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Neocortical layer 1 (L1) is a site of convergence between pyramidal-neuron dendrites and feedback axons where local inhibitory signaling can profoundly shape cortical processing. Evolutionary expansion of human neocortex is marked by distinctive pyramidal neurons with extensive L1 branching, but whether L1 interneurons are similarly diverse is underexplored. Using Patch-seq recordings from human neurosurgical tissue, we identified four transcriptomic subclasses with mouse L1 homologs, along with distinct subtypes and types unmatched in mouse L1. Subclass and subtype comparisons showed stronger transcriptomic differences in human L1 and were correlated with strong morphoelectric variability along dimensions distinct from mouse L1 variability. Accompanied by greater layer thickness and other cytoarchitecture changes, these findings suggest that L1 has diverged in evolution, reflecting the demands of regulating the expanded human neocortical circuit.
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Neocórtex , Animais , Humanos , Camundongos , Axônios/metabolismo , Interneurônios/metabolismo , Neocórtex/citologia , Neocórtex/metabolismo , Células Piramidais/metabolismo , TranscriptomaRESUMO
Human cortex transcriptomic studies have revealed a hierarchical organization of γ-aminobutyric acid-producing (GABAergic) neurons from subclasses to a high diversity of more granular types. Rapid GABAergic neuron viral genetic labeling plus Patch-seq (patch-clamp electrophysiology plus single-cell RNA sequencing) sampling in human brain slices was used to reliably target and analyze GABAergic neuron subclasses and individual transcriptomic types. This characterization elucidated transitions between PVALB and SST subclasses, revealed morphological heterogeneity within an abundant transcriptomic type, identified multiple spatially distinct types of the primate-specialized double bouquet cells (DBCs), and shed light on cellular differences between homologous mouse and human neocortical GABAergic neuron types. These results highlight the importance of multimodal phenotypic characterization for refinement of emerging transcriptomic cell type taxonomies and for understanding conserved and specialized cellular properties of human brain cell types.
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Neurônios GABAérgicos , Interneurônios , Neocórtex , Animais , Humanos , Camundongos , Fenômenos Eletrofisiológicos , Neurônios GABAérgicos/metabolismo , Ácido gama-Aminobutírico/metabolismo , Interneurônios/metabolismo , Neocórtex/citologia , Neocórtex/metabolismo , Técnicas de Patch-ClampRESUMO
As centre of energy production and key regulators of metabolic and cellular signaling pathways, the integrity of mitochondria is essential for mesenchymal stem cell function in tissue regeneration. Alterations in the size, shape and structural organization of mitochondria are correlated with the physiological state of the cell and its environment and could be used as diagnostic biomarkers. Therefore, high-throughput experimental and computational techniques are crucial to ensure adequate correlations between mitochondrial function and disease phenotypes. The emerge of microfluidic technologies can address the shortcomings of traditional methods to determine mitochondrial dimensions for diagnostic and therapeutic use. This review discusses optical detection methods compatible with microfluidics to measure mitochondrial dynamics and their potential for clinical stem cell research targeting mitochondrial dysfunction.
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OBJECTIVE: An estimated 5.5 million people in England have high-impact chronic pain, which is severe pain associated with significant disability. Current models of healthcare often fail to address their broad range of symptoms and address their complex non-medical needs. METHODS: A pilot project was designed with the aim of improving the quality of care and addressing unmet needs of patients high-impact musculoskeletal (MSK) pain through providing a personalised approach to their pain and wider psychosocial needs. The model comprised a longer initial appointment with a general practitioner, a later follow-appointment, and support of the multidisciplinary team (MDT) (informally and through a formal MDT meeting) with both primary care clinicians and specialists based in secondary care. RESULTS: Forty six patients were seen using this model, with prominent themes of consultations including self-management, social needs, mental health and understanding their diagnosis. Evaluation of the pilot demonstrated improvements in MSK and non-MSK symptoms, together with improved patient confidence in self-management and knowledge and understanding of their condition. Multidisciplinary working proved to be invaluable in addressing patients' wider needs but also upskilling and supporting primary care clinicians. Primary care staff also found it to be a satisfying way to care for patients, and developed increasing skills and confidence in supporting patients with chronic pain. CONCLUSION: This model of care appears to be an effective way to help primary care teams to provide more holistic personalised care to a group of patients who are highly complex and so often forgotten.
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Dor Crônica , Fibromialgia , Dor Musculoesquelética , Humanos , Fibromialgia/terapia , Atenção Secundária à Saúde , Dor Crônica/terapia , Projetos Piloto , Dor Musculoesquelética/terapia , Atenção Primária à SaúdeRESUMO
Owing to their self-renewal and multi-lineage differentiation capability, mesenchymal stem cells (MSCs) hold enormous potential in regenerative medicine. A prerequisite for a successful MSC therapy is the rigorous investigation of their function after in vitro cultivation. Damages introduced to mitochondria during cultivation adversely affect MSCs function and can determine their fate. While it has been shown that microtubules and vimentin intermediate filaments are important for mitochondrial dynamics and active mitochondrial transport within the cytoplasm of MSCs, the role of filamentous actin in this process has not been fully understood yet. To gain a deeper understanding of the interdependence between mitochondrial function and the cytoskeleton, we applied cytochalasin B to disturb the filamentous actin-based cytoskeleton of MSCs. In this study we combined conventional functional assays with a state-of-the-art oxygen sensor-integrated microfluidic device to investigate mitochondrial function. We demonstrated that cytochalasin B treatment at a dose of 16 µM led to a decrease in cell viability with high mitochondrial membrane potential, increased oxygen consumption rate, disturbed fusion and fission balance, nuclear extrusion and perinuclear accumulation of mitochondria. Treatment of MSCs for 48 h ultimately led to nuclear fragmentation, and activation of the intrinsic pathway of apoptotic cell death. Importantly, we could show that mitochondrial function of MSCs can efficiently recover from the damage to the filamentous actin-based cytoskeleton over a period of 24 h. As a result of our study, a causative connection between the filamentous actin-based cytoskeleton and mitochondrial dynamics was demonstrated.
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Células-Tronco Mesenquimais , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Células Cultivadas , Citocalasina B/metabolismo , Citocalasina B/farmacologia , Citoesqueleto/metabolismo , Células-Tronco Mesenquimais/metabolismo , Microtúbulos/metabolismo , MitocôndriasRESUMO
AIMS/HYPOTHESIS: Glucagon-like peptide 1 (GLP-1) receptors are expressed by pancreatic beta cells and GLP-1 receptor signalling promotes insulin secretion. GLP-1 receptor agonists have neural effects and are therapeutically promising for mild cognitive impairment and Alzheimer's disease. Our previous results showed that insulin is released by neurogliaform neurons in the cerebral cortex, but the expression of GLP-1 receptors on insulin-producing neocortical neurons has not been tested. In this study, we aimed to determine whether GLP-1 receptors are present in insulin-containing neurons. METHODS: We harvested the cytoplasm of electrophysiologically and anatomically identified neurogliaform interneurons during patch-clamp recordings performed in slices of rat neocortex. Using single-cell digital PCR, we determined copy numbers of Glp1r mRNA and other key genes in neurogliaform cells harvested in conditions corresponding to hypoglycaemia (0.5 mmol/l glucose) and hyperglycaemia (10 mmol/l glucose). In addition, we performed whole-cell patch-clamp recordings on neurogliaform cells to test the effects of GLP-1 receptor agonists for functional validation of single-cell digital PCR results. RESULTS: Single-cell digital PCR revealed GLP-1 receptor expression in neurogliaform cells and showed that copy numbers of mRNA of the Glp1r gene in hyperglycaemia exceeded those in hypoglycaemia by 9.6 times (p < 0.008). Moreover, single-cell digital PCR confirmed co-expression of Glp1r and Ins2 mRNA in neurogliaform cells. Functional expression of GLP-1 receptors was confirmed with whole-cell patch-clamp electrophysiology, showing a reversible effect of GLP-1 on neurogliaform cells. This effect was prevented by pre-treatment with the GLP-1 receptor-specific antagonist exendin-3(9-39) and was absent in hypoglycaemia. In addition, single-cell digital PCR of neurogliaform cells revealed that the expression of transcription factors (Pdx1, Isl1, Mafb) are important in beta cell development. CONCLUSIONS/INTERPRETATION: Our results provide evidence for the functional expression of GLP-1 receptors in neurons known to release insulin in the cerebral cortex. Hyperglycaemia increases the expression of GLP-1 receptors in neurogliaform cells, suggesting that endogenous incretins and therapeutic GLP-1 receptor agonists might have effects on these neurons, similar to those in pancreatic beta cells.
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Córtex Cerebral/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Insulina/metabolismo , Interneurônios/metabolismo , Animais , Citoplasma/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Hiperglicemia/metabolismo , Hipoglicemia/metabolismo , Masculino , Neocórtex/metabolismo , Ratos , Ratos Wistar , Transdução de SinaisRESUMO
Organ-on-a-chip technology has gained great interest in recent years given its ability to control the spatio-temporal microenvironments of cells and tissues precisely. While physical parameters of the respective niche such as microchannel network sizes, geometric features, flow rates, and shear forces, as well as oxygen tension and concentration gradients, have been optimized for stem cell cultures, little has been done to improve cell-matrix interactions in microphysiological systems. Specifically, detailed research on the effect of matrix elasticity and extracellular matrix (ECM) nanotopography on stem cell differentiation are still in its infancy, an aspect that is known to alter a stem cell's fate. Although a wide range of hydrogels such as gelatin, collagen, fibrin, and others are available for stem cell chip cultivations, only a limited number of elasticities are generally employed. Matrix elasticity and the corresponding nanotopography are key factors that guide stem cell differentiation. Given this, we investigated the addition of gold nanowires into hydrogels to create a tunable biointerface that could be readily integrated into any organ-on-a-chip and cell chip system. In the presented work, we investigated the matrix elasticity (Young's modulus, stiffness, adhesive force, and roughness) and nanotopography of gold nanowire loaded onto fibrin hydrogels using the bio-AFM (atomic force microscopy) method. Additionally, we investigated the capacity of human amniotic mesenchymal stem cells (hAMSCs) to differentiate into osteo- and chondrogenic lineages. Our results demonstrated that nanogold structured-hydrogels promoted differentiation of hAMSCs as shown by a significant increase in Collagen I and II production. Additionally, there was enhanced calcium mineralization activity and proteoglycans formation after a cultivation period of two weeks within microfluidic devices.
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We describe convergent evidence from transcriptomics, morphology, and physiology for a specialized GABAergic neuron subtype in human cortex. Using unbiased single-nucleus RNA sequencing, we identify ten GABAergic interneuron subtypes with combinatorial gene signatures in human cortical layer 1 and characterize a group of human interneurons with anatomical features never described in rodents, having large 'rosehip'-like axonal boutons and compact arborization. These rosehip cells show an immunohistochemical profile (GAD1+CCK+, CNR1-SST-CALB2-PVALB-) matching a single transcriptomically defined cell type whose specific molecular marker signature is not seen in mouse cortex. Rosehip cells in layer 1 make homotypic gap junctions, predominantly target apical dendritic shafts of layer 3 pyramidal neurons, and inhibit backpropagating pyramidal action potentials in microdomains of the dendritic tuft. These cells are therefore positioned for potent local control of distal dendritic computation in cortical pyramidal neurons.
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Córtex Cerebral/metabolismo , Córtex Cerebral/ultraestrutura , Neurônios GABAérgicos/metabolismo , Neurônios GABAérgicos/ultraestrutura , Transcriptoma , Adulto , Idoso , Axônios/ultraestrutura , Espinhas Dendríticas/metabolismo , Espinhas Dendríticas/ultraestrutura , Junções Comunicantes/metabolismo , Junções Comunicantes/ultraestrutura , Biblioteca Gênica , Humanos , Masculino , Reação em Cadeia da Polimerase , Terminações Pré-Sinápticas/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , Células Piramidais/metabolismo , Células Piramidais/ultraestrutura , RNA/análise , RNA/genética , Análise de Sequência de RNARESUMO
Outcome expectancy could mediate the psychological effects of exercise-related interventions, which implies that part of the psychological benefits of physical activity could be ascribed to placebo effects. In this framed field-experiment, 89 healthy participants were studied in three groups, (1) breathing-primed (deep-breathing with an exercise-related expectancy), (2) breathing-unprimed (deep-breathing with no exercise-related expectancy), and (3) control (no intervention). Deep-breathing lasted for three minutes. Before and after deep-breathing, or sitting quietly in the control group, participants completed two questionnaires assessing their positive- and negative affect (NA) and subjective well-being (WB). In contrast to the control group, both the breathing-primed and breathing-unprimed groups showed decreased NA and increased subjective WB. The breathing-primed group reported larger changes in WB than the breathing-unprimed group, in addition to also exhibiting significant increases in positive affect. These findings support the hypothesis of the work that expectations mediate the psychological effects of deep-breathing beyond the intervention's specific effects. Therefore, future research should control for expectations related to an intervention when gauging psychological changes.
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Atitude Frente a Saúde , Exercícios Respiratórios/psicologia , Adolescente , Exercício Físico/psicologia , Feminino , Voluntários Saudáveis , Humanos , Masculino , Efeito Placebo , Adulto JovemRESUMO
Functional and molecular changes associated with pathophysiological conditions are relatively easily detected based on tissue samples collected from patients. Population specific cellular responses to disease might remain undiscovered in samples taken from organs formed by a multitude of cell types. This is particularly apparent in the human cerebral cortex composed of a yet undefined number of neuron types with a potentially different involvement in disease processes. We combined cellular electrophysiology, anatomy and single cell digital PCR in human neurons identified in situ for the first time to assess mRNA expression and corresponding functional changes in response to edema and increased intracranial pressure. In single pyramidal cells, mRNA copy numbers of AQP1, AQP3, HMOX1, KCNN4, SCN3B and SOD2 increased, while CACNA1B, CRH decreased in edema. In addition, single pyramidal cells increased the copy number of AQP1, HTR5A and KCNS1 mRNAs in response to increased intracranial pressure. In contrast to pyramidal cells, AQP1, HMOX1and KCNN4 remained unchanged in single cell digital PCR performed on fast spiking cells in edema. Corroborating single cell digital PCR results, pharmacological and immunohistochemical results also suggested the presence of KCNN4 encoding the α-subunit of KCa3.1 channels in edema on pyramidal cells, but not on interneurons. We measured the frequency of spontaneous EPSPs on pyramidal cells in both pathophysiological conditions and on fast spiking interneurons in edema and found a significant decrease in each case, which was accompanied by an increase in input resistances on both cell types and by a drop in dendritic spine density on pyramidal cells consistent with a loss of excitatory synapses. Our results identify anatomical and/or physiological changes in human pyramidal and fast spiking cells in edema and increased intracranial pressure revealing cell type specific quantitative changes in gene expression. Some of the edema/increased intracranial pressure modulated and single human pyramidal cell verified gene products identified here might be considered as novel pharmacological targets in cell type specific neuroprotection.
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Edema Encefálico/metabolismo , Hipertensão Intracraniana/metabolismo , Neocórtex/metabolismo , Neurônios/metabolismo , Adulto , Edema Encefálico/patologia , Edema Encefálico/cirurgia , Feminino , Regulação da Expressão Gênica , Substância Cinzenta/metabolismo , Substância Cinzenta/patologia , Substância Cinzenta/cirurgia , Humanos , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/metabolismo , Hipertensão Intracraniana/patologia , Hipertensão Intracraniana/cirurgia , Pressão Intracraniana/fisiologia , Masculino , Potenciais da Membrana/fisiologia , Pessoa de Meia-Idade , Neocórtex/patologia , Neocórtex/cirurgia , Neurônios/patologia , RNA Mensageiro/metabolismo , Técnicas de Cultura de TecidosRESUMO
HTLV-1-associated myelopathy (HAM; HTLV-1 is human T-lymphotropic virus type 1) is a chronic debilitating neuroinflammatory disease with a predilection for the thoracic cord. Tissue damage is attributed to the cellular immune response to HTLV-1-infected lymphocytes. The brains of HTLV-1-infected patients, with and without HAM but no clinical evidence of brain involvement, were examined using a specific 18-kDa translocator protein ligand, 11C-PBR28, and T1-weighted and diffusion-weighted MRI. METHODS: Five subjects with HAM and 2 HTLV-1 asymptomatic carriers were studied. All underwent clinical neurologic assessment including cognitive function and objective measures of gait, quantification of HTLV-1 proviral load in peripheral blood mononuclear cells, and human leukocyte antigen-antigen D related expression on circulating CD8+ lymphocytes. 11C-PBR28 PET and MRI were performed on the same day. 11C-PBR28 PET total volume of distribution and distribution volume ratio (DVR) were estimated using 2-tissue-compartment modeling. MRI data were processed using tools from the FMRIB Software Library to estimate mean diffusivity (MD) and gray matter (GM) fraction changes. The results were compared with data from age-matched healthy volunteers. RESULTS: Across the whole brain, the total volume of distribution for the subjects with HAM (5.44 ± 0.84) was significantly greater than that of asymptomatic carriers (3.44 ± 0.80). The DVR of the thalamus in patients with severe and moderate HAM was higher than that in the healthy volunteers, suggesting increased translocator protein binding (z > 4.72). Subjects with more severe myelopathy and with high DR expression on CD8+ lymphocytes had increased DVR and MD (near-significant correlation found for the right thalamus MD: P = 0.06). On the T1-weighted MRI scans, the GM fraction of the brain stem was reduced in all HTLV-1-infected patients compared with controls (P < 0.001), whereas the thalamus GM fraction was decreased in patients with HAM and correlated with the disease severity. There was no correlation between neurocognitive function and these markers of central nervous system inflammation. CONCLUSION: This pilot study suggests that some patients with HAM have asymptomatic inflammation in the brain, which can be detected and monitored by 11C-PBR28 PET together with structural and diffusion-weighted MRI.
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Imagem de Difusão por Ressonância Magnética , Encefalite/complicações , Encefalite/diagnóstico por imagem , Imagem Multimodal , Paraparesia Espástica Tropical/complicações , Tomografia por Emissão de Pósitrons , Pirimidinas , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos PilotoRESUMO
OBJECTIVE: Men who have sex with men (MSM) are the key population most affected by HIV in Europe. We performed the first European multicenter, simple-randomized parallel-group study to test the effectiveness of a theory-guided computer-assisted intervention to improve safer sex among HIV-positive men who have sex with men. METHODS: Between February, 2011 and February, 2013, 112 participants were enrolled in 8 different European HIV-care settings. Intervention participants received 3 individual counseling sessions facilitated by trained service providers using computer-assisted tools. The control-group received sexual health advice delivered as part of regular HIV care. Outcome behavior (self-reported condom use at last intercourse; combined HIV transmission risk score), its influencing factors, and mediating variables were assessed at baseline, and at 3 and 6 months follow-up. Mixed effects models were used to compare primary outcomes (condom use at last intercourse, HIV transmission risk score), and mediation analysis to explore intervention effects. RESULTS: Condom use at last intercourse increased more among intervention than control participants at 3 months follow-up (odds ratio of 3.83; P = 0.03), but not significantly at 6 months follow-up. Intervention participants reported a lower transmission risk at 3 months follow-up than controls (odds ratio compared with baseline of 11.53 and 1.28, respectively; P = 0.008), but this effect became nonsignificant at 6 months. Intervention effects were mediated by the proximal variables, self-efficacy to negotiate condom use and condom attitudes. CONCLUSIONS: This intervention showed short-term effectiveness. The intervention should be replicated in other settings, eventually investigating if booster-counseling sessions would yield a longer lasting effect.
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Instrução por Computador , Infecções por HIV/prevenção & controle , Homossexualidade Masculina , Sexo Seguro , Adulto , Instrução por Computador/métodos , Instrução por Computador/normas , Preservativos/estatística & dados numéricos , Aconselhamento/métodos , Europa (Continente)/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Inquéritos e Questionários , Carga ViralRESUMO
Concentrations of insulin in the brain are severalfold higher than blood plasma levels. Insulin in the brain regulates the metabolism, molecular composition, and cognitive performance of microcircuits and reduces food intake; cerebral insulin levels are altered in diabetes, aging, obesity, and Alzheimer's disease. Released by pancreatic ß cells, insulin passes the blood-brain barrier, but sources of locally released insulin still remain unclear. We find that insulin is strongly expressed in GABAergic neurogliaform cells in the cerebral cortex of the rat detected by single-cell digital PCR. Focal application of glucose or glibenclamide to neurogliaform cells mimics the excitation suppressing effect of external insulin on local microcircuits via insulin receptors. Thus, neurogliaform cells might link GABAergic and insulinergic action in cortical microcircuits.
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Insulina/metabolismo , Neocórtex/citologia , Neocórtex/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Animais , Potenciais Pós-Sinápticos Excitadores/fisiologia , Secreção de Insulina , Masculino , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase , Radioimunoensaio , Ratos , Ratos Wistar , Ácido gama-Aminobutírico/metabolismoRESUMO
Whole-cell patch-clamp recording enables detection of electrophysiological signals from single neurons as well as harvesting of perisomatic RNA through the patch pipette for subsequent gene expression analysis. Amplification and profiling of RNA with traditional quantitative real-time PCR (qRT-PCR) do not provide exact quantitation due to experimental variation caused by the limited amount of nucleic acid in a single cell. Here we describe a protocol for quantifying mRNA or miRNA expression in individual neurons after patch-clamp recording using high-density nanocapillary digital PCR (dPCR). Expression of a known cell-type dependent marker gene (gabrd), as well as oxidative-stress related induction of hspb1 and hmox1 expression, was quantified in individual neurogliaform and pyramidal cells, respectively. The miRNA mir-132, which plays a role in neurodevelopment, was found to be equally expressed in three different types of neurons. The accuracy and sensitivity of this method were further validated using synthetic spike-in templates and by detecting genes with very low levels of expression.
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Neurônios/fisiologia , Técnicas de Patch-Clamp/métodos , Reação em Cadeia da Polimerase/métodos , RNA Mensageiro/metabolismo , Análise de Célula Única/métodos , Animais , Células Cultivadas , Perfilação da Expressão Gênica/métodos , Masculino , MicroRNAs/análise , MicroRNAs/genética , MicroRNAs/metabolismo , Neurônios/química , Neurônios/citologia , Neurônios/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/genética , Ratos , Ratos Wistar , Córtex Somatossensorial/citologiaRESUMO
Mannose-binding lectin (MBL) is a major, soluble, pattern-recognition molecule and an important component of the innate host defense. The role of MBL in inflammatory bowel diseases (IBDs) is controversial. We determined the prevalence of MBL deficiency in a Hungarian IBD patients' cohort, and whether it is associated with the antimicrobial antibody formation or particular clinical manifestations. Nine hundred ninety IBD patients and 225 healthy subjects were investigated. Sera were assayed for MBL and a panel of antimicrobial antibodies (anti-OMP, anti-Saccharomyces cerevisiae antibodies, and antiglycans) by ELISA. TLR4 and NOD2/CARD15 variants were tested by polymerase chain reaction/restriction fragment length polymorphism. Median MBL level was not significantly different between IBDs (Crohn's disease [CD]: 929; ulcerative colitis [UC]: 810 ng/ml) and the control group (1027 ng/ml), as well as the prevalence of absolute MBL deficiency (<100 ng/ml) (CD: 15.0%, UC: 18.4%, controls: 15.6%). The presence of a low MBL level (<500 ng/ml) was not associated with any of the examined serologic markers, or their combinations. In addition, there was no association with the clinical presentation, disease course, or response to treatment. TLR4 variant genotype was more common in CD patients without MBL deficiency (11% vs. 1.7%, OR: 7.29, 95% CI: 1.08-53.9, p = 0.02). We failed to confirm any association between MBL deficiency and serologic marker positivity. MBL deficiency was not predictive for clinical phenotype or disease activity in IBDs.
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Doenças Inflamatórias Intestinais/genética , Lectina de Ligação a Manose/genética , Proteínas de Transporte da Membrana Mitocondrial/imunologia , Proteínas de Saccharomyces cerevisiae/imunologia , Receptor 4 Toll-Like/genética , Adulto , Anticorpos Antifúngicos/sangue , Biomarcadores/sangue , Análise Mutacional de DNA , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Humanos , Hungria , Doenças Inflamatórias Intestinais/sangue , Doenças Inflamatórias Intestinais/epidemiologia , Doenças Inflamatórias Intestinais/fisiopatologia , Masculino , Lectina de Ligação a Manose/sangue , Pessoa de Meia-Idade , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Proteína Adaptadora de Sinalização NOD2/genética , Polimorfismo GenéticoRESUMO
The pathogenesis of multiple sclerosis (MS), a devastating neuroinflammatory disorder of the central nervous system, has been presumed to involve the possible importance of the receptor for advanced glycation end products (RAGE). The aim of this study was to investigate the relevance of the genetic polymorphisms of RAGE in MS patients. A total of 168 patients with MS were enrolled; 136 healthy blood donors served as controls. The -374 T/ A, -479 T/C, and the G82S polymorphisms of RAGE were determined by restriction fragment length polymorphism (RFLP). There was a significant difference in RAGE -374 T/A genotype distribution between the controls and the MS patients. The AA homozygote variants were detected in 8% of the patients with MS, as compared with 19% of healthy controls (OR=2.75; 95% CI=1.319-5.733, p = 0.007). No differences were observed between the MS patients and the controls, concerning the frequencies of the -479 T/C and G82S genotypes of the RAGE. Our results revealed an association between the -374 T/A polymorphism of the RAGE promoter and MS. The genetic variant -374 AA (which has previously been shown to exert significant effects on transcriptional activity) can be considered a preventive factor as regards the occurrence of MS. Our findings support the view that RAGE plays a role in the development of MS.
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Predisposição Genética para Doença/genética , Esclerose Múltipla/genética , Polimorfismo Genético/genética , Receptor para Produtos Finais de Glicação Avançada/genética , Adulto , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/fisiopatologia , Análise Mutacional de DNA , Feminino , Frequência do Gene/genética , Marcadores Genéticos/genética , Testes Genéticos , Variação Genética/genética , Genótipo , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla/metabolismo , Esclerose Múltipla/fisiopatologia , Fatores de Crescimento Neural/genética , Ligação Proteica/genética , Subunidade beta da Proteína Ligante de Cálcio S100 , Proteínas S100/genéticaRESUMO
OBJECTIVE: Helicobacter pylori-induced gastric inflammation is dependent on the persistence of the microorganism in the gastric epithelium. Modulation of the host epithelial antimicrobial responses may be a critical determinant in H. pylori-induced gastritis. Human beta-defensins (hBDs) are important components of the host defence at mucosal surfaces. The aim of the present study was to investigate the relevance of three single nucleotide polymorphisms (SNPs) of the human beta defensin-1 (hBD-1) gene in H. pylori-induced gastritis and to assess the mRNA expression of hBD-1 in H. pylori-infected AGS cells. MATERIAL AND METHODS: Three SNPs of the beta defensin DEFB1 gene, DEFB1 G-20A (rs11362), DEFB1 C-44G (rs1800972) and DEFB1 G-52A (rs1799946), were genotyped either by Custom TaqMan SNP genotyping assays or by restriction fragment length polymorphism (RFLP) in 150 patients with chronic active gastritis; 100 serologically H. pylori-positive subjects without gastric or duodenal symptoms served as controls. hBD-1 mRNA expression in AGS cells was measured by quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Significant differences in frequencies of the GA and AA genotypes of G-52A SNPs were observed between patients with chronic active gastritis and healthy controls. The maximum level of hBD-1 mRNA expression in AGS cells was observed at 24 h after infection with H. pylori, this not being dependent on the presence of the cag pathogenicity island (PAI). CONCLUSIONS: The results of these genetic and in vitro experiments suggest that not only the inducible, but also the constitutive form of hBD may be important in the pathogenesis of H. pylori-induced gastritis.
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Gastrite/imunologia , Gastrite/microbiologia , Infecções por Helicobacter/imunologia , Helicobacter pylori/imunologia , beta-Defensinas/genética , Adulto , Estudos de Casos e Controles , Distribuição de Qui-Quadrado , Feminino , Gastrite/metabolismo , Genótipo , Infecções por Helicobacter/genética , Humanos , Imunidade nas Mucosas , Masculino , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , beta-Defensinas/fisiologiaRESUMO
OBJECTIVE: It has been suggested that deficient defensin expression is associated with the chronic inflammation of Crohn's disease. The regional localization of Crohn's disease, ileal or colonic disease can be linked to different defensin profiles. As constitutive beta-defensin 1 has a colonic expression, we considered it of interest to investigate single nucleotide polymorphisms (SNPs) of the beta-defensin 1 gene (DEFB1) in Crohn's disease. MATERIAL AND METHODS: Three SNPs of the DEFB1 gene, DEFB1 G-20A (rs11362), DEFB1 C-44G (rs1800972) and DEFB1 G-52A (rs1799946), were genotyped either by Custom TaqMan SNP genotyping assays or by restriction fragment length polymorphisms (RFLP) in 190 patients with Crohn's disease and 95 Hungarian controls. RESULTS: It was found that the G-20A and C-44G SNPs had a strong association with the colonic and ileocolonic localizations of the disease, respectively, but no association was detected for the ileal localization. A significantly higher frequency of the GA genotype of G-20A was observed among patients with colonic localization (60%) as compared with healthy controls (39%), with an odds ratio (OR) of 2.39. The GG genotype of C-44G SNP, which is regarded as a protective genotype, was much less frequent (4%) among patients than among controls (12%), OR 3.367. CONCLUSIONS: These results indicate that genetic variations in the DEFB1 gene encoding constitutive human beta-defensin 1 may be associated with the risk for Crohn's disease and may determine disease phenotype, e.g. colonic localization.
Assuntos
Doença de Crohn/genética , DNA/genética , Polimorfismo de Fragmento de Restrição , beta-Defensinas/genética , Adulto , Doença de Crohn/epidemiologia , Doença de Crohn/metabolismo , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Hungria/epidemiologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Desequilíbrio de Ligação , Masculino , Prevalência , beta-Defensinas/metabolismoRESUMO
INTRODUCTION: Since the advent of Highly Active Anti-Retroviral Therapy (HAART), men with HIV experience good quality of life and expect to have normal sexual function. However, it appears that men infected with HIV commonly complain of sexual problems. There is evidence that men on HAART develop low sexual desire that is associated with raised estradiol levels. It has been postulated that abnormal metabolism seen in this group of men increases the aromatization of testosterone to estradiol. We hypothesized that letrozole, an aromatase inhibitor that inhibits the conversion of testosterone to estradiol, would be beneficial in these men. AIM: The aim of this study was to compare the effects of testosterone vs. an aromatase inhibitor, letrazole, in HIV-infected men with raised estradiol and low sexual desire. METHODS: Thirteen men who have sex with men on HAART with low sexual desire as well as raised estradiol levels (>120 pmol/L) were randomly allocated to receive either parenteral testosterone (Sustanon 250 intramuscular injection) (N = 6) or letrozole 2.5 mg orally daily (N = 7) for 6 weeks. MAIN OUTCOME MEASURES: Sex steroid hormone assays, sex hormone-binding globulin, virological, hematological, and biochemical parameters were measured before and after treatment. Each subject was given the Spector Sexual Desire Inventory and the Depression/Anxiety Stress Scale before and immediately after treatment. Subjects were also asked to estimate the number of actual sexual acts before and after treatment. Results. Inventory data showed a rise in dyadic desire in both treatment arms. Mean actual sexual acts rose from 0.33 to 1.5 in the testosterone group and from 0.43 to 1.29 for the letrozole group. Luteinizing hormone increased in seven of seven men on letrozole. Serum testosterone increased in seven of seven men on letrozole. There were no adverse events from either medication. CONCLUSION: Letrozole may be useful in the management of men on HAART who have low sexual desire.