UFGI publication round-up weeks 4/24, 5/1, 5/8/17

Versatile and precise gene-targeting strategies for functional studies in mammalian cell lines.

Author information: Wassef M¹, Luscan A², Battistella A³, Le Corre S³, Li H⁴, Wallace MR⁵, Vidaud M², Margueron R³.

Date of e-pub: May 2017

Abstract: The advent of programmable nucleases such as ZFNs, TALENs and CRISPR/Cas9 has brought the power of genetic manipulation to widely used model systems such as model cell lines. In mammalian cells, nuclease-mediated DNA double strand break is mainly repaired through the error-prone non-homologous end-joining (NHEJ) repair pathway, eventually leading to accumulation of small deletions or insertions (indels) that can inactivate gene function. However, due to the variable size of the indels and the polyploid status of many cell lines (e.g., cancer-derived cells), obtaining a knockout usually requires lengthy screening and characterization procedures. Given the more precise type of modifications that can be introduced upon homology-directed repair (HDR), we have developed HDR-based gene-targeting strategies that greatly facilitate the process of knockout generation in cell lines. To generate reversible knockouts (R-KO), a selectable promoter-less STOP cassette is inserted in an intron, interrupting transcription. Loss-of-function can be validated by RT-qPCR and is removable, enabling subsequent restoration of gene function. A variant of the R-KO procedure can be used to introduce point mutations. To generate constitutive knockouts (C-KO), an exon is targeted, which makes use of HDR-based gene disruption together with NHEJ-induced indels on non-HDR targeted allele(s). Hence the C-KO procedure greatly facilitates simultaneous inactivation of multiple alleles. Overall these genome-editing tools offer superior precision and efficiency for functional genetic approaches. We provide detailed protocols guiding in the design of targeting vectors and in the analysis and validation of gene targeting experiments.

 

 

Consequences of MEGF10 deficiency on myoblast function and Notch1 interactions.

Author information: Saha M¹, Mitsuhashi S², Jones MD¹, Manko K¹, Reddy HM¹, Bruels C¹, Cho KA^1,2, Pacak CA³, Draper I⁴, Kang PB^1,2,5,6.

Date of e-pub: May 2017

Abstract: Mutations in MEGF10 cause early onset myopathy, areflexia, respiratory distress, and dysphagia (EMARDD), a rare congenital muscle disease, but the pathogenic mechanisms remain largely unknown. We demonstrate that short hairpin RNA (shRNA)-mediated knockdown of Megf10, as well as overexpression of the pathogenic human p.C774R mutation, leads to impaired proliferation and migration of C2C12 cells. Myoblasts from Megf10-/- mice and Megf10-/-/mdx double knockout (dko) mice also show impaired proliferation and migration compared to myoblasts from wild type and mdx mice, whereas the dko mice show histological abnormalities that are not observed in either single mutant mouse. Cell proliferation and migration are known to be regulated by the Notch receptor, which plays an essential role in myogenesis. Reciprocal co-immunoprecipitation studies show that Megf10 and Notch1 interact via their respective intracellular domains. These interactions are impaired by the pathogenic p.C774R mutation. Megf10 regulation of myoblast function appears to be mediated at least in part via interactions with key components of the Notch signaling pathway, and defects in these interactions may contribute to the pathogenesis of EMARDD.

 

 

Authors’ Response to Jesse D. Riordan, Hum Gene Ther 2017;28:375-376; DOI: 10.1089/hum.2017.045.

Author information: Srivastava A¹, Carter BJ².

Date of e-pub: May 2017

Abstract: N/A

 

 

Target enrichment sequencing in cultivated peanut (Arachis hypogaea L.) using probes designed from transcript sequences.

Author information: Peng Z¹, Fan W¹, Wang L¹, Paudel D¹, Leventini D¹, Tillman BL¹, Wang J^2,3,4.

Date of e-pub: May 2017

Abstract: Enabled by the next generation sequencing, target enrichment sequencing (TES) is a powerful method to enrich genomic regions of interest and to identify sequence variations. The objective of this study was to explore the feasibility of probe design from transcript sequences for TES application in calling sequence variants in peanut, an important allotetraploid crop with a large genome size. In this study, we applied an in-solution hybridization method to enrich DNA sequences of seven peanut genotypes. Our results showed that it is feasible to apply TES with probes designed from transcript sequences in polyploid peanut. Using a set of 31,123 probes, a total of 5131 and 7521 genes were targeted in peanut A and B genomes, respectively. For each genotype used in this study, the probe target capture regions were efficiently covered with high depth. The average on-target rate of sequencing reads was 42.47%, with a significant amount of off-target reads coming from genomic regions homologous to target regions. In this study, when given predefined genomic regions of interest and the same amount of sequencing data, TES provided the highest coverage of target regions when compared to whole genome sequencing, RNA sequencing, and genotyping by sequencing. Single nucleotide polymorphism (SNP) calling and subsequent validation revealed a high validation rate (85.71%) of homozygous SNPs, providing valuable markers for peanut genotyping. This study demonstrated the success of applying TES for SNP identification in peanut, which shall provide valuable suggestions for TES application in other non-model species without a genome reference available.

 

 

Biopsychosocial influence on shoulder pain: Rationale and protocol for a pre-clinical trial.

Author information: George SZ¹, Staud R², Borsa PA³, Wu SS⁴, Wallace MR⁵, Greenfield WH⁶, Mackie LN⁷, Fillingim RB⁸.

Date of e-pub: May 2017

Abstract: Chronic musculoskeletal pain conditions are a prevalent and disabling problem. Preventing chronic musculoskeletal pain requires multifactorial treatment approaches that address its complex etiology. Prior cohort studies identified a high risk subgroup comprised of variation in COMT genotype and pain catastrophizing. This subgroup had increased chance of heightened pain responses (in a pre-clinical model) and higher 12month post-operatives pain intensity ratings (in a clinical model). This pre-clinical trial will test mechanisms and efficacy of personalized pain interventions matched to the genetic and psychological characteristics of the high-risk subgroup.

Potential participants will be screened for high risk subgroup membership, appropriateness for exercise-induced muscle injury protocol, and appropriateness for propranolol administration. Eligible participants that consent to the study will then be randomized into one of four treatment groups; 1) personalized pharmaceutical and psychological education; 2) personalized pharmaceutical and general education; 3) placebo pharmaceutical and psychological education; 4) placebo pharmaceutical and psychological education. Over the 5-day study period participants will complete an exercise-induced muscle injury protocol and receive study interventions. Pain and disability assessments will be completed daily, with primary outcomes being duration of shoulder pain (number of days until recovery), peak shoulder pain intensity, and peak shoulder disability. Secondary outcomes include inflammatory markers, psychological mediators, and measures of pain sensitivity regulation.

This pre-clinical trial builds on prior cohort studies and its completion will provide foundational data supporting efficacy and mechanisms of personalized interventions for individuals that may be at increased risk for developing chronic shoulder pain.

ClinicalTrials.gov registry, NCT02620579 (Registered on November 13, 2015).

 

 

Neonatal sepsis in rural India: timing, microbiology and antibiotic resistance in a population-based prospective study in the community setting.

Author information: Panigrahi P¹, Chandel DS², Hansen NI³, Sharma N⁴, Kandefer S⁵, Parida S⁶, Satpathy R⁷, Pradhan L⁸, Mohapatra A⁹, Mohapatra SS⁷, Misra PR¹⁰, Banaji N¹¹, Johnson JA¹², Morris JG Jr¹³, Gewolb IH¹⁴, Chaudhry R⁴.

Date of e-pub: May 2017

Abstract: To examine the timing and microbiology of neonatal sepsis in a population-based surveillance in the Indian community setting.

All live born infants in 223 villages of Odisha state were followed at home for 60 days. Suspect sepsis cases were referred to study hospitals for further evaluation including blood culture.

Of 12 622 births, 842 were admitted with suspected sepsis of whom 95% were 4 to 60 days old. Culture-confirmed incidence of sepsis was 6.7/1000 births with 51% Gram negatives (Klebsiella predominating) and 26% Gram positives (mostly Staphylococcus aureus). A very high level of resistance to penicillin and ampicillin, moderate resistance to cephalosporins and extremely low resistance to Gentamicin and Amikacin was observed.

The bacterial burden of sepsis in the Indian community is not high. Judicious choice of empiric antibiotics, antibiotic stewardship and alternate modalities should be considered for the management or prevention of neonatal sepsis in India.Journal of Perinatology advance online publication, 11 May 2017; doi:10.1038/jp.2017.67.

 

 

Benzotriazole ultraviolet stabilizers alter the expression of the thyroid hormone pathway in zebrafish (Danio rerio) embryos.

Author information: Liang X¹, Li J², Martyniuk CJ³, Wang J², Mao Y², Lu H², Zha J⁴.

Date of e-pub: May 2017

Abstract: Benzotriazole ultraviolet stabilizers (BUVSs) are widely used in industrial products as well as personal-hygiene products to protect the material or skin from harmful UV-radiation. Due to their persistence and bioaccumulation, BUVSs have been ubiquitously detected in aquatic environments. Although the toxicological effects of BUVSs in aquatic organisms have been previously examined, the effects of BUVSs on the thyroid system have not been adequately addressed. In this study, we assessed putative thyroid disrupting effects of BUVSs (UV-234, UV-326, UV-329 and UV-P) in zebrafish embryos at 1, 10 and 100 μg/L for 96 h. The heart rate was assessed in zebrafish and was observed to be decreased by 6.9%-21.4% in exposure of tested BUVSs. We also observed that the transcript levels of HPT axis-related genes were affected by the 4 BUVSs tested in different ways. Specifically, mRNA levels of thyroid hormone receptors (thraa and thrb) in zebrafish embryos were differentially expressed and the direction of change in these transcripts was isoform and BUVSs dependent. Pathway analysis of the targeted genes measured indicated that cellular processes putatively affected by BUVSs included response to organic substance, regulation of transcription from RNA polymerase II promoter, intracellular receptor signaling pathway, and hypothyroidism. Upon expansion of the network, novel genes involved in this predicted gene network may provide insight into the mechanisms of thyroid disrupting mechanisms of BUVSs. Taken together, our results indicate that BUVSs can potentially impact the thyroid system, and that this is dependent upon the type or structure of BUVSs.

 

 

Salinity Response in Chloroplasts: Insights from Gene Characterization.

Author information: Suo J¹, Zhao Q², David L³, Chen S⁴, Dai S^5,6.

Date of e-pub: May 2017

Abstract: Salinity is a severe abiotic stress limiting agricultural yield and productivity. Plants have evolved various strategies to cope with salt stress. Chloroplasts are important photosynthesis organelles, which are sensitive to salinity. An understanding of molecular mechanisms in chloroplast tolerance to salinity is of great importance for genetic modification and plant breeding. Previous studies have characterized more than 53 salt-responsive genes encoding important chloroplast-localized proteins, which imply multiple vital pathways in chloroplasts in response to salt stress, such as thylakoid membrane organization, the modulation of photosystem II (PS II) activity, carbon dioxide (CO₂) assimilation, photorespiration, reactive oxygen species (ROS) scavenging, osmotic and ion homeostasis, abscisic acid (ABA) biosynthesis and signaling, and gene expression regulation, as well as protein synthesis and turnover. This review presents an overview of salt response in chloroplasts revealed by gene characterization efforts.

 

 

Professor Yuichi Sugiyama: A Brilliant, Creative, Amicable, Charming and Humorous Pharmaceutical Scientist.

Author information: Kusuhara H¹, Obach RS², Rostami-Hodjegan A³, Pang KS⁴, Hammarlund-Udenaes M⁵, Derendorf H⁶, Artursson P⁷, Huang SM⁸, Suzuki H⁹, Terasaki T¹⁰.

Date of e-pub: May 2017

Abstract: N/A

 

 

Structural Insight into Ubiquitin-Like Protein Recognition and Oligomeric States of JAMM/MPN+ Proteases.

Author information: Cao S¹, Engilberge S², Girard E², Gabel F², Franzetti B³, Maupin-Furlow JA⁴.

Date of e-pub: May 2017

Abstract: JAMM/MPN+ metalloproteases cleave (iso)peptide bonds C-terminal to ubiquitin (Ub) and ubiquitin-like protein (Ubl) domains and typically require association with protein partners for activity, which has limited a molecular understanding of enzyme function. To provide an insight, we solved the X-ray crystal structures of a catalytically active Pyrococcus furiosus JAMM/MPN+ metalloprotease (PfJAMM1) alone and in complex with a Ubl (PfSAMP2) to 1.7- to 1.9-Å resolution. PfJAMM1 was found to have a redox sensitive dimer interface. In the PfJAMM1-bound state of the SAMP2, a Ubl-to-Ub conformational change was detected. Surprisingly, distant homologs of PfJAMM1 were found to be closely related in 3D structure, including the interface for Ubl/Ub binding. From this work, we infer the molecular basis of how JAMM/MPN+ proteases recognize and cleave Ubl/Ub tags from diverse proteins and highlight an α2-helix structural element that is conserved and crucial for binding and removing the Ubl SAMP2 tag.

 

 

De Novo Coding Variants Are Strongly Associated with Tourette Disorder.

Author information: Willsey AJ¹, Fernandez TV², Yu D³, King RA², Dietrich A⁴, Xing J⁵, Sanders SJ⁶, Mandell JD¹, Huang AY⁷, Richer P⁸, Smith L⁶, Dong S⁶, Samocha KE³; Tourette International Collaborative Genetics (TIC Genetics); Tourette Syndrome Association International Consortium for Genetics (TSAICG), Neale BM³, Coppola G⁷, Mathews CA⁹, Tischfield JA⁵, Scharf JM¹⁰, State MW¹¹, Heiman GA¹².

Date of e-pub: May 2017

Abstract: Whole-exome sequencing (WES) and de novo variant detection have proven a powerful approach to gene discovery in complex neurodevelopmental disorders. We have completed WES of 325 Tourette disorder trios from the Tourette International Collaborative Genetics cohort and a replication sample of 186 trios from the Tourette Syndrome Association International Consortium on Genetics (511 total). We observe strong and consistent evidence for the contribution of de novo likely gene-disrupting (LGD) variants (rate ratio [RR] 2.32, p = 0.002). Additionally, de novo damaging variants (LGD and probably damaging missense) are overrepresented in probands (RR 1.37, p = 0.003). We identify four likely risk genes with multiple de novo damaging variants in unrelated probands: WWC1 (WW and C2 domain containing 1), CELSR3 (Cadherin EGF LAG seven-pass G-type receptor 3), NIPBL (Nipped-B-like), and FN1 (fibronectin 1). Overall, we estimate that de novo damaging variants in approximately 400 genes contribute risk in 12% of clinical cases.

 

 

Resolving relationships among the megadiverse butterflies and moths with a novel pipeline for Anchored Phylogenomics.

Author information: Breinholt JW^1,2, Earl C1, Lemmon AR³, Moriarty Lemmon E³, Xiao L¹, Kawahara AY¹.

Date of e-pub: May 2017

Abstract: The advent of next-generation sequencing technology has allowed for the collection of large portions of the genome for phylogenetic analysis. Hybrid enrichment and transcriptomics are two techniques that leverage next-generation sequencing and have shown much promise. However, methods for processing hybrid enrichment data are still limited. We developed a pipeline for anchored hybrid enrichment (AHE) read assembly, orthology determination, contamination screening, and data processing for sequences flanking the target “probe” region. We apply this approach to study the phylogeny of butterflies and moths (Lepidoptera), a megadiverse group of more than 157,000 described species with poorly understood deep-level phylogenetic relationships. We introduce a new, 855 locus anchored hybrid enrichment kit for Lepidoptera phylogenetics and compare resulting trees to those from transcriptomes. The enrichment kit was designed from existing genomes, transcriptomes and expressed sequence tag (EST) data and was used to capture sequence data from 54 species from 23 lepidopteran families. Phylogenies estimated from AHE data were largely congruent with trees generated from transcriptomes, with strong support for relationships at all but the deepest taxonomic levels. We combine AHE and transcriptomic data to generate a new Lepidoptera phylogeny, representing 76 exemplar species in 42 families. The tree provides robust support for many relationships, including those among the seven butterfly families. The addition of AHE data to an existing transcriptomic dataset lowers node support along the Lepidoptera backbone, but firmly places taxa with AHE data on the phylogeny. To examine the efficacy of AHE at different taxonomic levels, phylogenetic analyses were also conducted on a sister group representing a more recent divergence, the Saturniidae and Sphingidae. These analyses utilized sequences from the probe region and data flanking it, nearly doubled the size of the dataset; all resulting trees were well supported. We hope that our data processing pipeline, hybrid enrichment gene set, and approach of combining AHE data with transcriptomes will be useful for the broader systematics community.

 

 

Phenotypic characterisation of cell populations in the brains of horses experimentally infected with West Nile Virus.

Author information: Delcambre GH¹, Liu J², Streit WJ³, Shaw GPJ³, Vallario K², Herrington J², Wenzlow N², Barr KL², Long MT².

Date of e-pub: May 2017

Abstract: West Nile Virus (WNV), a mosquito borne member of the Flaviviridae, is one of the most commonly diagnosed agents of viral encephalitis in horses and people worldwide.

A cassette of markers for formalin-fixed paraffin-embedded (FFPE) tissue and an archive of tissues from experimental infections in the horse were used to investigate the equine neuroimmune response to WNV meningoencephalomyelitis to phenotype the early response to WNV infection in the horse.

Quantitative analysis using archived tissue from experimentally infected horses.

The thalamus and hindbrain from two groups of six horses were compared and consisted of a culture positive tissues from WNV experimentally horses, in the other, normal horses. FFPE from the thalamus and hindbrain were immunolabeled for microglia, astrocytes, B cells, macrophages/neutrophils, CD3+ T cells. Fresh frozen tissues were immunolabeled for CD4+ and CD8+ T lymphocyte cell markers. Cell counts were obtained using a computer software program. Differences, after meeting assumptions of abnormality, were computed using a general linear model with a Tukey test (p<0.05) for pairwise comparisons.

In WNV-challenged horses, Iba-1+ microglia, CD3+ T lymphocyte, and MAC387+ macrophage staining were significantly increased. The T cell response for the WNV-challenged horses was mixed, composed of CD4+ and CD8+ T lymphocytes. A limited astrocyte response was also observed in WNV-challenged horses, and MAC387+ and B cells were the least abundant cell populations.

The results of this study were limited by a single collection time post-infection. Furthermore, a comprehensive analysis of cellular phenotypes is needed for naturally infected horses. Unfortunately, in clinical horses, there is high variability of sampling in terms of days post infection and tissue handling.

The data show that WNV-challenged horses recruit a mixed T cell population at the onset of neurologic disease. This article is protected by copyright. All rights reserved.

 

 

Plasmacytoid and conventional dendritic cells cooperate in cross-priming AAV capsid-specific CD8+ T cells.

Author information: Rogers GL¹, Shirley JL¹, Zolotukhin I¹, Kumar SRP¹, Sherman A¹, Perrin GQ¹, Hoffman BE¹, Srivastava A¹, Basner-Tschakarjan E², Wallet MA³, Terhorst C⁴, Biswas M¹, Herzog RW⁵.

Date of e-pub: May 2017

Abstract: Adeno-associated virus (AAV) is a replication-deficient parvovirus that is extensively used as a gene therapy vector. CD8+ T cell responses against the AAV capsid protein can, however, affect therapeutic efficacy. Little is known about the in vivo mechanism that leads to the cross-priming of CD8+ T cells against the input viral capsid antigen. Here we report that the TLR9-MyD88 pattern-recognition receptor pathway is uniquely capable of initiating this response. By contrast, the absence of TLR2, STING, or addition of TLR4 agonist has no effect. Surprisingly, both conventional (cDCs) and plasmacytoid dendritic cells (pDCs) are required for the cross-priming of capsid-specific CD8+ T cells, while other antigen presenting cells are not involved. TLR9 signaling is specifically essential in pDCs but not in cDCs, indicating that sensing of the viral genome by pDCs activates cDCs in trans to cross-present capsid antigen during CD8+ T cell activation. Cross-presentation and cross-priming depend not only on TLR9, but also on IFN I signaling, and both mechanisms can be inhibited by administering specific molecules to prevent induction of capsid-specific CD8+ T cells. Thus, these outcomes directly point to therapeutic interventions and demonstrate that innate immune blockade can eliminate unwanted immune responses in gene therapy.

 

 

Redox signaling in neurotransmission and cognition during aging.

Author information: Kumar A^1,2, Yegla B³, Foster TC⁴.

Date of e-pub: May 2017

Abstract: Oxidative stress increases in the brain with aging and neurodegenerative diseases. Previous work emphasized irreversible oxidative damage in relation to cognitive impairment. This research has evolved to consider a continuum of alterations, from redox signaling to oxidative damage, which provides a basis for understanding the onset and progression of cognitive impairment. This review provides an update on research linking redox signaling to altered function of neural circuits involved in information processing and memory.

Starting in middle-age, redox signaling triggers changes in nervous system physiology described as senescent physiology. Recent studies indicates N-methyl-D-aspartate and ryanodine receptors, and Ca2+ signaling molecules as molecular substrates of redox-mediated senescent physiology.

We review redox homeostasis mechanisms and consider the chemical character of reactive oxygen and nitrogen species, and their role in regulating different transmitter systems. In this regard, senescent physiology may represent the co-opting of pathways normally responsible for feedback regulation of synaptic transmission.

It will be important to identify the intrinsic mechanisms for the shift in oxidative/reductive processes. Intrinsic mechanism will depend on the transmitter system, the oxidative stressors, and the expression/activity of antioxidant enzymes. In addition, it will be important to identify how intrinsic processes interact with other aging factors including changes in inflammatory or hormonal signals.

Results suggest that physiological rather than pathological mechanisms underlie for the initial diagnosis of cognitive impairment. Because redox signaling is reversible, it provides hope for early identification and treatment of cognitive decline.

 

 

A KCNC3 mutation causes a neurodevelopmental, non-progressive SCA13 subtype associated with dominant negative effects and aberrant EGFR trafficking.

Author information: Khare S^1,2,3, Nick JA^1,2, Zhang Y⁴, Galeano K^1,2, Butler B^2,5, Khoshbouei H^2,5, Rayaprolu S⁵, Hathorn T⁶, Ranum LPW⁶, Smithson L^2,5, Golde TE^2,5, Paucar M^7,8, Morse R⁹, Raff M¹⁰, Simon J¹⁰, Nordenskjöld M^11,12, Wirdefeldt K^8,13, Rincon-Limas DE^1,2, Lewis J^2,5, Kaczmarek LK⁴, Fernandez-Funez P^1,2, Nick HS^2,5, Waters MF^1,2,3,5.

Date of e-pub: May 2017

Abstract: The autosomal dominant spinocerebellar ataxias (SCAs) are a diverse group of neurological disorders anchored by the phenotypes of motor incoordination and cerebellar atrophy. Disease heterogeneity is appreciated through varying comorbidities: dysarthria, dysphagia, oculomotor and/or retinal abnormalities, motor neuron pathology, epilepsy, cognitive impairment, autonomic dysfunction, and psychiatric manifestations. Our study focuses on SCA13, which is caused by several allelic variants in the voltage-gated potassium channel KCNC3 (Kv3.3). We detail the clinical phenotype of four SCA13 kindreds that confirm causation of the KCNC3R423H allele. The heralding features demonstrate congenital onset with non-progressive, neurodevelopmental cerebellar hypoplasia and lifetime improvement in motor and cognitive function that implicate compensatory neural mechanisms. Targeted expression of human KCNC3R423H in Drosophila triggers aberrant wing veins, maldeveloped eyes, and fused ommatidia consistent with the neurodevelopmental presentation of patients. Furthermore, human KCNC3R423H expression in mammalian cells results in altered glycosylation and aberrant retention of the channel in anterograde and/or endosomal vesicles. Confirmation of the absence of plasma membrane targeting was based on the loss of current conductance in cells expressing the mutant channel. Mechanistically, genetic studies in Drosophila, along with cellular and biophysical studies in mammalian systems, demonstrate the dominant negative effect exerted by the mutant on the wild-type (WT) protein, which explains dominant inheritance. We demonstrate that ocular co-expression of KCNC3R423H with Drosophila epidermal growth factor receptor (dEgfr) results in striking rescue of the eye phenotype, whereas KCNC3R423H expression in mammalian cells results in aberrant intracellular retention of human epidermal growth factor receptor (EGFR). Together, these results indicate that the neurodevelopmental consequences of KCNC3R423H may be mediated through indirect effects on EGFR signaling in the developing cerebellum. Our results therefore confirm the KCNC3R423H allele as causative for SCA13, through a dominant negative effect on KCNC3WT and links with EGFR that account for dominant inheritance, congenital onset, and disease pathology.

 

 

Preemptive Panel-Based Pharmacogenetic Testing: The Time is Now.

Author information: Weitzel KW^1,2, Cavallari LH^3,4, Lesko LJ⁵.

Date of e-pub: May 2017

Abstract: While recent discoveries have paved the way for the use of genotype-guided prescribing in some clinical environments, significant debate persists among clinicians and researchers about the optimal approach to pharmacogenetic testing in clinical practice. One crucial factor in this debate surrounds the timing and methodology of genotyping, specifically whether genotyping should be performed reactively for targeted genes when a single drug is prescribed, or preemptively using a panel-based approach prior to drug prescribing. While early clinical models that employed a preemptive approach were largely developed in academic health centers through multidisciplinary efforts, increasing examples of pharmacogenetic testing are emerging in community-based and primary care practice environments. However, educational and practice-based resources for these clinicians remain largely nonexistent. As such, there is a need for the health care system to shift its focus from debating about preemptive genotyping to developing and disseminating needed resources to equip frontline clinicians for clinical implementation of pharmacogenetics. Providing tools and guidance to support these emerging models of care will be essential to support the thoughtful, evidence-based use of pharmacogenetic information in diverse clinical practice environments. Specifically, the creation of efficient and accurate point-of-care resources, practice-based tools, and clinical models is needed, along with identification and dissemination of sustainable avenues for pharmacogenetic test reimbursement.

 

 

The bovine embryo hatches from the zona pellucida through either the embryonic or abembryonic pole.

Author information: Negrón-Pérez VM¹, Hansen PJ².

Date of e-pub: May 2017

Abstract: Implantation of the mammalian embryo in the uterus is preceded by escape from the zona pellucida. In some species, hatching from the zona occurs preferentially from one or the other poles of the embryo. The situation for the bovine embryo, in which hatching precedes attachment to the uterus by more than a week, is unclear. The purpose was to describe whether hatching of the bovine embryo from the zona pellucida occurs preferentially from the embryonic or abembryonic pole.

Bovine blastocysts undergoing hatching were examined by light microscopy (n = 84) and epifluorescence imaging using antibodies for markers of epiblast, hypoblast, and trophectoderm (TE) (n = 26). The location of hatching was classified as being at the embryonic pole, if hatching occurred ipsilateral to the inner cell mass (ICM), or abembryonic, if hatching occurred contralateral to the ICM.

A total of 55% of blastocysts exited the zona pellucida through an opening at the embryonic pole. In these cases, 68% of the cells emerging through the zona pellucida were derived from the ICM. The remainder of blastocysts hatched from an opening either contralateral or to the side of the ICM. In these cases, 87% of hatched cells were TE.

For the bovine embryo, there is nearly equal probability of hatching from the embryonic or abembryonic poles. Given that the surface area of the zona pellucida in contact with the TE overlying the ICM is less than for the remainder of the blastocyst, there is some preference for hatching through the embryonic pole. Thus, the bovine embryo is distinct from the mouse and human, where hatching occurs preferentially at the abembryonic pole.

 

 

Old Plants, New Tricks: Phenological Research Using Herbarium Specimens.

Author information: Willis CG¹, Ellwood ER², Primack RB³, Davis CC⁴, Pearson KD⁵, Gallinat AS³, Yost JM⁶, Nelson G⁵, Mazer SJ⁷, Rossington NL⁷, Sparks TH⁸, Soltis PS⁹.

Date of e-pub: April 2017

Abstract: The timing of phenological events, such as leaf-out and flowering, strongly influence plant success and their study is vital to understanding how plants will respond to climate change. Phenological research, however, is often limited by the temporal, geographic, or phylogenetic scope of available data. Hundreds of millions of plant specimens in herbaria worldwide offer a potential solution to this problem, especially as digitization efforts drastically improve access to collections. Herbarium specimens represent snapshots of phenological events and have been reliably used to characterize phenological responses to climate. We review the current state of herbarium-based phenological research, identify potential biases and limitations in the collection, digitization, and interpretation of specimen data, and discuss future opportunities for phenological investigations using herbarium specimens.

 

 

Fatal Systemic Salmonellosis in a Florida Manatee ( Trichechus manatus latirostris).

Author information: Vorbach BS^1,2, Rotstein DS³, Stacy NI², Mavian C⁴, Salemi M⁴, Waltzek TB⁵, de Wit M¹.

Date of e-pub: May 2017

Abstract: A subadult male Florida manatee ( Trichechus manatus latirostris) stranded dead on Florida’s Atlantic coast in January 2015. Necropsy and histopathologic findings confirmed chronic systemic bacterial infection caused by Salmonella enterica serotype IV 50:z4,z23,:- involving renal, respiratory, lymphatic, and skeletal systems. This was a unique case of systemic salmonellosis in a Florida manatee.

 

 

Genetic and Small Molecule Disruption of the AID/RAD51 Axis Similarly Protects Nonobese Diabetic Mice from Type 1 Diabetes through Expansion of Regulatory B Lymphocytes.

Author information: Ratiu JJ¹, Racine JJ¹, Hasham MG¹, Wang Q^1,2, Branca JA¹, Chapman HD¹, Zhu J³, Donghia N¹, Philip V¹, Schott WH¹, Wasserfall C⁴, Atkinson MA⁴, Mills KD⁵, Leeth CM⁶, Serreze DV⁷.

Date of e-pub: May 2017

Abstract: B lymphocytes play a key role in type 1 diabetes (T1D) development by serving as a subset of APCs preferentially supporting the expansion of autoreactive pathogenic T cells. As a result of their pathogenic importance, B lymphocyte-targeted therapies have received considerable interest as potential T1D interventions. Unfortunately, the B lymphocyte-directed T1D interventions tested to date failed to halt β cell demise. IgG autoantibodies marking humans at future risk for T1D indicate that B lymphocytes producing them have undergone the affinity-maturation processes of class switch recombination and, possibly, somatic hypermutation. This study found that CRISPR/Cas9-mediated ablation of the activation-induced cytidine deaminase gene required for class switch recombination/somatic hypermutation induction inhibits T1D development in the NOD mouse model. The activation-induced cytidine deaminase protein induces genome-wide DNA breaks that, if not repaired through RAD51-mediated homologous recombination, result in B lymphocyte death. Treatment with the RAD51 inhibitor 4,4′-diisothiocyanatostilbene-2, 2′-disulfonic acid also strongly inhibited T1D development in NOD mice. The genetic and small molecule-targeting approaches expanded CD73+ B lymphocytes that exert regulatory activity suppressing diabetogenic T cell responses. Hence, an initial CRISPR/Cas9-mediated genetic modification approach has identified the AID/RAD51 axis as a target for a potentially clinically translatable pharmacological approach that can block T1D development by converting B lymphocytes to a disease-inhibitory CD73+ regulatory state.

 

 

Evaluation of engineered AAV capsids for hepatic factor IX gene transfer in murine and canine models.

Author information: Markusic DM¹, Nichols TC², Merricks EP², Palaschak B³, Zolotukhin I³, Marsic D³, Zolotukhin S³, Srivastava A³, Herzog RW⁴.

Date of e-pub: May 2017

Abstract: Adeno-associated virus (AAV) gene therapy vectors have shown the best outcomes in human clinical studies for the treatment of genetic diseases such as hemophilia. However, these pivotal investigations have also identified several challenges. For example, high vector doses are often used for hepatic gene transfer, and cytotoxic T lymphocyte responses against viral capsid may occur. Therefore, achieving therapy at reduced vector doses and other strategies to reduce capsid antigen presentation are desirable.

We tested several engineered AAV capsids for factor IX (FIX) expression for the treatment of hemophilia B by hepatic gene transfer. These capsids lack potential phosphorylation or ubiquitination sites, or had been generated through molecular evolution.

AAV2 capsids lacking either a single lysine residue or 3 tyrosine residues directed substantially higher coagulation FIX expression in mice compared to wild-type sequence or other mutations. In hemophilia B dogs, however, expression from the tyrosine-mutant vector was merely comparable to historical data on AAV2. Evolved AAV2-LiC capsid was highly efficient in hemophilia B mice but lacked efficacy in a hemophilia B dog.

Several alternative strategies for capsid modification improve the in vivo performance of AAV vectors in hepatic gene transfer for correction of hemophilia. However, capsid optimization solely in mouse liver may not predict efficacy in other species and thus is of limited translational utility.

 

 

Comparison between an exclusive in vitro-produced embryo transfer system and artificial insemination for genetic, technical, and financial herd performance.

Author information: Kaniyamattam K¹, Block J¹, Hansen PJ¹, De Vries A².

Date of e-pub: April 2017

Abstract: The objective of this study was to implement an in vitro produced embryo transfer (IVP-ET) system in an existing stochastic dynamic dairy simulation model with multitrait genetics to evaluate the genetic, technical, and financial performance of a dairy herd implementing an exclusive IVP-ET or artificial insemination (AI) system. In the AI system, sexed semen was used on the genetically best heifers only. In the IVP-ET system, all of the animals in the herd were impregnated with female sexed embryos created through in vitro fertilization of oocytes collected from animals of superior genetics for different traits of interest. Each donor was assumed to yield on average 4.25 transferable embryos per collection. The remaining animals in the herd were used as recipients and received either a fresh embryo or a frozen embryo when fresh embryos were not available. Selection of donors was random or based on the greatest estimated breeding value (EBV) of lifetime net merit (NM$), milk yield, or daughter pregnancy rate. For both the IVP-ET and AI systems, culling of surplus heifer calves not needed to replace culled cows was based on the lowest EBV for the same traits. A herd of 1,000 milking cows was simulated 15 yr over time after the start of the IVP-ET system. The default cost to produce and transfer 1 embryo was set at $165. Prices of fresh embryos at which an exclusive IVP-ET system financially breaks even with the comparable AI system in yr 15 and for an investment period of 15 yr were also estimated. More surplus heifer calves were sold from the IVP-ET systems than from the comparable AI systems. The surplus calves from the IVP-ET systems were also genetically superior to the surplus calves from the comparable AI systems, which might be reflected in their market value as a premium price. The most profitable scenario among the 4 IVP-ET scenarios in yr 15 was the one in which NM$ was maximized in the herd. This scenario had an additional profit of $8/cow compared with a similar AI scenario that maximized NM$, provided that surplus heifer calves could be sold at a premium price based on the superiority of the EBV of NM$. For the IVP-ET system to be at least as profitable as the comparable AI system during a 15-yr investment period, the surplus calves from the IVP-ET system needed to be sold at the premium prices. The break-even price of fresh embryos was estimated to be $84 for the exclusive IVP-ET system. This resulted in the same profit as the AI system, which maximized NM$ for a 15-yr investment period and in which heifer calves were sold at a premium price.

 

 

Postnatal phenotype of dairy cows is altered by in vitro embryo production using reverse X-sorted semen.

Author information: Siqueira LGB¹, Dikmen S², Ortega MS³, Hansen PJ⁴.

Date of e-pub: April 2017

Abstract: Abnormal fetuses, neonates, and adult offspring derived by assisted reproductive technologies have been reported in humans and mice and have been associated with increased likelihood of certain adult diseases. To test the hypothesis that bovine females derived by assisted reproductive technologies have altered postnatal growth and adult function, a retrospective cohort study evaluated survival, growth, and production traits of offspring derived by in vitro embryo production (IVP) with conventional (IVP-conv) or reverse X-sorted semen (IVP-sexed), multiple ovulation and embryo transfer, and artificial insemination (AI) in a large dairy herd. Live calves produced by IVP were born slightly heavier compared with AI calves. In addition, IVP-sexed calves had a higher cumulative mortality from 90 to 180 d of age compared with AI offspring. Mortality of IVP-conv and multiple ovulation and embryo transfer offspring was intermediate and not different from AI or IVP-sexed offspring. The altered phenotype of offspring from IVP-sexed extended to adult milk production. Cows derived by IVP-sexed produced less milk, fat, and protein in their first lactation compared with dairy cows derived by AI. Additionally, females born to nulliparous dams had a distinct postnatal phenotype compared with offspring from parous dams even when data were restricted to offspring of surrogate females. In conclusion, procedures associated with in vitro production of embryos involving use of reverse-sorted spermatozoa for fertilization result in an alteration of embryonic programming that persists postnatally and causes an effect on milk production in adulthood. Thus, some benefits of reverse-sorted semen for genetic improvement may be offset by adverse programming events.

 

 

Recent Advances in Nanomaterials for Gene Delivery-A Review.

Author information: Riley MK^1,2, Vermerris W^3,4,5.

Date of e-pub: April 2017

Abstract: With the rapid development of nanotechnology in the recent decade, novel DNA and RNA delivery systems for gene therapy have become available that can be used instead of viral vectors. These non-viral vectors can be made of a variety of materials, including inorganic nanoparticles, carbon nanotubes, liposomes, protein and peptide-based nanoparticles, as well as nanoscale polymeric materials. They have as advantages over viral vectors a decreased immune response, and additionally offer flexibility in design, allowing them to be functionalized and targeted to specific sites in a biological system with low cytotoxicity. The focus of this review is to provide an overview of novel nanotechnology-based methods to deliver DNA and small interfering RNAs into biological systems.

 

 

Complete Genome Sequences of Identical Zika virus Isolates in a Nursing Mother and Her Infant.

Author information: Blohm GM^1,2, Lednicky JA^3,4, Márquez M^2,5, White SK^3,4, Loeb JC⁴, Pacheco CA⁶, Nolan DJ^3,7, Paisie T^3,7, Salemi M^3,7, Rodríguez-Morales AJ⁸, Morris JG Jr^3,9, Pulliam JRC^1,3,10, Carrillo AS¹¹, Plaza JD¹¹, Paniz-Mondolfi AE¹².

Date of e-pub: April 2017

Abstract: Complete genome sequences were obtained for Zika viruses isolated from the breast milk of a Venezuelan patient and her child, who was exclusively breastfeeding at the time. These sequences are the first to be reported from a presumptive autochthonous postnatal transmission case from mother to child in Venezuela.

 

 

Quercetin, a natural product supplement, impairs mitochondrial bioenergetics and locomotor behavior in larval zebrafish (Danio rerio).

Author information: Zhang JL¹, Laurence Souders C 2nd², Denslow ND², Martyniuk CJ³.

Date of e-pub: April 2017

Abstract: Quercetin is a natural product that is sold as a supplement in health food stores. While there are reported benefits for this flavonoid as a dietary supplement due to antioxidant properties, the full scope of its biological interactions has not been fully addressed. To learn more about the mechanisms of action related to quercetin, we exposed zebrafish (Danio rerio) embryos to 1 and 10μg/L quercetin for 96h starting at 3h post fertilization. Quercetin up to 10μg/L did not induce significant mortality in developing fish, but did increase prevalence of an upward-curved dorsal plane in hatched larvae. To determine whether this developmental defect was potentially related to mitochondrial bioenergetics during development, we measured oxygen consumption rate in whole embryos following a 24-hour exposure to quercetin. Basal mitochondrial and ATP-linked respiration were decreased at 1 and 10μg/L quercetin, and maximal respiration was decreased at 10μg/L quercetin, suggesting that quercetin impairs mitochondrial bioenergetics. This is proposed to be related to the deformities observed during development. Due to the fact that ATP production was affected by quercetin, larval behaviors related to locomotion were investigated, as well as transcriptional responses of six myogenesis transcripts. Quercetin at 10μg/L significantly reduced the swimming velocity of zebrafish larvae. The expression levels of both myostatin A (mstna) and myogenic differentiation (myoD) were also altered by quercetin. Mstna, an inhibitory factor for myogenesis, was significantly increased at 1μg/L quercetin exposure, while myoD, a stimulatory factor for myogenesis, was significantly increased at 10μg/L quercetin exposure. There were no changes in transcripts related to apoptosis (bcl2, bax, casp3, casp7), but we did observe a decrease in mRNA levels for catalase (cat) in fish exposed to each dose, supporting an oxidative stress response. Our data support the hypothesis that quercetin may affect locomotion and induce deformities in zebrafish larvae by diminishing ATP production and by altering the expression of transcripts related to muscle formation and activity.

 

 

Role of CC Cytokines in Spatial Arrangement of the Inner Cell Mass of the Bovine Embryo.

Author information: Negrón-Pérez VM¹, Vargas-Franco D², Hansen PJ¹.

Date of e-pub: April 2017

Abstract: The process of spatial rearrangement of cells of the inner cell mass (ICM) that are destined to become hypoblast is not well understood. The observation that the chemokine CCL24 and several other genes involved in chemokine signaling are expressed more in the ICM than the trophectoderm of the bovine embryo resulted in the hypothesis that CCL24 participates in spatial organization of the ICM. Temporally, expression of CCL24 in the bovine embryo occurs coincident with blastocyst formation: transcript abundance was low until the late morula stage, peaked in the blastocyst at Day 7 of development and declined by Day 9. Treatment of embryos with two separate antagonists of CCR3 (the prototypical receptor for CCL24) decreased the percent of GATA6+ cells (hypoblast precursors) that were located in the outside of the ICM. Similarly, injection of zygotes with a CCL24-specific morpholino decreased the percent of GATA6+ cells in the outside of the ICM. In conclusion, CCL24 assists in spatial arrangement of the ICM in the bovine embryo. This experiment points to new functions of chemokine signaling in the bovine embryo and is consistent with the idea that cell migration is involved in the spatial organization of hypoblast cells in the blastocyst.

 

 

Insights into the evolution of hydroxyproline rich glycoproteins from 1000 plant transcriptomes.

Author information: Johnson KL¹, Cassin AM², Lonsdale A³, Wong GK⁴, Soltis D⁵, Miles NW⁶, Melkonian M⁷, Melkonian B⁷, Deyholos MK⁸, Leebens-Mack J⁹, Rothfels CJ¹⁰, Stevenson DW¹¹, Graham SW¹², Wang X¹³, Wu S¹³, Pires JC¹⁴, Edger PP¹⁵, Carpenter EJ¹⁶, Bacic A¹⁷, Doblin MS¹⁸, Schultz CJ¹⁹.

Date of e-pub: April 2017

Abstract: The carbohydrate-rich cell walls of land plants and algae have been the focus of much interest given the value of cell wall based products to our current and future economies. Hydroxyproline-rich glycoproteins (HRGPs), a major group of wall glycoproteins, play important roles in plant growth and development, yet little is known about how they have evolved in parallel with the polysaccharide components of walls. We investigate the origins and evolution of the HRGP superfamily, which is commonly divided into three major multigene families: the arabinogalactan-proteins (AGPs), extensins (EXTs) and proline-rich proteins (PRPs). Using MAAB, a newly developed bioinformatics pipeline, we identified HRGPs in sequences from the 1000 plants (1KP) transcriptome project (www.onekp.com). Our analyses provide new insights into the evolution of HRGPs across major evolutionary milestones, including the transition to land and early radiation of angiosperms. Significantly, data mining reveals the origin of GPI-anchored AGPs in green algae and a 3-4 fold increase in GPI-AGPs in liverworts and mosses. The first detection of cross-linking (CL)-EXTs is observed in bryophytes, which suggest that CL-EXTs arose though the juxtaposition of pre-existing SPn EXT glycomotifs with refined Y-based motifs. We also detected the loss of CL-EXT in a few lineages, including the grass family (Poaceae), that have a cell wall composition distinct from other monocots and eudicots. A key challenge in HRGP research is tracking individual HRGPs throughout evolution. Using the 1KP output we were able to find putative orthologs of Arabidopsis pollen-specific GPI-AGPs in basal eudicots.

 

 

Whole genome sequencing reveals a novel deletion variant in the KIT gene in horses with white spotted coat colour phenotypes.

Author information: Dürig N^1,2, Jude R^1,2,3, Holl H^4,5, Brooks SA⁴, Lafayette C⁵, Jagannathan V^1,2, Leeb T^1,2.

Date of e-pub: April 2017

Abstract: White spotting phenotypes in horses can range in severity from the common white markings up to completely white horses. EDNRB, KIT, MITF, PAX3 and TRPM1 represent known candidate genes for such phenotypes in horses. For the present study, we re-investigated a large horse family segregating a variable white spotting phenotype, for which conventional Sanger sequencing of the candidate genes’ individual exons had failed to reveal the causative variant. We obtained whole genome sequence data from an affected horse and specifically searched for structural variants in the known candidate genes. This analysis revealed a heterozygous ~1.9-kb deletion spanning exons 10-13 of the KIT gene (chr3:77,740,239_77,742,136del1898insTATAT). In continuity with previously named equine KIT variants we propose to designate the newly identified deletion variant W22. We had access to 21 horses carrying the W22 allele. Four of them were compound heterozygous W20/W22 and had a completely white phenotype. Our data suggest that W22 represents a true null allele of the KIT gene, whereas the previously identified W20 leads to a partial loss of function. These findings will enable more precise genetic testing for depigmentation phenotypes in horses.

 

 

Tet2 loss leads to hypermutagenicity in haematopoietic stem/progenitor cells.

Author information: Pan F^1,2,3, Wingo TS^4,5,6, Zhao Z⁷, Gao R¹, Makishima H⁸, Qu G^1,2, Lin L⁴, Yu M⁹, Ortega JR¹⁰, Wang J², Nazha A⁸, Chen L⁴, Yao B⁴, Liu C¹, Chen S¹, Weeks O³, Ni H¹¹, Phillips BL¹², Huang S¹³, Wang J¹⁴, He C⁹, Li GM¹⁰, Radivoyevitch T⁸, Aifantis I^15,16, Maciejewski JP⁸, Yang FC^1,2, Jin P⁴, Xu M^1,2.

Date of e-pub: April 2017

Abstract: TET2 is a dioxygenase that catalyses multiple steps of 5-methylcytosine oxidation. Although TET2 mutations frequently occur in various types of haematological malignancies, the mechanism by which they increase risk for these cancers remains poorly understood. Here we show that Tet2-/- mice develop spontaneous myeloid, T- and B-cell malignancies after long latencies. Exome sequencing of Tet2-/- tumours reveals accumulation of numerous mutations, including Apc, Nf1, Flt3, Cbl, Notch1 and Mll2, which are recurrently deleted/mutated in human haematological malignancies. Single-cell-targeted sequencing of wild-type and premalignant Tet2-/- Lin-c-Kit+ cells shows higher mutation frequencies in Tet2-/- cells. We further show that the increased mutational burden is particularly high at genomic sites that gained 5-hydroxymethylcytosine, where TET2 normally binds. Furthermore, TET2-mutated myeloid malignancy patients have significantly more mutational events than patients with wild-type TET2. Thus, Tet2 loss leads to hypermutagenicity in haematopoietic stem/progenitor cells, suggesting a novel TET2 loss-mediated mechanism of haematological malignancy pathogenesis.

 

 

ESPRIT-Forest: Parallel clustering of massive amplicon sequence data in subquadratic time.

Author information: Cai Y¹, Zheng W², Yao J³, Yang Y¹, Mai V⁴, Mao Q³, Sun Y^2,3,5.

Date of e-pub: April 2017

Abstract: The rapid development of sequencing technology has led to an explosive accumulation of genomic sequence data. Clustering is often the first step to perform in sequence analysis, and hierarchical clustering is one of the most commonly used approaches for this purpose. However, it is currently computationally expensive to perform hierarchical clustering of extremely large sequence datasets due to its quadratic time and space complexities. In this paper we developed a new algorithm called ESPRIT-Forest for parallel hierarchical clustering of sequences. The algorithm achieves subquadratic time and space complexity and maintains a high clustering accuracy comparable to the standard method. The basic idea is to organize sequences into a pseudo-metric based partitioning tree for sub-linear time searching of nearest neighbors, and then use a new multiple-pair merging criterion to construct clusters in parallel using multiple threads. The new algorithm was tested on the human microbiome project (HMP) dataset, currently one of the largest published microbial 16S rRNA sequence dataset. Our experiment demonstrated that with the power of parallel computing it is now compu- tationally feasible to perform hierarchical clustering analysis of tens of millions of sequences. The software is available at http://www.acsu.buffalo.edu/∼yijunsun/lab/ESPRIT-Forest.html.

 

 

Insulitis in Autoantibody-Positive Pancreatic Donor With History of Gestational Diabetes Mellitus.

Author information: Jackson J^1,2, Posgai A¹, Campbell-Thompson M¹, Kusmartseva I³.

Date of e-pub: May 2017

Abstract: N/A

 

 

Disulfide bond disrupting agents activate the unfolded protein response in EGFR- and HER2-positive breast tumor cells.

Author information: Ferreira RB¹, Wang M^2,3, Law ME^2,3, Davis BJ^2,3, Bartley AN¹, Higgins PJ⁴, Kilberg MS⁵, Santostefano KE⁶, Terada N⁶, Heldermon CD⁷, Castellano RK¹, Law BK^2,3.

Date of e-pub: April 2017

Abstract: Many breast cancer deaths result from tumors acquiring resistance to available therapies. Thus, new therapeutic agents are needed for targeting drug-resistant breast cancers. Drug-refractory breast cancers include HER2+ tumors that have acquired resistance to HER2-targeted antibodies and kinase inhibitors, and “Triple-Negative” Breast Cancers (TNBCs) that lack the therapeutic targets Estrogen Receptor, Progesterone Receptor, and HER2. A significant fraction of TNBCs overexpress the HER2 family member Epidermal Growth Factor Receptor (EGFR). Thus agents that selectively kill EGFR+ and HER2+ tumors would provide new options for breast cancer therapy. We previously identified a class of compounds we termed Disulfide bond Disrupting Agents (DDAs) that selectively kill EGFR+ and HER2+ breast cancer cells in vitro and blocked the growth of HER2+ breast tumors in an animal model. DDA-dependent cytotoxicity was found to correlate with downregulation of HER1-3 and Akt dephosphorylation. Here we demonstrate that DDAs activate the Unfolded Protein Response (UPR) and that this plays a role in their ability to kill EGFR+ and HER2+ cancer cells. The use of breast cancer cell lines ectopically expressing EGFR or HER2 and pharmacological probes of UPR revealed all three DDA responses: HER1-3 downregulation, Akt dephosphorylation, and UPR activation, contribute to DDA-mediated cytotoxicity. Significantly, EGFR overexpression potentiates each of these responses. Combination studies with DDAs suggest that they may be complementary with EGFR/HER2-specific receptor tyrosine kinase inhibitors and mTORC1 inhibitors to overcome drug resistance.

 

 

LPS Stimulation of Cord Blood Reveals a Newborn-Specific Neutrophil Transcriptomic Response and Cytokine Production.

Author information: Mathias B¹, Mira JC, Rehfuss JP, Rincon JC, Ungaro R, Nacionales DC, Lopez MC, Baker HV, Moldawer LL, Larson SD.

Date of e-pub: May 2017

Abstract: The neonatal innate immune system differs to microbial infection both quantitatively and qualitatively when compared with adults. Here, we provide the first genome-wide ex-vivo expression profile of umbilical cord blood (UCB) neutrophils from full-term infants prior to and in response to whole-blood lipopolysaccharide (LPS) stimulation. Additionally, we provide cytokine expression prior to and following LPS stimulation. The genomic expression and cytokine profile are compared with LPS-stimulated whole blood from healthy adult subjects (HC).

Whole blood from UCB (n = 6) and HC (n = 6) was studied at baseline or was stimulated for 24 h with 100 ngs/mL of LPS. CD66b neutrophils were subsequently isolated with microfluidic techniques and genome-wide expression analyses were performed. Ingenuity Pathway Analysis (IPA) software was utilized to predict downstream functional effects. Additionally, cytokine concentrations in whole blood prior to and after 24 h of LPS incubation were determined.

LPS stimulated whole blood from UCB demonstrated significant differences in both ex-vivo cytokine production and PMN gene expression. Mixed-effect modeling identified 1,153 genes whose expression changed significantly in UCB and HC after exposure to LPS (P < 0.001 with a minimum 1.5-fold change). IPA downstream predictions suggest that PMNs from UCB fail to effectively upregulate genes associated with activation, phagocytosis, and chemotaxis in response to LPS stimulation. Furthermore, whole blood from UCB showed increased interleukin (IL)-10 production to LPS, but failed to significantly increase several pro-inflammatory cytokines.

LPS-stimulated whole blood from UCB exhibited a markedly suppressed inflammatory cytokine production and PMN innate immune genome response. These differences in gene expression and cytokine production may be an adaptive response to a prior fetal environment, but may also explain their increased susceptibility to infections. Characterization of these deficits is the first step toward developing prophylactic and therapeutic interventions.

 

 

Semipermeable Functional DNA-Encapsulated Nanocapsules as Protective Bioreactors for Biosensing in Living Cells.

Author information: Hong CY^1,2, Wu SX¹, Li SH¹, Liang H¹, Chen S¹, Li J^1,3, Yang HH¹, Tan W^3,2.

Date of e-pub: May 2017

Abstract: The development of functional DNA-based nanosensors in living cells has experienced some design challenges, including, for example, poor cellular uptake, rapid nuclease degradation, and high false positives. Herein, we designed selectively permeable poly(methacrylic acid) (PMA) nanocapsules to encapsulate functional DNAs for metal ions and small-molecules sensing in living cells. Since functional DNAs are concentrated in the nanocapsules, an increasing reaction rate is obtained in vitro. During endocytosis, polymeric capsules simultaneously improve cellular uptake of functional DNAs and preserve their structural integrity inside the confined capsule space. More importantly, selective shell permeability allows for the free diffusion of small molecular targets through capsule shells but limits the diffusion of large biomolecules, such as nuclease and nonspecific protein. Compared to the free DNAzyme, PMA nanocapsules could reduce false positives and enhance detection accuracy. Furthermore, PMA nanocapsules are biocompatible and biodegradable. Through the controllability of wall thickness, permeability, and size distribution, these nanocapsules could be expanded easily to other targets, such as microRNAs, small peptides, and metabolites. These nanocapsules will pave the way for in situ monitoring of various biological processes in living cells and in vivo.

 

 

Functional characterization of LotP from Liberibacter asiaticus.

Author information: Loto F^1,2, Coyle JF¹, Padgett KA^1,3, Pagliai FA¹, Gardner CL¹, Lorca GL¹, Gonzalez CF¹.

Date of e-pub: May 2017

Abstract: Liberibacter asiaticus is an unculturable parasitic bacterium of the alphaproteobacteria group hosted by both citrus plants and a psyllid insect vector (Diaphorina citri). In the citrus tree, the bacteria thrive only inside the phloem, causing a systemically incurable and deadly plant disease named citrus greening or Huanglongbing. Currently, all commercial citrus cultivars in production are susceptible to L. asiaticus, representing a serious threat to the citrus industry worldwide. The technical inability to isolate and culture L. asiaticus has hindered progress in understanding the biology of this bacterium directly. Consequently, a deep understanding of the biological pathways involved in the regulation of host-pathogen interactions becomes critical to rationally design future and necessary strategies of control. In this work, we used surrogate strains to evaluate the biochemical characteristics and biological significance of CLIBASIA_03135. This gene, highly induced during early stages of plant infection, encodes a 23 kDa protein and was renamed in this work as LotP. This protein belongs to an uncharacterized family of proteins with an overall structure resembling the LON protease N-terminus. Co-immunoprecipitation assays allowed us to identify the Liberibacter chaperonin GroEL as the main LotP-interacting protein. The specific interaction between LotP and GroEL was reconstructed and confirmed using a two-hybrid system in Escherichia coli. Furthermore, it was demonstrated that LotP has a native molecular weight of 44 kDa, corresponding to a dimer in solution with ATPase activity in vitro. In Liberibacter crescens, LotP is strongly induced in response to conditions with high osmolarity but repressed at high temperatures. Electrophoretic mobility shift assay (EMSA) results suggest that LotP is a member of the LdtR regulon and could play an important role in tolerance to osmotic stress.

 

 

Association of the HLA-B*53:01 Allele With Drug Reaction With Eosinophilia and Systemic Symptoms (DRESS) Syndrome During Treatment of HIV Infection With Raltegravir.

Author information: Thomas M^1,2, Hopkins C², Duffy E², Lee D³, Loulergue P⁴, Ripamonti D⁵, Ostrov DA⁶, Phillips E^7,8,9.

Date of e-pub: May 2017

Abstract: Drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome is a rare, severe adverse event during treatment with raltegravir. The occurrence of DRESS syndrome during treatment with other drugs is strongly associated with particular HLA alleles.

We performed HLA testing in 3 of the 5 patients previously reported to have developed raltegravir-induced DRESS syndrome and in 1 previously unreported patient. We then used virtual modeling to visualize interactions between raltegravir and the imputed HLA molecule.

Five of the 6 patients who developed raltegravir-induced DRESS syndrome were African, and 1 was Hispanic. HLA typing was performed in 4 patients, all of whom carried both the HLA-B*53 allele and the HLA-C*04 allele to which it is commonly haplotypic. No other HLA alleles were shared by all of the tested patients. Given the approximate prevalence of HLA-B*53 carriage in African (20%) and Hispanic (6%) populations, the probability of all 4 patients being HLA-B*53 carriers, and 2 of 3 African patients being homozygous for HLA-B*53:01, is approximately 0.00002.

These data implicate the prevalent African allele HLA-B*53:01 in the immunopathogenesis of raltegravir-induced DRESS syndrome. Although the immunopathogenic mechanisms are currently unknown, virtual modeling suggests that raltegravir may bind within the antigen binding cleft of the HLA-B*53:01 molecule, but not within the closely related HLA-B*35:01 molecule. Further studies are necessary to confirm the strength of the association between carriage of the HLA-B*53:01 allele and raltegravir-induced DRESS syndrome, and the potential utility of HLA screening before raltegravir treatment.

 

 

Dynamic Gene Regulatory Networks of Human Myeloid Differentiation.

Author information: Ramirez RN¹, El-Ali NC¹, Mager MA¹, Wyman D², Conesa A³, Mortazavi A⁴.

Date of e-pub: April 2017

Abstract: The reconstruction of gene regulatory networks underlying cell differentiation from high-throughput gene expression and chromatin data remains a challenge. Here, we derive dynamic gene regulatory networks for human myeloid differentiation using a 5-day time series of RNA-seq and ATAC-seq data. We profile HL-60 promyelocytes differentiating into macrophages, neutrophils, monocytes, and monocyte-derived macrophages. We find a rapid response in the expression of key transcription factors and lineage markers that only regulate a subset of their targets at a given time, which is followed by chromatin accessibility changes that occur later along with further gene expression changes. We observe differences between promyelocyte- and monocyte-derived macrophages at both the transcriptional and chromatin landscape level, despite using the same differentiation stimulus, which suggest that the path taken by cells in the differentiation landscape defines their end cell state. More generally, our approach of combining neighboring time points and replicates to achieve greater sequencing depth can efficiently infer footprint-based regulatory networks from long series data.

 

 

Pharmacogenetic Associations of β1-Adrenergic Receptor Polymorphisms With Cardiovascular Outcomes in the SPS3 Trial (Secondary Prevention of Small Subcortical Strokes).

Author information: Magvanjav O¹, McDonough CW¹, Gong Y¹, McClure LA¹, Talbert RL¹, Horenstein RB¹, Shuldiner AR¹, Benavente OR¹, Mitchell BD¹, Johnson JA²; NINDS SiGN (Stroke Genetics Network).

Date of e-pub: May 2017

Abstract: Functional polymorphisms (Ser49Gly and Arg389Gly) in ADRB1 have been associated with cardiovascular and β-blocker response outcomes. Herein we examined associations of these polymorphisms with major adverse cardiovascular events (MACE), with and without stratification by β-blocker treatment in patients with a history of stroke.

Nine hundred and twenty-six participants of the SPS3 trial’s (Secondary Prevention of Small Subcortical Strokes) genetic substudy with hypertension were included. MACE included stroke, myocardial infarction, and all-cause death. Kaplan-Meier and multivariable Cox regression analyses were used. Because the primary component of MACE was ischemic stroke, we tested the association of Ser49Gly with ischemic stroke among 41 475 individuals of European and African ancestry in the NINDS (National Institute of Neurological Disorders and Stroke) SiGN (Stroke Genetics Network).

MACE was higher in carriers of the Gly49 allele than in those with the Ser49Ser genotype (10.5% versus 5.4%, log-rank P=0.005). Gly49 carrier status was associated with MACE (hazard ratio, 1.62; 95% confidence interval, 1.00-2.68) and ischemic stroke (hazard ratio, 1.81; 95% confidence interval, 1.01-3.23) in SPS3 and with small artery ischemic stroke (odds ratio, 1.14; 95% confidence interval, 1.03-1.26) in SiGN. In SPS3, β-blocker-treated Gly49 carriers had increased MACE versus non-β-blocker-treated individuals and noncarriers (hazard ratio, 2.03; 95% confidence interval, 1.20-3.45). No associations were observed with the Arg389Gly polymorphism.

Among individuals with previous small artery ischemic stroke, the ADRB1 Gly49 polymorphism was associated with MACE, particularly small artery ischemic stroke, a risk that may be increased among β-blocker-treated individuals. Further research is needed to define β-blocker benefit among ischemic stroke patients by ADRB1 genotype.

 

 

Structure Determination of Mycobacterium tuberculosis Serine Protease Hip1 (Rv2224c).

Author information: Naffin-Olivos JL¹, Daab A^1,2, White A¹, Goldfarb NE³, Milne AC¹, Liu D⁴, Baikovitz J⁵, Dunn BM⁶, Rengarajan J⁷, Petsko GA⁸, Ringe D¹.

Date of e-pub: May 2017

Abstract: The Mycobacterium tuberculosis (Mtb) serine protease Hip1 (hydrolase important for pathogenesis; Rv2224c) promotes tuberculosis (TB) pathogenesis by impairing host immune responses through proteolysis of a protein substrate, Mtb GroEL2. The cell surface localization of Hip1 and its immunomodulatory functions make Hip1 a good drug target for new adjunctive immune therapies for TB. Here, we report the crystal structure of Hip1 to a resolution of 2.6 Å and the kinetic studies of the enzyme against model substrates and the protein GroEL2. The structure shows a two-domain protein, one of which contains the catalytic residues that are the signature of a serine protease. Surprisingly, a threonine is located within the active site close enough to hydrogen bond with the catalytic residues Asp463 and His490. Mutation of this residue, Thr466, to alanine established its importance for function. Our studies provide insights into the structure of a member of a novel family of proteases. Knowledge of the Hip1 structure will aid in designing inhibitors that could block Hip1 activity.

 

 

Development of a QTL-environment-based predictive model for node addition rate in common bean.

Author information: Zhang L¹, Gezan SA², Eduardo Vallejos C³, Jones JW¹, Boote KJ⁴, Clavijo-Michelangeli JA⁴, Bhakta M³, Osorno JM⁵, Rao I⁶, Beebe S⁶, Roman-Paoli E⁷, Gonzalez A⁷, Beaver J⁷, Ricaurte J⁶, Colbert R⁵, Correll MJ⁸.

Date of e-pub: May 2017

Abstract: This work reports the effects of the genetic makeup, the environment and the genotype by environment interactions for node addition rate in an RIL population of common bean. This information was used to build a predictive model for node addition rate. To select a plant genotype that will thrive in targeted environments it is critical to understand the genotype by environment interaction (GEI). In this study, multi-environment QTL analysis was used to characterize node addition rate (NAR, node day- 1) on the main stem of the common bean (Phaseolus vulgaris L). This analysis was carried out with field data of 171 recombinant inbred lines that were grown at five sites (Florida, Puerto Rico, 2 sites in Colombia, and North Dakota). Four QTLs (Nar1, Nar2, Nar3 and Nar4) were identified, one of which had significant QTL by environment interactions (QEI), that is, Nar2 with temperature. Temperature was identified as the main environmental factor affecting NAR while day length and solar radiation played a minor role. Integration of sites as covariates into a QTL mixed site-effect model, and further replacing the site component with explanatory environmental covariates (i.e., temperature, day length and solar radiation) yielded a model that explained 73% of the phenotypic variation for NAR with root mean square error of 16.25% of the mean. The QTL consistency and stability was examined through a tenfold cross validation with different sets of genotypes and these four QTLs were always detected with 50-90% probability. The final model was evaluated using leave-one-site-out method to assess the influence of site on node addition rate. These analyses provided a quantitative measure of the effects on NAR of common beans exerted by the genetic makeup, the environment and their interactions.

 

 

Duvoglustat HCl Increases Systemic and Tissue Exposure of Active Acid α-Glucosidase in Pompe Patients Co-administered with Alglucosidase α.

Author information: Kishnani P¹, Tarnopolsky M², Roberts M³, Sivakumar K⁴, Dasouki M⁵, Dimachkie MM⁵, Finanger E⁶, Goker-Alpan O⁷, Guter KA⁸, Mozaffar T⁹, Pervaiz MA¹⁰, Laforet P¹¹, Levine T¹², Adera M¹³, Lazauskas R¹⁴, Sitaraman S¹⁴, Khanna R¹⁴, Benjamin E¹⁴, Feng J¹⁴, Flanagan JJ¹⁵, Barth J¹⁴, Barlow C¹⁶, Lockhart DJ¹⁷, Valenzano KJ¹⁴, Boudes P¹⁸, Johnson FK¹⁹, Byrne B²⁰.

Date of e-pub: May 2017

Abstract: Duvoglustat HCl (AT2220, 1-deoxynojirimycin) is an investigational pharmacological chaperone for the treatment of acid α-glucosidase (GAA) deficiency, which leads to the lysosomal storage disorder Pompe disease, which is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. The current standard of care is enzyme replacement therapy with recombinant human GAA (alglucosidase alfa [AA], Genzyme). Based on preclinical data, oral co-administration of duvoglustat HCl with AA increases exposure of active levels in plasma and skeletal muscles, leading to greater substrate reduction in muscle. This phase 2a study consisted of an open-label, fixed-treatment sequence that evaluated the effect of single oral doses of 50 mg, 100 mg, 250 mg, or 600 mg duvoglustat HCl on the pharmacokinetics and tissue levels of intravenously infused AA (20 mg/kg) in Pompe patients. AA alone resulted in increases in total GAA activity and protein in plasma compared to baseline. Following co-administration with duvoglustat HCl, total GAA activity and protein in plasma were further increased 1.2- to 2.8-fold compared to AA alone in all 25 Pompe patients; importantly, muscle GAA activity was increased for all co-administration treatments from day 3 biopsy specimens. No duvoglustat-related adverse events or drug-related tolerability issues were identified.

 

 

Structural Insights into Human Bocaparvoviruses.

Author information: Mietzsch M¹, Kailasan S¹, Garrison J¹, Ilyas M¹, Chipman P¹, Kantola K², Janssen ME³, Spear J⁴, Sousa D⁴, McKenna R¹, Brown K⁵, Söderlund-Venermo M², Baker T³, Agbandje-McKenna M⁶.

Date of e-pub: May 2017

Abstract: Bocaparvoviruses are emerging pathogens of the Parvoviridae family. Human bocavirus 1 (HBoV1) causes severe respiratory infections and HBoV2 to HBoV4 cause gastrointestinal infections in young children. Recent reports of life-threatening cases, lack of direct treatment or vaccination, and a limited understanding of their disease mechanisms highlight the need to study these pathogens on a molecular and structural level for the development of therapeutics. Toward this end, the capsid structures of HBoV1, HBoV3, and HBoV4 were determined to a resolution of 2.8 to 3.0 Å by cryo-electron microscopy and three-dimensional image reconstruction. The bocaparvovirus capsids, which display different tissue tropisms, have features in common with other parvoviruses, such as depressions at the icosahedral 2-fold symmetry axis and surrounding the 5-fold symmetry axis, protrusions surrounding the 3-fold symmetry axis, and a channel at the 5-fold symmetry axis. However, unlike other parvoviruses, densities extending the 5-fold channel into the capsid interior are conserved among the bocaparvoviruses and are suggestive of a genus-specific function. Additionally, their major viral protein 3 contains loops with variable regions at their apexes conferring capsid surface topologies different from those of other parvoviruses. Structural comparisons at the strain (HBoV) and genus (bovine parvovirus and HBoV) levels identified differences in surface loops that are functionally important in host/tissue tropism, pathogenicity, and antigenicity in other parvoviruses and likely play similar roles in these viruses. This study thus provides a structural framework to characterize determinants of host/tissue tropism, pathogenicity, and antigenicity for the development of antiviral strategies to control human bocavirus infections.IMPORTANCE Human bocaviruses are one of only a few members of the Parvoviridae family pathogenic to humans, especially young children and immunocompromised adults. There are currently no treatments or vaccines for these viruses or the related enteric bocaviruses. This study obtained the first high-resolution structures of three human bocaparvoviruses determined by cryo-reconstruction. HBoV1 infects the respiratory tract, and HBoV3 and HBoV4 infect the gastrointestinal tract, tissues that are likely targeted by the capsid. Comparison of these viruses provides information on conserved bocaparvovirus-specific features and variable regions resulting in unique surface topologies that can serve as guides to characterize HBoV determinants of tissue tropism and antigenicity in future experiments. Based on the comparison to other existing parvovirus capsid structures, this study suggests capsid regions that likely control successful infection, including determinants of receptor attachment, host cell trafficking, and antigenic reactivity. Overall, these observations could impact efforts to design antiviral strategies and vaccines for HBoVs.

 

 

Fluorescence Resonance Energy Transfer-Based DNA Tetrahedron Nanotweezer for Highly Reliable Detection of Tumor-Related mRNA in Living Cells.

Author information: He L¹, Lu DQ¹, Liang H¹, Xie S¹, Luo C¹, Hu M¹, Xu L¹, Zhang X¹, Tan W^1,2.

Date of e-pub: April 2017

Abstract: Accurate detection and imaging of tumor-related mRNA in living cells hold great promise for early cancer detection. However, currently, most probes designed to image intracellular mRNA confront intrinsic interferences arising from complex biological matrices and resulting in inevitable false-positive signals. To circumvent this problem, an intracellular DNA nanoprobe, termed DNA tetrahedron nanotweezer (DTNT), was developed to reliably image tumor-related mRNA in living cells based on the FRET (fluorescence resonance energy transfer) “off” to “on” signal readout mode. DTNT was self-assembled from four single-stranded DNAs. In the absence of target mRNA, the respectively labeled donor and acceptor fluorophores are separated, thus inducing low FRET efficiency. However, in the presence of target mRNA, DTNT alters its structure from the open to closed state, thus bringing the dual fluorophores into close proximity for high FRET efficiency. The DTNT exhibited high cellular permeability, fast response and excellent biocompatibility. Moreover, intracellular imaging experiments showed that DTNT could effectively distinguish cancer cells from normal cells and, moreover, distinguish among changes of mRNA expression levels in living cells. The DTNT nanoprobe also exhibits minimal effect of probe concentration, distribution and laser power as other ratiometric probe. More importantly, as a result of the FRET “off” to “on” signal readout mode, the DTNT nanoprobe almost entirely avoids false-positive signals due to intrinsic interferences, such as nuclease digestion, protein binding and thermodynamic fluctuations in complex biological matrices. This design blueprint can be applied to the development of powerful DNA nanomachines for biomedical research and clinical early diagnosis.

 

 

Electrophysiologic features of ulnar neuropathy in childhood and adolescence.

Author information: Karakis I¹, Liew W², Fournier HS³, Jones HR Jr⁴, Darras BT³, Kang PB⁵.

Date of e-pub: May 2017

Abstract: To analyze patterns of nerve injury in pediatric ulnar neuropathy (PUN).

Retrospective analysis of 49 children with PUN.

Sensory loss in digit V was the prevailing complaint (89%). Predominant localization was at the elbow (55%). Diminished ulnar SNAP was the most common abnormality (71%) with median axon loss estimate (MAXE) of 62%. Dorsal ulnar cutaneous (DUC) sensory nerve action potential (SNAP) was reduced in 55% with MAXE of 43%. Abductor digiti minimi (ADM) and first dorsal interosseous (FDI) compound muscle action potential (CMAP) were reduced half of the time, with MAXE of 30% and 28% respectively. There was high correlation between ulnar sensory MAXE and ADM MAXE (r=0.76, p<0.0001), FDI MAXE (r=0.81, p<0.0001) and DUC MAXE (r=0.60, p=0.0048). Neurogenic changes were seen in the ADM, FDI, flexor carpi ulnaris (FCU) and flexor digitorum profundus IV (FDP IV) in 79%, 77%, 25% and 35% respectively. Pathophysiology was demyelinating in 27%, axonal in 59% and mixed in 14%.

In proximal axonal lesions, sensory fibers to digit V and motor fibers to distal muscles are predominantly affected, whereas in demyelinating lesions, slowing occurs twice as frequently as conduction block.

There is frequent axonal and fascicular injury in PUN.

 

 

DNA probes for monitoring dynamic and transient molecular encounters on live cell membranes.

Author information: You M^1,2,3,4, Lyu Y^1,2, Han D^1,2, Qiu L^1,2, Liu Q¹, Chen T^1,2, Sam Wu C^1,2, Peng L², Zhang L^1,2, Bao G⁵, Tan W^1,2.

Date of e-pub: May 2017

Abstract: Cells interact with the extracellular environment through molecules expressed on the membrane. Disruption of these membrane-bound interactions (or encounters) can result in disease progression. Advances in super-resolution microscopy have allowed membrane encounters to be examined, however, these methods cannot image entire membranes and cannot provide information on the dynamic interactions between membrane-bound molecules. Here, we show a novel DNA probe that can transduce transient membrane encounter events into readable cumulative fluorescence signals. The probe, which translocates from one anchor site to another, mimicking motor proteins, is realized through a toehold-mediated DNA strand displacement reaction. Using this probe, we successfully monitored rapid encounter events of membrane lipid domains using flow cytometry and fluorescence microscopy. Our results show a preference for encounters within the same lipid domains.

 

 

Impact of the CYP2C19 genotype on voriconazole exposure in adults with invasive fungal infections.

Author information: Hamadeh IS¹, Klinker KP, Borgert SJ, Richards AI, Li W, Mangal N, Hiemenz JW, Schmidt S, Langaee TY, Peloquin CA, Johnson JA, Cavallari LH.

Date of e-pub: May 2017

Abstract: Voriconazole, a first-line agent for the treatment of invasive fungal infections (IFIs), is metabolized by CYP2C19. A significant proportion of patients fail to achieve therapeutic trough concentrations with standard weight-based voriconazole dosing, placing them at increased risk for treatment failure, which can be life threatening. We sought to test the association between the CYP2C19 genotype and subtherapeutic voriconazole concentrations in adults with IFIs.

Adults receiving weight-based voriconazole dosing for the treatment of IFIs were genotyped for the CYP2C19*2, *3, and *17 polymorphisms, and CYP2C19 metabolizer phenotypes were inferred. Steady-state voriconazole trough plasma concentrations and the prevalence of subtherapeutic troughs (

Of 70 patients included (mean age 52.5±18 years), 39% were RMs or UMs. Compared with patients with the other phenotypes, RMs/UMs had a lower steady-state trough concentration (4.26±2.2 vs. 2.86±2.3, P=0.0093) and a higher prevalence of subtherapeutic troughs (16 vs. 52%, P=0.0028), with an odds ratio of 5.6 (95% confidence interval: 1.64-19.24, P=0.0044).

Our findings indicate that adults with the CYP2C19 RM or UM phenotype are more likely to have subtherapeutic concentrations with weight-based voriconazole dosing. These results corroborate previous findings in children and support the potential clinical utility of CYP2C19 genotype-guided voriconazole dosing to avoid underexposure in RMs and UMs.

 

 

Electroacupuncture Promotes Central Nervous System-Dependent Release of Mesenchymal Stem Cells.

Author information: Salazar TE^1,2,3, Richardson MR⁴, Beli E³, Ripsch MS^5,6, George J², Kim Y⁵, Duan Y³, Moldovan L³, Yan Y¹, Bhatwadekar A³, Jadhav V⁵, Smith JA⁵, McGorray S⁷, Bertone AL⁸, Traktuev DO^9,10, March KL^9,10, Colon-Perez LM¹¹, Avin KG¹², Sims E¹³, Mund JA^4,13, Case J^13,14, Deng X¹⁵, Kim MS¹⁶, McDavitt B¹⁷, Boulton ME³, Thinschmidt J¹⁸, Li Calzi S³, Fitz SD¹⁹, Fuchs RK¹², Warden SJ¹², McKinley T²⁰, Shekhar A¹⁹, Febo M¹¹, Johnson PL²¹, Chang LJ^22,23, Gao Z²⁴, Kolonin MG²⁴, Lai S²⁵, Ma J²⁵, Dong X²⁶, White FA^5,6, Xie H²⁷, Yoder MC^4,13, Grant MB³.

Date of e-pub: May 2017

Abstract: Electroacupuncture (EA) performed in rats and humans using limb acupuncture sites, LI-4 and LI-11, and GV-14 and GV-20 (humans) and Bai-hui (rats) increased functional connectivity between the anterior hypothalamus and the amygdala and mobilized mesenchymal stem cells (MSCs) into the systemic circulation. In human subjects, the source of the MSC was found to be primarily adipose tissue, whereas in rodents the tissue sources were considered more heterogeneous. Pharmacological disinhibition of rat hypothalamus enhanced sympathetic nervous system (SNS) activation and similarly resulted in a release of MSC into the circulation. EA-mediated SNS activation was further supported by browning of white adipose tissue in rats. EA treatment of rats undergoing partial rupture of the Achilles tendon resulted in reduced mechanical hyperalgesia, increased serum interleukin-10 levels and tendon remodeling, effects blocked in propranolol-treated rodents. To distinguish the afferent role of the peripheral nervous system, phosphoinositide-interacting regulator of transient receptor potential channels (Pirt)-GCaMP3 (genetically encoded calcium sensor) mice were treated with EA acupuncture points, ST-36 and LIV-3, and GV-14 and Bai-hui and resulted in a rapid activation of primary sensory neurons. EA activated sensory ganglia and SNS centers to mediate the release of MSC that can enhance tissue repair, increase anti-inflammatory cytokine production and provide pronounced analgesic relief. Stem Cells 2017;35:1303-1315.

 

 

The IGNITE Pharmacogenetics Working Group: An Opportunity for Building Evidence with Pharmacogenetic Implementation in a Real-World Setting.

Author information: Cavallari LH¹, Beitelshees AL², Blake KV³, Dressler LG⁴, Duarte JD¹, Elsey A¹, Eichmeyer JN⁵, Empey PE⁶, Franciosi JP⁷, Hicks JK⁸, Holmes AM⁹, Jeng L¹⁰, Lee CR¹¹, Lima JJ³, Limdi NA¹², Modlin J⁵, Obeng AO¹³, Petry N¹⁴, Pratt VM¹⁵, Skaar TC¹⁶, Tuteja S¹⁷, Voora D¹⁸, Wagner M⁵, Weitzel KW¹, Wilke RA¹⁹, Peterson JF²⁰, Johnson JA¹.

Date of e-pub: May 2017

Abstract: N/A

 

 

Aptasensor with Expanded Nucleotide Using DNA Nanotetrahedra for Electrochemical Detection of Cancerous Exosomes.

Author information: Wang S^1,2, Zhang L^3,2, Wan S², Cansiz S², Cui C², Liu Y^3,2, Cai R², Hong C², Teng IT², Shi M^3,2, Wu Y^3,2, Dong Y¹, Tan W^3,2.

Date of e-pub: April 2017

Abstract: Exosomes are extracellular vesicles (50-100 nm) circulating in biofluids as intercellular signal transmitters. Although the potential of cancerous exosomes as tumor biomarkers is promising, sensitive and rapid detection of exosomes remains challenging. Herein, we combined the strengths of advanced aptamer technology, DNA-based nanostructure, and portable electrochemical devices to develop a nanotetrahedron (NTH)-assisted aptasensor for direct capture and detection of hepatocellular exosomes. The oriented immobilization of aptamers significantly improved the accessibility of an artificial nucleobase-containing aptamer to suspended exosomes, and the NTH-assisted aptasensor could detect exosomes with 100-fold higher sensitivity when compared to the single-stranded aptamer-functionalized aptasensor. The present study provides a proof-of-concept for sensitive and efficient quantification of tumor-derived exosomes. We thus expect the NTH-assisted electrochemical aptasensor to become a powerful tool for comprehensive exosome studies.

 

 

tRNA N6-adenosine threonylcarbamoyltransferase defect due to KAE1/TCS3 (OSGEP) mutation manifest by neurodegeneration and renal tubulopathy.

Author information: Edvardson S^1,2, Prunetti L³, Arraf A⁴, Haas D³, Bacusmo JM³, Hu JF⁵, Ta-Shma A¹, Dedon PC^5,6, de Crécy-Lagard V³, Elpeleg O¹.

Date of e-pub: May 2017

Abstract: Post-transcriptional tRNA modifications are numerous and require a large set of highly conserved enzymes in humans and other organisms. In yeast, the loss of many modifications is tolerated under unstressed conditions; one exception is the N6-threonyl-carbamoyl-adenosine (t6A) modification, loss of which causes a severe growth phenotype. Here we aimed at a molecular diagnosis in a brother and sister from a consanguineous family who presented with global developmental delay, failure to thrive and a renal defect manifesting in proteinuria and hypomagnesemia. Using exome sequencing, the patients were found to be homozygous for the c.974G>A (p.(Arg325Gln)) variant of the KAE1 gene. KAE1 is a constituent of the KEOPS complex, a five-subunit complex that catalyzes the second biosynthetic step of t6A in the cytosol. The yeast KAE1 allele carrying the equivalent mutation did not rescue the t6A deficiency of the kae1Δ yeast strain as efficiently as the WT allele; furthermore, t6A levels quantified by LC-MS/MS were lower in the kae1Δ strain which was complemented by the mutation than in the kae1Δ strain, which was complemented by the WT allele. We conclude that homozygosity for c.974G>A (p.(Arg325Gln)) in KAE1 likely exerts its pathogenic effect by perturbing t6A synthesis, thereby interfering with global protein production. This is the first report of t6A biosynthesis defect in human. KAE1 joins the growing list of cytoplasmic tRNA modification enzymes, all associated with severe neurological disorders.

 

 

Association of single nucleotide polymorphisms in candidate genes previously related to genetic variation in fertility with phenotypic measurements of reproductive function in Holstein cows.

Author information: Ortega MS¹, Denicol AC¹, Cole JB², Null DJ², Taylor JF³, Schnabel RD⁴, Hansen PJ⁵.

Date of e-pub: May 2017

Abstract: Many genetic markers related to health or production traits are not evaluated in populations independent of the discovery population or related to phenotype. Here we evaluated 68 single nucleotide polymorphisms (SNP) in candidate genes previously associated with genetic merit for fertility and production traits for association with phenotypic measurements of fertility in a population of Holstein cows that was selected based on predicted transmitting ability (PTA) for daughter pregnancy rate (DPR; high, ≥1, n = 989; low, ≤ -1.0, n = 1,285). Cows with a high PTA for DPR had higher pregnancy rate at first service, fewer services per conception, and fewer days open than cows with a low PTA for DPR. Of the 68 SNP, 11 were associated with pregnancy rate at first service, 16 with services per conception, and 19 with days open. Single nucleotide polymorphisms in 12 genes (BDH2, BSP3, CAST, CD2, CD14, FUT1, FYB, GCNT3, HSD17B7, IBSP, OCLN, and PCCB) had significant associations with 2 fertility traits, and SNP in 4 genes (CSPP1, FCER1G, PMM2, and TBC1D24) had significant associations with each of the 3 traits. Results from this experiment were compared with results from 2 earlier studies in which the SNP were associated with genetic estimates of fertility. One study involved the same animals as used here, and the other study was of an independent population of bulls. A total of 13 SNP associated with 1 or more phenotypic estimates of fertility were directionally associated with genetic estimates of fertility in the same cow population. Moreover, 14 SNP associated with reproductive phenotype were directionally associated with genetic estimates of fertility in the bull population. Nine SNP (located in BCAS, BSP3, CAST, FUT1, HSD17B7, OCLN, PCCB, PMM2, and TBC1D24) had a directional association with fertility in all 3 studies. Examination of the function of the genes with SNP associated with reproduction in more than one study indicates the importance of steroid hormones and immune function as determinants of reproductive function. All but 1 of the 68 evaluated SNP were variable in 11 breeds besides Holstein, indicating the potential effects of these SNP on reproductive function across breeds of cattle.

 

 

Effects of supplementation with docosahexaenoic acid on reproduction of dairy cows.

Author information: Sinedino LD^1,2, Honda PM¹, Souza LR¹, Lock AL³, Boland MP⁴, Staples CR¹, Thatcher WW^1,2, Santos JE^5,2.

Date of e-pub: May 2017

Abstract: The objectives were to determine the effects of supplementing docosahexaenoic acid (DHA)-rich algae on reproduction of dairy cows. Holstein cows were assigned randomly to either a control (n = 373) or the same diet supplemented daily with 100 g/cow of an algae product containing 10% DHA (algae, n = 366) from 27 to 147 days postpartum. Measurements included yields of milk and milk components, fatty acids (FA) profiles in milk fat and plasma phospholipids, resumption of ovulation by 57 days postpartum, pregnancy per artificial insemination (AI) and expression of interferon-stimulated genes in leukocytes. Feeding algae increased resumption of estrous cyclicity (77.6 vs 65.9%) and pregnancy at first AI (47.6 vs 32.8%) in primiparous cows. Algae increased pregnancy per AI in all AI in both primiparous and multiparous cows (41.6 vs 30.7%), which reduced days to pregnancy by 22 days (102 vs 124 days) compared with control cows. Pregnant cows fed algae had greater expression of RTP4 in blood leukocytes compared with those in pregnant control cows. Feeding algae increased the incorporation of DHA, eicosapentaenoic acid, conjugated linoleic acid isomers cis-9 trans-11, trans-10 cis-12 and total n-3 FA in phospholipids in plasma and milk fat. Yields of milk and true protein increased by 1.1 kg/day and 30 g/day respectively, whereas fat yield decreased 40 g/day in algae compared with that in control. Supplementing DHA-rich algae altered the FA composition of lipid fractions and improved reproduction in dairy cows. The benefits on reproduction might be mediated by enhanced embryo development based on changes in interferon-stimulated gene expression.

 

 

Conditional knockout of activin like kinase-1 (ALK-1) leads to heart failure without maladaptive remodeling.

Author information: Morine KJ¹, Qiao X¹, Paruchuri V¹, Aronovitz MJ¹, Mackey EE¹, Buiten L¹, Levine J¹, Ughreja K¹, Nepali P¹, Blanton RM¹, Karas RH¹, Oh SP², Kapur NK³.

Date of e-pub: May 2017

Abstract: Activin like kinase-1 (AlK-1) mediates signaling via the transforming growth factor beta (TGFβ) family of ligands. AlK-1 activity promotes endothelial proliferation and migration. Reduced AlK-1 activity is associated with arteriovenous malformations. No studies have examined the effect of global AlK-1 deletion on indices of cardiac remodeling. We hypothesized that reduced levels of AlK-1 promote maladaptive cardiac remodeling. To test this hypothesis, we employed AlK-1 conditional knockout mice (cKO) harboring the ROSA26-CreER knock-in allele, whereby a single dose of intraperitoneal tamoxifen triggered ubiquitous Cre recombinase-mediated excision of floxed AlK-1 alleles. Tamoxifen treated wild-type (WT-TAM; n = 5) and vehicle treated AlK-1-cKO mice (cKO-CON; n = 5) served as controls for tamoxifen treated AlK-1-cKO mice (cKO-TAM; n = 15). AlK-1 cKO-TAM mice demonstrated reduced 14-day survival compared to cKO-CON controls (13 vs 100%, respectively, p < 0.01). Seven days after treatment, cKO-TAM mice exhibited reduced left ventricular (LV) fractional shortening, progressive LV dilation, and gastrointestinal bleeding. After 14 days total body mass was reduced, but LV and lung mass increased in cKO-TAM not cKO-CON mice. Peak LV systolic pressure, contractility, and arterial elastance were reduced, but LV end-diastolic pressure and stroke volume were increased in cKO-TAM, not cKO-CON mice. LV AlK-1 mRNA levels were reduced in cKO-TAM, not cKO-CON mice. LV levels of other TGFβ-family ligands and receptors (AlK5, TBRII, BMPRII, Endoglin, BMP7, BMP9, and TGFβ1) were unchanged between groups. Cardiomyocyte area and LV levels of BNP were increased in cKO-TAM mice, but LV levels of β-MHC and SERCA were unchanged. No increase in markers of cardiac fibrosis, Type I collagen, CTGF, or PAI-1, were observed between groups. No differences were observed for any variable studied between cKO-CON and WT-TAM mice. Global deletion of AlK-1 is associated with the development of high output heart failure without maladaptive remodeling. Future studies exploring the functional role of AlK-1 in cardiac remodeling independent of systemic AVMs are required.

 

 

Diversification of Rosaceae since the Late Cretaceous based on plastid phylogenomics.

Author information: Zhang SD¹, Jin JJ^1,2, Chen SY¹, Chase MW^3,4, Soltis DE^5,6,7, Li HT¹, Yang JB¹, Li DZ¹, Yi TS¹.

Date of e-pub: May 2017

Abstract: Phylogenetic relationships in Rosaceae have long been problematic because of frequent hybridisation, apomixis and presumed rapid radiation, and their historical diversification has not been clarified. With 87 genera representing all subfamilies and tribes of Rosaceae and six of the other eight families of Rosales (outgroups), we analysed 130 newly sequenced plastomes together with 12 from GenBank in an attempt to reconstruct deep relationships and reveal temporal diversification of this family. Our results highlight the importance of improving sequence alignment and the use of appropriate substitution models in plastid phylogenomics. Three subfamilies and 16 tribes (as previously delimited) were strongly supported as monophyletic, and their relationships were fully resolved and strongly supported at most nodes. Rosaceae were estimated to have originated during the Late Cretaceous with evidence for rapid diversification events during several geological periods. The major lineages rapidly diversified in warm and wet habits during the Late Cretaceous, and the rapid diversification of genera from the early Oligocene onwards occurred in colder and drier environments. Plastid phylogenomics offers new and important insights into deep phylogenetic relationships and the diversification history of Rosaceae. The robust phylogenetic backbone and time estimates we provide establish a framework for future comparative studies on rosaceous evolution.

 

 

Seasonal resource value and male size influence male aggressive interactions in the leaf footed cactus bug, Narnia femorata.

Author information: Nolen ZJ¹, Allen PE², Miller CW³.

Date of e-pub: May 2017

Abstract: In animal contests, resource value (the quality of a given resource) and resource holding potential (a male’s absolute fighting ability) are two important factors determining the level of engagement and outcome of contests. Few studies have tested these factors simultaneously. Here, we investigated whether natural, seasonal differences in cactus phenology (fruit quality) influence interactions between males in the leaf-footed cactus bug, Narnia femorata (Hemiptera: Coreidae). We also considered whether males were more likely to interact when they were similar in size, as predicted by theory. Finally, we examined if male size relative to the size of an opponent predicted competitive success. We found that males have more interactions on cactus with high value ripe fruit, as we predicted. Further, we found that males that were closer in size were more likely to interact, and larger males were more likely to become dominant.

 

 

Structure and Origin of the White Cap Locus and Its Role in Evolution of Grain Color in Maize.

Author information: Tan BC¹, Guan JC², Ding S¹, Wu S², Saunders JW², Koch KE², McCarty DR³.

Date of e-pub: May 2017

Abstract: Selection for yellow- and white-grain types has been central to postdomestication improvement of maize. While genetic control of carotenoid biosynthesis in endosperm is attributed primarily to the Yellow1 (Y1) phytoene synthase gene, less is known about the role of the dominant white endosperm factor White Cap (Wc). We show that the Wc locus contains multiple, tandem copies of a Carotenoid cleavage dioxygenase 1 (Ccd1) gene that encodes a carotenoid-degrading enzyme. A survey of 111 maize inbreds and landraces, together with 22 teosinte accessions, reveals that Wc is exclusive to maize, where it is prevalent in white-grain (y1) varieties. Moreover, Ccd1 copy number varies extensively among Wc alleles (from 1 to 23 copies), and confers a proportional range of Ccd1 expression in diverse organs. We propose that this dynamic source of quantitative variation in Ccd1 expression was created in maize shortly after domestication by a two-step, Tam3L transposon-mediated process. First, a chromosome segment containing Ccd1 and several nearby genes duplicated at a position 1.9 Mb proximal to the progenitor Ccd1r locus on chromosome 9. Second, a subsequent interaction of Tam3L transposons at the new locus created a 28-kb tandem duplication, setting up expansion of Ccd1 copy number by unequal crossing over. In this way, transposon-mediated variation in copy number at the Wc locus generated phenotypic variation that provided a foundation for breeding and selection of white-grain color in maize.

 

 

The Arabidopsis Elongator complex is required for nonhost resistance against the bacterial pathogens Xanthomonas citri subsp. citri and Pseudomonas syringae pv. phaseolicola NPS3121.

Author information: An C¹, Wang C¹, Mou Z¹.

Date of e-pub: May 2017

Abstract: Although in recent years nonhost resistance has attracted considerable attention for its broad spectrum and durability, the genetic and mechanistic components of nonhost resistance have not been fully understood. We used molecular and histochemical approaches including quantitative PCR, chromatin immunoprecipitation, and 3,3′-diaminobenzidine and aniline blue staining. The evolutionarily conserved histone acetyltransferase complex Elongator was identified as a major component of nonhost resistance against Xanthomonas citri subsp. citri (Xcc) and Pseudomonas syringae pv. phaseolicola (Psp) NPS3121. Mutations in Elongator genes inhibit Xcc-, Psp NPS3121- and/or flg22-induced defense responses including defense gene expression, callose deposition, and reactive oxygen species (ROS) and salicylic acid (SA) accumulation. Mutations in Elongator also attenuate the ROS-SA amplification loop. We show that suppressed ROS and SA accumulation in Elongator mutants is correlated with reduced expression of the Arabidopsis respiratory burst oxidase homologue AtrbohD and the SA biosynthesis gene ISOCHORISMATE SYNTHASE1 (ICS1). Furthermore, we found that the Elongator subunit ELP2 is associated with the chromatin of AtrbohD and ICS1 and is required for maintaining basal histone H3 acetylation levels in these key defense genes. As both AtrbohD and ICS1 contribute to nonhost resistance against Xcc, our results reveal an epigenetic mechanism by which Elongator regulates nonhost resistance in Arabidopsis.

 

 

Anticoagulation endpoints with clinical implementation of warfarin pharmacogenetic dosing in a real-world setting: A proposal for a new pharmacogenetic dosing approach.

Author information: Arwood MJ^1,2, Deng J³, Drozda K⁴, Pugach O⁵, Nutescu EA^6,7,8, Schmidt S³, Duarte JD^1,2, Cavallari LH^1,2.

Date of e-pub: May 2017

Abstract: Achieving therapeutic anticoagulation efficiently with warfarin is important to reduce thrombotic and bleeding risks and is influenced by genotype. Utilizing data from a diverse population of 257 patients who received VKORC1 and CYP2C9 genotype-guided warfarin dosing, we aimed to examine genotype-associated differences in anticoagulation endpoints and derive a novel pharmacogenetic nomogram to more optimally dose warfarin. We observed significant differences across patients with 0, 1, or ≥2 reduced-function VKORC1 or CYP2C9 alleles, respectively, in time to achieve therapeutic international normalized ratio (INR) (7.8 ± 5.8, 7.2 ± 4.7, and 5.4 ± 4.6 days, P = 0.0004) and mean percentage of time in therapeutic range in the first 28 days (22.2, 27.8, and 32.2%, P = 0.0127) with use of existing pharmacogenetic algorithms. These data suggest that more aggressive dosing is necessary for patients with 0 to 1 VKORC1/CYP2C9 variants to more efficiently achieve therapeutic anticoagulation. Herein, we provide a novel kinetic/pharmacodynamic-derived dosing nomogram optimized for a heterogeneous patient population.

 

 

Prevalence of Escherichia coli O157:H7 From House Flies (Diptera: Muscidae) and Dairy Samples in North Central Florida1.

Author information: Burrus RG¹, Hogsette JA², Kaufman PE³, Maruniak JE³, Simonne AH⁴, Mai V^5,6.

Date of e-pub: May 2017

Abstract:  La detección de Escherichia coli O157:H7 en las lecherías es importante para mejorar la seguridad de los productos lácteos, y se ha llevado a cabo principalmente mediante el aislamiento de las bacterias a partir de las muestras de estiércol. Sin embargo, los componentes biliares presentes en el estiércol complica la identificación genética utilizando la técnica del PCR, y el aislamiento microbiológico se dificulta por la presencia de bacterias competidoras que comparten características microbiológicas similares. El aislamiento de E. coli O157:H7 a partir de la mosca doméstica evita las dificultades asociadas con el estiércol del ganado. El aislamiento de patógenos a partir de las moscas domésticas proporciona información adicional sobre el potencial impacto epidemiológico de la dispersión de la mosca doméstica en la distribución de patógenos, ya que las moscas domésticas se dispersan desde las lecherías donde la E. coli O157:H7 existe en forma endémica en el ganado. En este estudio, se encontró que las moscas domésticas son 2,6 veces más sensibles para la detección de E. coli O157:H7 en las lecherías. Las moscas son más fáciles de capturar y manejar que el estiércol, y deberían ser utilizadas en cualquier ensayo para detectar E. coli O157:H7 en las lecherías y otros establecimientos.

 

 

Advances in the development of aptamer drug conjugates for targeted drug delivery.

Author information: Chen K¹, Liu B¹, Yu B¹, Zhong W¹, Lu Y¹, Zhang J¹, Liao J¹, Liu J^1,2,3,4,5,6, Pu Y¹, Qiu L^2,3,4,5,6, Zhang L^2,3,4,5,6,7, Liu H¹, Tan W^2,3,4,5,6,7.

Date of e-pub: May 2017

Abstract:  A key goal of modern medicine is target-specific therapeutic intervention. However, most drugs lack selectivity, resulting in ‘off-target’ side effects. To address the requirements of ‘targeted therapy,’ aptamers, which are artificial oligonucleotides, have been used as novel targeting ligands to construct aptamer drug conjugates (ApDC) that can specifically bind to a broad spectrum of targets, including diseased cells. Accordingly, the application of aptamers in targeted drug delivery has attracted broad interest due to their impressive selectivity and affinity, low immunogenicity, easy synthesis with high reproducibility, facile modification, and relatively rapid tissue penetration with no toxicity. Functionally, aptamers themselves can be used as macromolecular drugs, and they are also commonly used in biomarker discovery and targeted drug delivery. In this review, we will highlight the most recent advances in the development of aptamers and aptamer conjugates, and discuss their potential in targeted therapy. WIREs Nanomed Nanobiotechnol 2017, 9:e1438. doi: 10.1002/wnan.1438 For further resources related to this article, please visit the WIREs website.

 

 

Electrophysiologic features of fibular neuropathy in childhood and adolescence.

Author information: Karakis I^1,2,3, Khoshnoodi M^3,4, Liew W^1,5, Nguyen ES¹, Jones HR^1,2, Darras BT¹, Kang PB^1,6.

Date of e-pub: May 2017

Abstract: We studied patterns of nerve injury in pediatric common fibular (peroneal) neuropathy (CFN).

A retrospective analysis was performed on data from 53 children with CFN at a pediatric electromyography laboratory.

Conduction block at the fibular head was present in 35% of patients. Deep fibular axonal loss was identified in 77%, while superficial fibular axonal loss was identified in 45%. The pathophysiology was predominantly axonal in 72%, mostly demyelinating in 6%, and mixed in 22%. Predominantly demyelinating lesions at the fibular head demonstrated sparing of the superficial fibular sensory nerve (P = 0.01, Fischer exact test). Predominantly axonal lesions had a moderate correlation between superficial and deep fibular axonal loss (Spearman r = 0.52; P = 0.0001).

There is frequent axonal and fascicular injury in pediatric CFN, similar to adults. Deep and superficial fibular nerve involvements correlate in axonal lesions, whereas superficial fibular sensory fibers are often spared in demyelinating lesions. Muscle Nerve, 2016 Muscle Nerve 55: 693-697, 2017.

 

 

Life-extending Dietary Restriction Reduces Oxidative Damage of Proteins in Grasshoppers but Does Not Alter Allocation of Ingested Nitrogen to Somatic Tissues.

Author information: Heck MJ¹, Pehlivanovic M^1,2, Purcell JU^1,3, Hahn DA⁴, Hatle JD¹.

Date of e-pub: May 2017

Abstract:  Dietary restriction (DR) extends life span and reduces reproduction in most animals. The disposable soma hypothesis suggests that this longevity is the result of reduced investment in reproduction and increased nutrient allocation to the soma, permitting an increase in cellular maintenance. To investigate the role of nutrient allocation upon life-extending DR, tissue-specific nitrogen allocation was tracked in grasshoppers (Romalea microptera) upon a full or restricted (60% of full) diet. In addition, carbonyl (oxidized protein) assays addressed tissue maintenance. To develop a labeled diet on which grasshoppers could thrive, hydroponically grown Romaine lettuce was enriched with 15N. This allowed quantification of nitrogen allocation upon a normal or restricted diet. There was a 50% decrease in reproductive investment upon DR. At the same time, relative allocation of 15N to the ovary did not change. Most important, relative allocation was similar between restricted and full diet grasshoppers for somatic tissues (ie, mandibular and femur muscle, dried hemolymph, gut, and fat body). Carbonyl assays of muscles, hemolymph, and gut revealed an overall reduction in protein oxidation upon DR. These data suggest that DR does not alter nutrient allocation but does reduce protein oxidation, an observation that is inconsistent with the basic predictions of the disposable soma hypothesis.
NOTE: These abstracts were retrieved from the U.S. National Library of Medicine website managed in collaboration with the U.S. National Library of Medicine