RESUMO
Dedicator of cytokinesis 8 (DOCK8) immunodeficiency syndrome is characterized by a failure of the germinal center response, a process involving the proliferation and positive selection of antigen-specific B cells. Here, we describe how DOCK8-deficient B cells are blocked at a light-zone checkpoint in the germinal centers of immunized mice, where they are unable to respond to T cell-dependent survival and selection signals and consequently differentiate into plasma cells or memory B cells. Although DOCK8-deficient B cells can acquire and present antigen to initiate activation of cognate T cells, integrin up-regulation, B cell-T cell conjugate formation, and costimulation are insufficient for sustained B cell and T cell activation when antigen availability is limited. Our findings provide an explanation for the failure of the humoral response in DOCK8 immunodeficiency syndrome and insight into how the level of available antigen modulates B cell-T cell cross-talk to fine-tune humoral immune responses and immunological memory.
Assuntos
Linfócitos B , Fatores de Troca do Nucleotídeo Guanina , Camundongos Endogâmicos C57BL , Linfócitos T , Animais , Fatores de Troca do Nucleotídeo Guanina/imunologia , Fatores de Troca do Nucleotídeo Guanina/deficiência , Fatores de Troca do Nucleotídeo Guanina/genética , Linfócitos B/imunologia , Camundongos , Linfócitos T/imunologia , Ativação Linfocitária/imunologia , Camundongos Knockout , Antígenos/imunologia , Centro Germinativo/imunologiaRESUMO
Spatial transcriptomics measures in situ gene expression at millions of locations within a tissue1, hitherto with some trade-off between transcriptome depth, spatial resolution and sample size2. Although integration of image-based segmentation has enabled impactful work in this context, it is limited by imaging quality and tissue heterogeneity. By contrast, recent array-based technologies offer the ability to measure the entire transcriptome at subcellular resolution across large samples3-6. Presently, there exist no approaches for cell type identification that directly leverage this information to annotate individual cells. Here we propose a multiscale approach to automatically classify cell types at this subcellular level, using both transcriptomic information and spatial context. We showcase this on both targeted and whole-transcriptome spatial platforms, improving cell classification and morphology for human kidney tissue and pinpointing individual sparsely distributed renal mouse immune cells without reliance on image data. By integrating these predictions into a topological pipeline based on multiparameter persistent homology7-9, we identify cell spatial relationships characteristic of a mouse model of lupus nephritis, which we validate experimentally by immunofluorescence. The proposed framework readily generalizes to new platforms, providing a comprehensive pipeline bridging different levels of biological organization from genes through to tissues.
Assuntos
Células , Perfilação da Expressão Gênica , Espaço Intracelular , Rim , Transcriptoma , Animais , Feminino , Humanos , Camundongos , Células/classificação , Células/metabolismo , Modelos Animais de Doenças , Imunofluorescência , Perfilação da Expressão Gênica/métodos , Rim/citologia , Rim/imunologia , Rim/metabolismo , Rim/patologia , Nefrite Lúpica/genética , Nefrite Lúpica/imunologia , Nefrite Lúpica/metabolismo , Nefrite Lúpica/patologia , Reprodutibilidade dos Testes , Espaço Intracelular/genética , Espaço Intracelular/metabolismoRESUMO
Motivation: Pooled designs for single-cell RNA sequencing, where many cells from distinct samples are processed jointly, offer increased throughput and reduced batch variation. This study describes expression-aware demultiplexing (EAD), a computational method that employs differential co-expression patterns between individuals to demultiplex pooled samples without any extra experimental steps. Results: We use synthetic sample pools and show that the top interindividual differentially co-expressed genes provide a distinct cluster of cells per individual, significantly enriching the regulation of metabolism. Our application of EAD to samples of six isogenic inbred mice demonstrated that controlling genetic and environmental effects can solve interindividual variations related to metabolic pathways. We utilized 30 samples from both sepsis and healthy individuals in six batches to assess the performance of classification approaches. The results indicate that combining genetic and EAD results can enhance the accuracy of assignments (Min. 0.94, Mean 0.98, Max. 1). The results were enhanced by an average of 1.4% when EAD and barcoding techniques were combined (Min. 1.25%, Median 1.33%, Max. 1.74%). Furthermore, we demonstrate that interindividual differential co-expression analysis within the same cell type can be used to identify cells from the same donor in different activation states. By analysing single-nuclei transcriptome profiles from the brain, we demonstrate that our method can be applied to nonimmune cells. Availability and implementation: EAD workflow is available at https://isarnassiri.github.io/scDIV/ as an R package called scDIV (acronym for single-cell RNA-sequencing data demultiplexing using interindividual variations).
RESUMO
BACKGROUND: Whole genome sequencing is increasingly being used for the diagnosis of patients with rare diseases. However, the diagnostic yields of many studies, particularly those conducted in a healthcare setting, are often disappointingly low, at 25-30%. This is in part because although entire genomes are sequenced, analysis is often confined to in silico gene panels or coding regions of the genome. METHODS: We undertook WGS on a cohort of 122 unrelated rare disease patients and their relatives (300 genomes) who had been pre-screened by gene panels or arrays. Patients were recruited from a broad spectrum of clinical specialties. We applied a bioinformatics pipeline that would allow comprehensive analysis of all variant types. We combined established bioinformatics tools for phenotypic and genomic analysis with our novel algorithms (SVRare, ALTSPLICE and GREEN-DB) to detect and annotate structural, splice site and non-coding variants. RESULTS: Our diagnostic yield was 43/122 cases (35%), although 47/122 cases (39%) were considered solved when considering novel candidate genes with supporting functional data into account. Structural, splice site and deep intronic variants contributed to 20/47 (43%) of our solved cases. Five genes that are novel, or were novel at the time of discovery, were identified, whilst a further three genes are putative novel disease genes with evidence of causality. We identified variants of uncertain significance in a further fourteen candidate genes. The phenotypic spectrum associated with RMND1 was expanded to include polymicrogyria. Two patients with secondary findings in FBN1 and KCNQ1 were confirmed to have previously unidentified Marfan and long QT syndromes, respectively, and were referred for further clinical interventions. Clinical diagnoses were changed in six patients and treatment adjustments made for eight individuals, which for five patients was considered life-saving. CONCLUSIONS: Genome sequencing is increasingly being considered as a first-line genetic test in routine clinical settings and can make a substantial contribution to rapidly identifying a causal aetiology for many patients, shortening their diagnostic odyssey. We have demonstrated that structural, splice site and intronic variants make a significant contribution to diagnostic yield and that comprehensive analysis of the entire genome is essential to maximise the value of clinical genome sequencing.
Assuntos
Variação Genética , Doenças Raras , Humanos , Doenças Raras/diagnóstico , Doenças Raras/genética , Sequenciamento Completo do Genoma , Testes Genéticos , Mutação , Proteínas de Ciclo CelularRESUMO
Prolidase deficiency (PD) is a multisystem disorder caused by mutations in the PEPD gene, which encodes a ubiquitously expressed metallopeptidase essential for the hydrolysis of dipeptides containing C-terminal proline or hydroxyproline. PD typically presents in childhood with developmental delay, skin ulcers, recurrent infections, and, in some patients, autoimmune features that can mimic systemic lupus erythematosus. The basis for the autoimmune association is uncertain, but might be due to self-antigen exposure with tissue damage, or indirectly driven by chronic infection and microbial burden. In this study, we address the question of causation and show that Pepd-null mice have increased antinuclear autoantibodies and raised serum IgA, accompanied by kidney immune complex deposition, consistent with a systemic lupus erythematosus-like disease. These features are associated with an accumulation of CD4 and CD8 effector T cells in the spleen and liver. Pepd deficiency leads to spontaneous T cell activation and proliferation into the effector subset, which is cell intrinsic and independent of Ag receptor specificity or antigenic stimulation. However, an increase in KLRG1+ effector CD8 cells is not observed in mixed chimeras, in which the autoimmune phenotype is also absent. Our findings link autoimmune susceptibility in PD to spontaneous T cell dysfunction, likely to be acting in combination with immune activators that lie outside the hemopoietic system but result from the abnormal metabolism or loss of nonenzymatic prolidase function. This knowledge provides insight into the role of prolidase in the maintenance of self-tolerance and highlights the importance of treatment to control T cell activation.
Assuntos
Lúpus Eritematoso Sistêmico , Deficiência de Prolidase , Animais , Camundongos , Autoimunidade , Ativação Linfocitária , AutoantígenosRESUMO
Peripheral tolerance prevents the initiation of damaging immune responses by autoreactive lymphocytes. While tolerogenic mechanisms are tightly regulated by antigen-dependent and independent signals, downstream pathways are incompletely understood. N-myc downstream-regulated gene 1 (NDRG1), an anti-cancer therapeutic target, has previously been implicated as a CD4+ T cell clonal anergy factor. By RNA-sequencing, we identified Ndrg1 as the third most upregulated gene in anergic, compared to naïve follicular, B cells. Ndrg1 is upregulated by B cell receptor activation (signal one) and suppressed by co-stimulation (signal two), suggesting that NDRG1 may be important in B cell tolerance. However, though Ndrg1-/- mice have a neurological defect mimicking NDRG1-associated Charcot-Marie-Tooth (CMT4d) disease, primary and secondary immune responses were normal. We find that B cell tolerance is maintained, and NDRG1 does not play a role in downstream responses during re-stimulation of in vivo antigen-experienced CD4+ T cells, demonstrating that NDGR1 is functionally redundant for lymphocyte anergy.
Assuntos
Doença de Charcot-Marie-Tooth , Doença de Refsum , Camundongos , Animais , Linfócitos T , Doença de Refsum/genética , Doença de Refsum/metabolismo , Doença de Charcot-Marie-Tooth/genética , Tolerância Imunológica , Ativação LinfocitáriaRESUMO
BACKGROUND: It is well established that decreased kidney function can increase blood pressure (BP), but it is unproven whether moderately elevated BP causes chronic kidney disease (CKD) or glomerular hyperfiltration. METHODS: 311 119 White British UK Biobank participants were included in logistic regression analyses to estimate the odds of CKD (defined as long-term kidney replacement therapy, estimated glomerular filtration rate [eGFR]< 60mL/min/1.73m2, or urinary albumin:creatinine ratio ≥3 mg/mmol) associated with higher genetically predicted BP using genetic risk scores comprising 219 systolic and 223 diastolic BP loci. Analyses estimating associations with clinical categories of eGFR and urinary albumin:creatinine ratio were also conducted, with an eGFR ≥120 mL (min·1.73m2) considered evidence of glomerular hyperfiltration. RESULTS: 21 623 participants had CKD: 7781 with reduced eGFR and 15 500 with albuminuria. 1828 participants had an eGFR ≥120 mL/min/1.73m2. Each genetically predicted 10 mmHg higher systolic BP and 5 mmHg higher diastolic BP were associated with a 37% (95% CI, 1.29-1.45) and 19% (1.14-1.25) higher odds of CKD, respectively. Associations were evident for both the reduced eGFR and albuminuria components of the CKD outcome. The odds of hyperfiltration (versus an eGFR ≥60 and <90 mL/min/1.73m2 were 49% higher (95% CI, 1.21-1.84) for each genetically predicted 10 mmHg higher systolic BP. Associations with CKD and hyperfiltration were similar irrespective of preexisting diabetes, vascular disease, or different levels of adiposity. CONCLUSIONS: In this general population, genetic epidemiological evidence supports a causal role of life-long differences in BP for decreased kidney function, glomerular hyperfiltration, and albuminuria. Physiological autoregulation may not afford complete renal protection against the moderate BP elevations.
Assuntos
Hipertensão , Insuficiência Renal Crônica , Humanos , Albuminúria/epidemiologia , Albuminúria/genética , Pressão Sanguínea/genética , Creatinina/urina , Epidemiologia Molecular , Insuficiência Renal Crônica/epidemiologia , Insuficiência Renal Crônica/genética , Insuficiência Renal Crônica/complicações , Taxa de Filtração Glomerular , Rim , Hipertensão/epidemiologia , Hipertensão/genética , Hipertensão/complicações , AlbuminasRESUMO
BACKGROUND: Variants in genes encoding nuclear pore complex (NPC) proteins are a newly identified cause of paediatric steroid-resistant nephrotic syndrome (SRNS). Recent reports describing NUP93 variants suggest these could be a significant cause of paediatric onset SRNS. We report NUP93 cases in the UK and demonstrate in vivo functional effects of Nup93 depletion in a fly (Drosophila melanogaster) nephrocyte model. METHODS: Three hundred thirty-seven paediatric SRNS patients from the National cohort of patients with Nephrotic Syndrome (NephroS) were whole exome and/or whole genome sequenced. Patients were screened for over 70 genes known to be associated with Nephrotic Syndrome (NS). D. melanogaster Nup93 knockdown was achieved by RNA interference using nephrocyte-restricted drivers. RESULTS: Six novel homozygous and compound heterozygous NUP93 variants were detected in 3 sporadic and 2 familial paediatric onset SRNS characterised histologically by focal segmental glomerulosclerosis (FSGS) and progressing to kidney failure by 12 months from clinical diagnosis. Silencing of the two orthologs of human NUP93 expressed in D. melanogaster, Nup93-1, and Nup93-2 resulted in significant signal reduction of up to 82% in adult pericardial nephrocytes with concomitant disruption of NPC protein expression. Additionally, nephrocyte morphology was highly abnormal in Nup93-1 and Nup93-2 silenced flies surviving to adulthood. CONCLUSION: We expand the spectrum of NUP93 variants detected in paediatric onset SRNS and demonstrate its incidence within a national cohort. Silencing of either D. melanogaster Nup93 ortholog caused a severe nephrocyte phenotype, signaling an important role for the nucleoporin complex in podocyte biology. A higher resolution version of the Graphical abstract is available as Supplementary information.
Assuntos
Drosophila melanogaster , Síndrome Nefrótica , Complexo de Proteínas Formadoras de Poros Nucleares , Podócitos , Adulto , Animais , Criança , Modelos Animais de Doenças , Drosophila melanogaster/genética , Resistência a Medicamentos/genética , Glucocorticoides/efeitos adversos , Glucocorticoides/uso terapêutico , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mutação , Síndrome Nefrótica/tratamento farmacológico , Síndrome Nefrótica/genética , Síndrome Nefrótica/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Podócitos/metabolismoAssuntos
Vacinas contra COVID-19 , COVID-19 , Anticorpos Neutralizantes , Anticorpos Antivirais , Humanos , Diálise Renal , SARS-CoV-2 , Reino Unido , VacinaçãoRESUMO
Developing B cells can be positively or negatively selected by self-antigens, but the mechanisms that determine these outcomes are incompletely understood. Here, we show that a B cell intrinsic switch between positive and negative selection during ontogeny is determined by a change from Lin28b to let-7 gene expression. Ectopic expression of a Lin28b transgene in murine B cells restored the positive selection of autoreactive B-1 B cells by self-antigen in adult bone marrow. Analysis of antigen-specific immature B cells in early and late ontogeny identified Lin28b-dependent genes associated with B-1 B cell development, including Arid3a and Bhleh41, and Lin28b-independent effects are associated with the presence or absence of self-antigen. These findings identify cell intrinsic and extrinsic determinants of B cell fate during ontogeny and reconcile lineage and selection theories of B cell development. They explain how changes in the balance of positive and negative selection may be able to adapt to meet the immunological needs of an individual during its lifetime.
Assuntos
Linfócitos B/imunologia , Proteínas de Ligação a RNA/imunologia , Animais , Linfócitos B/citologia , Proliferação de Células , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/imunologia , Proteínas de Ligação a RNA/genéticaRESUMO
By developing a high-density murine immunophenotyping platform compatible with high-throughput genetic screening, we have established profound contributions of genetics and structure to immune variation (http://www.immunophenotype.org). Specifically, high-throughput phenotyping of 530 unique mouse gene knockouts identified 140 monogenic 'hits', of which most had no previous immunologic association. Furthermore, hits were collectively enriched in genes for which humans show poor tolerance to loss of function. The immunophenotyping platform also exposed dense correlation networks linking immune parameters with each other and with specific physiologic traits. Such linkages limit freedom of movement for individual immune parameters, thereby imposing genetically regulated 'immunologic structures', the integrity of which was associated with immunocompetence. Hence, we provide an expanded genetic resource and structural perspective for understanding and monitoring immune variation in health and disease.
Assuntos
Infecções por Enterobacteriaceae/imunologia , Variação Genética/genética , Ensaios de Triagem em Larga Escala/métodos , Imunofenotipagem/métodos , Infecções por Salmonella/imunologia , Animais , Citrobacter/imunologia , Infecções por Enterobacteriaceae/microbiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Animais , Salmonella/imunologia , Infecções por Salmonella/microbiologiaRESUMO
Physiological effects of cellular hypoxia are sensed by prolyl hydroxylase (PHD) enzymes which regulate HIFs. Genetic interventions on HIF/PHD pathways reveal multiple phenotypes that extend the known biology of hypoxia. Recent studies unexpectedly implicate HIF in aspects of multiple immune and inflammatory pathways. However such studies are often limited by systemic lethal effects and/or use tissue-specific recombination systems, which are inherently irreversible, un-physiologically restricted and difficult to time. To study these processes better we developed recombinant mice which express tetracycline-regulated shRNAs broadly targeting the main components of the HIF/PHD pathway, permitting timed bi-directional intervention. We have shown that stabilization of HIF levels in adult mice through PHD2 enzyme silencing by RNA interference, or inducible recombination of floxed alleles, results in multi-lineage leukocytosis and features of autoimmunity. This phenotype was rapidly normalized on re-establishment of the hypoxia-sensing machinery when shRNA expression was discontinued. In both situations these effects were mediated principally through the Hif2a isoform. Assessment of cells bearing regulatory T cell markers from these mice revealed defective function and pro-inflammatory effects in vivo. We believe our findings have shown a new role for the PHD2/Hif2a couple in the reversible regulation of T cell and immune activity.
Assuntos
Prolina Dioxigenases do Fator Induzível por Hipóxia , Interferência de RNA/imunologia , Transdução de Sinais , Linfócitos T Reguladores , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/imunologia , Prolina Dioxigenases do Fator Induzível por Hipóxia/genética , Prolina Dioxigenases do Fator Induzível por Hipóxia/imunologia , Camundongos , Camundongos Transgênicos , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismoRESUMO
Despite the known importance of zinc for human immunity, molecular insights into its roles have remained limited. Here we report a novel autosomal recessive disease characterized by absent B cells, agammaglobulinemia and early onset infections in five unrelated families. The immunodeficiency results from hypomorphic mutations of SLC39A7, which encodes the endoplasmic reticulum-to-cytoplasm zinc transporter ZIP7. Using CRISPR-Cas9 mutagenesis we have precisely modeled ZIP7 deficiency in mice. Homozygosity for a null allele caused embryonic death, but hypomorphic alleles reproduced the block in B cell development seen in patients. B cells from mutant mice exhibited a diminished concentration of cytoplasmic free zinc, increased phosphatase activity and decreased phosphorylation of signaling molecules downstream of the pre-B cell and B cell receptors. Our findings highlight a specific role for cytosolic Zn2+ in modulating B cell receptor signal strength and positive selection.
Assuntos
Agamaglobulinemia/imunologia , Linfócitos B/imunologia , Proteínas de Transporte de Cátions/imunologia , Zinco/imunologia , Agamaglobulinemia/genética , Agamaglobulinemia/metabolismo , Animais , Linfócitos B/metabolismo , Proteínas de Transporte de Cátions/deficiência , Proteínas de Transporte de Cátions/genética , Pré-Escolar , Citosol/imunologia , Citosol/metabolismo , Modelos Animais de Doenças , Retículo Endoplasmático/imunologia , Retículo Endoplasmático/metabolismo , Feminino , Perfilação da Expressão Gênica , Humanos , Lactente , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação , Linhagem , Zinco/metabolismoRESUMO
BACKGROUND: Sickle cell disease (SCD) is one of the commonest severe monogenic disorders in the world, due to the inheritance of two abnormal haemoglobin (beta-globin) genes. SCD can cause severe pain, significant end-organ damage, pulmonary complications, and premature death. Kidney disease is a frequent and potentially severe complication in people with SCD.Chronic kidney disease is defined as abnormalities of kidney structure or function, present for more than three months. Sickle cell nephropathy refers to the spectrum of kidney complications in SCD.Glomerular damage is a cause of microalbuminuria and can develop at an early age in children with SCD, and increases in prevalence in adulthood. In people with sickle cell nephropathy, outcomes are poor as a result of the progression to proteinuria and chronic kidney insufficiency. Up to 12% of people who develop sickle cell nephropathy will develop end-stage renal disease. OBJECTIVES: To assess the effectiveness of any intervention in preventing or reducing kidney complications or chronic kidney disease in people with SCD (including red blood cell transfusions, hydroxyurea and angiotensin-converting enzyme inhibitor (ACEI)), either alone or in combination with each other. SEARCH METHODS: We searched for relevant trials in the Cochrane Library, MEDLINE (from 1946), Embase (from 1974), the Transfusion Evidence Library (from 1980), and ongoing trial databases; all searches current to 05 April 2016. We searched the Cochrane Cystic Fibrosis and Genetic Disorders Group Trials Register: 13 April 2017. SELECTION CRITERIA: Randomised controlled trials comparing interventions to prevent or reduce kidney complications or chronic kidney disease in people with SCD. There were no restrictions by outcomes examined, language or publication status. DATA COLLECTION AND ANALYSIS: Two authors independently assessed trial eligibility, extracted data and assessed the risk of bias. MAIN RESULTS: We included two trials with 215 participants. One trial was published in 2011 and included 193 children aged 9 months to 18 months, and compared treatment with hydroxyurea to placebo. The second trial was published in 1998 and included 22 adults with normal blood pressure and microalbuminuria and compared ACEI to placebo.We rated the quality of evidence as low to very low across different outcomes according to GRADE methodology. This was due to trials having: a high or unclear risk of bias including attrition and detection bias; indirectness (the available evidence was for children aged 9 months to 18 months in one trial and a small and select adult sample size in a second trial); and imprecise outcome effect estimates of significant benefit or harm. Hydroxyurea versus placebo We are very uncertain if hydroxyurea reduces or prevents progression of kidney disease (assessed by change in glomerular filtration rate), or reduces hyperfiltration in children aged 9 to 18 months, mean difference (MD) 0.58 (95% confidence interval (CI) -14.60 to 15.76 (mL/min per 1.73 m²)) (one study; 142 participants; very low-quality evidence).In children aged 9 to 18 months, hydroxyurea may improve the ability to concentrate urine, MD 42.23 (95% CI 12.14 to 72.32 (mOsm/kg)) (one study; 178 participants; low-quality evidence).Hydroxyurea may make little or no difference to SCD-related serious adverse events including: incidence of acute chest syndrome, risk ratio (RR) 0.39 (99% CI 0.13 to 1.16); painful crisis, RR 0.68 (99% CI 0.45 to 1.02); and hospitalisations, RR 0.83 (99% CI 0.68 to 1.01) (one study, 193 participants; low-quality evidence).No deaths occurred in the trial. Quality of life was not reported. ACEI versus placeboWe are very uncertain if ACEI reduces proteinuria in adults with SCD who have normal blood pressure and microalbuminuria, MD -49.00 (95% CI -124.10 to 26.10 (mg per day)) (one study; 22 participants; very low-quality evidence). We are very uncertain if ACEI reduce or prevent kidney disease as measured by creatinine clearance. The authors state that creatinine clearance remained constant over six months in both groups, but no comparative data were provided (very low-quality evidence).All-cause mortality, serious adverse events and quality of life were not reported. AUTHORS' CONCLUSIONS: In young children aged 9 months to 18 months, we are very uncertain if hydroxyurea improves glomerular filtration rate or reduces hyperfiltration, but it may improve young children's ability to concentrate urine and may make little or no difference on the incidence of acute chest syndrome, painful crises and hospitalisations.We are very uncertain if giving ACEI to adults with normal blood pressure and microalbuminuria has any effect on preventing or reducing kidney complications.This review identified no trials that looked at red cell transfusions nor any combinations of interventions to prevent or reduce kidney complications.Due to lack of evidence this review cannot comment on the management of either children aged over 18 months or adults with any known genotype of SCD.We have identified a lack of adequately-designed and powered studies, and no ongoing trials which address this critical question. Trials of hydroxyurea, ACEI or red blood cell transfusion in older children and adults are urgently needed to determine any effect on prevention or reduction kidney complications in people with SCD.
Assuntos
Anemia Falciforme/complicações , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Antidrepanocíticos/uso terapêutico , Hidroxiureia/uso terapêutico , Falência Renal Crônica/prevenção & controle , Adulto , Albuminúria/complicações , Anemia Falciforme/tratamento farmacológico , Creatinina/metabolismo , Taxa de Filtração Glomerular/efeitos dos fármacos , Hospitalização/estatística & dados numéricos , Humanos , Lactente , Falência Renal Crônica/tratamento farmacológico , Falência Renal Crônica/etiologia , Placebos , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
Outer retinal and renal glomerular functions rely on specialized vasculature maintained by VEGF that is produced by neighboring epithelial cells, the retinal pigment epithelium (RPE) and podocytes, respectively. Dysregulation of RPE- and podocyte-derived VEGF is associated with neovascularization in wet age-related macular degeneration (ARMD), choriocapillaris degeneration, and glomerular thrombotic microangiopathy (TMA). Since complement activation and genetic variants in inhibitory complement factor H (CFH) are also features of both ARMD and TMA, we hypothesized that VEGF and CFH interact. Here, we demonstrated that VEGF inhibition decreases local CFH and other complement regulators in the eye and kidney through reduced VEGFR2/PKC-α/CREB signaling. Patient podocytes and RPE cells carrying disease-associated CFH genetic variants had more alternative complement pathway deposits than controls. These deposits were increased by VEGF antagonism, a common wet ARMD treatment, suggesting that VEGF inhibition could reduce cellular complement regulatory capacity. VEGF antagonism also increased markers of endothelial cell activation, which was partially reduced by genetic complement inhibition. Together, these results suggest that VEGF protects the retinal and glomerular microvasculature, not only through VEGFR2-mediated vasculotrophism, but also through modulation of local complement proteins that could protect against complement-mediated damage. Though further study is warranted, these findings could be relevant for patients receiving VEGF antagonists.
Assuntos
Fator H do Complemento/metabolismo , Proteínas do Olho/metabolismo , Podócitos/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Idoso , Animais , Fator H do Complemento/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas do Olho/antagonistas & inibidores , Proteínas do Olho/genética , Feminino , Humanos , Nefropatias/genética , Nefropatias/metabolismo , Nefropatias/patologia , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino , Camundongos , Camundongos Knockout , Podócitos/patologia , Proteína Quinase C-alfa/genética , Proteína Quinase C-alfa/metabolismo , Epitélio Pigmentado da Retina/patologia , Microangiopatias Trombóticas/genética , Microangiopatias Trombóticas/metabolismo , Microangiopatias Trombóticas/patologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
The positive and negative selection of lymphocytes by antigen is central to adaptive immunity and self-tolerance, yet how this is determined by different antigens is not completely understood. We found that thymocyte-selection-associated family member 2 (Themis2) increased the positive selection of B1 cells and germinal center B cells by self and foreign antigens. Themis2 lowered the threshold for B-cell activation by low-avidity, but not high-avidity, antigens. Themis2 constitutively bound the adaptor protein Grb2, src-kinase Lyn and signal transducer phospholipase γ2 (PLC-γ2), and increased activation of PLC-γ2 and its downstream pathways following B cell receptor stimulation. Our findings identify a unique function for Themis2 in differential signaling and provide insight into how B cells discriminate between antigens of different quantity and quality.
Assuntos
Linfócitos B/fisiologia , Seleção Clonal Mediada por Antígeno , Centro Germinativo/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Ativação Linfocitária , Imunidade Adaptativa , Animais , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Proteína Adaptadora GRB2/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Fosfolipase C gama/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Tolerância a Antígenos Próprios , Quinases da Família src/metabolismoRESUMO
Folliculin (FLCN) is a tumor-suppressor protein mutated in the Birt-Hogg-Dubé (BHD) syndrome, which associates with two paralogous proteins, folliculin-interacting protein (FNIP)1 and FNIP2, forming a complex that interacts with the AMP-activated protein kinase (AMPK). Although it is clear that this complex influences AMPK and other metabolic regulators, reports of its effects have been inconsistent. To address this issue, we created a recessive loss-of-function variant of Fnip1 Homozygous FNIP1 deficiency resulted in profound B-cell deficiency, partially restored by overexpression of the antiapoptotic protein BCL2, whereas heterozygous deficiency caused a loss of marginal zone B cells. FNIP1-deficient mice developed cardiomyopathy characterized by left ventricular hypertrophy and glycogen accumulation, with close parallels to mice and humans bearing gain-of-function mutations in the γ2 subunit of AMPK. Concordantly, γ2-specific AMPK activity was elevated in neonatal FNIP1-deficient myocardium, whereas AMPK-dependent unc-51-like autophagy activating kinase 1 (ULK1) phosphorylation and autophagy were increased in FNIP1-deficient B-cell progenitors. These data support a role for FNIP1 as a negative regulator of AMPK.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Linfócitos B/citologia , Cardiomiopatias/metabolismo , Proteínas de Transporte/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Linfócitos B/enzimologia , Linfócitos B/metabolismo , Cardiomiopatias/genética , Proteínas de Transporte/metabolismo , Contagem de Células , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genéticaRESUMO
This is the protocol for a review and there is no abstract. The objectives are as follows: To assess the effectiveness of any intervention in preventing or reducing kidney complications or CKD in people with SCD (including red blood cell transfusions, hydroxyurea and ACEI (either alone or in combination with each other)).