sold in the United States have 3 safety features (riving knife, blade guard, and antikickback device) designed to prevent personal injury; however, these features can hinder the user's movements and are often disabled or removed. Despite the frequency of table saw injuries, there is relatively limited literature regarding them.

Methods: We performed a retrospective, observational study of characteristics associated with injuries incurred while using a table saw. Electronic medical records from Bassett Medical Center were reviewed from 2002 to 2014 to identify patients who sustained table saw injury, and surveys were sent to the patients. Tests of association between survey variables (eg, presence/absence of safety device vs severity of injury) were carried out using chi-square or Fisher exact test.

Results: Seventy-three percent of those injured had disabled their safety features or had no safety features present; 27% of those injured had one or more safety features enabled when they sustained the injury. Of those injured, 18% sustained a "mild" injury (skin laceration), 39% sustained a "moderate" injury (nerve or tendon injury), and 43% sustained a "severe" injury (partial or complete amputation). There was no statistically significant association between severity of the injury and presence/absence of safety features.

Conclusions: This study highlights 2 major problems with the current safety features: People are dissatisfied with current safety features, disabling them and sustaining injuries, and even when they are enabled, people are still getting injured-suggesting inadequacy of the current features.
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http://dx.doi.org/10.1177/1558944717695754DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5950965PMC
March 2018

Epidemiology of Pediatric Multidrug-Resistant Tuberculosis in the United States, 1993-2014.

Clin Infect Dis 2017 Oct;65(9):1437-1443

Division of Global HIV and Tuberculosis, Center for Global Health.

Background: Multidrug-resistant tuberculosis (MDR-TB) is an important global public health threat, but accurate estimates of MDR-TB burden among children are lacking.

Methods: We analyzed demographic, clinical, and laboratory data for newly diagnosed pediatric (age <15 years) TB cases reported to the US National TB Surveillance System during 1993-2014. MDR-TB was defined as culture-confirmed TB disease with resistance to at least isoniazid and rifampicin. To ascertain potential underestimation of pediatric MDR-TB, we surveyed high-burden states for clinically diagnosed cases treated for MDR-TB.

Results: Of 20789 pediatric TB cases, 5162 (24.8%) had bacteriologically confirmed TB. Among 4826 (93.5%) with drug susceptibility testing, 82 (1.7%) had MDR-TB. Most pediatric MDR-TB cases were female (n = 51 [62%]), median age was 5 years (interquartile range, 1-12 years), one-third were Hispanic (n = 28 [34%]), and two-thirds (n = 55 [67%]) were born in the United States. Most cases had additional resistance to ≥1 other first-line drug (n = 66 [81%]) and one-third had resistance to ≥1 second-line drug (24/73 tested). Of 77 who started treatment prior to 2013, 66 (86%) completed treatment and 4 (5%) died. Among the 4 high-TB-burden states/jurisdictions surveyed, there was 42%-55% underestimation of pediatric MDR-TB cases when using only culture-confirmed case definitions.

Conclusions: Only one-quarter of pediatric TB cases had culture-confirmed TB, likely resulting in underestimation of true pediatric MDR-TB burden in the United States using strictly bacteriologic criteria. Better estimates of pediatric MDR-TB burden in the United States are needed and should include clinical diagnoses based on epidemiologic criteria.
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http://dx.doi.org/10.1093/cid/cix561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5801661PMC
October 2017

Long-Term Engraftment and Fetal Globin Induction upon Gene Editing in Bone-Marrow-Derived CD34 Hematopoietic Stem and Progenitor Cells.

Mol Ther Methods Clin Dev 2017 Mar 11;4:137-148. Epub 2017 Jan 11.

Biogen, Cambridge, MA 02142, USA.

To develop an effective and sustainable cell therapy for sickle cell disease (SCD), we investigated the feasibility of targeted disruption of the gene, either within exon 2 or at the GATAA motif in the intronic erythroid-specific enhancer, using zinc finger nucleases in human bone marrow (BM) CD34 hematopoietic stem and progenitor cells (HSPCs). Both targeting strategies upregulated fetal globin expression in erythroid cells to levels predicted to inhibit hemoglobin S polymerization. However, complete inactivation of resulting from bi-allelic frameshift mutations in exon 2 adversely affected erythroid enucleation. In contrast, bi-allelic disruption of the GATAA motif in the erythroid enhancer of did not negatively impact enucleation. Furthermore, exon 2-edited BM-CD34 cells demonstrated a significantly reduced engraftment potential in immunodeficient mice. Such an adverse effect on HSPC function was not observed upon erythroid-enhancer GATAA motif editing, because enhancer-edited CD34 cells achieved robust long-term engraftment and gave rise to erythroid cells with elevated levels of fetal globin expression when chimeric BM was cultured ex vivo. Altogether, our results support further clinical development of the erythroid-specific enhancer editing in BM-CD34 HSPCs as an autologous stem cell therapy in SCD patients.
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http://dx.doi.org/10.1016/j.omtm.2016.12.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5363298PMC
March 2017
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