Search

G protein-coupled receptor 17 (GPR17) and the WNT pathway are critical players of oligodendrocyte (OL) differentiation acting as essential timers in developing brain to achieve fully-myelinating cells. However, whether and how these two systems are related to each other is still unknown. Of interest, both factors are dysregulated in developing and adult brain diseases, including white matter injury and cancer, making the understanding of their reciprocal interactions of potential importance for identifying new targets and strategies for myelin repair. Here, by a combined pharmacological and biotechnological approach, we examined regulatory mechanisms linking WNT signaling to GPR17 expression in OLs. We first analyzed the relative expression of mRNAs encoding for GPR17 and the T cell factor/Lymphoid enhancer-binding factor-1 (TCF/LEF) transcription factors of the canonical WNT/β-CATENIN pathway, in PDGFRα+ and O4+ OLs during mouse post-natal development. In O4+ cells, Gpr17 mRNA level peaked at post-natal day 14 and then decreased concomitantly to the physiological uprise of WNT tone, as shown by increased Lef1 mRNA level. The link between WNT signaling and GPR17 expression was further reinforced in vitro in primary PDGFRα+ cells and in Oli-neu cells. High WNT tone impaired OL differentiation and drastically reduced GPR17 mRNA and protein levels. In Oli-neu cells, WNT/β-CATENIN activation repressed Gpr17 promoter activity through both putative WNT response elements (WRE) and upregulation of the inhibitor of DNA-binding protein 2 (Id2). We conclude that the WNT pathway influences OL maturation by repressing GPR17, which could have implications in pathologies characterized by dysregulations of the OL lineage including multiple sclerosis and oligodendroglioma.

Abstract Aim Attention‐deficit/hyperactivity disorder is a heterogeneous neurobiological disorder that is characterized by inattention, impulsivity, and an increase in motor activity. Although methylphenidate has been used as a medication for decades, unknown is whether methylphenidate treatment can cause drug dependence in patients with attention‐deficit/hyperactivity disorder. This study investigated the reward‐enhancing effects of methylphenidate using intracranial self‐stimulation in an animal model of attention‐deficit/hyperactivity disorder, dopamine transporter knockout mice. Methods For the intracranial self‐stimulation procedures, the mice were trained to nosepoke to receive direct electrical stimulation via an electrode that was implanted in the lateral hypothalamus. After the acquisition of nosepoke responding for intracranial self‐stimulation, the effects of methylphenidate on intracranial self‐stimulation were investigated. Results In the progressive‐ratio procedure, dopamine transporter knockout mice exhibited an increase in intracranial self‐stimulation compared with wild‐type mice. Treatment with 5 and 10 mg/kg methylphenidate increased intracranial self‐stimulation responding in wild‐type mice. Methylphenidate at the same doses did not affect intracranial self‐stimulation responding in dopamine transporter knockout mice. We then investigated the effects of high‐dose methylphenidate (60 mg/kg) in a rate‐frequency procedure. High‐dose methylphenidate significantly decreased intracranial self‐stimulation responding in both wild‐type and dopamine transporter knockout mice. Conclusions These results suggest that low‐dose methylphenidate alters the reward system (ie, increases intracranial self‐stimulation responding) in wild‐type mice via dopamine transporter inhibition, whereas dopamine transporter knockout mice do not exhibit such alterations. High‐dose methylphenidate appears to suppress intracranial self‐stimulation responding not through dopamine transporter inhibition but rather through other mechanisms. These results support the possibility that methylphenidate treatment for attention‐deficit/hyperactivity disorder does not increase the risk of drug dependence, in attention‐deficit/hyperactivity disorder patients with dopamine transporter dysfunction.

Adult hippocampal neurogenesis is strongly impaired in Alzheimer's disease (AD). In several mouse models of AD, it was shown that adult-born neurons exhibit reduced survival and altered synaptic integration due to a severe lack of dendritic spines. In the present work, using the APPxPS1 mouse model of AD, we reveal that this reduced number of spines is concomitant of a marked deficit in their neuronal mitochondrial content. Remarkably, we show that targeting the overexpression of the pro-neural transcription factor Neurod1 into APPxPS1 adult-born neurons restores not only their dendritic spine density, but also their mitochondrial content and the proportion of spines associated with mitochondria. Using primary neurons, a bona fide model of neuronal maturation, we identified that increases of mitochondrial respiration accompany the stimulating effect of Neurod1 overexpression on dendritic growth and spine formation. Reciprocally, pharmacologically impairing mitochondria prevented Neurod1-dependent trophic effects. Thus, since overexpression of Neurod1 into new neurons of APPxPS1 mice rescues spatial memory, our present data suggest that manipulating the mitochondrial system of adult-born hippocampal neurons provides neuronal plasticity to the AD brain. These findings open new avenues for far-reaching therapeutic implications towards neurodegenerative diseases associated with cognitive impairment.

The P2X4 channel is involved in different physiological and pathological conditions and functions in the nervous system. Despite the existence of several mouse models for which the expression of the gene was manipulated, there is still little information on the expression of the protein at the cellular level. In particular, supposedly specific available antibodies have often proved to recognize unrelated proteins in P2X4-deficient mice. Here, we used an in vivo DNA vaccine approach to generate a series of monoclonal antibodies and nanobodies specific for human, mouse, and rat P2X4 channels. We further characterized these antibodies and show that they solely recognize the native form of the proteins both in biochemical and cytometric applications. Some of these antibodies prove to specifically recognize P2X4 channels by immunostaining in brain or sensory ganglia slices, as well as at the cellular and subcellular levels. Due to their clonality, these different antibodies should represent versatile tools for further characterizing the cellular functions of P2X4 in the nervous system as well as at the periphery.

Microglia are the resident macrophages of the central nervous system. They play key roles in brain development, and physiology during life and aging. Equipped with a variety of molecular sensors and through the various functions they can fulfill, they are critically involved in maintaining the brain’s homeostasis. In Alzheimer disease (AD), microglia reaction was initially thought to be incidental and triggered by amyloid deposits and dystrophic neurites. However, recent genome-wide association studies have established that the majority of AD risk loci are found in or near genes that are highly and sometimes uniquely expressed in microglia. This leads to the concept of microglia being critically involved in the early steps of the disease and identified them as important potential therapeutic targets. Whether microglia reaction is beneficial, detrimental or both to AD progression is still unclear and the subject of intense debate. In this review, we are presenting a state-of-knowledge report intended to highlight the variety of microglial functions and pathways shown to be critically involved in AD progression. We first address both the acquisition of new functions and the alteration of their homeostatic roles by reactive microglia. Second, we propose a summary of new important parameters currently emerging in the field that need to be considered to identify relevant microglial targets. Finally, we discuss the many obstacles in designing efficient therapeutic strategies for AD and present innovative technologies that may foster our understanding of microglia roles in the pathology. Ultimately, this work aims to fly over various microglial functions to make a general and reliable report of the current knowledge regarding microglia’s involvement in AD and of the new research opportunities in the field.

BACKGROUND:Monitoring adherence to national recommendations for annual chlamydia screening of female adolescents and young adult women is important for targeting quality improvement interventions to improve low screening rates. However, accurate measurement of rates may vary depending on the data source used to determine eligible sexually-active women. METHODS:The 2001-2004 NHANES data linked with Medicaid administrative data by respondent's unique identifier, the 2011-2012 NHANES data, and the 2004 and 2010 Medicaid data were used in this cross-sectional analysis. We defined self-reported sexual activity by self-reported sexual behaviors, claim-identified sexual activity by reproductive-related claims among women who had ≥ one healthcare claim, HEDIS-defined sexual activity by reproductive-related claims among women who were enrolled in Medicaid for ≥330 days and had ≥ one healthcare claim, and chlamydia tests by claims submitted in the 12 months prior to the survey interview. RESULTS:Of Medicaid women aged 18-25 years, 91.5% self-reported to be sexually-active. Of self-reported sexually-active women aged 18-25 years, 92.0% had ≥ one healthcare claim in the 12 months prior to the survey interview; of this subpopulation, only 58.8% were enrolled in Medicaid for ≥ 330 days in the 12 months prior to the survey interview; of this further subpopulation, 74.1% had healthcare claims identifying them as sexually-active in the 12 months prior to the survey interview. Of HEDIS-defined sexually-active women, 42.4% had chlamydia testing. CONCLUSION:Our study suggests that the number of sexually-active women aged 18-25 years used as the denominator in the chlamydia testing measure could be significantly different, depending upon the definition applied and the data used. Our data highlight the limited representativeness of Medicaid population in the current HEDIS measure on chlamydia testing when a high proportion of women who were enrolled in Medicaid for

CRISPR-Cas9 technology has become an essential tool for plant genome editing. Recent advancements have significantly improved the ability to target multiple genes simultaneously within the same genetic background through various strategies. Additionally, there has been significant progress in developing methods for inducible or tissue-specific editing. These advancements offer numerous possibilities for tailored genome modifications. Building upon existing research, we have developed an optimized and modular strategy allowing the targeting of several genes simultaneously in combination with the synchronized expression of the Cas9 endonuclease in the egg cell. This system allows significant editing efficiency while avoiding mosaicism. In addition, the versatile system we propose allows adaptation to inducible and/or tissue-specific edition according to the promoter chosen to drive the expression of the Cas9 gene. Here, we describe a step-by-step protocol for generating the binary vector necessary for establishing Arabidopsis edited lines using a versatile cloning strategy that combines Gateway® and Golden Gate technologies. We describe a versatile system that allows the cloning of as many guides as needed to target DNA, which can be multiplexed into a polycistronic gene and combined in the same construct with sequences for the expression of the Cas9 endonuclease. The expression of Cas9 is controlled by selecting from among a collection of promoters, including constitutive, inducible, ubiquitous, or tissue-specific promoters. Only one vector containing the polycistronic gene (tRNA-sgRNA) needs to be constructed. For that, sgRNA (composed of protospacers chosen to target the gene of interest and sgRNA scaffold) is cloned in tandem with the pre-tRNA sequence. Then, a single recombination reaction is required to assemble the promoter, the zCas9 coding sequence, and the tRNA-gRNA polycistronic gene. Each element is cloned in an entry vector and finally assembled according to the Multisite Gateway® Technology. Here, we detail the process to express zCas9 under the control of egg cell promoter fused to enhancer sequence (EC1.2en-EC1.1p) and to simultaneously target two multiple C2 domains and transmembrane region protein genes (MCTP3 and MCTP4, respectively at3g57880 and at1g51570), using one or two sgRNA per gene.

Certain sociodemographic factors are associated with low technology access and digital healthy literacy.

Context: Quality assurance in cervical cytology is based on the cyto-histological correlation that is performed in several countries even without standardized protocols. Aims: To evaluate the quality of the Pap smear with the Clinical and Laboratory Standards Institute (CLSI) EP12-A2 guideline in a Peruvian hospital. Settings and Design: This prospective study was carried out at tertiary care national hospital. Methods and Material: The 156 cyto-histological results were collected and coded according to the Bethesda 2014 and FIGO system. The evaluation with the CLSI EP12-A2 guide allowed estimating the performance and quality of the test. Statistical Analysis Used: We performed a descriptive analysis of the cytological and histological data and correlation with the weight Kappa test. From the calculation of the likelihood ratios, the post-test probability was estimated using Bayes' theorem. Results: In cytology, 57 (36.5%) were undetermined abnormalities, 34 (21.8%) low-grade squamous intraepithelial lesion (SIL), and 42 (26.9%) high-grade SIL. Of the total biopsies, 56 (36.9%) were cervical intraepithelial neoplasia (CIN) grade 1, 23 (14.7%) were both CIN grade 2 and 3. We determined sensitivity, specificity, a positive and negative predictive value of 94%, 74.6%, 58%, and 97.1%, respectively. We determined a moderate cyto-histological agreement (κ = 0.57). Atypical squamous cells of undetermined significance (40%), and cannot exclude high-grade squamous intraepithelial lesions (42.1%) that showed higher overdiagnosis results. Conclusions: The quality and performance of the Papanicolaou test show high sensitivity and moderate specificity. The concordance found was moderate and the proportion of underdiagnosis was higher in abnormalities of undetermined significance.

Background: Educating medical students to better understand the complexities of cultural competence, the social determinants and environmental determinants of health that are important and integral components of the medical school curriculum. Methods: In 2014, Weill Cornell Medicine (WCM) implemented a new curriculum, the adoption of which provided the means to enhance an existing global health program, informally introduced in 2009, and to address the issues of cultural competency. In this article, we share WCM’s experience in building and expanding its global health curriculum. Results: A hallmark of our program is the successful collaboration between students and faculty to create a multi-disciplinary global health program that incorporates electives, clinical field placement, and collaborative research. Conclusion: Key lessons learned through our experience include the necessity for strong faculty-student collaboration, full support from the administration, and building global partnerships. Our example could be a useful guide for other medical schools seeking to establish a global health education curriculum.

OBJECTIVES:. Severe acute brain injury (SABI) from cardiac arrest and traumatic brain injury happens suddenly and unexpectedly, carrying high potential for lifelong disability with substantial prognostic uncertainty. Comprehensive assessments of family experiences and support needs after SABI are lacking. Our objective is to elicit “on-the-ground” perspectives about the experiences and needs of families of patients with SABI. DESIGN:. Two-phase qualitative study of families and multidisciplinary U.S. healthcare professionals (mHCPs) with expertise in SABI: Phase 1 included semistructured interviews to generate formative findings; phase 2 entailed facilitated discussions to confirm and expand initial findings. SETTING:. Phase 1: academic medical center; phase 2: virtual workshop. SUBJECTS:. Phase 1 included seven family members and 12 mHCPs. Phase 2 included nationally recruited stakeholders (17 family members and 12 mHCPs). INTERVENTION:. None. MEASUREMENTS AND RESULTS:. We explored: 1) what are families’ needs in the first 48 hours? 2) How are these needs addressed? and 3) How can hospitals better meet these needs? Qualitative analysis included inductive and deductive approaches guided by a conceptual ecological model. Four major needs were identified: 1) challenges in coping with uncertainty in early prognostication, 2) inattention to physical needs of family, 3) deficits in compassionate and consistent communication, and 4) need for engagement with families as stakeholders in improving future practices. Participants’ recommendations included: 1) ways to communicate more clearly and consistently, 2) better assistance with navigating resources and access to places for families to care for themselves, and 3) opportunities for families to remain connected with their loved ones, social support networks, and the clinical team. CONCLUSIONS:. Stakeholders identified novel insights regarding families’ experiences during the hospitalization of comatose SABI patients and factors that can contribute to improved decision-making and physical/emotional outcomes. Interventions to address these unmet needs are promising targets to improve outcomes.

Urinary tract infections (UTIs) are associated with changes in the gut microbiome. Here, the authors evaluate the relationship between the gut microbiome and development of UTI in kidney transplant patients and show that uropathogenic gut abundance might represent a risk factor for development of bacteriuria and UTI.

Abstract: Low-dose interleukin-2 (IL-2) therapy for chronic graft-versus-host disease (cGVHD) generates a rapid rise in plasma IL-2 levels and CD4+CD25+CD127−Foxp3+ regulatory T-cell (CD4Treg) proliferation, but both decrease over time despite continued daily administration. To test whether IL-2 dose escalation at the time of anticipated falls in plasma levels could circumvent tachyphylaxis and enhance CD4Treg expansion, we conducted a phase 1 trial in 10 adult and 11 pediatric patients with steroid-refractory cGVHD (www.clinicaltrials.gov: NCT02318082). Daily IL-2 was initiated in children and adults (0.33 × 106 and 0.67 × 106 IU/m2 per day, respectively). Dose escalations were scheduled at weeks 2 and 4 to a maximum dose of 1 × 106 IU/m2 per day in children and 2 × 106 IU/m2 per day in adults. Patients continued at their maximum tolerated dose (MTD) until week 8. Children tolerated IL-2 dose escalation with partial responses (PRs) in 9 of 11 patients (82%) at multiple cGVHD sites, including lung. Patient-reported outcome scores for skin and lung improved significantly in pediatric patients. In contrast, 5 of 10 adults required dose reduction, and only 2 of 7 evaluable patients (29%) had PRs at week 8. CD4Tregs and natural killer cells expanded in both cohorts without significant changes in conventional CD4+ T cells (Tcons) or CD8+ T cells. Children achieved a higher median CD4Treg/Tcon ratio at week 8 (0.4 vs 0.18, P = .02) despite lower IL-2 doses. We show for the first time that low-dose IL-2 is safe and effective in children with advanced cGVHD. In adults, escalation above the previously defined MTD did not improve CD4Treg expansion or clinical response.

The vast bacteriophage population harbors an immense reservoir of genetic information. Almost 2000 phage genomes have been sequenced from phages infecting hosts in the phylum Actinobacteria, and analysis of these genomes reveals substantial diversity, pervasive mosaicism, and novel mechanisms for phage replication and lysogeny. Here, we describe the isolation and genomic characterization of 46 phages from environmental samples at various geographic locations in the U.S. infecting a single Arthrobacter sp. strain. These phages include representatives of all three virion morphologies, and Jasmine is the first sequenced podovirus of an actinobacterial host. The phages also span considerable sequence diversity, and can be grouped into 10 clusters according to their nucleotide diversity, and two singletons each with no close relatives. However, the clusters/singletons appear to be genomically well separated from each other, and relatively few genes are shared between clusters. Genome size varies from among the smallest of siphoviral phages (15,319 bp) to over 70 kbp, and G+C contents range from 45-68%, compared to 63.4% for the host genome. Although temperate phages are common among other actinobacterial hosts, these Arthrobacter phages are primarily lytic, and only the singleton Galaxy is likely temperate.