Search

Small regulatory RNAs can move between organisms during pathogenic interactions and regulate gene expression in the recipient. If and how such “trans-species” small RNAs are distinguished from normal small RNAs is not known. The parasitic plant Cuscuta campestris produces a number of microRNAs that specifically accumulate at the interface between parasite and host, several of which have been demonstrated to have trans-species activity. We find that induction of C. campestris interface-induced microRNAs was similar regardless of host species, and can be replicated in haustoria stimulated to develop in the complete absence of a host. We also find that the loci encoding C. campestris interface-induced microRNAs are distinguished by a common 10 base-pair cis-regulatory element. This element is identical to a previously described upstream sequence element used by all plant small nuclear RNA loci. The sequence context of this element strongly suggests U6-like transcription by RNA polymerase III. The element promotes accumulation of interface-induced miRNAs in a heterologous system. This common promoter element distinguishes C. campestris interface-induced microRNA loci from other plant small RNAs; other plant small RNA loci are transcribed by polymerases II or IV, and lack any common promoter motifs. Our data suggest that C. campestris interface-induced miRNAs are produced in a manner distinct from canonical miRNAs. All confirmed C. campestris microRNAs with confirmed trans-species activity are interface-induced and possess these features. We speculate that this distinct production may allow these miRNAs to be exported to hosts.

Objective A meta-analysis to determine if patients with varicose veins are at an increased risk of venous thromboembolism (VTE) when undergoing major lower limb arthroplasty. Methods Medline, Embase, and Cochrane Library databases were searched using appropriate terms for studies that reported post-operative VTE in patients who had lower limb arthroplasty with any history of varicose veins. Methodological quality of included studies was quantified using the Risk of Bias (ROB) assessment tools. Findings were reported using the meta-analysis of observational studies in epidemiology (MOOSE) checklist. Results A total of 129 studies were identified with 11 observational studies being eligible for inclusion. This consisted of 265,194 patients who underwent lower limb arthroplasty, 2188 of which had pre-existing varicose veins. Overall, VTE occurred in 1845 patients, and 122 cases had varicose veins present at time of arthroplasty. Meta-analysis indicates that patients undergoing lower limb arthroplasty with varicose veins are at increased risk of having a VTE, OR 2.37, 95% CI 1.54–3.63, ( p < 0.001). One study evaluated if previous varicose veins surgery influenced the risk of VTE in arthroplasty patients, OR 0.96 (95% CI 0.7–1.28), p = 0.429. Conclusions Varicose veins and lower limb arthroplasty are known independent risk factors for VTE. There is a paucity of data regarding the risk of VTE in patients undergoing lower limb arthroplasty who have co-existing varicose veins. This meta-analysis shows that patients with varicose veins are at an increased risk of VTE when undergoing major lower limb arthroplasty. Further studies are required in order to determine if such patients should undergo varicose vein surgery before undertaking major lower limb joint replacement.

The IQ Consortium NHP Reuse Working Group (WG) comprises members from 15 pharmaceutical and biotechnology companies. In 2020, the WG developed and distributed a detailed questionnaire on protein non-naïve NHP reuse to the WG member companies. The WG received responses from key stakeholders including principal investigators, facility managers, animal welfare officers and research scientists. This paper’s content reflects the consolidated opinion of the WG members and the questionnaire responses on the subject of NHP reuse within nonclinical programs at all stages of research and development. Many of the pharmaceutical companies represented in the working group or participating in the questionnaire have already achieved some level of NHP reuse in their nonclinical programs, but the survey results suggested that there is significant potential to increase NHP reuse further and a need to understand the considerations involved in reuse more clearly. The WG has also focused carefully on the inherent concerns and risks of implementing protein non-naive NHP reuse and has evaluated the best methods of risk assessment and decision-making. This paper presents a discussion on the challenges and opportunities surrounding protein non-naïve NHP reuse and aims to stimulate further industry dialogue on the subject and provide guidance for pharmaceutical companies to establish roadmaps and decision trees enabling increased protein non-naïve NHP reuse. In addition, this paper represents a solid basis for collaborative engagement between pharmaceutical and biotechnology companies with contract research organizations (CROs) to discuss how the availability of protein non-naïve NHP within CROs can be better leveraged for their use within nonclinical studies.

Purpose: There remains an unmet need to provide effective treatment therapy for patients with primary and metastatic brain tumors; lack of drug penetration across the blood brain barrier (BBB) is a key factor. Synthetic lethality is an attractive mechanism in treating brain tumors post radiotherapy. Here we investigate the brain penetration of niraparib and olaparib in healthy monkeys to understand the potential of each to cross the intact BBB. Experimental Procedures: Four healthy male Rhesus macaque monkeys were dosed daily via oral gavage for five days with either niraparib (n=2; 6 mg/kg) or olaparib (n=2; 10 mg/kg). Pre-dose blood was collected daily and terminal blood, cerebrospinal fluid (CSF), and brain tissue was collected at necropsy. Coronal brain sections were analyzed by matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) to quantitatively assess the tissue distribution of the dosed compounds. Blood, CSF, and bulk homogenate of brain tissue were analyzed by LC-MS bioanalysis. Summary of Data: Greater brain penetration was observed for niraparib when compared to olaparib in healthy Rhesus macaque monkeys following five days oral administration. The unbound brain-to-plasma partition coefficient (Kp,uu,brain) was 15x higher for niraparib compared to olaparib. Quantitative MALDI IMS of coronal brain sections from monkeys administered niraparib showed consistent concentrations distributed throughout the brain parenchyma. Olaparib was not detected by MALDI IMS in any of the coronal brain sections collected from the monkeys administered olaparib. Similar plasma and CSF concentrations were observed between the monkey’s administered niraparib vs olaparib, highlighting the unique ability of niraparib to cross the intact BBB of monkeys and distribute throughout the brain. Conclusions: Niraparib showed markedly higher brain penetration than olaparib in healthy Rhesus macaque monkeys demonstrating enhanced ability to cross intact BBB. Summary of Quantification Results Displayed as Total Conc. with Adjusted Free Conc. in Parentheses Compound Animal LC-MS Pre-Dose Plasma Days 1-5 (ng/mL) LC-MS Pre-Dose Plasma Days 1-5 (ng/mL) LC-MS Terminal CSF (ng/mL) LC-MS Bulk Brain Homogenate (ng/g) MALDI IMS Brain Section Average (ng/g) Kp,uu Brain/Plasma Mean Kp, uu Brain/Plasma Niraparib Monkey 1 BLQ, 28 (10), 49 (18), 59 (22), 72 (26) 84 (31) 9 378 (14) 283 (10) 0.45 0.30 Niraparib Monkey 2 BLQ, 58 (21), 87 (32), 74 (27), 66 (24) 436 (160) 25 797 (29) 684 (25) 0.16 Olaparib Monkey 3 BLQ, 27 (8), 26 (8), 33 (10), 33 (10) 322 (99) 13 12 (2) BLQ 0.02 0.02 Olaparib Monkey 4 BLQ, 51 (16), 17 (5), 53 (16), 45 (14) 254 (78) 6 11 (2) BLQ 0.03 Citation Format: Mark Reid Groseclose, Jeremy Barry, Yongle Pang, Jennifer Deutsch, Shannon Berry, Elaina McCormick, David K. Lim, Hoang Tran, Sean Maguire, Hasan Alsaid, Amine Aziez, Elaine Paul, Keyur Gada. Differentiation of niraparib and olaparib brain penetration in healthy rhesus macaque monkeys. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3581.

Hydroxyurea induces production of fetal hemoglobin (HbF), a tetramer of α and γ globin proteins and corresponding heme molecules, normally found in less than 1% of adult RBC. Increases in circulating HbF are correlated with clinical improvement of patients with hemoglobinopathies, and hydroxyurea, as a daily medication, is the standard treatment for sickle cell anemia. Although olive baboons (Papio anubis) are considered a key model species for HbF induction, cynomolgus macaques (Macaca fasicularis) are another species that conserves the ability to produce HbF into maturity. In this study, moderate anemia was experimentally induced in cynomolgus macaques by phlebotomy, to stimulate accelerated erythropoiesis and HbF production. In contrast to previous studies, vascular access ports were implanted for phlebotomy of conscious monkeys, followed by fluid replacement. As total Hgb levels dropped, reticulocyte counts and the percentage of HbF-expressing cells increased. Once total Hgb levels declined to less than 8 g/dL, 2 courses of oral hydroxyurea (once daily for 5 d) were completed, with a 9-d interval between courses. After hydroxyurea dosing, the percentage of HbF-expressing cells and total HbF were increased significantly. In addition, a significant but transient decrease in reticulocyte count and a transient increase in MCV occurred, replicating the characteristic response of patients receiving hydroxyurea. Daily clinical observations revealed no serious health issues or decreases in food consumption or activity levels. Methods were established for assessing the patency of vascular access ports. This study details a new protocol for the safe and routine induction of moderate anemia in cynomolgus macaques and validates its use in the investigation of novel pharmacologic entities to induce the production of HbF.

Long-lasting, latently infected resting CD4+ T cells are the greatest obstacle to obtaining a cure for HIV infection, as these cells can persist despite decades of treatment with antiretroviral therapy (ART). Estimates indicate that more than 70 years of continuous, fully suppressive ART are needed to eliminate the HIV reservoir1. Alternatively, induction of HIV from its latent state could accelerate the decrease in the reservoir, thus reducing the time to eradication. Previous attempts to reactivate latent HIV in preclinical animal models and in clinical trials have measured HIV induction in the peripheral blood with minimal focus on tissue reservoirs and have had limited effect2–9. Here we show that activation of the non-canonical NF-κB signalling pathway by AZD5582 results in the induction of HIV and SIV RNA expression in the blood and tissues of ART-suppressed bone-marrow–liver–thymus (BLT) humanized mice and rhesus macaques infected with HIV and SIV, respectively. Analysis of resting CD4+ T cells from tissues after AZD5582 treatment revealed increased SIV RNA expression in the lymph nodes of macaques and robust induction of HIV in almost all tissues analysed in humanized mice, including the lymph nodes, thymus, bone marrow, liver and lung. This promising approach to latency reversal—in combination with appropriate tools for systemic clearance of persistent HIV infection—greatly increases opportunities for HIV eradication. Activation of the non-canonical NF-κB signalling pathway by AZD5582 results in the induction of HIV and SIV RNA expression in the blood and tissues of antiretroviral-therapy-treated humanized mice and rhesus macaques.

The recent Scientific Committee on Health, Environmental and Emerging Risks Final Opinion on “The need for nonhuman primates in biomedical research, production and testing of products and devices” (2017 SCHEER) highlights approaches that could significantly contribute to the replacement, reduction, and refinement of nonhuman primate (NHP) studies. Initiatives that have the potential to affect NHP welfare and/or their use are expected to be appropriate, fair, and objective and publicly disseminated information focused on NHPs in biomedical research, which includes toxicologic and pathologic research and testing, should be objectively evaluated by stakeholder scientists, researchers, and veterinarians. Thus, IQ Consortium member companies convened to develop an informed and objective response, focusing on identifying areas of agreement, potential gaps, or missing information in 2017 SCHEER. Overall, the authors agree that many positions in the 2017 SCHEER Opinion generally align with industry views on the use of NHPs in research and testing, including the ongoing need of NHPs in many areas of research. From the perspective of the IQ Consortium, there are several topics in the 2017 SCHEER that merit additional comment, attention, or research, as well as consideration in future opinions.

GlaxoSmithKline has recently made significant organizational changes to its nonclinical safety, drug metabolism and pharmacokinetic, and laboratory animal science/veterinary functions, with the goal to increase our focus on scientific partnership with the discovery part of the organization. One specific change was bringing together pathologists and comparative medicine veterinarians and scientists into a single functional unit. We describe our early activities (assessing our capabilities and gaps, external benchmarking, listening to our discovery partners, redesigning some of our working practices) aimed at implementing these changes. In addition, early on we held a Discovery Engagement Workshop attended by all pathologists and comparative medicine veterinarians and scientists, as well as selected discovery scientists. The purpose of this workshop was to share learnings from the above activities and devise plans aimed at achieving our overall goal of functional integration: driving pathobiology expertise into drug discovery and increasing the human (translational) relevance of experimental data. This review describes the new organizational structure, the workshop activities, and implementation plans; updates our progress; and considers the opportunity for a pan-industry network of discovery-focused pathologists and comparative medicine veterinarians and scientists.

Hydroxyurea induces production of fetal hemoglobin (HbF), a tetramer of α and γ globin proteins and corresponding heme molecules, normally found in less than 1% of adult RBC. Increases in circulating HbF are correlated with clinical improvement of patients with hemoglobinopathies, and hydroxyurea, as a daily medication, is the standard treatment for sickle cell anemia. Although olive baboons (Papio anubis) are considered a key model species for HbF induction, cynomolgus macaques (Macaca fasicularis) are another species that conserves the ability to produce HbF into maturity. In this study, moderate anemia was experimentally induced in cynomolgus macaques by phlebotomy, to stimulate accelerated erythropoiesis and HbF production. In contrast to previous studies, vascular access ports were implanted for phlebotomy of conscious monkeys, followed by fluid replacement. As total Hgb levels dropped, reticulocyte counts and the percentage of HbF-expressing cells increased. Once total Hgb levels declined to less than 8 g/dL, 2 courses of oral hydroxyurea (once daily for 5 d) were completed, with a 9-d interval between courses. After hydroxyurea dosing, the percentage of HbF-expressing cells and total HbF were increased significantly. In addition, a significant but transient decrease in reticulocyte count and a transient increase in MCV occurred, replicating the characteristic response of patients receiving hydroxyurea. Daily clinical observations revealed no serious health issues or decreases in food consumption or activity levels. Methods were established for assessing the patency of vascular access ports. This study details a new protocol for the safe and routine induction of moderate anemia in cynomolgus macaques and validates its use in the investigation of novel pharmacologic entities to induce the production of HbF.

Data is rapidly changing how companies operate, offering them new business opportunities as they generate increasingly sophisticated insights from the analysis of an ever-increasing pool of information. Businesses have clearly moved beyond a focus on data collection to data use, but users have an inadequate model of notice and consent at the point of data collection to limit inappropriate use. An interoperable context-aware metadata-based architecture that allows permissions and policies to be bound to data, and is flexible enough to allow for changing trust norms, help balance the tension between users and business, satisfy regulators’ desire for increased transparency and greater accountability, and still enable data to flow in ways that provide value to all participants in the ecosystem.

Coccidioides spp. are saprophytic, dimorphic fungi that are endemic to arid climates, are capable of infecting many species, and result in diverse clinical presentations. An indoor-housed laboratory rhesus macaque presented with weight loss and decreased activity and appetite. During the diagnostic evaluation, a bronchiolar–alveolar pattern in the cranial lung lobes, consistent with bronchopneumonia, was noted on radiographs. Given the poor prognosis, the macaque was euthanized. Confirming the radiographic assessment, gross necropsy findings included multifocal to coalescing areas of consolidation in the right and left cranial lung lobes. Microscopically, the consolidated regions were consistent with a pyogranulomatous bronchopneumonia and contained round, nonbudding, fungal yeast structures considered to be morphologically consistent with Coccidioides immitis. Culture and colony morphology results were confirmed through additional diagnostic testing. Sequencing of the D1–D2 domain of the 28S large ribosomal subunit positively matched with a known sequence specific to C. immitis. Serology for Coccidioides spp. by both latex agglutination (IgM) and immunodiffusion (IgG) was positive. In this rhesus macaque, the concordant results from histology, culture, DNA sequencing, and serology were collectively used to confirm the diagnosis of coccidioidomycosis. This animal likely acquired a latent pulmonary infection with Coccidioides months prior to arrival, when housed outdoors in a Coccidioides-endemic area. The nonspecific clinical presentation in this macaque, coupled with the recent history of indoor housing and lag between clinical presentation and outdoor housing, can make similar diagnostic cases challenging and highlights the need for awareness regarding animal source when making an accurate diagnosis in an institutional laboratory setting.

Therapies that suppress RIPK1 kinase activity are emerging as promising therapeutic agents for the treatment of multiple inflammatory disorders. The ability to directly measure drug binding of a RIPK1 inhibitor to its target is critical for providing insight into pharmacokinetics, pharmacodynamics, safety and clinical efficacy, especially for a first‐in‐class small‐molecule inhibitor where the mechanism has yet to be explored. Here, we report a novel method for measuring drug binding to RIPK1 protein in cells and tissues. This TEAR1 (Target Engagement Assessment for RIPK 1) assay is a pair of immunoassays developed on the principle of competition, whereby a first molecule (ie, drug) prevents the binding of a second molecule (ie, antibody) to the target protein. Using the TEAR1 assay, we have validated the direct binding of specific RIPK1 inhibitors in cells, blood and tissues following treatment with benzoxazepinone (BOAz) RIPK1 inhibitors. The TEAR1 assay is a valuable tool for facilitating the clinical development of the lead RIPK1 clinical candidate compound, GSK2982772, as a first‐in‐class RIPK1 inhibitor for the treatment of inflammatory disease.

Silencing and variegated transgene expression are poorly understood problems that can interfere with gene function studies in human embryonic stem cells (hESCs). We show that transgene expression (enhanced green fluorescent protein [EGFP]) from random integration sites in hESCs is affected by variegation and silencing, with only half of hESCs expressing the transgene, which is gradually lost after withdrawal of selection and differentiation. We tested the hypothesis that a transgene integrated into the adeno-associated virus type 2 (AAV2) target region on chromosome 19, known as the AAVS1 locus, would maintain transgene expression in hESCs. When we used AAV2 technology to target the AAVS1 locus, 4.16% of hESC clones achieved AAVS1-targeted integration. Targeted clones expressed Oct-4, stage-specific embryonic antigen-3 (SSEA3), and Tra-1–60 and differentiated into all three primary germ layers. EGFP expression from the AAVS1 locus showed significantly reduced variegated expression when in selection, with 90% ± 4% of cells expressing EGFP compared with 57% ± 32% for randomly integrated controls, and reduced tendency to undergo silencing, with 86% ± 7% hESCs expressing EGFP 25 days after withdrawal of selection compared with 39% ± 31% for randomly integrated clones. In addition, quantitative polymerase chain reaction analysis of hESCs also indicated significantly higher levels of EGFP mRNA in AAVS1-targeted clones as compared with randomly integrated clones. Transgene expression from the AAVS1 locus was shown to be stable during hESC differentiation, with more than 90% of cells expressing EGFP after 15 days of differentiation, as compared with ∼30% for randomly integrated clones. These results demonstrate the utility of transgene integration at the AAVS1 locus in hESCs and its potential clinical application. Disclosure of potential conflicts of interest is found at the end of this article.

This paper presents a hybrid technique for measuring conversion gain that blends spatial and temporal information, allowing users to calculate an accurate conversion gain with little knowledge of sensor defects. It blends a single pixel method with multiple pixel methods. We present measured data from a visible CMOS image sensor using two multiple pixel methods and the hybrid method. Additionally, we provide arguments for validity of the hybrid method. To our knowledge, this is the first report of this technique. Conversion gain (e-/DN) directly relates measured digital numbers (DN) to input-referred electrons (e-) for an image sensor. Conversion gain can be directly measured by considering the sensor under varying illumination states in coordination with Poisson statistics. Typically, there are two approaches: measure a single pixel over time or measure a group of pixels at one point in time after correcting for gain non-uniformity. The plotted statistics from these measurements are called either mean-variance or photon-transfer curves. The measurement of a single pixel is relatively straightforward and requires collection of many consecutive frames to get meaningful statistics not dominated by thermal noise. The data volume for an accurate single-pixel measurement can become unwieldy in terms of number of frames required. This is especially true for large format image sensors. In contrast, the measurement of a group of pixels requires fewer consecutive frames, but needs non-uniformity adjustments to correctly calculate statistics.

Over a 3 month period blood was collected in the Phlebotomy Department from adult diabetics attending at outpatients. Further blood samples were obtained from adult diabetic patients in the hospital's diabetic ward. From the total of 262 blood samples, 82 were fasting and 180 were non-fasting. Capillary blood samples were analysed with the Medisense Precision G system using a new biosensor electrode (G2b). Venous blood samples were taken at the same time from the patients and were assayed using the laboratory's plasma glucose method on the Synchron CX7 (Beckman, CA, USA). Comparison of the Precision G results with the laboratory Synchron CX7 plasma assay was performed using comparison of means and examination of the differences between the two methods. Despite differences in specimen source, specimen type and test methodology, the Precision G system gave results on capillary samples almost equivalent to plasma glucose results in the same patients on the Synchron CX7. Mean bias was −4.2% in 180 non-fasting specimens and −3.0% in 82 fasting specimens. Blood glucose results were not affected between haematocrit values of 0.34 and 0.50 in a study on 62 diabetic patients. We conclude that the Precision G system for blood glucose analysis is rapid, precise, accurate, easy to use and clinically acceptable. Copyright © 2000 John Wiley & Sons, Ltd.

Down syndrome (DS) is caused by trisomy of chromosome 21 (Hsa21) and presents a complex phenotype that arises from abnormal dosage of genes on this chromosome. However, the individual dosage-sensitive genes underlying each phenotype remain largely unknown. To help dissect genotype – phenotype correlations in this complex syndrome, the first fully transchromosomic mouse model, the Tc1 mouse, which carries a copy of human chromosome 21 was produced in 2005. The Tc1 strain is trisomic for the majority of genes that cause phenotypes associated with DS, and this freely available mouse strain has become used widely to study DS, the effects of gene dosage abnormalities, and the effect on the basic biology of cells when a mouse carries a freely segregating human chromosome. Tc1 mice were created by a process that included irradiation microcell-mediated chromosome transfer of Hsa21 into recipient mouse embryonic stem cells. Here, the combination of next generation sequencing, array-CGH and fluorescence in situ hybridization technologies has enabled us to identify unsuspected rearrangements of Hsa21 in this mouse model; revealing one deletion, six duplications and more than 25 de novo structural rearrangements. Our study is not only essential for informing functional studies of the Tc1 mouse but also (1) presents for the first time a detailed sequence analysis of the effects of gamma radiation on an entire human chromosome, which gives some mechanistic insight into the effects of radiation damage on DNA, and (2) overcomes specific technical difficulties of assaying a human chromosome on a mouse background where highly conserved sequences may confound the analysis. Sequence data generated in this study is deposited in the ENA database, Study Accession number: ERP000439.

Publisher Summary This chapter develops a formulary that includes drugs in common use within the laboratory animal community. This formulary is limited to the species relevant to this volume, with particular emphasis on rabbits, guinea pigs, hamsters, and degus. There are many other excellent published sources that cover laboratory animals and other exotic species. This formulary is divided into five major areas within each species. These areas include analgesics/sedatives anesthetics, antiinfectives, parasiticides, and a miscellaneous category. It is also important to note that few of the drugs have undergone formal studies to determine their efficacy, pharmacokinetics, or pharmacodynamics. Most drug dosages listed in this formulary are based on clinical observation and experience, or extrapolated from limited scientific data. Extrapolation to other species may be considered using allometric scaling. Before any of the referenced doses are used in animals, it is the responsibility of the reader to study the source citation to understand how the listed dose was determined.

ash LIDAR, called the Vision Navigation Sensor (VNS), will be the primary navigation instrument used by the Orion vehicle during rendezvous, proximity operations, and docking. The DC will be used by the Orion crew for piloting cues during docking. This paper provides an overview of the STORRM test objectives and the concept of operations. It continues with a description of STORRM’s major hardware components, which include the VNS, docking camera, and supporting avionics. Next, an overview of crew and analyst training activities will describe how the STORRM team prepared for ight. Then an overview of in-ight data collection and analysis is presented. Key ndings and results from this project are summarized. Finally, the paper concludes with lessons learned from the STORRM DTO.

Stroke-prone rat strains exhibit an increased stroke risk and sensitivity, and reduced endogenous mechanisms of ischemic brain tolerance. This experiment provides a comparative, serial evaluation of neurological deficits and brain injury following middle cerebral artery occlusion/permanent focal stroke in this high-risk strain. Stroke-prone spontaneously hypertensive (SHR-SP), spontaneously hypertensive (SHR) and Wistar Kyoto (WKY) rats were evaluated over 28 days using magnetic resonance imaging (MRI), histopathology, and neurobehavioral testing. T2- and diffusion weighted-MRI was performed after 1, 10 and 28 days to measure the degree of stroke-induced brain injury. Normotensive WKY rats receiving the same stroke and other SHR-SP rats receiving sham surgery were used for control comparisons. Functional deficits were scored after 1, 4, 11, 18 and 28 days. The degree of brain infarction/injury was practically identical in hypertensive and stroke-prone rats. WKY rats exhibited significantly smaller infarcts (P

The last 10 years have seen growing recognition of the significance of the genes encoding enzymes responsible for hepatic alcohol metabolism as protective factors in the development of alcoholism.We have developed DNA sequencing assays for measuring genetic variation at the alcohol dehydrogenase 2 (ADH2), ADH3, and aldehyde dehydrogenase 2 (ALDH2) loci. These have been used to survey volunteer control subjects from three New Zealand ethnic groups (white, Asian, and Polynesian) and young male alcoholics recruited from white and New Zealand Maori patients in a local treatment program.The allele frequency values for whites and Asians obtained in our study closely match those obtained previously in other laboratories. Our data (the first for Polynesians) are 0.42 for ADH2*2, 0.78 for ADH3*1, and 0.00 for ALDH2*2. In the New Zealand Maori alcoholic patients, the ADH2*2 frequency is significantly lower (0.15; p0.01). The frequency of ADH3*1 is also lower in this group (0.60), but this value is not significant (0.05p0.06).In young male New Zealand Maori, the ADH2*2 allele is a protective factor against alcoholism even in the absence of ALDH2*2.

Nature 496, 498–503 (2013); doi:10.1038/nature12111 In this Letter, five authors were inadvertently omitted: Sharmin Begum and Christine Lloyd from the Wellcome Trust Sanger Institute, and Christa Lanz, Gunter Raddatz and Stephan C. Schuster from the Max Planck Institute for Developmental Biology. David Elliot was incorrectly listed as David Eliot, Beverley Mortimore was incorrectly listed as Beverly Mortimer, and James D.

BackgroundThe zebrafish (Danio rerio) is an important vertebrate model organism system for biomedical research. The syntenic conservation between the zebrafish and human genome allows one to investigate the function of human genes using the zebrafish model. To facilitate analysis of the zebrafish genome, genetic maps have been constructed and sequence annotation of a reference zebrafish genome is ongoing. However, the duplicative nature of teleost genomes, including the zebrafish, complicates accurate assembly and annotation of a representative genome sequence. Cytogenetic approaches provide "anchors" that can be integrated with accumulating genomic data.ResultsHere, we cytogenetically define the zebrafish genome by first estimating the size of each linkage group (LG) chromosome using flow cytometry, followed by the cytogenetic mapping of 575 bacterial artificial chromosome (BAC) clones onto metaphase chromosomes. Of the 575 BAC clones, 544 clones localized to apparently unique chromosomal locations. 93.8% of these clones were assigned to a specific LG chromosome location using fluorescencein situhybridization (FISH) and compared to the LG chromosome assignment reported in the zebrafish genome databases. Thirty-one BAC clones localized to multiple chromosomal locations in several different hybridization patterns. From these data, a refined second generation probe panel for each LG chromosome was also constructed.ConclusionThe chromosomal mapping of the 575 large-insert DNA clones allows for these clones to be integrated into existing zebrafish mapping data. An accurately annotated zebrafish reference genome serves as a valuable resource for investigating the molecular basis of human diseases using zebrafish mutant models.