RESUMO
The risk of developing Alzheimer's disease (AD) significantly increases in individuals carrying the APOEε4 allele. Elderly cognitively healthy individuals with APOEε4 also exist, suggesting the presence of cellular mechanisms that counteract the pathological effects of APOEε4; however, these mechanisms are unknown. We hypothesized that APOEε4 carriers without dementia might carry genetic variations that could protect them from developing APOEε4-mediated AD pathology. To test this, we leveraged whole-genome sequencing (WGS) data in the National Institute on Aging Alzheimer's Disease Family Based Study (NIA-AD FBS), Washington Heights/Inwood Columbia Aging Project (WHICAP), and Estudio Familiar de Influencia Genetica en Alzheimer (EFIGA) cohorts and identified potentially protective variants segregating exclusively among unaffected APOEε4 carriers. In homozygous unaffected carriers above 70 years old, we identified 510 rare coding variants. Pathway analysis of the genes harboring these variants showed significant enrichment in extracellular matrix (ECM)-related processes, suggesting protective effects of functional modifications in ECM proteins. We prioritized two genes that were highly represented in the ECM-related gene ontology terms, (FN1) and collagen type VI alpha 2 chain (COL6A2) and are known to be expressed at the blood-brain barrier (BBB), for postmortem validation and in vivo functional studies. An independent analysis in a large cohort of 7185 APOEε4 homozygous carriers found that rs140926439 variant in FN1 was protective of AD (OR = 0.29; 95% CI [0.11, 0.78], P = 0.014) and delayed age at onset of disease by 3.37 years (95% CI [0.42, 6.32], P = 0.025). The FN1 and COL6A2 protein levels were increased at the BBB in APOEε4 carriers with AD. Brain expression of cognitively unaffected homozygous APOEε4 carriers had significantly lower FN1 deposition and less reactive gliosis compared to homozygous APOEε4 carriers with AD, suggesting that FN1 might be a downstream driver of APOEε4-mediated AD-related pathology and cognitive decline. To validate our findings, we used zebrafish models with loss-of-function (LOF) mutations in fn1b-the ortholog for human FN1. We found that fibronectin LOF reduced gliosis, enhanced gliovascular remodeling, and potentiated the microglial response, suggesting that pathological accumulation of FN1 could impair toxic protein clearance, which is ameliorated with FN1 LOF. Our study suggests that vascular deposition of FN1 is related to the pathogenicity of APOEε4, and LOF variants in FN1 may reduce APOEε4-related AD risk, providing novel clues to potential therapeutic interventions targeting the ECM to mitigate AD risk.
Assuntos
Doença de Alzheimer , Fibronectinas , Idoso , Animais , Humanos , Doença de Alzheimer/genética , Fibronectinas/genética , Variação Genética/genética , Gliose , Peixe-ZebraRESUMO
Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) is a rare primary cutaneous non-Hodgkin lymphoma involving CD8+ T cells, the genetic underpinnings of which remain incompletely understood. Here we report two unrelated patients with B cell Expansion with NF-κB and T cell Anergy (BENTA) disease and a novel presentation of SPTCL. Patient 1 presented early in life with recurrent infections and B cell lymphocytosis, linked to a novel gain-of-function (GOF) CARD11 mutation (p.Lys238del). He developed SPTCL-like lesions and membranoproliferative glomerulonephritis by age 2, treated successfully with cyclosporine. Patient 2 presented at 13 months with splenomegaly, lymphadenopathy, and SPTCL with evidence of hemophagocytic lymphohistiocytosis. Genetic analysis revealed two in cis germline GOF CARD11 variants (p.Glu121Asp/p.Gly126Ser). Autologous bone marrow transplant resulted in SPTCL remission despite persistent B cell lymphocytosis. These cases illuminate an unusual pathological manifestation for BENTA disease, suggesting that CARD11 GOF mutations can manifest in cutaneous CD4+and CD8+ T cell malignancies.
Assuntos
Síndromes de Imunodeficiência , Linfocitose , Linfoma de Células T , Paniculite , Masculino , Humanos , Pré-Escolar , Linfócitos T CD8-Positivos/patologia , Paniculite/genética , Paniculite/patologia , Paniculite/terapia , Linfoma de Células T/genética , Linfoma de Células T/terapiaRESUMO
Ionizing radiation-induced tissue damage recruits monocytes into the exposed area where they are differentiated to macrophages. These implement phagocytic removal of dying cells and elicit an acute inflammatory response, but can also facilitate tumorigenesis due to production of anti-inflammatory cytokines. Using primary human monocyte-derived macrophages (MDMs) and the THP1 monocytic cell line, we demonstrate that gamma radiation triggers monocyte differentiation toward the macrophage phenotype with increased expression of type I interferons (IFN-I) and both pro- and anti-inflammatory macrophage activation markers. We found that these changes correlate with significantly upregulated expression of 622 retroelements from various groups, particularly of several clades of human endogenous retroviruses (HERVs). Elevated transcription was detected in both sense and antisense directions in the HERV subgroups tested, including the most genetically homogeneous clade HML-2. The level of antisense transcription was three- to five-fold higher than of the sense strand levels. Using a proximity ligation assay and immunoprecipitation followed by RNA quantification, we identified an increased amount of the dsRNA receptors MDA-5 and TLR3 bound to an equivalent number of copies of sense and antisense chains of HERVK HML-2 RNA. This binding triggered MAVS-associated signaling pathways resulting in increased expression of IFN-I and inflammation related genes that enhanced the cumulative inflammatory effect of radiation-induced senescence. HML-2 knockdown was accompanied with reduced expression and secretion of IFNα, pro-inflammatory (IL-1ß, IL-6, CCL2, CCL3, CCL8, and CCL20) and anti-inflammatory (IL10) modulators in irradiated monocytes and MDMs. Taken together, our data indicate that radiation stress-induced HERV expression enhances the IFN-I and cytokine response and results in increased levels of pro-inflammatory modulators along with expression of anti-inflammatory factors associated with the macrophage tumorigenic phenotype.
Assuntos
Retrovirus Endógenos/genética , Raios gama , Inflamação/imunologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Retroelementos/genética , Diferenciação Celular , Citocinas/metabolismo , Humanos , Inflamação/metabolismo , Inflamação/patologia , Macrófagos/metabolismo , Macrófagos/efeitos da radiação , Monócitos/metabolismo , Monócitos/efeitos da radiação , TranscriptomaRESUMO
Traumatic brain injury (TBI) causes diffuse axonal injury which can produce chronic white matter pathology and subsequent post-traumatic neurodegeneration with poor patient outcomes. Tau modulates axon cytoskeletal functions and undergoes phosphorylation and mis-localization in neurodegenerative disorders. The effects of tau pathology on neurodegeneration after TBI are unclear. We used mice with neuronal expression of human mutant tau to examine effects of pathological tau on white matter pathology after TBI. Adult male and female hTau.P301S (Tg2541) transgenic and wild-type (Wt) mice received either moderate single TBI (s-TBI) or repetitive mild TBI (r-mTBI; once daily × 5), or sham procedures. Acutely, s-TBI produced more extensive axon damage in the corpus callosum (CC) as compared to r-mTBI. After s-TBI, significant CC thinning was present at 6 weeks and 4 months post-injury in Wt and transgenic mice, with homozygous tau expression producing additional pathology of late demyelination. In contrast, r-mTBI did not produce significant CC thinning except at the chronic time point of 4 months in homozygous mice, which exhibited significant CC atrophy (- 29.7%) with increased microgliosis. Serum neurofilament light quantification detected traumatic axonal injury at 1 day post-TBI in Wt and homozygous mice. At 4 months, high tau and neurofilament in homozygous mice implicated tau in chronic axon pathology. These findings did not have sex differences detected. Conclusions: Neuronal tau pathology differentially exacerbated CC pathology based on injury severity and chronicity. Ongoing CC atrophy from s-TBI became accompanied by late demyelination. Pathological tau significantly worsened CC atrophy during the chronic phase after r-mTBI.
Assuntos
Lesões Encefálicas Traumáticas , Doenças Desmielinizantes , Tauopatias , Substância Branca , Adulto , Animais , Feminino , Humanos , Masculino , Camundongos , Atrofia/patologia , Lesões Encefálicas Traumáticas/patologia , Doenças Desmielinizantes/patologia , Camundongos Transgênicos , Proteínas tau/genética , Proteínas tau/metabolismo , Substância Branca/patologiaRESUMO
Background and Objectives: Stress can overload adaptive mechanisms, leading to epigenetic effects harmful to health. Research on the reversal of these effects is in its infancy. Early results suggest some meditation techniques have health benefits that grow with repeated practice. This study focused on possible transcriptomic effects of 38 years of twice-daily Transcendental Meditation® (TM®) practice. Materials and Methods: First, using Illumina® BeadChip microarray technology, differences in global gene expression in peripheral blood mononuclear cells (PBMCs) were sought between healthy practitioners and tightly matched controls (n = 12, age 65). Second, these microarray results were verified on a subset of genes using quantitative polymerase chain reaction (qPCR) and were validated using qPCR in larger TM and control groups (n = 45, age 63). Bioinformatics investigation employed Ingenuity® Pathway Analysis (IPA®), DAVID, Genomatix, and R packages. Results: The 200 genes and loci found to meet strict criteria for differential expression in the microarray experiment showed contrasting patterns of expression that distinguished the two groups. Differential expression relating to immune function and energy efficiency were most apparent. In the TM group, relative to the control, all 49 genes associated with inflammation were downregulated, while genes associated with antiviral and antibody components of the defense response were upregulated. The largest expression differences were shown by six genes related to erythrocyte function that appeared to reflect a condition of lower energy efficiency in the control group. Results supporting these gene expression differences were obtained with qPCR-measured expression both in the well-matched microarray groups and in the larger, less well-matched groups. Conclusions: These findings are consistent with predictions based on results from earlier randomized trials of meditation and may provide evidence for stress-related molecular mechanisms underlying reductions in anxiety, post-traumatic stress disorder (PTSD), cardiovascular disease (CVD), and other chronic disorders and diseases.
Assuntos
Meditação , Biologia Computacional , Leucócitos Mononucleares , Estresse Psicológico/prevenção & controle , TranscriptomaRESUMO
BACKGROUND: Astrocytes respond to central nervous system (CNS) injury and disease by transforming to a reactive astrogliosis cell state that can contribute to either CNS dysfunction or repair. Neuroinflammation is a powerful driver of a harmful A1 astrogliosis phenotype associated with in vitro neurotoxicity and histopathology in human neurodegenerative diseases. Here we report a protocol for the rapid development of a human cell culture model of neuroinflammatory astrogliosis using induced pluripotent stem cells (iPSCs). METHODS: Using RNA sequencing and in vitro cell assays, we measured transcriptional and cellular effects of chronic exposure of human iPSC-derived astrocytes to the cytokines TNFα (tumor necrosis factor alpha) or IL-1ß (interleukin-1 beta). RESULTS: We show TNFα and IL-1ß induce pro-inflammatory gene signatures but by widely different magnitudes. TNFα treatment results in 606 differential expressed genes, the suppression of glutamate-uptake, and increased phagocytic activity in astrocyte cultures. In contrast, IL-1ß effects are attenuated to 33 differential expressed genes and no significant effects on glutamate-uptake or increased phagocytic activity. CONCLUSION: Our approach demonstrates a rapid tool for modeling neuroinflammatory human astrocytic responses in nervous system trauma and disease. In particular, we reveal a model for robust TNFα-induced human astrogliosis suitable for the study of neurotoxic A1 astrocytes.
Assuntos
Astrócitos/metabolismo , Sangue Fetal/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Mediadores da Inflamação/metabolismo , Fagocitose/fisiologia , Astrócitos/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Sangue Fetal/citologia , Sangue Fetal/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Mediadores da Inflamação/farmacologia , Fagocitose/efeitos dos fármacosRESUMO
Human induced pluripotent stem (iPS) cell-derived neurons and astrocytes are attractive cellular tools for nervous system disease modeling and drug screening. Optimal utilization of these tools requires differentiation protocols that efficiently generate functional cell phenotypes in vitro. As nervous system function is dependent on networked neuronal activity involving both neuronal and astrocytic synaptic functions, we examined astrocyte effects on the functional maturation of neurons from human iPS cell-derived neural stem cells (NSCs). We first demonstrate human iPS cell-derived NSCs can be rapidly differentiated in culture to either neurons or astrocytes with characteristic cellular, molecular and physiological features. Although differentiated neurons were capable of firing multiple action potentials (APs), few cells developed spontaneous electrical activity in culture. We show spontaneous electrical activity was significantly increased by neuronal differentiation of human NSCs on feeder layers of neonatal mouse cortical astrocytes. In contrast, co-culture on feeder layers of isogenic human iPS cell-derived astrocytes had no positive effect on spontaneous neuronal activity. Spontaneous electrical activity was dependent on glutamate receptor-channel function and occurred without changes in INa , IK , Vm , and AP properties of iPS cell-derived neurons. These data demonstrate co-culture with neonatal mouse cortical astrocytes but not human isogenic iPS cell-derived astrocytes stimulates glutamatergic synaptic transmission between iPS cell-derived neurons in culture. We present RNA-sequencing data for an immature, fetal-like status of our human iPS cell-derived astrocytes as one possible explanation for their failure to enhance synaptic activity in our co-culture system.
Assuntos
Astrócitos/fisiologia , Córtex Cerebral/fisiologia , Células Alimentadoras/fisiologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Neurogênese/fisiologia , Neurônios/fisiologia , Potenciais de Ação , Animais , Astrócitos/citologia , Linhagem Celular , Córtex Cerebral/citologia , Técnicas de Cocultura , Células Alimentadoras/citologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/fisiologia , Neurônios/citologia , Receptores de Glutamato/metabolismo , TranscriptomaRESUMO
Neuroinflammation plays a critical role in the pathogenesis of traumatic brain injury (TBI). TBI induces rapid activation of astrocytes and microglia, infiltration of peripheral leukocytes, and secretion of inflammatory cytokines. In the context of modest or severe TBI, such inflammation contributes to tissue destruction and permanent brain damage. However, it is clear that the inflammatory response is also necessary to promote post-injury healing. To date, anti-inflammatory therapies, including the broad class of non-steroidal anti-inflammatory drugs (NSAIDs), have met with little success in treatment of TBI, perhaps because these drugs have inhibited both the tissue-damaging and repair-promoting aspects of the inflammatory response, or because inhibition of inflammation alone is insufficient to yield therapeutic benefit. Salsalate is an unacetylated salicylate with long history of use in limiting inflammation. This drug is known to block activation of NF-κB, and recent data suggest that salsalate has a number of additional biological activities, which may also contribute to its efficacy in treatment of human disease. Here, we show that salsalate potently blocks pro-inflammatory gene expression and nitrite secretion by microglia in vitro. Using the controlled cortical impact (CCI) model in mice, we find that salsalate has a broad anti-inflammatory effect on in vivo TBI-induced gene expression, when administered post-injury. Interestingly, salsalate also elevates expression of genes associated with neuroprotection and neurogenesis, including the neuropeptides, oxytocin and thyrotropin releasing hormone. Histological analysis reveals salsalate-dependent decreases in numbers and activation-associated morphological changes in microglia/macrophages, proximal to the injury site. Flow cytometry data show that salsalate changes the kinetics of CCI-induced accumulation of various populations of CD11b-positive myeloid cells in the injured brain. Behavioral assays demonstrate that salsalate treatment promotes significant recovery of function following CCI. These pre-clinical data suggest that salsalate may show promise as a TBI therapy with a multifactorial mechanism of action to enhance functional recovery.
Assuntos
Lesões Encefálicas Traumáticas/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Inflamação/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Recuperação de Função Fisiológica/efeitos dos fármacos , Salicilatos/uso terapêutico , Animais , Encéfalo/patologia , Lesões Encefálicas Traumáticas/patologia , Linhagem Celular , Fluxo Gênico/efeitos dos fármacos , Inflamação/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/patologia , Fármacos Neuroprotetores/farmacologia , Ocitocina/genética , Salicilatos/farmacologia , Hormônio Liberador de Tireotropina/genéticaRESUMO
PURPOSE: Activating germline mutations in CARD11 have recently been linked to a rare genetic disorder associated with congenital B cell lymphocytosis. We describe a patient with a similar clinical phenotype who had a de novo germline G123D CARD11 mutation. METHODS: Whole exome sequencing was performed on DNA from the patient and his biological parents. Laboratory studies examined characteristics of the patient's B and T lymphocytes. A CARD11 cDNA containing the mutation was transfected into a lymphocyte cell line to gain an understanding of its function. RNA sequencing was performed on samples from the patient and from patients with alternate germline CARD11 mutations and differential gene expression analysis was performed. RESULTS: The patient had a decade-long history of severe polyclonal B lymphocytosis in the 20,000-90,000 lymphocytes/mm(3) range, which was markedly exacerbated by EBV infection and splenectomy at different times. He had a heterozygous germline CARD11 mutation causing a G123D amino acid substitution, which was demonstrated to induce NF-κB activation in unstimulated lymphocytes. In contrast to previous patients with CARD11 mutations, this patient's B cells exhibited higher expression of several cell cycle progression genes, as well as enhanced proliferation and improved survival following B cell receptor stimulation. CONCLUSIONS: This is the third reported germline and first de novo CARD11 mutation shown to cause congenital B cell lymphocytosis. The mutation was associated with a dramatically greater lymphocytosis than in previously described cases, disproportionate to the level of constitutive NF-κB activation. However, comparative review of the patient's clinical history, combined with additional genomic and functional analyses, underscore other important variables that may affect pathophysiology or regulate mutant CARD11 function in B cell proliferation and disease. We now refer to these patients as having BENTA disease (B cell Expansion with NF-κB and T cell Anergy).
Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Mutação em Linhagem Germinativa , Guanilato Ciclase/genética , Linfocitose/congênito , Linfocitose/genética , Substituição de Aminoácidos , Apoptose/genética , Linfócitos B/imunologia , Linfócitos B/metabolismo , Linfócitos B/patologia , Sequência de Bases , Proteínas Adaptadoras de Sinalização CARD/química , Proliferação de Células/genética , Criança , DNA/genética , Perfilação da Expressão Gênica , Guanilato Ciclase/química , Humanos , Ativação Linfocitária/genética , Linfocitose/imunologia , Masculino , Proteínas Mutantes/química , Proteínas Mutantes/genética , Mutação de Sentido Incorreto , NF-kappa B/metabolismoRESUMO
Wound research has typically been performed without regard for where the wounds are located on the body, despite well-known heterogeneities in physical and biological properties between different skin areas. The skin covering the palms and soles is highly specialized, and plantar ulcers are one of the most challenging and costly wound types to manage. Using primarily the porcine model, we show that plantar skin is molecularly and functionally more distinct from nonplantar skin than previously recognized, with unique gene and protein expression profiles, broad alterations in cellular functions, constitutive activation of many wound-associated phenotypes, and inherently delayed healing. This unusual physiology is likely to play a significant but underappreciated role in the pathogenesis of plantar ulcers as well as the last 25+ years of futility in therapy development efforts. By revealing this critical yet unrecognized pitfall, we hope to contribute to the development of more effective therapies for these devastating nonhealing wounds.
Assuntos
Fenótipo , Pele , Cicatrização , Animais , Cicatrização/fisiologia , Suínos , Pele/patologia , Pele/lesões , Pele/metabolismo , Modelos Animais de Doenças , Úlcera do Pé/fisiopatologia , Úlcera do Pé/patologia , Humanos , Feminino , Fenômenos Fisiológicos da Pele , PéRESUMO
BACKGROUND: Suicide is a societal and public health concern of global scale. Identifying genetic risk factors for suicide attempt can characterize underlying biology and enable early interventions to prevent deaths. Recent studies have described common genetic variants for suicide-related behaviors. Here, we advance this search for genetic risk by analyzing the association between suicide attempt and uncommon variation exome-wide in a large, ancestrally diverse sample. METHODS: We sequenced whole genomes of 13,584 soldiers from the Army STARRS (Army Study to Assess Risk and Resilience in Servicemembers), including 979 individuals with a history of suicide attempt. Uncommon, nonsilent protein-coding variants were analyzed exome-wide for association with suicide attempt using gene-collapsed and single-variant analyses. RESULTS: We identified 19 genes with variants enriched in individuals with history of suicide attempt, either through gene-collapsed or single-variant analysis (Bonferroni padjusted < .05). These genes were CIB2, MLF1, HERC1, YWHAE, RCN2, VWA5B1, ATAD3A, NACA, EP400, ZNF585A, LYST, RC3H2, PSD3, STARD9, SGMS1, ACTR6, RGS7BP, DIRAS2, and KRTAP10-1. Most genes had variants across multiple genomic ancestry groups. Seventeen of these genes were expressed in healthy brain tissue, with 9 genes expressed at the highest levels in the brain versus other tissues. Brains from individuals deceased from suicide aberrantly expressed RGS7BP (padjusted = .035) in addition to nominally significant genes including YWHAE and ACTR6, all of which have reported associations with other mental disorders. CONCLUSIONS: These results advance the molecular characterization of suicide attempt behavior and support the utility of whole-genome sequencing for complementing the findings of genome-wide association studies in suicide research.
Assuntos
Militares , Tentativa de Suicídio , Humanos , Militares/psicologia , Masculino , Estados Unidos/epidemiologia , Feminino , Adulto , Adulto Jovem , Predisposição Genética para Doença/genética , Variação Genética/genética , Estudo de Associação Genômica Ampla , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Fetal Alcohol Spectrum Disorder (FASD) encompasses an array of effects of prenatal alcohol exposure (PAE), including physical abnormalities and cognitive and behavioral deficits. Disruptions of cortical development have been implicated in multiple PAE studies, with deficits including decreased progenitor proliferation, disrupted neuronal differentiation, aberrant radial migration of pyramidal neurons, and decreased cortical thickness. While several mechanisms of alcohol teratogenicity have been explored, how specific cell types in the brain at different developmental time points may be differentially affected by PAE is still poorly understood. In this study, we used single nucleus RNA sequencing (snRNAseq) to investigate whether moderate PAE from neurulation through peak cortical neurogenesis induces cell type-specific transcriptomic changes in the developing murine brain. Cluster analysis identified 25 neuronal cell types, including subtypes of radial glial cells (RGCs), intermediate progenitor cells (IPCs), projection neurons, and interneurons. Only Wnt-expressing cortical hem RGCs showed a significant decrease in the percentage of cells after PAE, with no cell types showing PAE-induced apoptosis as measured by caspase expression. Cell cycle analysis revealed only a subtype of RGCs expressing the downstream Wnt signaling transcription factor Tcf7l2 had a decreased percentage of cells in the G2/M phase of the cell cycle, suggesting decreased proliferation in this RGC subtype and further implicating disrupted Wnt signaling after PAE at this early developmental timepoint. An increased pseudotime score in IPC and projection neuron cell types indicated that PAE led to increased or premature differentiation of these cells. Biological processes affected by PAE included the upregulation of pathways related to synaptic activity and neuronal differentiation and downregulation of pathways related to chromosome structure and the cell cycle. Several cell types showed a decrease in Wnt signaling pathways, with several genes related to Wnt signaling altered by PAE in multiple cell types. As Wnt has been shown to promote proliferation and inhibit differentiation at earlier stages in development, the downregulation of Wnt signaling may have resulted in premature neuronal maturation of projection neurons and their intermediate progenitors. Overall, these findings provide further insight into the cell type-specific effects of PAE during early corticogenesis.
RESUMO
Interferon γ (IFNγ) is an essential and pleiotropic activator of human monocytes, but little is known about the changes in cellular metabolism required for IFNγ-induced activation. We sought to elucidate the mechanisms by which IFNγ reprograms monocyte metabolism to support its immunologic activities. We found that IFNγ increased oxygen consumption rates (OCR) in monocytes, indicative of reactive oxygen species generation by both mitochondria and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Transcriptional profiling revealed that this oxidative phenotype was driven by IFNγ-induced reprogramming of NAD+ metabolism, which is dependent on nicotinamide phosphoribosyltransferase (NAMPT)-mediated NAD+ salvage to generate NADH and NADPH for oxidation by mitochondrial complex I and NADPH oxidase, respectively. Consistent with this pathway, monocytes from patients with gain-of-function mutations in STAT1 demonstrated higher-than-normal OCR, whereas chemical or genetic disruption of mitochondrial complex I (rotenone treatment or Leigh syndrome patient monocytes) or NADPH oxidase (diphenyleneiodonium treatment or chronic granulomatous disease [CGD] patient monocytes) reduced OCR. Interestingly, inhibition of NAMPT in healthy monocytes completely abrogated the IFNγ-induced oxygen consumption, comparable to levels observed in CGD monocytes. These data identify an IFNγ-induced, NAMPT-dependent, NAD+ salvage pathway that is critical for IFNγ activation of human monocytes.
Assuntos
Doença Granulomatosa Crônica , Monócitos , Doença Granulomatosa Crônica/metabolismo , Humanos , Interferon gama/farmacologia , Monócitos/metabolismo , NAD/metabolismo , NADP/metabolismo , NADPH Oxidases/metabolismo , Explosão RespiratóriaRESUMO
While BRCA1 and BRCA2 mutations are known to confer the largest risk of breast cancer and ovarian cancer, the incomplete penetrance of the mutations and the substantial variability in age at cancer onset among carriers suggest additional factors modifying the risk of cancer in BRCA1/2 mutation carriers. To identify genetic modifiers of BRCA1/2, we carried out a whole-genome sequencing study of 66 ovarian cancer patients that were enriched with BRCA carriers, followed by validation using data from the Pan-Cancer Analysis of Whole Genomes Consortium. We found PPARGC1A, a master regulator of mitochondrial biogenesis and function, to be highly mutated in BRCA carriers, and patients with both PPARGC1A and BRCA1/2 mutations were diagnosed with breast or ovarian cancer at significantly younger ages, while the mutation status of each gene alone did not significantly associate with age of onset. Our study suggests PPARGC1A as a possible BRCA modifier gene. Upon further validation, this finding can help improve cancer risk prediction and provide personalized preventive care for BRCA carriers.
RESUMO
In prostate cancer, emerging data highlight the role of DNA damage repair genes (DDRGs) in aggressive forms of the disease. However, DDRG mutations in African American men are not yet fully defined. Here, we profile germline mutations in all known DDRGs (N = 276) using whole genome sequences from blood DNA of a matched cohort of patients with primary prostate cancer comprising of 300 African American and 300 European Ancestry prostate cancer patients, to determine whether the mutation status can enhance patient stratification for specific targeted therapies. Here, we show that only 13 of the 46 DDRGs identified with pathogenic/likely pathogenic mutations are present in both African American and European ancestry patients. Importantly, RAD family genes (RAD51, RAD54L, RAD54B), which are potentially targetable, as well as PMS2 and BRCA1, are among the most frequently mutated DDRGs in African American, but not in European Ancestry patients.
Assuntos
Negro ou Afro-Americano , Neoplasias da Próstata , Negro ou Afro-Americano/genética , Dano ao DNA/genética , Mutação em Linhagem Germinativa , Humanos , Masculino , Mutação , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologiaAssuntos
Linfócitos B/imunologia , Proteínas Adaptadoras de Sinalização CARD/genética , Guanilato Ciclase/genética , Linfocitose/genética , Transtornos Linfoproliferativos/genética , Mutação de Sentido Incorreto , Adolescente , Linfócitos B/patologia , Proteínas Adaptadoras de Sinalização CARD/imunologia , Proliferação de Células , Análise Mutacional de DNA , Feminino , Guanilato Ciclase/imunologia , Humanos , Memória Imunológica , Linfocitose/congênito , Linfocitose/imunologia , Linfocitose/patologia , Transtornos Linfoproliferativos/congênito , Transtornos Linfoproliferativos/imunologia , Transtornos Linfoproliferativos/patologia , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de DoençaRESUMO
In females, estrogens have two main modes of action relating to gonadotropin secretion: positive feedback and negative feedback. Estrogen positive and negative feedback are controlled by different regions of the hypothalamus: the preoptic area/anterior portion (mainly the anteroventral periventricular nucleus, AVPV) of the hypothalamus is associated with estrogen positive feedback while the mediobasal hypothalamus (mainly the arcuate nucleus of the hypothalamus, ARH), is associated with estrogen negative feedback. In this study, we examined the temporal pattern of gene transcription in these two regions following estrogen treatment. Adult, ovariectomized, Long Evans rats received doses of estradiol benzoate (EB) or oil every 4 days for 3 cycles. On the last EB priming cycle, hypothalamic tissues were dissected into the AVPV+ and ARH+ at 0 hrs (baseline/oil control), 6 hrs, or 24 hrs after EB treatment. RNA was extracted and sequenced using bulk RNA sequencing. Differential gene analysis, gene ontology, and weighted correlation network analysis (WGCNA) was performed. Overall, we found that the AVPV+ and ARH+ respond differently to estradiol stimulation. In both regions, estradiol treatment resulted in more gene up-regulation than down-regulation. S100g was very strongly up-regulated by estradiol in both regions at 6 and 24 hrs after EB treatment. In the AVPV+ the highest number of differentially expressed genes occurred 24 hrs after EB. In the ARH+, the highest number of genes differentially expressed by EB occurred between 6 and 24 hrs after EB, while in the AVPV+, the fewest genes changed their expression between these time points, demonstrating a temporal difference in the way that EB regulates transcription these two areas. Several genes strongly implicated in gonadotropin release were differentially affected by estradiol including Esr1, encoding estrogen receptor-α and Kiss1, encoding kisspeptin. As an internal validation, Kiss1 was up-regulated in the AVPV+ and down-regulated in the ARH+. Gene network analysis revealed the vastly different clustering of genes modulated by estradiol in the AVPV+ compared with the ARH+. These results indicate that gene expression in these two hypothalamic regions have specific responses to estradiol in timing and direction.
Assuntos
Núcleo Arqueado do Hipotálamo/metabolismo , Estradiol/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônio Liberador de Gonadotropina/metabolismo , Hipotálamo Anterior/metabolismo , Hipotálamo/metabolismo , Análise de Sequência de RNA/métodos , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Receptor alfa de Estrogênio/metabolismo , Estrogênios/farmacologia , Feminino , Hipotálamo/efeitos dos fármacos , Hipotálamo Anterior/efeitos dos fármacos , Kisspeptinas/metabolismo , Modelos Animais , Ovariectomia/métodos , Ratos , Ratos Long-EvansRESUMO
The rising incidence and mortality of endometrial cancer (EC) in the United States calls for an improved understanding of the disease's progression. Current methodologies for diagnosis and treatment rely on the use of cell lines as models for tumor biology. However, due to inherent heterogeneity and differential growing environments between cell lines and tumors, these comparative studies have found little parallels in molecular signatures. As a consequence, the development and discovery of preclinical models and reliable drug targets are delayed. In this study, we established transcriptome parallels between cell lines and tumors from The Cancer Genome Atlas (TCGA) with the use of optimized normalization methods. We identified genes and signaling pathways associated with regulating the transformation and progression of EC. Specifically, the LXR/RXR activation, neuroprotective role for THOP1 in Alzheimer's disease, and glutamate receptor signaling pathways were observed to be mostly downregulated in advanced cancer stage. While some of these highlighted markers and signaling pathways are commonly found in the central nervous system (CNS), our results suggest a novel function of these genes in the periphery. Finally, our study underscores the value of implementing appropriate normalization methods in comparative studies to improve the identification of accurate and reliable markers.
RESUMO
The adaptive immune response relies on specific apoptotic programs to maintain homeostasis. Conventional effector T cell (Tcon) expansion is constrained by both forkhead box P3 (FOXP3)+-regulatory T cells (Tregs) and restimulation-induced cell death (RICD), a propriocidal apoptosis pathway triggered by repeated stimulation through the T-cell receptor (TCR). Constitutive FOXP3 expression protects Tregs from RICD by suppressing SLAM-associated protein (SAP), a key adaptor protein that amplifies TCR signaling strength. The role of transient FOXP3 induction in activated human CD4 and CD8 Tcons remains unresolved, but its expression is inversely correlated with acquired RICD sensitivity. Here, we describe a novel role for FOXP3 in protecting human Tcons from premature RICD during expansion. Unlike FOXP3-mediated protection from RICD in Tregs, FOXP3 protects Tcons through a distinct mechanism requiring de novo transcription that does not require SAP suppression. Transcriptome profiling and functional analyses of expanding Tcons revealed that FOXP3 enhances expression of the SLAM family receptor CD48, which in turn sustains basal autophagy and suppresses pro-apoptotic p53 signaling. Both CD48 and FOXP3 expression reduced p53 accumulation upon TCR restimulation. Furthermore, silencing FOXP3 expression or blocking CD48 decreased the mitochondrial membrane potential in expanding Tcons with a concomitant reduction in basal autophagy. Our findings suggest that FOXP3 governs a distinct transcriptional program in early-stage effector Tcons that maintains RICD resistance via CD48-dependent protective autophagy and p53 suppression.
Assuntos
Antígeno CD48/metabolismo , Morte Celular , Fatores de Transcrição Forkhead/metabolismo , Ativação Linfocitária , Receptores de Antígenos de Linfócitos T/metabolismo , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária/metabolismo , Linfócitos T Reguladores/imunologia , Apoptose , Autofagia , Antígeno CD48/genética , Fatores de Transcrição Forkhead/genética , Humanos , Receptores de Antígenos de Linfócitos T/genética , Transdução de Sinais , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
BACKGROUND: Chronic tobacco smoke exposure results in a broad range of lung pathologies including emphysema, airway disease and parenchymal fibrosis as well as a multitude of extra-pulmonary comorbidities. Prior work using CT imaging has identified several clinically relevant subgroups of smoking related lung disease, but these investigations have generally lacked organ specific molecular correlates. RESEARCH QUESTION: Can CT imaging be used to identify clinical phenotypes of smoking related lung disease that have specific bronchial epithelial gene expression patterns to better understand disease pathogenesis? STUDY DESIGN AND METHODS: Using K-means clustering, we clustered participants from the COPDGene study (n = 5,273) based on CT imaging characteristics and then evaluated their clinical phenotypes. These clusters were replicated in the Detection of Early Lung Cancer Among Military Personnel (DECAMP) cohort (n = 360), and were further characterized using bronchial epithelial gene expression. RESULTS: Three clusters (preserved, interstitial predominant and emphysema predominant) were identified. Compared to the preserved cluster, the interstitial and emphysema clusters had worse lung function, exercise capacity and quality of life. In longitudinal follow-up, individuals from the emphysema group had greater declines in exercise capacity and lung function, more emphysema, more exacerbations, and higher mortality. Similarly, genes involved in inflammatory pathways (tumor necrosis factor-α, interferon-ß) are more highly expressed in bronchial epithelial cells from individuals in the emphysema cluster, while genes associated with T-cell related biology are decreased in these samples. Samples from individuals in the interstitial cluster generally had intermediate levels of expression of these genes. INTERPRETATION: Using quantitative CT imaging, we identified three groups of individuals in older ever-smokers that replicate in two cohorts. Airway gene expression differences between the three groups suggests increased levels of inflammation in the most severe clinical phenotype, possibly mediated by the tumor necrosis factor-α and interferon-ß pathways. CLINICAL TRIAL REGISTRATION: COPDGene (NCT00608764), DECAMP-1 (NCT01785342), DECAMP-2 (NCT02504697).