RESUMEN
Evolution of the mammalian brain encompassed a remarkable increase in size of the cerebral cortex, which includes tangential and radial expansion. However, the mechanisms underlying these key features are still largely unknown. Here, we identified the DNA-associated protein Trnp1 as a regulator of cerebral cortex expansion in both of these dimensions. Gain- and loss-of-function experiments in the mouse cerebral cortex in vivo demonstrate that high Trnp1 levels promote neural stem cell self-renewal and tangential expansion. In contrast, lower levels promote radial expansion, with a potent increase of the number of intermediate progenitors and basal radial glial cells leading to folding of the otherwise smooth murine cerebral cortex. Remarkably, TRNP1 expression levels exhibit regional differences in the cerebral cortex of human fetuses, anticipating radial or tangential expansion. Thus, the dynamic regulation of Trnp1 is critical to control tangential and radial expansion of the cerebral cortex in mammals.
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Corteza Cerebral/crecimiento & desarrollo , Proteínas Nucleares/metabolismo , Secuencia de Aminoácidos , Animales , Proteínas de Ciclo Celular , Corteza Cerebral/citología , Proteínas de Unión al ADN , Embrión de Mamíferos/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Datos de Secuencia Molecular , Células-Madre Neurales/metabolismo , Neuroglía/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/genética , Activación TranscripcionalRESUMEN
BACKGROUND: Mental disorders are the leading global cause of health burden among adolescents. However, prevalence data for mental disorders among adolescents in low-income and middle-income countries are scarce with often limited generalisability. This study aimed to generate nationally representative prevalence estimates for mental disorders in adolescents in Kenya, Indonesia, and Viet Nam. METHODS: As part of the National Adolescent Mental Health Surveys (NAMHS), a multinational cross-sectional study, nationally representative household surveys were conducted in Kenya, Indonesia, and Viet Nam between March and December, 2021. Adolescents aged 10-17 years and their primary caregiver were interviewed from households selected randomly according to sampling frames specifically designed to elicit nationally representative results. Six mental disorders (social phobia, generalised anxiety disorder, major depressive disorder, post-traumatic stress disorder, conduct disorder, and attention-deficit hyperactivity disorder) were assessed with the Diagnostic Interview Schedule for Children, Version 5. Suicidal behaviours and self-harm in the past 12 months were also assessed. Prevalence in the past 12 months and past 4 weeks was calculated for each mental disorder and collectively for any mental disorder (ie, of the six mental disorders assessed). Prevalence of suicidal behaviours (ie, ideation, planning, and attempt) and self-harm in the past 12 months was calculated, along with adjusted odds ratios (aORs) to show the association with prevalence of any mental disorder in the past 12 months. Inverse probability weighting was applied to generate national estimates with corresponding 95% CIs. FINDINGS: Final samples consisted of 5155 households (ie, adolescent and primary caregiver pairs) from Kenya, 5664 households from Indonesia, and 5996 households from Viet Nam. In Kenya, 2416 (46·9%) adolescents were male and 2739 (53·1%) were female; in Indonesia, 2803 (49·5%) adolescents were male and 2861 (50·5%) were female; and in Viet Nam, 3151 (52·5%) were male and 2845 (47·4%) were female. Prevalence of any mental disorder in the past 12 months was 12·1% (95% CI 10·9-13·5) in Kenya, 5·5% (4·3-6·9) in Indonesia, and 3·3% (2·7-4·1) in Viet Nam. Prevalence in the past 4 weeks was 9·4% (8·3-10·6) in Kenya, 4·4% (3·4-5·6) in Indonesia, and 2·7% (2·2-3·3) in Viet Nam. The prevalence of suicidal behaviours in the past 12 months was low in all three countries, with suicide ideation ranging from 1·4% in Indonesia (1·0-2·0) and Viet Nam (1·0-1·9) to 4·6% (3·9-5·3) in Kenya, suicide planning ranging from 0·4% in Indonesia (0·3-0·8) and Viet Nam (0·2-0·6) to 2·4% (1·9-2·9) in Kenya, and suicide attempts ranging from 0·2% in Indonesia (0·1-0·4) and Viet Nam (0·1-0·3) to 1·0% (0·7-1·4) in Kenya. The prevalence of self-harm in the past 12 months was also low in all three countries, ranging from 0·9% (0·6-1·3) in Indonesia to 1·2% (0·9-1·7) in Kenya. However, the prevalence of suicidal behaviours and self-harm in the past 12 months was significantly higher among those with any mental disorder in the past 12 months than those without (eg, aORs for suicidal ideation ranged from 7·1 [3·1-15·9] in Indonesia to 14·7 [7·5-28·6] in Viet Nam). INTERPRETATION: NAMHS provides the first national adolescent mental disorders prevalence estimates for Kenya, Indonesia, and Viet Nam. These data can inform mental health and broader health policies in low-income and middle-income countries. FUNDING: The University of Queensland in America (TUQIA) through support from Pivotal Ventures, a Melinda French Gates company.
Asunto(s)
Trastornos Mentales , Humanos , Adolescente , Indonesia/epidemiología , Femenino , Estudios Transversales , Masculino , Kenia/epidemiología , Prevalencia , Vietnam/epidemiología , Niño , Trastornos Mentales/epidemiología , Encuestas EpidemiológicasRESUMEN
Neuroinflammation has been suggested as a pathogenetic mechanism contributing to Parkinson's disease (PD). However, anti-inflammatory treatment strategies have not yet been established as a therapeutic option for PD patients. We have used a human α-synuclein mouse model of progressive PD to examine the anti-inflammatory and neuroprotective effects of inflammasome inhibition on dopaminergic (DA) neurons in the substantia nigra (SN). As the NLRP3 (NOD-, LRR- and pyrin domain-containing 3)-inflammasome is a core interface for both adaptive and innate inflammation and is also highly druggable, we investigated the implications of its inhibition. Repeat administration of MCC950, an inhibitor of NLRP3, in a PD model with ongoing pathology reduced CD4+ and CD8+ T cell infiltration into the SN. Furthermore, the anti-inflammasome treatment mitigated microglial activation and modified the aggregation of α-synuclein protein in DA neurons. MCC950-treated mice showed significantly less neurodegeneration of DA neurons and a reduction in PD-related motor behavior. In summary, early inflammasome inhibition can reduce neuroinflammation and prevent DA cell death in an α-synuclein mouse model for progressive PD.
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Inflamasomas , Enfermedad de Parkinson , Humanos , Ratones , Animales , Inflamasomas/metabolismo , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/etiología , Enfermedad de Parkinson/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , alfa-Sinucleína/metabolismo , Neuronas Dopaminérgicas , Enfermedades Neuroinflamatorias , Microglía/metabolismo , Ratones Endogámicos NOD , Sulfonamidas/farmacología , Modelos Animales de Enfermedad , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: To explore the cross-sectional and longitudinal association between vitamin D and depressive symptoms across early adolescence. METHODS: This longitudinal study included 1607 early adolescents [mean (s.d.) age, 12.49 years; 972 (60.5%) males] from the Chinese Early Adolescents Cohort, recruited from a middle school in Anhui Province and followed up annually (2019-2021). Serum 25(OH)D levels were measured in both 2019 and 2021. Self-reports on depression were assessed at each of three time points from 2019 to 2021. RESULTS: In the whole sample, higher baseline serum 25(OH)D levels were linked with a lower risk of cumulative incident depression within two-year follow-ups (adjusted RR = 0.97, 95% CI 0.94-0.99) and the increasing trajectory of depression symptoms across the three waves (adjusted RR = 0.97, 95% CI 0.95-0.99). Baseline vitamin D deficiency (VDD) (adjusted RR = 1.50, 95% CI 1.10-2.05) were associated with an increased risk for the increasing trajectory of depression symptoms across the three waves. Remitted VDD was positively related to one dichotomous depression symptoms across three waves (adjusted OR 2.15, 95% CI 1.15-4.01). The above-mentioned significant association was also found in males. Additionally, baseline VDD (adjusted OR 1.59, 95% CI 1.04-2.44) and persistent VDD (adjusted OR 1.58, 95% CI 1.02-2.60) were linked to an increased risk of having two dichotomous depression symptoms only in males. CONCLUSIONS: Our results highlight a prospective association between baseline vitamin D and depression risk in early adolescents. Additionally, a male-specific association between vitamin D and depression risk was observed. Our findings support a potential beneficial effect of vitamin D supplementation in reducing depression risk in early adolescents.
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Deficiencia de Vitamina D , Vitamina D , Masculino , Humanos , Adolescente , Niño , Femenino , Depresión/epidemiología , Estudios Longitudinales , Estudios Transversales , Deficiencia de Vitamina D/epidemiologíaRESUMEN
Early adolescents (ages 10-14) living in low- and middle-income countries have heightened vulnerability to psychosocial risks, but available evidence from these settings is limited. This study used data from the Global Early Adolescent Study to characterize prototypical patterns of emotional and behavioral problems among 10,437 early adolescents (51% female) living in the Democratic Republic of Congo (DRC), Malawi, Indonesia, and China, and explore the extent to which these patterns varied by country and sex. LCA was used to identify and classify patterns of emotional and behavioral problems separately by country. Within each country, measurement invariance by sex was evaluated. LCA supported a four-class solution in DRC, Malawi, and Indonesia, and a three-class solution in China. Across countries, early adolescents fell into the following subgroups: Well-Adjusted (40-62%), Emotional Problems (14-29%), Behavioral Problems (15-22%; not present in China), and Maladjusted (4-15%). Despite the consistency of these patterns, there were notable contextual differences. Further, tests of measurement invariance indicated that the prevalence and nature of these classes differed by sex. Findings can be used to support the tailoring of interventions targeting psychosocial adjustment, and suggest that such programs may have utility across diverse cross-national settings.
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Trastornos Mentales , Problema de Conducta , Humanos , Adolescente , Femenino , Masculino , Países en Desarrollo , Emociones , ChinaRESUMEN
BACKGROUND: Hoffman syndrome is a syndromic, inborn error of immunity due to autosomal-dominant mutations in TOP2B, an essential gene required to alleviate topological stress during DNA replication and gene transcription. Although mutations identified in patients lead to a block in B-cell development and the absence of circulating B cells, an effect on natural killer (NK) cells was not previously examined. OBJECTIVE: We sought to determine whether disease-associated mutations in TOP2B impact NK-cell development and function. METHODS: Using a knockin murine model and patient-derived induced pluripotent stem cells (iPSCs), we investigated NK-cell development in mouse bone marrow and spleen, and performed immunophenotyping by flow cytometry, gene expression, and functional assessment of cytotoxic activity in murine NK cells, and human IPSC-derived NK cells. RESULTS: Mature NK cells were reduced in the periphery of TOP2B knockin mice consistent with patient reports, with reduced cytotoxicity toward target cell lines. IPSCs were successfully derived from patients with Hoffman syndrome, but under optimal conditions showed reduced cytotoxicity compared with iPSC-derived NK cells from healthy controls. CONCLUSIONS: Hoffman syndrome-associated mutations in TOP2B impact NK-cell development and function in murine and human models.
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Células Madre Pluripotentes Inducidas , Células Asesinas Naturales , Animales , Línea Celular , Anomalías Craneofaciales , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Deformidades Congénitas de las Extremidades , Ratones , Mutación , Enfermedades de Inmunodeficiencia Primaria , Anomalías UrogenitalesRESUMEN
Dysregulation of proteins involved in synaptic plasticity is associated with pathologies in the CNS, including psychiatric disorders. The bed nucleus of the stria terminalis (BNST), a brain region of the extended amygdala circuit, has been identified as the critical hub responsible for fear responses related to stress coping and pathologic systems states. Here, we report that one particular nucleus, the oval nucleus of the BNST (ovBNST), is rich in brain-derived neurotrophic factor (BDNF) and tropomyosin receptor kinase B (TrkB) receptor. Whole-cell patch-clamp recordings of neurons from male mouse ovBNST in vitro showed that the BDNF/TrkB interaction causes a hyperpolarizing shift of the membrane potential from resting value, mediated by an inwardly rectifying potassium current, resulting in reduced neuronal excitability in all major types of ovBNST neurons. Furthermore, BDNF/TrkB signaling mediated long-term depression (LTD) at postsynaptic sites in ovBNST neurons. LTD of ovBNST neurons was prevented by a BDNF scavenger or in the presence of TrkB inhibitors, indicating the contribution to LTD induction. Our data identify BDNF/TrkB signaling as a critical regulator of synaptic activity in ovBNST, which acts at postsynaptic sites to dampen excitability at short and long time scales. Given the central role of ovBNST in mediating maladaptive behaviors associated with stress exposure, our findings suggest a synaptic entry point of the BDNF/TrkB system for adaptation to stressful environmental encounters.
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Factor Neurotrófico Derivado del Encéfalo/fisiología , Depresión Sináptica a Largo Plazo/fisiología , Glicoproteínas de Membrana/fisiología , Proteínas Tirosina Quinasas/fisiología , Núcleos Septales/fisiología , Animales , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Masculino , Glicoproteínas de Membrana/antagonistas & inhibidores , Glicoproteínas de Membrana/metabolismo , Potenciales de la Membrana/fisiología , Ratones , Ratones Endogámicos C57BL , Técnicas de Placa-Clamp , Canales de Potasio de Rectificación Interna/fisiología , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/metabolismo , Núcleos Septales/metabolismo , Estrés Psicológico/fisiopatología , Sinapsis/fisiología , Transmisión Sináptica/fisiologíaRESUMEN
Antibody-dependent cellular cytotoxicity (ADCC) is a key effector mechanism of natural killer (NK) cells that is mediated by therapeutic monoclonal antibodies (mAbs). This process is facilitated by the Fc receptor CD16a on human NK cells. CD16a appears to be the only activating receptor on NK cells that is cleaved by the metalloprotease a disintegrin and metalloproteinase-17 upon stimulation. We previously demonstrated that a point mutation of CD16a prevents this activation-induced surface cleavage. This noncleavable CD16a variant is now further modified to include the high-affinity noncleavable variant of CD16a (hnCD16) and was engineered into human induced pluripotent stem cells (iPSCs) to create a renewable source for human induced pluripotent stem cell-derived NK (hnCD16-iNK) cells. Compared with unmodified iNK cells and peripheral blood-derived NK (PB-NK) cells, hnCD16-iNK cells proved to be highly resistant to activation-induced cleavage of CD16a. We found that hnCD16-iNK cells were functionally mature and exhibited enhanced ADCC against multiple tumor targets. In vivo xenograft studies using a human B-cell lymphoma demonstrated that treatment with hnCD16-iNK cells and anti-CD20 mAb led to significantly improved regression of B-cell lymphoma compared with treatment utilizing anti-CD20 mAb with PB-NK cells or unmodified iNK cells. hnCD16-iNK cells, combined with anti-HER2 mAb, also mediated improved survival in an ovarian cancer xenograft model. Together, these findings show that hnCD16-iNK cells combined with mAbs are highly effective against hematologic malignancies and solid tumors that are typically resistant to NK cell-mediated killing, demonstrating the feasibility of producing a standardized off-the-shelf engineered NK cell therapy with improved ADCC properties to treat malignancies that are otherwise refractory.
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Anticuerpos Monoclonales/uso terapéutico , Citotoxicidad Celular Dependiente de Anticuerpos , Células Asesinas Naturales/trasplante , Linfoma de Células B/terapia , Neoplasias Ováricas/terapia , Receptores de IgG/inmunología , Animales , Antígenos CD20/inmunología , Antineoplásicos Inmunológicos/uso terapéutico , Línea Celular , Línea Celular Tumoral , Femenino , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/inmunología , Células Asesinas Naturales/citología , Células Asesinas Naturales/inmunología , Linfoma de Células B/inmunología , Ratones Endogámicos NOD , Ratones SCID , Neoplasias Ováricas/inmunologíaRESUMEN
BACKGROUND: Despite of a multimodal approach, recurrences can hardly be prevented in glioblastoma. This may be in part due to so called glioma stem cells. However, there is no established marker to identify these stem cells. METHODS: Paired samples from glioma patients were analyzed by immunohistochemistry for expression of the following stem cell markers: CD133, Musashi, Nanog, Nestin, octamer-binding transcription factor 4 (Oct4), and sex determining region Y-box 2 (Sox2). In addition, the expression of osteopontin (OPN) was investigated. The relative number of positively stained cells was determined. By means of Kaplan-Meier analysis, a possible association with overall survival by marker expression was investigated. RESULTS: Sixty tissue samples from 30 patients (17 male, 13 female) were available for analysis. For Nestin, Musashi and OPN a significant increase was seen. There was also an increase (not significant) for CD133 and Oct4. Patients with mutated Isocitrate Dehydrogenase-1/2 (IDH-1/2) status had a reduced expression for CD133 and Nestin in their recurrent tumors. Significant correlations were seen for CD133 and Nanog between OPN in the primary and recurrent tumor and between CD133 and Nestin in recurrent tumors. By confocal imaging we could demonstrate a co-expression of CD133 and Nestin within recurrent glioma cells. Patients with high CD133 expression had a worse prognosis (22.6 vs 41.1 months, p = 0.013). A similar trend was seen for elevated Nestin levels (24.9 vs 41.1 months, p = 0.08). CONCLUSIONS: Most of the evaluated markers showed an increased expression in their recurrent tumor. CD133 and Nestin were associated with survival and are candidate markers for further clinical investigation.
RESUMEN
BACKGROUND: Antigen-specific neuroinflammation and neurodegeneration are characteristic for neuroimmunological diseases. In Parkinson's disease (PD) pathogenesis, α-synuclein is a known culprit. Evidence for α-synuclein-specific T cell responses was recently obtained in PD. Still, a causative link between these α-synuclein responses and dopaminergic neurodegeneration had been lacking. We thus addressed the functional relevance of α-synuclein-specific immune responses in PD in a mouse model. METHODS: We utilized a mouse model of PD in which an Adeno-associated Vector 1/2 serotype (AAV1/2) expressing human mutated A53T-α-Synuclein was stereotactically injected into the substantia nigra (SN) of either wildtype C57BL/6 or Recombination-activating gene 1 (RAG1)-/- mice. Brain, spleen, and lymph node tissues from different time points following injection were then analyzed via FACS, cytokine bead assay, immunohistochemistry and RNA-sequencing to determine the role of T cells and inflammation in this model. Bone marrow transfer from either CD4+/CD8-, CD4-/CD8+, or CD4+/CD8+ (JHD-/-) mice into the RAG-1-/- mice was also employed. In addition to the in vivo studies, a newly developed A53T-α-synuclein-expressing neuronal cell culture/immune cell assay was utilized. RESULTS: AAV-based overexpression of pathogenic human A53T-α-synuclein in dopaminergic neurons of the SN stimulated T cell infiltration. RNA-sequencing of immune cells from PD mouse brains confirmed a pro-inflammatory gene profile. T cell responses were directed against A53T-α-synuclein-peptides in the vicinity of position 53 (68-78) and surrounding the pathogenically relevant S129 (120-134). T cells were required for α-synuclein-induced neurodegeneration in vivo and in vitro, while B cell deficiency did not protect from dopaminergic neurodegeneration. CONCLUSIONS: Using T cell and/or B cell deficient mice and a newly developed A53T-α-synuclein-expressing neuronal cell culture/immune cell assay, we confirmed in vivo and in vitro that pathogenic α-synuclein peptide-specific T cell responses can cause dopaminergic neurodegeneration and thereby contribute to PD-like pathology.
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Enfermedad de Parkinson , alfa-Sinucleína , Animales , Modelos Animales de Enfermedad , Dopamina , Neuronas Dopaminérgicas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Enfermedad de Parkinson/patología , ARN , Sustancia Negra/metabolismo , Linfocitos T/metabolismo , alfa-Sinucleína/metabolismoRESUMEN
PURPOSE: To report 12-month visual outcomes, incidence of intraocular pressure (IOP) changes and postoperative complications after pars plana vitrectomy with retropupillary implantation of an iris claw intraocular lens (IOL) in aphakic eyes after complicated cataract surgery and eyes with a dislocation of the IOL. METHODS: This is a retrospective analysis of eyes undergoing implantation of an iris claw IOL combined with pars plana vitrectomy from 1st of January 2009 until 30th of June 2018 after complicated cataract extraction with capsular loss (Group A) or dislocation of an IOL (Group B). Corrected distance visual acuity was analyzed in logarithm of the minimum angle of resolution (logMAR) units, IOP was recorded in mmHg. RESULTS: Eyes in Group A (n = 49) improved from a preoperative median visual acuity of 0.523 logMAR (Snellen 20/65) to 0.201 logMAR (Snellen 20/30), P < 0.01. Eyes in Group B (n = 126) showed stable median visual acuity, preoperative 0.301 logMAR (Snellen 20/40) versus postoperative 0.222 logMAR (Snellen 20/30), P > 0.05. During 12 months in Group A, IOP >21 mmHg occurred in 9 (18.4%) eyes; no eye had an IOP <6 mmHg. In Group B, IOP >21 mmHg occurred in 15 (11.9%) eyes, IOP <6 mmHg in 5 (4%) cases. None of the eyes in Group A and B had IOP >21 mmHg or <6 mmHg at 12 months follow-up. CONCLUSION: The retropupillary implantation of an iris claw IOL with pars plana vitrectomy provides adequate visual rehabilitation and seems to be safe in IOP changes.
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Lentes Intraoculares , Vitrectomía , Humanos , Implantación de Lentes Intraoculares , Complicaciones Posoperatorias , Estudios Retrospectivos , Agudeza VisualRESUMEN
Motor learning depends on synaptic plasticity between corticostriatal projections and striatal medium spiny neurons. Retrograde tracing from the dorsolateral striatum reveals that both layer II/III and V neurons in the motor cortex express BDNF as a potential regulator of plasticity in corticostriatal projections in male and female mice. The number of these BDNF-expressing cortical neurons and levels of BDNF protein are highest in juvenile mice when adult motor patterns are shaped, while BDNF levels in the adult are low. When mice are trained by physical exercise in the adult, BDNF expression in motor cortex is reinduced, especially in layer II/III projection neurons. Reduced expression of cortical BDNF in 3-month-old mice results in impaired motor learning while space memory is preserved. These findings suggest that activity regulates BDNF expression differentially in layers II/III and V striatal afferents from motor cortex and that cortical BDNF is essential for motor learning.SIGNIFICANCE STATEMENT Motor learning in mice depends on corticostriatal BDNF supply, and regulation of BDNF expression during motor learning is highest in corticostriatal projection neurons in cortical layer II/III.
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Factor Neurotrófico Derivado del Encéfalo/fisiología , Aprendizaje/fisiología , Actividad Motora , Corteza Motora/fisiología , Neuronas/fisiología , Animales , Factor Neurotrófico Derivado del Encéfalo/genética , Cuerpo Estriado/fisiología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Vías Nerviosas/fisiología , Plasticidad Neuronal , Condicionamiento Físico AnimalRESUMEN
Glycine receptors (GlyRs) are the major mediators of fast synaptic inhibition in the adult human spinal cord and brainstem. Hereditary mutations to GlyRs can lead to the rare, but potentially fatal, neuromotor disorder hyperekplexia. Most mutations located in the large intracellular domain (TM3-4 loop) of the GlyRα1 impair surface expression levels of the receptors. The novel GLRA1 mutation P366L, located in the TM3-4 loop, showed normal surface expression but reduced chloride currents, and accelerated whole-cell desensitization observed in whole-cell recordings. At the single-channel level, we observed reduced unitary conductance accompanied by spontaneous opening events in the absence of extracellular glycine. Using peptide microarrays and tandem MS-based analysis methods, we show that the proline-rich stretch surrounding P366 mediates binding to syndapin I, an F-BAR domain protein involved in membrane remodeling. The disruption of the noncanonical Src homology 3 recognition motif by P366L reduces syndapin I binding. These data suggest that the GlyRα1 subunit interacts with intracellular binding partners and may therefore play a role in receptor trafficking or synaptic anchoring, a function thus far only ascribed to the GlyRß subunit. Hence, the P366L GlyRα1 variant exhibits a unique set of properties that cumulatively affect GlyR functionality and thus might explain the neuropathological mechanism underlying hyperekplexia in the mutant carriers. P366L is the first dominant GLRA1 mutation identified within the GlyRα1 TM3-4 loop that affects GlyR physiology without altering protein expression at the whole-cell and surface levels.SIGNIFICANCE STATEMENT We show that the intracellular domain of the inhibitory glycine receptor α1 subunit contributes to trafficking and synaptic anchoring. A proline-rich stretch in this receptor domain forms a noncanonical recognition motif important for the interaction with syndapin I (PACSIN1). The disruption of this motif, as present in a human patient with hyperekplexia led to impaired syndapin I binding. Functional analysis revealed that the altered proline-rich stretch determines several functional physiological parameters of the ion channel (e.g., faster whole-cell desensitization) reduced unitary conductance and spontaneous opening events. Thus, the proline-rich stretch from the glycine receptor α1 subunit represents a multifunctional intracellular protein motif.
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Receptores de Glicina/genética , Receptores de Glicina/metabolismo , Síndrome de la Persona Rígida/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Secuencias de Aminoácidos , Animales , Humanos , Mutación , Unión Proteica/genética , Estructura Cuaternaria de Proteína , Transporte de Proteínas/genética , Receptores de Glicina/químicaRESUMEN
BACKGROUND: Lung cancer management is characterised by a high disease burden, poor survival and substantial variation in management and outcomes. Service redesign provides opportunities for quality improvement (QI) and this improvement may be leveraged across multiple sites using QI collaboration. AIM: This initiative targeted Quality Improvement (QI) in lung cancer management, engaging a QI collaborative using service redesign methodologies in five Victorian hospitals. QI targets included timeliness from referral and diagnosis to treatment, multi-disciplinary meeting (MDM) presentation and supportive care screening. Redesign strategies targeted process sustainability through enhanced team capability. METHODS: This study engaged a prospective quality improvement cohort design targeting newly diagnosed tissue confirmed lung cancer with 6-month pre-intervention period and 6-month redesign implementation period, between September 2016 and August 2017, evaluated using Interrupted Time Series Analysis. Hospital sites included three regional and two metropolitan hospitals in Victoria. QI redesign targeted time intervals from referral to first specialist appointment (FSA), referral to diagnosis, diagnosis to first treatment (any intent), MDM documented in medical records and Supportive Care Screening Tool documented in medical records. RESULTS: There was a marked reduction in referral to FSA interval across all sites, with median (interquartile range) falling from 6 (0-15) to 4 (1-10) days, and proportion seen by a specialist within 14 days increased from 74.3% to 84.2%. The interval between diagnosis and treatment was not substantively changed in the 6-month implementation period. The proportion of subjects with documented presentation to the MDM increased from 61% to 67%. The proportion for which Supportive Care Screening documentation remained low at 26.3% post-intervention. CONCLUSIONS: Data-driven redesign initiatives enable identification and analysis of clinical practice variation and may be utilised to enhance timeliness of cancer care and improve local data service capabilities.
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Neoplasias Pulmonares , Mejoramiento de la Calidad , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/epidemiología , Neoplasias Pulmonares/terapia , Estudios Prospectivos , Derivación y ConsultaRESUMEN
Inflammation is crucial in the pathophysiology of stroke and thus a promising therapeutic target. High-frequency stimulation (HFS) of the mesencephalic locomotor region (MLR) reduces perilesional inflammation after photothrombotic stroke (PTS). However, the underlying mechanism is not completely understood. Since distinct neural and immune cells respond to electrical stimulation by releasing acetylcholine, we hypothesize that HFS might trigger the cholinergic anti-inflammatory pathway via activation of the α7 nicotinic acetylcholine receptor (α7nAchR). To test this hypothesis, rats underwent PTS and implantation of a microelectrode into the MLR. Three hours after intervention, either HFS or sham-stimulation of the MLR was applied for 24 h. IFN-γ, TNF-α, and IL-1α were quantified by cytometric bead array. Choline acetyltransferase (ChAT)+ CD4+-cells and α7nAchR+-cells were quantified visually using immunohistochemistry. Phosphorylation of NFĸB, ERK1/2, Akt, and Stat3 was determined by Western blot analyses. IFN-γ, TNF-α, and IL-1α were decreased in the perilesional area of stimulated rats compared to controls. The number of ChAT+ CD4+-cells increased after MLR-HFS, whereas the amount of α7nAchR+-cells was similar in both groups. Phospho-ERK1/2 was reduced significantly in stimulated rats. The present study suggests that MLR-HFS may trigger anti-inflammatory processes within the perilesional area by modulating the cholinergic system, probably via activation of the α7nAchR.
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Inflamación/terapia , Neuroinmunomodulación/genética , Accidente Cerebrovascular/terapia , Receptor Nicotínico de Acetilcolina alfa 7/genética , Acetilcolina/metabolismo , Animales , Colina O-Acetiltransferasa/genética , Modelos Animales de Enfermedad , Estimulación Eléctrica , Humanos , Inflamación/genética , Inflamación/patología , Mesencéfalo/patología , Mesencéfalo/efectos de la radiación , Neuroinmunomodulación/efectos de la radiación , Ratas , Accidente Cerebrovascular/genética , Accidente Cerebrovascular/patología , Factor de Necrosis Tumoral alfa/genética , Receptor Nicotínico de Acetilcolina alfa 7/metabolismoRESUMEN
NMR relaxation dispersion experiments play a central role in exploring molecular motion over an important range of timescales, and are an example of a broader class of multidimensional NMR experiments that probe important biomolecules. However, resolving the spectral features of these experiments using the Fourier transform requires sampling the full Nyquist grid of data, making these experiments very costly in time. Practitioners often reduce the experiment time by omitting 1D experiments in the indirectly observed dimensions, and reconstructing the spectra using one of a variety of post-processing algorithms. In prior work, we described a fast, Fourier-based reconstruction method using iterated maps according to the Difference Map algorithm of Veit Elser (DiffMap). Here we describe coDiffMap, a new reconstruction method that is based on DiffMap, but which exploits the strong correlations between 2D data slices in a pseudo-3D experiment. We apply coDiffMap to reconstruct dispersion curves from an [Formula: see text] relaxation dispersion experiment, and demonstrate that the method provides fast reconstructions and accurate relaxation curves down to very low numbers of sparsely-sampled data points.
Asunto(s)
Análisis de Fourier , Procesamiento de Imagen Asistido por Computador/métodos , Simulación de Dinámica Molecular , Resonancia Magnética Nuclear Biomolecular/métodos , Algoritmos , Factores de TiempoRESUMEN
Many of the ubiquitous experiments of biomolecular NMR, including [Formula: see text], [Formula: see text], and CEST, involve acquiring repeated 2D spectra under slightly different conditions. Such experiments are amenable to acceleration using non-uniform sampling spectral reconstruction methods that take advantage of prior information. We previously developed one such technique, an iterated maps method (DiffMap) that we successfully applied to 2D NMR spectra, including [Formula: see text] relaxation dispersion data. In that prior work, we took a top-down approach to reconstructing the 2D spectrum with a minimal number of sparse samples, reaching an undersampling fraction that appeared to leave some room for improvement. In this study, we develop an in-depth understanding of the action of the DiffMap algorithm, identifying the factors that cause reconstruction errors for different undersampling fractions. This improved understanding allows us to formulate a bottom-up approach to finding the lowest number of sparse samples required to accurately reconstruct individual spectral features with DiffMap. We also discuss the difficulty of extending this method to reconstructing many peaks at once, and suggest a way forward.
Asunto(s)
Algoritmos , Resonancia Magnética Nuclear Biomolecular/métodos , Tamaño de la Muestra , Manejo de Especímenes/métodosRESUMEN
The aryl hydrocarbon receptor (AHR) plays an important physiological role in hematopoiesis. AHR is highly expressed in hematopoietic stem and progenitor cells (HSPCs) and inhibition of AHR results in a marked expansion of human umbilical cord blood-derived HSPCs following cytokine stimulation. It is unknown whether AHR also contributes earlier in human hematopoietic development. To model hematopoiesis, human embryonic stem cells (hESCs) were allowed to differentiate in defined conditions in the presence of the AHR antagonist StemReginin-1 (SR-1) or the AHR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). We demonstrate a significant increase in CD34+CD31+ hematoendothelial cells in SR-1-treated hESCs, as well as a twofold expansion of CD34+CD45+ hematopoietic progenitor cells. Hematopoietic progenitor cells were also significantly increased by SR-1 as quantified by standard hematopoietic colony-forming assays. Using a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-engineered hESC-RUNX1c-tdTomato reporter cell line with AHR deletion, we further demonstrate a marked enhancement of hematopoietic differentiation relative to wild-type hESCs. We also evaluated whether AHR antagonism could promote innate lymphoid cell differentiation from hESCs. SR-1 increased conventional natural killer (cNK) cell differentiation, whereas TCDD treatment blocked cNK development and supported group 3 innate lymphoid cell (ILC3) differentiation. Collectively, these results demonstrate that AHR regulates early human hematolymphoid cell development and may be targeted to enhance production of specific cell populations derived from human pluripotent stem cells.
Asunto(s)
Hematopoyesis , Células Madre Pluripotentes/citología , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , Diferenciación Celular , Células Cultivadas , Células Madre Embrionarias/citología , Células Madre Hematopoyéticas/citología , Humanos , Subgrupos Linfocitarios/citología , Receptores de Hidrocarburo de Aril/agonistasRESUMEN
Human pluripotent stem cells (PSCs) provide a promising resource to produce immune cells for adoptive cellular immunotherapy to better treat and potentially cure otherwise lethal cancers. Cytotoxic T cells and natural killer (NK) cells can now be routinely produced from human PSCs. These PSC-derived lymphocytes have phenotype and function similar to primary lymphocytes isolated from peripheral blood. PSC-derived T and NK cells have advantages compared with primary immune cells, as they can be precisely engineered to introduce improved anti-tumor activity and produced in essentially unlimited numbers. Stem Cells 2018;36:134-145.
Asunto(s)
Inmunoterapia/métodos , Células Madre Pluripotentes/citología , Humanos , Células Asesinas Naturales/citología , Células Asesinas Naturales/fisiología , Linfocitos/citología , Linfocitos/fisiología , Células Madre Pluripotentes/fisiología , Linfocitos T Citotóxicos/citología , Linfocitos T Citotóxicos/fisiologíaRESUMEN
Endothelial-to-hematopoietic transition (EHT) is an important stage in definitive hematopoietic development. However, the genetic mechanisms underlying human EHT remain poorly characterized. We performed single cell RNA-seq using 55 hemogenic endothelial cells (HECs: CD31+ CD144+ CD41- CD43- CD45- CD73- RUNX1c+ ), 47 vascular endothelial cells without hematopoietic potential (non-HE: CD31+ CD144+ CD41- CD43- CD45- CD73- RUNX1c- ), and 35 hematopoietic progenitor cells (HPCs: CD34+ CD43+ RUNX1c+ ) derived from human embryonic stem cells (hESCs). HE and HP were enriched in genes implicated in hemogenic endothelial transcriptional networks, such as ERG, GATA2, and FLI. We found transcriptional overlap between individual HECs and HPCs; however, these populations were distinct from non-HE. Further analysis revealed novel biomarkers for human HEC/HPCs, including TIMP3, ESAM, RHOJ, and DLL4. Collectively, we demonstrate that hESC-derived HE and HP share a common developmental pathway, while non-HE are more heterogeneous and transcriptionally distinct. Our findings provide a novel strategy to test new genetic targets and optimize the production of definitive hematopoietic cells from human pluripotent stem cells. Stem Cells 2018;36:206-217.