RESUMO
Despite the advancements in modern medicine, there are still difficulties in diagnosing common illnesses. The invasiveness and price of the tests used to follow up certain diseases can be a barrier to proper patient follow-up. Sphingolipids are a diverse category of lipids. They are structural molecules in cell membranes and signaling molecules involved in the regulation of crucial cell functions, including cell growth, differentiation, proliferation and apoptosis. Recent research has shown that abnormal sphingolipid metabolism is associated with genetic and metabolic disease processes. Given their crucial role to maintain homeostasis within the body, sphingolipids have been investigated as potential biomarkers to predict disease in the population. Here we discuss how sphingolipids levels are altered in different diseases, thus illustrating their possible use as diagnostic and prognostic biomarkers for disease.
Assuntos
Biomarcadores , Transdução de Sinais , Esfingolipídeos , Ciclo Celular , Membrana Celular , Homeostase , HumanosRESUMO
Fibulin-1 (FBLN1) is a member of a growing family of extracellular matrix glycoproteins that includes eight members and is involved in cellular functions such as adhesion, migration, and differentiation. FBLN1 has also been implicated in embryonic heart and valve development and in the formation of neural crest-derived structures, including aortic arch, thymus, and cranial nerves. Fibroblast growth factor 8 (FGF8) is a member of a large family of growth factors, and its functions include neural crest cell (NCC) maintenance, specifically NCC migration as well as patterning of structures formed from NCC such as outflow tract and cranial nerves. In this report, we sought to investigate whether FBLN1 and FGF8 have cooperative roles in vivo given their influence on the development of the same NCC-derived structures. Surface plasmon resonance binding data showed that FBLN1 binds tightly to FGF8 and prevents its enzymatic degradation by ADAM17. Moreover, overexpression of FBLN1 up-regulates FGF8 gene expression, and down-regulation of FBLN1 by siRNA inhibits FGF8 expression. The generation of a double mutant Fbln1 and Fgf8 mice (Fbln1(-/-) and Fgf8(-/-)) showed that haplo-insufficiency (Fbln1(+/-) and Fgf8(+/-)) resulted in increased embryonic mortality compared with single heterozygote crosses. The mortality of the FGF8/Fbln1 double heterozygote embryos occurred between 14.5 and 16.5 days post-coitus. In conclusion, FBLN1/FGF8 interaction plays a role in survival of vertebrate embryos, and reduced levels of both proteins resulted in added mortality in utero The FBLN1/FGF8 interaction may also be involved in the survival of neural crest cell population during development.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Embrião de Mamíferos/metabolismo , Fator 8 de Crescimento de Fibroblasto/metabolismo , Crista Neural/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Perda do Embrião/genética , Perda do Embrião/metabolismo , Feminino , Fator 8 de Crescimento de Fibroblasto/genética , Humanos , Camundongos , Camundongos Knockout , Células NIH 3T3 , Gravidez , Ressonância de Plasmônio de SuperfícieRESUMO
BACKGROUND: Self-report measures indicate that Yoga practices are perceived to reduce stress; however, molecular mechanisms through which YB affects stress are just beginning to be understood. While invasive sampling such as blood has been widely used to measure biological indicators such as pro-inflammatory biomarkers, the use of saliva to measure changes in various biomolecules has been increasingly recognized. As Yoga practice stimulates salivary secretion, and saliva is considered a source of biomarkers, changes in salivary cytokines before and after Yogic breathing exercise as specified in an ancient Tamil script, Thirumanthiram, were examined using a Cytokine Multiplex to compare to Attention Control (AC) group. METHODS: Twenty healthy volunteers were randomized into two groups stratified by gender (N = 10 per YB and AC groups); The YB group performed two YB exercises, each for ten minutes, for a total of twenty minutes in a single session as directed by a trained Yoga instructor. The AC group read a text of their choice for 20 min. Saliva was collected immediately after YB training at 0, 5, 10, 15 and 20 min and analyzed by Multiplex enzyme linked immunosorbent assay (ELISA). RESULTS: The levels of interleukin (IL)-1ß, IL-8, and monocyte chemotactic protein -1 (MCP-1) were significantly reduced in YB group when compared to AC group. The level of reduction of IL-8 was significant at all time points tested, whereas IL-1ß showed reduction at 15 and 20 min time points (p < 0.05), and MCP-1 level was marginally different at 5-20 min. There were no significant differences between YB and AC groups in the salivary levels of IL-1RA, IL-6, IL-10, IL-17, IP-10, MIP-1b, and TNF-α. CONCLUSIONS: These data are the first to demonstrate the feasibility of detecting salivary cytokines using multiplex assay in response to a Yoga practice. This study was registered in Clinical Trials.gov # NCT02108769.
Assuntos
Biomarcadores/análise , Exercícios Respiratórios , Inflamação/metabolismo , Meditação , Saliva/química , Yoga , Biomarcadores/metabolismo , Citocinas/análise , Citocinas/metabolismo , Feminino , Humanos , Masculino , Projetos PilotoRESUMO
We sought to examine the relationship between elevated transferrin saturation (TS) and measures of health status (telomere length and patient-reported health-related quality of life) to assess whether elevated TS is associated with negative patient outcomes beyond increased risk for morbidity and mortality, using a cross-sectional analysis of the Hemochromatosis and Iron Overload Screening Study supplemented with assays for leukocyte telomere length in adults ≥25 years old (n = 669). Among individuals with elevated TS (≥45 % for women and ≥50 % for men), who also had a usual source of care, only 5.2 % reported ever being told by a doctor that they had an elevated iron condition. In a fully adjusted general linear regression model controlling for demographic characteristics as well as health conditions associated with iron overload, elevated TS versus non-elevated TS was associated with worse general health status (60.4 vs. 63.8, P < 0.05), mental health status (76.5 vs. 82.2, P < 0.0001) and shorter telomere length (241.4 vs. 261.3, P < 0.05). Increased surveillance of elevated TS may be in order as elevated TS is associated with decreased health status and very few patients with elevated TS are aware of their condition.
Assuntos
Qualidade de Vida , Telômero/metabolismo , Transferrina/análise , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Telômero/genética , Transferrina/metabolismoRESUMO
Elevated body iron stores are associated with morbidity and mortality due to oxidative stress. Hereditary hemochromatosis, a common condition caused by HFE gene mutations, can lead to excess iron storage and disease but clinical penetrance of HFE gene mutations is low and many people with elevated iron stores lack HFE mutations. We analyzed data from the Hemochromatosis and Iron Overload Screening Study to assess the relationship among HFE genotype (individuals with either homozygous or compound heterozygous status for C282Y and/or H63D HFE mutations were defined as genotype positive, or G+), elevated iron phenotype (individuals exceeding gender-specific transferrin saturation and serum ferritin threshold levels were considered phenotype positive, or P+), and leukocyte telomere length, a marker of biological aging and cumulative oxidative stress. In unadjusted analyses in comparison to individuals who were G-P-, G+P- were not significantly different (OR 0.74; 95% CI 0.26-2.04), while the G+P+ (OR 2.03; 95% CI 1.15-3.56), and G-P+ (OR 2.24; 95% CI 1.5-3.29) had increased risk of short telomeres (<=25th percentile) rather than long telomeres (>=75th percentile). In analyses adjusting for age, gender, and race/ethnicity, the effect of individuals with elevated iron phenotypes having short telomeres persisted with G+P+ individuals (OR 1.94; 95% CI 1.02-3.72), and G-P+ individuals (OR 2.17; 95% CI 1.39-3.39) being significantly different from the G-P- group. In conclusion, elevated iron phenotype, but not HFE genotype, was associated with shortened telomeres. Further studies will be needed to determine whether telomere length provides a marker for morbidities specifically associated with iron overload.
Assuntos
Hemocromatose/genética , Antígenos de Histocompatibilidade Classe I/genética , Ferro/metabolismo , Proteínas de Membrana/genética , Telômero/ultraestrutura , Adulto , Feminino , Genótipo , Hemocromatose/sangue , Proteína da Hemocromatose , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Ferro/sangue , Sobrecarga de Ferro/genética , Sobrecarga de Ferro/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Mutação , Fenótipo , Telômero/química , Telômero/metabolismoRESUMO
BACKGROUND: Trabeculation is an integral component of cardiac ventricular morphogenesis and is dependent on the matrix metalloproteinase, ADAMTS1. A substrate of ADAMTS1 is the proteoglycan versican which is expressed in the developing ventricle and which has been implicated in trabeculation. Fibulin-1 is a versican and ADAMTS1-binding extracellular matrix protein required for ventricular morphogenesis. Here we investigated the involvement of fibulin-1 in ADAMTS1-mediated cleavage of versican in vitro, and the involvement of fibulin-1 in versican cleavage in ventricular morphogenesis. RESULTS: We show that fibulin-1 is a cofactor for ADAMTS1-dependent in vitro cleavage of versican V1, yielding a 70-kDa amino-terminal fragment. Furthermore, fibulin-1-deficiency in mice was found to cause a significant reduction (>90%) in ventricular levels of the 70-kDa versican V1 cleavage product and a 2-fold increase in trabecular cardiomyocyte proliferation. Decreased versican V1 cleavage and augmented trabecular cardiomyocyte proliferation in fibulin-1 null hearts is accompanied by increased ventricular activation of ErbB2 and Erk1/2. By contrast, versican deficiency was found to lead to decreased cardiomyocyte proliferation and reduced ventricular trabeculation. CONCLUSION: We conclude that fibulin-1 regulates versican-dependent events in ventricular morphogenesis by promoting ADAMTS1 cleavage of versican leading to suppression of trabecular cardiomyocyte proliferation mediated by the ErbB2-Map kinase pathway.
Assuntos
Proteínas ADAM/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proliferação de Células , Ventrículos do Coração/embriologia , Morfogênese , Miócitos Cardíacos/fisiologia , Proteína ADAMTS1 , Animais , Proteínas de Ligação ao Cálcio/genética , Ventrículos do Coração/citologia , Ventrículos do Coração/metabolismo , Camundongos , Camundongos Mutantes , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Receptor ErbB-2/metabolismoRESUMO
Familial hypercholesterolemia (FH) patients suffer from excessively high levels of Low Density Lipoprotein Cholesterol (LDL-C), which can cause severe cardiovascular disease. Statins, bile acid sequestrants, PCSK9 inhibitors, and cholesterol absorption inhibitors are all inefficient at treating FH patients with homozygous LDLR gene mutations (hoFH). Drugs approved for hoFH treatment control lipoprotein production by regulating steady-state Apolipoprotein B (apoB) levels. Unfortunately, these drugs have side effects including accumulation of liver triglycerides, hepatic steatosis, and elevated liver enzyme levels. To identify safer compounds, we used an iPSC-derived hepatocyte platform to screen a structurally representative set of 10,000 small molecules from a proprietary library of 130,000 compounds. The screen revealed molecules that could reduce the secretion of apoB from cultured hepatocytes and from humanized livers in mice. These small molecules are highly effective, do not cause abnormal lipid accumulation, and share a chemical structure that is distinct from any known cholesterol lowering drug.
Assuntos
Anticolesterolemiantes , Hipercolesterolemia Familiar Homozigota , Hiperlipoproteinemia Tipo II , Células-Tronco Pluripotentes Induzidas , Humanos , Animais , Camundongos , Pró-Proteína Convertase 9/genética , Pró-Proteína Convertase 9/farmacologia , Pró-Proteína Convertase 9/uso terapêutico , LDL-Colesterol , Hiperlipoproteinemia Tipo II/tratamento farmacológico , Hiperlipoproteinemia Tipo II/genética , Anticolesterolemiantes/farmacologia , Apolipoproteínas B/genética , Apolipoproteínas B/farmacologia , Apolipoproteínas B/uso terapêutico , HepatócitosRESUMO
Systemic lupus erythematosus (SLE) patients display impaired endothelial nitric oxide synthase (eNOS) function required for normal vasodilatation. SLE patients express increased compensatory activity of inducible nitric oxide synthase (iNOS) generating excess nitric oxide that may result in inflammation. We examined the effects of genetic deletion of NOS2 and NOS3, encoding iNOS and eNOS respectively, on accelerated vascular disease in MRL/lpr lupus mouse model. NOS2 and NOS3 knockout (KO) MRL/lpr mice had higher plasma levels of triglycerides (23% and 35%, respectively), ceramide (45% and 21%, respectively), and sphingosine 1-phosphate (S1P) (21%) compared to counterpart MRL/lpr controls. Plasma levels of the anti-inflammatory cytokine interleukin 10 (IL-10) in NOS2 and NOS3 KO MRL/lpr mice were lower (53% and 80%, respectively) than counterpart controls. Nodule-like lesions in the adventitia were detected in aortas from both NOS2 and NOS3 KO MRL/lpr mice. Immunohistochemical evaluation of the lesions revealed activated endothelial cells and lipid-laden macrophages (foam cells), elevated sphingosine kinase 1 expression, and oxidized low-density lipoprotein immune complexes (oxLDL-IC). The findings suggest that advanced vascular disease in NOS2 and NOS3 KO MRL/lpr mice maybe mediated by increased plasma triglycerides, ceramide and S1P; decreased plasma IL-10; and accumulation of oxLDL-IC in the vessel wall. The results expose possible new targets to mitigate lupus-associated complications.
Assuntos
Aorta/imunologia , Lipoproteínas LDL/imunologia , Lúpus Eritematoso Sistêmico/enzimologia , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Esfingolipídeos/sangue , Animais , Aorta/enzimologia , Aorta/patologia , Lipoproteínas LDL/metabolismo , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/deficiência , Óxido Nítrico Sintase Tipo III/deficiênciaRESUMO
Heat shock proteins (HSPs) have been implicated in the activation and survival of macrophages. This study examined the role of HSP70B', a poorly characterized member of the HSP70 family, in response to oxidatively modified LDL (oxLDL) and immune complexes prepared with human oxLDL and purified human antibodies to oxLDL (oxLDL-IC) in monocytic and macrophage cell lines. Immunoblot analysis of cell lysates and conditioned medium from U937 cells treated with oxLDL alone revealed an increase in intracellular HSP70B' protein levels accompanied by a concomitant increase in HSP70B' extracellular levels. Fluorescence immunohistochemistry and confocal microscopy, however, demonstrated that oxLDL-IC stimulated the release of HSP70B', which co-localized with cell-associated oxLDL-IC. In HSP70B'-green fluorescent protein-transfected mouse RAW 264.7 cells, oxLDL-IC-induced HSP70B' co-localized with membrane-associated oxLDL-IC as well as the lipid moiety of internalized oxLDL-IC. Furthermore, the data demonstrated that HSP70B' is involved in cell survival, and this effect could be mediated by sphingosine kinase 1 (SK1) activation. An examination of regularly implicated cytokines revealed a significant relationship between HSP70B' and the release of the anti-inflammatory cytokine interleukin-10 (IL-10). Small interfering RNA knockdown of HSP70B' resulted in a corresponding decrease in SK1 mRNA levels and SK1 phosphorylation as well as increased release of IL-10. In conclusion, these findings suggest that oxLDL-IC induce the synthesis and release of HSP70B', and once stimulated, HSP70B' binds to the cell-associated and internalized lipid moiety of oxLDL-IC. The data also implicate HSP70B' in key cellular functions, such as regulation of SK1 activity and release of IL-10, which influence macrophage activation and survival.
Assuntos
Complexo Antígeno-Anticorpo/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Lipoproteínas LDL/metabolismo , Ativação de Macrófagos/fisiologia , Macrófagos/metabolismo , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Complexo Antígeno-Anticorpo/genética , Complexo Antígeno-Anticorpo/imunologia , Sobrevivência Celular , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/imunologia , Humanos , Interleucina-10/biossíntese , Interleucina-10/genética , Interleucina-10/imunologia , Lipoproteínas LDL/genética , Lipoproteínas LDL/imunologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Fosfotransferases (Aceptor do Grupo Álcool)/biossíntese , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/imunologia , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/genética , Transporte Proteico/imunologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Mensageiro/imunologia , RNA Interferente Pequeno , Células U937RESUMO
BACKGROUND: Extracellular matrix alterations are important elements in the arterial changes seen in diabetes, being associated with increased vascular stiffness and the development of cardiovascular diseases. However, no biomarkers for diabetes-related arterial changes have been defined. METHODS: Mammary artery specimens from 17 men with type 2 diabetes and 18 nondiabetic individuals were used for microarray expression profiling, quantitative real-time PCR, immunoassay, and immunohistochemical analyses. A derived candidate marker, fibulin-1, which is an elastin-associated matrix molecule, was measured immunochemically in plasma from (a) 70 patients scheduled for vascular surgery, (b) 305 patients with type 2 diabetes examined with carotid ultrasonography and echocardiography, and (c) 308 patients with type 2 diabetes, followed for 15 years. RESULTS: The most upregulated transcript in nonatherosclerotic arterial tissue from patients with type 2 diabetes encoded the extracellular matrix protein, fibulin-1. Higher concentrations of fibulin-1-protein were present in artery extracts from patients with diabetes than extracts from individuals without diabetes, and increased fibulin-1 immunostaining was apparent around the external elastic lamina of diabetic arteries. Patients with diabetes displayed increased plasma concentrations of fibulin-1 (P = 0.006). Plasma fibulin-1 concentrations correlated with hemoglobin A(1c) (P < 0.001), arterial stiffness indices including pulse pressure (P < 0.001), and carotid compliance (P = 0.004), as well as plasma N-terminal pro-B-type natriuretic peptide concentrations (P < 0.001) and were predictive of 15-year mortality (P = 0.013). CONCLUSIONS: Fibulin-1 accumulates in the arterial wall and in plasma of patients with type 2 diabetes, and appears to be a factor associated with arterial extracellular matrix changes in type 2 diabetes.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Matriz Extracelular/metabolismo , Artéria Torácica Interna/metabolismo , Idoso , Biomarcadores/metabolismo , Proteínas de Ligação ao Cálcio/sangue , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/mortalidade , Espessura Intima-Media Carotídea , Estudos de Casos e Controles , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/mortalidade , Feminino , Seguimentos , Humanos , Imunoensaio , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Valor Preditivo dos Testes , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that affects females more than males, with African Americans developing more severe manifestation of the disease. SLE patients are at increased risk for cardiovascular disease (CVD), and SLE women 35-44 years old have 50 fold the incidence rate of CVD. Because SLE patients do not follow the typical age and gender pattern for CVD, but instead an accelerated disease course, the traditional biomarkers of elevated LDL and total cholesterol levels do not accurately assess their CVD risk. Recently, we have reported that African American SLE patients had higher ceramide, hexosylceramide, sphingosine and dihydrosphingosine 1-phosphate levels compared to their healthy controls, and those with atherosclerosis had higher sphingomyelin and sphingoid bases levels than those without (PLoS One. 2019; e0224496). In the current study, we sought to identify sphingolipid species that correlate with and pose the potential to predict atherosclerosis severity in African American SLE patients. Plasma samples from a group of African American predominantly female SLE patients with well-defined carotid atherosclerotic plaque burden were analyzed for sphingolipidomics using targeted mass spectroscopy. The data demonstrated that at baseline, plaque area and C3 values correlated inversely with most lactoceramide species. After one-year follow-up visit, values of the change of plaque area correlated positively with the lactoceramide species. There was no correlation between LDL-C concentrations and lactoceramide species. Taken together, lactocylcermide levels may have a 'predictive' value and sphingolipidomics have an added benefit to currently available tools in early diagnosis and prognosis of African American SLE patients with CVD.
Assuntos
Doenças das Artérias Carótidas/sangue , Lúpus Eritematoso Sistêmico/sangue , Esfingolipídeos/sangue , Adulto , Negro ou Afro-Americano , Doenças Assintomáticas , Biomarcadores/sangue , Doenças das Artérias Carótidas/diagnóstico por imagem , Doenças das Artérias Carótidas/etnologia , Estudos Transversais , Feminino , Humanos , Lipidômica , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/etnologia , Masculino , Pessoa de Meia-Idade , Placa Aterosclerótica , Valor Preditivo dos Testes , Prognóstico , Estudos Prospectivos , Fatores Raciais , Fatores de Tempo , UltrassonografiaRESUMO
Sphingolipids are bioactive lipids associated with cellular membranes and plasma lipoproteins, and their synthesis and degradation are tightly regulated. We have previously determined that low plasma concentrations of certain ceramide species predict the development of nephropathy in diabetes patients with normal albumin excretion rates at baseline. Herein, we tested the hypothesis that altering the sphingolipid content of circulating lipoproteins can alter the metabolic and signaling pathways in podocytes, whose dysfunction leads to an impairment of glomerular filtration. Cultured human podocytes were treated with lipoproteins from healthy subjects enriched in vitro with C16 ceramide, or D-erythro 2-hydroxy C16 ceramide, a ceramide naturally found in skin. The RNA-Seq data demonstrated differential expression of genes regulating sphingolipid metabolism, sphingolipid signaling, and mTOR signaling pathways. A multiplex analysis of mTOR signaling pathway intermediates showed that the majority (eight) of the pathway phosphorylated proteins measured (eleven) were significantly downregulated in response to C16 ceramide-enriched HDL2 compared to HDL2 alone and hydroxy ceramide-enriched HDL2. In contrast, C16 ceramide-enriched HDL3 upregulated the phosphorylation of four intermediates in the mTOR pathway. These findings highlight a possible role for lipoprotein-associated sphingolipids in regulating metabolic and signaling pathways in podocytes and could lead to novel therapeutic targets in glomerular kidney diseases.
Assuntos
Ceramidas/metabolismo , Lipoproteínas/farmacologia , Podócitos/metabolismo , Esfingolipídeos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Transcriptoma/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Isótopos de Carbono , Linhagem Celular , Ceramidas/genética , HDL-Colesterol/farmacologia , Adesões Focais/efeitos dos fármacos , Adesões Focais/genética , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Fosforilação , Podócitos/efeitos dos fármacos , RNA-Seq , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Esfingolipídeos/genética , Serina-Treonina Quinases TOR/genética , Transcriptoma/efeitos dos fármacosRESUMO
Here we report that mouse embryos homozygous for a gene trap insertion in the fibulin-1 (Fbln1) gene are deficient in Fbln1 and exhibit cardiac ventricular wall thinning and ventricular septal defects with double outlet right ventricle or overriding aorta. Fbln1 nulls also display anomalies of aortic arch arteries, hypoplasia of the thymus and thyroid, underdeveloped skull bones, malformations of cranial nerves and hemorrhagic blood vessels in the head and neck. The spectrum of malformations is consistent with Fbln1 influencing neural crest cell (NCC)-dependent development of these tissues. This is supported by evidence that Fbln1 expression is associated with streams of cranial NCCs migrating adjacent to rhombomeres 2-7 and that Fbln1-deficient embryos display patterning anomalies of NCCs forming cranial nerves IX and X, which derive from rhombomeres 6 and 7. Additionally, Fbln1-deficient embryos show increased apoptosis in areas populated by NCCs derived from rhombomeres 4, 6 and 7. Based on these findings, it is concluded that Fbln1 is required for the directed migration and survival of cranial NCCs contributing to the development of pharyngeal glands, craniofacial skeleton, cranial nerves, aortic arch arteries, cardiac outflow tract and cephalic blood vessels.
Assuntos
Proteínas de Ligação ao Cálcio/genética , Morfogênese/fisiologia , Crista Neural/fisiologia , Animais , Antígenos CD4/genética , Proteínas de Ligação ao Cálcio/deficiência , Circulação Cerebrovascular/genética , Mapeamento Cromossômico , Cruzamentos Genéticos , Retículo Endoplasmático/fisiologia , Coração Fetal/patologia , Coração Fetal/fisiologia , Genótipo , Ventrículos do Coração/embriologia , Ventrículos do Coração/patologia , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase , beta-Galactosidase/genéticaRESUMO
Fibulin-1 is a fibrinogen-binding blood protein and a component of many extracellular matrices (ECM) including those of blood vessels. In this study, the deposition patterns of fibulin-1 and fibrinogen were examined in human coronary artery atherosclerotic lesions excised by atherectomy from 20 patients. Fibulin-1 deposition was found to be closely overlapping with fibrinogen located within the atherosclerotic lesions and in regions containing fresh thrombi. Pronounced intracellular fibulin-1 immunostaining was apparent in lesion areas rich in macrophages and foam cells, although THP-1 macrophages and foam cells were found not to express fibulin-1. Strong ECM deposition of fibulin-1 was observed in acellular atheromatous and myxomatous regions. By contrast, fibulin-1 was present at relatively low levels in the ECM associated with smooth muscle cells within and outside of lesions and was not detected in sclerotic regions. These results reveal the pattern of fibulin-1 within human atherosclerotic lesions and highlight the potential for fibulin-1, perhaps derived from the blood and acting in conjunction with fibrinogen, to play a role in the etiology and cardiovascular disease progression, particularly with respect to thrombotic aspects of atherosclerosis.
Assuntos
Aterosclerose/metabolismo , Aterosclerose/patologia , Proteínas de Ligação ao Cálcio/metabolismo , Fibrinogênio/metabolismo , Adulto , Idoso , Especificidade de Anticorpos , Proteínas de Ligação ao Cálcio/imunologia , Células Cultivadas , Doença da Artéria Coronariana/metabolismo , Doença da Artéria Coronariana/patologia , Fibrinogênio/imunologia , Humanos , Imuno-Histoquímica , Pessoa de Meia-IdadeRESUMO
Systemic lupus erythematous (SLE) is a chronic multi-organ autoimmune disease. Genetic and environmental factors contribute to disease onset and severity. Sphingolipids are signaling molecules involved in regulating cell functions and have been associated with multiple genetic disease processes. African-Americans are more likely to suffer from SLE morbidity than Whites. The Medical University of South Carolina has banked plasma samples from a well-characterized lupus cohort that includes African-Americans and Whites. This study examined the influence of race on plasma sphingolipid profiles in SLE patients and association of sphingolipid levels with comorbid atherosclerosis and SLE disease activity. Mass spectrometry revealed that healthy African-Americans had higher sphingomyelin levels and lower lactosylcermide levels compared to healthy Whites. SLE patients, irrespective of race, had higher levels of ceramides, and sphingoid bases (sphingosine and dihydrosphingosine) and their phosphates compared to healthy subjects. Compared to African-American controls, African-American SLE patients had higher levels of ceramides, hexosylceramides, sphingosine and dihydrosphingosine 1-phosphate. Compared to White controls, White SLE patients exhibited higher levels of sphingoid bases and their phosphates, but lower ratios of C16:0 ceramide/sphingosine 1-phosphate and C24:1 ceramide/sphingosine 1-phosphate. White SLE patients with atherosclerosis exhibited lower levels of sphingoid bases compared to White SLE patients without atherosclerosis. In contrast, African-American SLE patients with atherosclerosis had higher levels of sphingoid bases and sphingomyelins compared to African-American SLE patients without atherosclerosis. Compared to White SLE patients with atherosclerosis, African-American SLE patients with atherosclerosis had higher levels of select sphingolipids. Plasma levels of sphingosine, C16:0 ceramide/sphingosine 1-phosphate ratio and C24:1 ceramide/sphingosine 1-phosphate ratio significantly correlated with SLEDAI in the African-American but not White SLE patients. The C16:0 ceramide/sphingosine 1-phosphate ratio in SLE patients, and levels of C18:1 and C26:1 lactosylcermides, C20:0 hexosylceramide, and sphingoid bases in SLE patients with atherosclerosis could be dependent on race. Further ethnic studies in SLE cohorts are necessary to verify use of sphingolipidomics as complementary diagnostic tool.
Assuntos
Doenças Cardiovasculares/sangue , Disparidades nos Níveis de Saúde , Lipidômica/estatística & dados numéricos , Lúpus Eritematoso Sistêmico/sangue , Esfingolipídeos/sangue , Adulto , Negro ou Afro-Americano/estatística & dados numéricos , Doenças Cardiovasculares/epidemiologia , Estudos de Casos e Controles , Comorbidade , Feminino , Humanos , Lúpus Eritematoso Sistêmico/epidemiologia , Espectrometria de Massas , Pessoa de Meia-Idade , População Branca/estatística & dados numéricos , Adulto JovemRESUMO
High aspect ratio zinc oxide nanowires (ZnONWs) have become one of the most important products in nanotechnology. The wide range applications of ZnONWs have heightened the need for evaluating the risks and biological consequences to these particles. In this study, we investigated inflammatory pathways activated by ZnONWs in cultured cells as well as the consequences of systemic exposure in mouse models. Confocal microscopy showed rapid phagocytic uptake of FITC-ZnONWs by macrophages. Exposure of macrophages or lung epithelial cells to ZnONWs induced the production of CCL2 and CCL11. Moreover, ZnONWs exposure induced both IL-6 and TNF-α production only in macrophages but not in LKR13 cells. Intratracheal instillation of ZnONWs in C57BL/6 mice induced a significant increase in the total numbers of immune cells in the broncho alveolar lavage fluid (BALFs) 2 days after instillation. Macrophages and eosinophils were the predominant cellular infiltrates of ZnONWs exposed mouse lungs. Similar cellular infiltrates were also observed in a mouse air-pouch model. Pro-inflammatory cytokines IL-6 and TNF-α as well as chemokines CCL11, and CCL2 were increased both in BALFs and air-pouch lavage fluids. These results suggest that exposure to ZnONWs may induce distinct inflammatory responses through phagocytic uptake and formation of soluble Zn2+ ions.
Assuntos
Quimiocina CCL11/imunologia , Eosinófilos/efeitos dos fármacos , Eosinófilos/imunologia , Inflamação/etiologia , Nanofios/efeitos adversos , Óxido de Zinco/efeitos adversos , Animais , Quimiocina CCL11/genética , Quimiocina CCL2/genética , Modelos Animais de Doenças , Técnicas In Vitro , Inflamação/genética , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Interleucina-6/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Nanofios/química , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Células RAW 264.7 , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Necrose Tumoral alfa/genética , Regulação para Cima/efeitos dos fármacos , Óxido de Zinco/químicaRESUMO
Aberrant activation of phosphatidylinosito-4,5-bisphosphate 3-kinase/protein kinase B (PI3K/AKT) signaling in cancer has led to pursuit of inhibitors for targeting this pathway. However, inhibitors of PI3K and AKT have failed to yield efficacious results without adverse effects. Here, we screened a library containing 441 authenticated traditional chinese medicine (TCM) plant extracts by examining their effect on cell viability of a human mammary epithelial cell line HMEC-PIK3CAH1047R, which expresses mutant PIK3CAH1047R and has constitutively active AKT signaling. We found that Oridonin, an extract from Rabdosia rubescens, reduced cell viability to the greatest extent. Oridonin binds to AKT1 and potentially functions as an ATP-competitive AKT inhibitor. Importantly, Oridonin selectively impaired tumor growth of human breast cancer cells with hyperactivation of PI3K/AKT signaling. Moreover, Oridonin prevented the initiation of mouse mammary tumors driven by PIK3CAH1047R. Our results suggest that Oridonin may serve as a potent and durable therapeutic agent for the treatment of breast cancers with hyperactivation of PI3K/AKT signaling.
RESUMO
Tissue engineering is a fast-evolving field of biomedical science and technology with future promise to manufacture living tissues and organs for replacement, repair, and regeneration of diseased organs. Owing to the specific role of hemodynamics in the development, maintenance, and functioning of the cardiovascular system, bioreactors are a fundamental of cardiovascular tissue engineering. The development of perfusion bioreactor technology for cardiovascular tissue engineering is a direct sequence of previous historic successes in extracorporeal circulation techniques. Bioreactors provide a fluidic environment for tissue engineered tissue and organs, and guarantee their viability, maturation, biomonitoring, testing, storage, and transportation. There are different types of bioreactors and they vary greatly in their size, complexity, and functional capabilities. Although progress in design and functional properties of perfusion bioreactors for tissue engineered blood vessels, heart valves, and myocardial patches is obvious, there are some challenges and insufficiently addressed issues, and room for bioreactor design improvement and performance optimization. These challenges include creating a triple perfusion bioreactor for vascularized tubular tissue engineered cardiac construct; designing and manufacturing fluidics-based perfused minibioreactors; incorporation of systematic mathematical modeling and computer simulation based on computational fluid dynamics into the bioreactor designing process; and development of automatic systems of hydrodynamic regime control. Designing and engineering of built-in noninvasive biomonitoring systems is another important challenge. The optimal and most efficient perfusion and conditioning regime, which accelerates tissue maturation of tissue-engineered constructs also remains to be determined. This is a first article in a series of reviews on critical elements of cardiovascular tissue engineering technology describing the current status, unsolved problems, and challenges of bioreactor technology in cardiovascular tissue engineering and outlining future trends and developments.
Assuntos
Reatores Biológicos , Procedimentos Cirúrgicos Cardiovasculares/instrumentação , Engenharia Tecidual/métodos , Desenho de Equipamento , Humanos , Perfusão , Engenharia Tecidual/instrumentaçãoRESUMO
The Myo1c motor functions as a cargo transporter supporting various cellular events, including vesicular trafficking, cell migration, and stereociliary movements of hair cells. Although its partial crystal structures were recently described, the structural details of its interaction with cargo proteins remain unknown. This study presents the first structural demonstration of a cargo protein, Neph1, attached to Myo1c, providing novel insights into the role of Myo1c in intracellular movements of this critical slit diaphragm protein. Using small angle X-ray scattering studies, models of predominant solution conformation of unliganded full-length Myo1c and Myo1c bound to Neph1 were constructed. The resulting structures show an extended S-shaped Myo1c with Neph1 attached to its C-terminal tail. Importantly, binding of Neph1 did not induce a significant shape change in Myo1c, indicating this as a spontaneous process or event. Analysis of interaction surfaces led to the identification of a critical residue in Neph1 involved in binding to Myo1c. Indeed, a point mutant from this site abolished interaction between Neph1 and Myo1c when tested in the in vitro and in live-cell binding assays. Live-cell imaging, including fluorescence recovery after photobleaching, provided further support for the role of Myo1c in intracellular vesicular movement of Neph1 and its turnover at the membrane.
Assuntos
Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Miosina Tipo I/química , Miosina Tipo I/metabolismo , Sítios de Ligação , Células Cultivadas , Humanos , Proteínas de Membrana/genética , Modelos Moleculares , Podócitos/metabolismo , Podócitos/ultraestrutura , Mutação Puntual , Ligação Proteica , Transporte Proteico , Espalhamento a Baixo Ângulo , Difração de Raios XRESUMO
Fibulin-1 (Fbln-1) is an immunogenic breast cancer-related glycoprotein identified by serological analysis of cDNA expression library (SEREX) strategy. Here, we show that dendritic cells from two breast cancer patients elicited a CD4(+)-mediated T-cell response to Fbln-1 presentation. In both patients, an antibody response to Fbln-1 was also found. By contrast, a Fbln-1-seronegative patient and a weakly seropositive patient demonstrated no such T-cell response. Analysis of human breast cancers for Fbln-1 RNA and protein expression revealed the presence of Fbln-1C and -1D variants. Fbln-1 was detected in the cytoplasm and at the cell surface of different human breast carcinoma cell lines. Immunohistochemical analysis of 528 archival primary breast carcinomas showed the expression of Fbln-1 in 35% of the cases. When the immunohistochemical findings were compared against pathobiological information associated with each specimen, an inverse relationship between Fbln-1 and cathepsin D expression was observed (P=0.04). Furthermore, even though long-term survival was similar between Fbln-1-positive and -negative cases, the survival of Fbln-1-positive cases improved when a lymphoid infiltrate was present at the tumour site. Taken together, our findings of an Fbln-1-specific immunity and the improved survival associated with Fbln-1 expression in the presence of lymphoid infiltration point to a role of Fbln-1 in tumour immunosurveillance.