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1.
Nutr Metab Cardiovasc Dis ; 31(4): 1148-1155, 2021 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-33618923

RESUMO

BACKGROUND AND AIMS: Iron deficiency is prevalent, but there is limited data about the relationship between iron status and poor outcomes in chronic kidney disease patients undergoing peritoneal dialysis (PD). We aimed to investigate the association between iron status and mortality in PD patients. METHODS AND RESULTS: This retrospective study was conducted on incident PD patients from January 2006 to December 2016 and followed up until December 2018. Patients were categorized into four groups according to baseline serum transferrin saturation (percent) and ferritin levels (ng/ml): reference (20-30%, 100-500 ng/ml), absolute iron deficiency (<20%, <100 ng/ml), function iron deficiency (FID) (<20%, >100 ng/ml), and high iron (>30%, >500 ng/ml). Among the 1173 patients, 77.5% had iron deficiency. During a median follow-up period of 43.7 months, compared with the reference group, the FID group was associated with increased risk for all-cause [adjusted hazard ratio (aHR) 1.87, 95% confidence interval (95% CI) 1.05-3.31, P = 0.032], but not cardiovascular (CV) mortality. Additionally, the high iron group had a more than four-fold increased risk of both all-cause and CV mortality [aHR 4.32 (95% CI 1.90-9.81), P < 0.001; aHR 4.41 (95% CI 1.47-13.27), P = 0.008; respectively]. CONCLUSION: FID and high iron predict worse prognosis of patients on PD.


Assuntos
Distúrbios do Metabolismo do Ferro/sangue , Ferro/sangue , Nefropatias/terapia , Diálise Peritoneal/mortalidade , Adulto , Biomarcadores/sangue , China/epidemiologia , Feminino , Ferritinas/sangue , Humanos , Ferro/deficiência , Distúrbios do Metabolismo do Ferro/diagnóstico , Distúrbios do Metabolismo do Ferro/mortalidade , Nefropatias/sangue , Nefropatias/diagnóstico , Nefropatias/mortalidade , Masculino , Pessoa de Meia-Idade , Diálise Peritoneal/efeitos adversos , Prevalência , Estudos Retrospectivos , Medição de Risco , Fatores de Risco , Fatores de Tempo , Transferrina/metabolismo , Resultado do Tratamento
2.
Int J Nanomedicine ; 16: 283-296, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33469287

RESUMO

Methods: In this study, we used MTT assays to demonstrate that a combination of SPIO-Serum and wild-type p53 overexpression can reduce ovarian cancer cell viability in vitro. Prussian blue staining and iron assays were used to determine changes in intracellular iron concentration following SPIO-Serum treatment. TEM was used to evaluate any mitochondrial damage induced by SPIO-Serum treatment, and Western blot was used to evaluate the expression of the iron transporter and lipid peroxidation regulator proteins. JC-1 was used to measure mitochondrial membrane potential, and ROS levels were estimated by flow cytometry. Finally, xCT protein expression and mitochondrial ROS levels were confirmed using fluorescence microscopy. Results: SPIO-Serum effectively induced lipid peroxidation and generated abundant toxic ROS. It also facilitated the downregulation of GPX4 and xCT, ultimately resulting in iron-dependent oxidative death. These effects could be reversed by iron chelator DFO and lipid peroxidation inhibitor Fer-1. SPIO-Serum treatment disrupted intracellular iron homeostasis by regulating iron uptake and the cells presented with missing mitochondrial cristae and ruptured outer mitochondrial membranes. Moreover, we were able to show that p53 contributed to SPIO-Serum-induced ferroptosis in ovarian cancer cells. Conclusion: SPIO-Serum induced ferroptosis and overexpressed p53 contributed to ferroptosis in ovarian cancer cells. Our data provide a theoretical basis for ferroptosis as a novel cell death phenotype induced by nanomaterials.


Assuntos
Ferroptose , Neoplasias Ovarianas/patologia , Soro/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Feminino , Humanos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Modelos Biológicos , Neoplasias Ovarianas/metabolismo , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Receptores da Transferrina/metabolismo , Transferrina/metabolismo
3.
Int J Mol Sci ; 22(2)2021 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-33467196

RESUMO

In ß-thalassemia, ineffective erythropoiesis leads to anemia and systemic iron overload. The management of iron overload by chelation therapy is a standard of care. However, iron chelation does not improve the ineffective erythropoiesis. We recently showed that the oral ferroportin inhibitor VIT-2763 ameliorates anemia and erythropoiesis in the Hbbth3/+ mouse model of ß-thalassemia. In this study, we investigated whether concurrent use of the iron chelator deferasirox (DFX) and the ferroportin inhibitor VIT-2763 causes any pharmacodynamic interactions in the Hbbth3/+ mouse model of ß-thalassemia. Mice were treated with VIT-2763 or DFX alone or with the combination of both drugs once daily for three weeks. VIT-2763 alone or in combination with DFX improved anemia and erythropoiesis. VIT-2763 alone decreased serum iron and transferrin saturation (TSAT) but was not able to reduce the liver iron concentration. While DFX alone had no effect on TSAT and erythropoiesis, it significantly reduced the liver iron concentration alone and in the presence of VIT-2763. Our results clearly show that VIT-2763 does not interfere with the iron chelation efficacy of DFX. Furthermore, VIT-2763 retains its beneficial effects on improving ineffective erythropoiesis when combined with DFX in the Hbbth3/+ mouse model. In conclusion, co-administration of the oral ferroportin inhibitor VIT-2763 and the iron chelator DFX is feasible and might offer an opportunity to improve both ineffective erythropoiesis and iron overload in ß-thalassemia.


Assuntos
Benzimidazóis/farmacologia , Eritropoese/efeitos dos fármacos , Oxazóis/farmacologia , Piridinas/farmacologia , Talassemia beta/tratamento farmacológico , Administração Oral , Animais , Benzimidazóis/administração & dosagem , Benzimidazóis/efeitos adversos , Benzimidazóis/uso terapêutico , Proteínas de Transporte de Cátions/antagonistas & inibidores , Células Cultivadas , Deferasirox/administração & dosagem , Deferasirox/farmacologia , Deferasirox/uso terapêutico , Combinação de Medicamentos , Interações Medicamentosas , Feminino , Ferro/sangue , Quelantes de Ferro/administração & dosagem , Quelantes de Ferro/farmacologia , Quelantes de Ferro/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxazóis/administração & dosagem , Oxazóis/efeitos adversos , Oxazóis/uso terapêutico , Piridinas/administração & dosagem , Piridinas/efeitos adversos , Piridinas/uso terapêutico , Transferrina/metabolismo
4.
Diabetes Res Clin Pract ; 172: 108653, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33422582

RESUMO

AIMS: The relationship between iron status, obesity and type 2 diabetes mellitus (T2DM) has scarcely been tested. This study hypothesizes that patients with obesity and T2DM have altered iron metabolism. METHODS: 537 T2DM patients were selected from the cross-sectional DICARIVA study excluding patients with high-sensitivity-C-reactive-protein (hs-CRP) ≥  10 mg/L. Three groups according to body mass index (BMI) and waist perimeter (WP) were analysed: a) BMI < 30 kg/m2, non-high WP (n = 105); b) BMI < 30 kg/m2, high WP (n = 202); and c) diabesity, BMI ≥  30 kg/m2, high WP (n = 230). Group differences on cardiometabolic and iron status markers were tested. RESULTS: Women had significantly lower iron, ferritin, and transferrin saturation (TSAT) but higher transferrin and total iron binding capacity than men. Triglycerides/HDL-c ratio, as insulin-resistance (IR) marker, was higher in men while hs-CRP in women. TSAT was inversely related to BMI and hs-CRP. The diabesity group showed the highest hs-CRP (p < 0.001) and IR (p < 0.001) with the lowest TSAT (p = 0.003). CONCLUSIONS: Low TSAT was highly prevalent in diabesity, mainly in women, suggesting that IR, inflammation, and abdominal adiposity alter iron transport and accumulation. The convenience of iron supplementation in diabesity patients with low TSAT should be urgently assessed, due the pro-oxidant effects of excess iron.


Assuntos
Diabetes Mellitus Tipo 2/sangue , Ferro/metabolismo , Obesidade/complicações , Transferrina/metabolismo , Idoso , Estudos Transversais , Feminino , Humanos , Masculino
5.
Anal Chem ; 93(4): 1944-1950, 2021 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-33399445

RESUMO

Carboxyl-group specific chemical cross-linking is gaining an increased interest as a structural mass spectrometry/structural proteomics technique that is complementary to the more commonly used amine-specific chemistry using succinimide esters. One of these protocols uses a combination of dihydrazide linkers and the coupling reagent DMTMM [4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium] chloride, which allows performing the reaction at neutral pH. The reaction yields two types of products, carboxyl-carboxyl cross-links that incorporate the dihydrazide linker and zero-length carboxyl-amine cross-links induced by DMTMM alone. Until now, it has not been systematically investigated how the balance between the two products is affected by experimental conditions. Here, we studied the role of the ratios of the two reagents (using pimelic dihydrazide and DMTMM) and demonstrate that the concentration of the two reagents can be systematically adjusted to favor one reaction product over the other. Using a set of five model proteins, we observed that the number of identified cross-linked peptides could be more than doubled by a combination of three different reaction conditions. We also applied this strategy to the bovine 20S proteasome and the Escherichia coli 70S ribosome, again demonstrating complementarity and increased cross-link coverage.


Assuntos
Reagentes para Ligações Cruzadas/química , Proteínas/química , Proteômica , Animais , Catalase/química , Catalase/metabolismo , Conalbumina/química , Conalbumina/metabolismo , Creatina Quinase/química , Creatina Quinase/metabolismo , Espectrometria de Massas/métodos , Proteínas/metabolismo , Albumina Sérica/química , Albumina Sérica/metabolismo , Transferrina/química , Transferrina/metabolismo
6.
Int J Biol Macromol ; 172: 589-596, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33454336

RESUMO

Agarose native gel electrophoresis has been developed to separate proteins and protein complexes in the native state. Here, we applied this technology to analyze proteins that undergo degradation, post-translational modification or chemical/physical changes. Antibodies showed aggregation/association upon acid or heat treatment. Limited reduction of disulfide bonds resulted in non-covalent aggregation of bovine serum albumin and cleavage of only inter-chain linkages of an antibody that had no effects on its overall structure. Native agarose gel analysis showed changes in mobility of human transferrin upon Fe3+ binding. Analysis of a commercial glycated human hemoglobin A1c showed no difference in electrophoretic pattern from un-modified hemoglobin. Native agarose gel showed aggregation of a virus upon acid or heat treatment. We have extracted bands of bovine serum albumin from the agarose native gel for sodium dodecylsulfate gel electrophoresis analysis, showing degradation of aged sample. Lastly, we analyzed phosphorylation of Zap70 kinase by native gel and Western blotting. These applications should expand the utility of this native gel electrophoresis technology.


Assuntos
Eletroforese em Gel de Poliacrilamida/métodos , Hemoglobina A Glicada/metabolismo , Processamento de Proteína Pós-Traducional , Soroalbumina Bovina/metabolismo , Transferrina/metabolismo , Proteína-Tirosina Quinase ZAP-70/metabolismo , Animais , Bovinos , Dependovirus/genética , Dependovirus/metabolismo , Hemoglobina A Glicada/genética , Humanos , Peróxido de Hidrogênio/farmacologia , Fosforilação , Agregados Proteicos , Desnaturação Proteica , Proteólise , Soroalbumina Bovina/genética , Dodecilsulfato de Sódio/química , Transferrina/genética , Proteínas Virais/genética , Proteínas Virais/metabolismo , Proteína-Tirosina Quinase ZAP-70/genética
7.
Postepy Biochem ; 66(3): 213-228, 2020 09 30.
Artigo em Polonês | MEDLINE | ID: mdl-33315314

RESUMO

Congenital disorders of glycosylation (CDG) are a group of genetic disorders caused by abnormal N- and O-glycosylation pathway of proteins and lipids. The glycosylation process plays an important role in the proper functioning of the body and its disorder leads to serious clinical defects. The clinical picture is extremely heterogeneous, including symptoms involving many organs or systems with predominantly neurological manifestation.A broad clinical phenotype poses a challenge in CDG diagnosis. A large group among CDG are defects associated with protein N-hypoglycosylation. A simple test its diagnosis is isoelectrofocusing (IEF) of serum transferrin which is still the "gold standard" in the diagnostics. Normal isoform transferrin profile does not rule out all glycosylation defects. Molecular diagnostics play an important role and the dissemination of next generation sequencing (NGS) has allowed new disorders to be identified.


Assuntos
Defeitos Congênitos da Glicosilação , Defeitos Congênitos da Glicosilação/diagnóstico , Glicosilação , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Patologia Molecular , Isoformas de Proteínas , Transferrina/metabolismo
8.
Eur J Epidemiol ; 35(8): 763-773, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32816244

RESUMO

Iron metabolism and anemia may play an important role in multiple organ dysfunction syndrome in Coronavirus disease 2019 (COVID-19). We conducted a systematic review and meta-analysis to evaluate biomarkers of anemia and iron metabolism (hemoglobin, ferritin, transferrin, soluble transferrin receptor, hepcidin, haptoglobin, unsaturated iron-binding capacity, erythropoietin, free erythrocyte protoporphyrine, and erythrocyte indices) in patients diagnosed with COVID-19, and explored their prognostic value. Six bibliographic databases were searched up to August 3rd 2020. We included 189 unique studies, with data from 57,563 COVID-19 patients. Pooled mean hemoglobin and ferritin levels in COVID-19 patients across all ages were 129.7 g/L (95% Confidence Interval (CI), 128.51; 130.88) and 777.33 ng/mL (95% CI, 701.33; 852.77), respectively. Hemoglobin levels were lower with older age, higher percentage of subjects with diabetes, hypertension and overall comorbidities, and admitted to intensive care. Ferritin level increased with older age, increasing proportion of hypertensive study participants, and increasing proportion of mortality. Compared to moderate cases, severe COVID-19 cases had lower hemoglobin [weighted mean difference (WMD), - 4.08 g/L (95% CI - 5.12; - 3.05)] and red blood cell count [WMD, - 0.16 × 1012 /L (95% CI - 0.31; - 0.014)], and higher ferritin [WMD, - 473.25 ng/mL (95% CI 382.52; 563.98)] and red cell distribution width [WMD, 1.82% (95% CI 0.10; 3.55)]. A significant difference in mean ferritin levels of 606.37 ng/mL (95% CI 461.86; 750.88) was found between survivors and non-survivors, but not in hemoglobin levels. Future studies should explore the impact of iron metabolism and anemia in the pathophysiology, prognosis, and treatment of COVID-19.


Assuntos
Anemia/diagnóstico , Infecções por Coronavirus , Coronavirus/metabolismo , Ferro/metabolismo , Pandemias , Pneumonia Viral , Betacoronavirus , Biomarcadores/análise , Biomarcadores/sangue , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Eritropoetina , Ferritinas/sangue , Hemoglobinas/análise , Hemoglobinas/metabolismo , Hepcidinas/sangue , Hepcidinas/metabolismo , Humanos , Ferro/sangue , Pneumonia Viral/epidemiologia , Receptores da Transferrina/sangue , Transferrina/análise , Transferrina/metabolismo
9.
PLoS One ; 15(8): e0235551, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833964

RESUMO

VPS34 is a key regulator of endomembrane dynamics and cargo trafficking, and is essential in cultured cell lines and in mice. To better characterize the role of VPS34 in cell growth, we performed unbiased cell line profiling studies with the selective VPS34 inhibitor PIK-III and identified RKO as a VPS34-dependent cellular model. Pooled CRISPR screen in the presence of PIK-III revealed endolysosomal genes as genetic suppressors. Dissecting VPS34-dependent alterations with transcriptional profiling, we found the induction of hypoxia response and cholesterol biosynthesis as key signatures. Mechanistically, acute VPS34 inhibition enhanced lysosomal degradation of transferrin and low-density lipoprotein receptors leading to impaired iron and cholesterol uptake. Excess soluble iron, but not cholesterol, was sufficient to partially rescue the effects of VPS34 inhibition on mitochondrial respiration and cell growth, indicating that iron limitation is the primary driver of VPS34-dependency in RKO cells. Loss of RAB7A, an endolysosomal marker and top suppressor in our genetic screen, blocked transferrin receptor degradation, restored iron homeostasis and reversed the growth defect as well as metabolic alterations due to VPS34 inhibition. Altogether, our findings suggest that impaired iron mobilization via the VPS34-RAB7A axis drive VPS34-dependence in certain cancer cells.


Assuntos
Classe III de Fosfatidilinositol 3-Quinases/metabolismo , Ferro/metabolismo , Neoplasias/metabolismo , Hipóxia Celular , Linhagem Celular Tumoral , Proliferação de Células , Colesterol/biossíntese , Colesterol/genética , Classe III de Fosfatidilinositol 3-Quinases/genética , Endossomos/metabolismo , Células HEK293 , Humanos , Lisossomos/metabolismo , Receptores de LDL/metabolismo , Transferrina/metabolismo , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
10.
Nature ; 583(7816): 425-430, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32612231

RESUMO

The vascular interface of the brain, known as the blood-brain barrier (BBB), is understood to maintain brain function in part via its low transcellular permeability1-3. Yet, recent studies have demonstrated that brain ageing is sensitive to circulatory proteins4,5. Thus, it is unclear whether permeability to individually injected exogenous tracers-as is standard in BBB studies-fully represents blood-to-brain transport. Here we label hundreds of proteins constituting the mouse blood plasma proteome, and upon their systemic administration, study the BBB with its physiological ligand. We find that plasma proteins readily permeate the healthy brain parenchyma, with transport maintained by BBB-specific transcriptional programmes. Unlike IgG antibody, plasma protein uptake diminishes in the aged brain, driven by an age-related shift in transport from ligand-specific receptor-mediated to non-specific caveolar transcytosis. This age-related shift occurs alongside a specific loss of pericyte coverage. Pharmacological inhibition of the age-upregulated phosphatase ALPL, a predicted negative regulator of transport, enhances brain uptake of therapeutically relevant transferrin, transferrin receptor antibody and plasma. These findings reveal the extent of physiological protein transcytosis to the healthy brain, a mechanism of widespread BBB dysfunction with age and a strategy for enhanced drug delivery.


Assuntos
Envelhecimento/metabolismo , Envelhecimento/patologia , Barreira Hematoencefálica/metabolismo , Transcitose , Fosfatase Alcalina/metabolismo , Animais , Anticorpos/metabolismo , Transporte Biológico , Proteínas Sanguíneas/administração & dosagem , Proteínas Sanguíneas/metabolismo , Proteínas Sanguíneas/farmacocinética , Encéfalo/irrigação sanguínea , Encéfalo/metabolismo , Sistemas de Liberação de Medicamentos , Saúde , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plasma/metabolismo , Proteoma/administração & dosagem , Proteoma/metabolismo , Proteoma/farmacocinética , Receptores da Transferrina/imunologia , Transcrição Genética , Transferrina/metabolismo
11.
Nat Commun ; 11(1): 2332, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393788

RESUMO

Fasting-mimicking diets delay tumor progression and sensitize a wide range of tumors to chemotherapy, but their therapeutic potential in combination with non-cytotoxic compounds is poorly understood. Here we show that vitamin C anticancer activity is limited by the up-regulation of the stress-inducible protein heme-oxygenase-1. The fasting-mimicking diet selectivity reverses vitamin C-induced up-regulation of heme-oxygenase-1 and ferritin in KRAS-mutant cancer cells, consequently increasing reactive iron, oxygen species, and cell death; an effect further potentiated by chemotherapy. In support of a potential role of ferritin in colorectal cancer progression, an analysis of The Cancer Genome Atlas Database indicates that KRAS mutated colorectal cancer patients with low intratumor ferritin mRNA levels display longer 3- and 5-year overall survival. Collectively, our data indicate that the combination of a fasting-mimicking diet and vitamin C represents a promising low toxicity intervention to be tested in randomized clinical trials against colorectal cancer and possibly other KRAS mutated tumors.


Assuntos
Ácido Ascórbico/farmacologia , Dieta , Jejum/fisiologia , Mutação/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Progressão da Doença , Heme Oxigenase-1/metabolismo , Humanos , Ferro/metabolismo , Camundongos Endogâmicos BALB C , Oxaliplatina/farmacologia , Fosforilação/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Análise de Sobrevida , Transferrina/metabolismo
12.
Diabetes Res Clin Pract ; 163: 108149, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32304796

RESUMO

AIMS: To assess the role of serum ferritin and transferrin with prevalent and incident type 2 diabetes mellitus (T2DM) and metabolic syndrome (MetS) and whether these associations are independent of inflammatory markers and hepatic enzymes. METHODS: We analyzed data from 3,232 participants aged 20-81 years of the population-based Study of Health in Pomerania (SHIP) from Northeast Germany with a median follow-up time of 10.6 years. Logistic and Cox regression analyses were performed. RESULTS: Serum ferritin concentrations were associated with a higher prevalence of T2DM (total population OR: 1.16 [95% CI: 1.07, 1.26]; men OR: 1.18 [95% CI: 1.08, 1.30) and MetS (total population OR: 1.27 [95% CI: 1.16, 1.38]; men OR: 1.26 [95% CI: 1.15, 1.38]) in the total population and men independently of inflammatory markers and hepatic enzymes. In longitudinal analyses, baseline ferritin concentrations were associated with a higher risk of incident T2DM in women (HR: 1.38 [95% CI: 1.10, 1.71]), but not in men or in the total population and also with a higher risk of incident MetS (HR: 1.09 [95% CI: 1.01, 1.17]) in the total population. These longitudinal associations attenuated considerably after adjustment for hepatic enzymes but not inflammatory markers. Transferrin was not associated with any of the outcomes. CONCLUSIONS: Our results suggest a link between ferritin and T2DM and MetS, which might be partially explained by hepatic dysfunction.


Assuntos
Biomarcadores/sangue , Diabetes Mellitus Tipo 2/sangue , Ferritinas/sangue , Ferro/metabolismo , Síndrome Metabólica/diagnóstico , Transferrina/metabolismo , Adulto , Feminino , Humanos , Masculino , Síndrome Metabólica/epidemiologia , Pessoa de Meia-Idade , Estudos Prospectivos
13.
Fluids Barriers CNS ; 17(1): 28, 2020 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-32295615

RESUMO

BACKGROUND: Iron is crucial for proper functioning of all organs including the brain. Deficiencies and excess of iron are common and contribute to substantial morbidity and mortality. Whereas iron's involvement in erythropoiesis drives clinical practice, the guidelines informing interventional strategies for iron repletion in neurological disorders are poorly defined. The objective of this study was to determine if peripheral iron status is communicated to the brain. METHODS: We used a bi-chamber cell culture model of the blood-brain-barrier to determine transcytosis of iron delivered by transferrin as a metric of iron transport. In the apical chamber (representative of the blood) we placed transferrin complexed with iron59 and in the basal chamber (representative of the brain) we placed human cerebrospinal fluid. Cerebrospinal fluid (CSF) samples (N = 24) were collected via lumbar puncture. The integrity of the tight junctions were monitored throughout the experiments using RITC-Dextran. RESULTS: We demonstrate that iron transport correlates positively with plasma hemoglobin concentrations but not serum ferritin levels. CONCLUSIONS: The clinical ramifications of these findings are several- fold. They suggest that erythropoietic demands for iron take precedence over brain requirements, and that the metric traditionally considered to be the most specific test reflecting total body iron stores and relied upon to inform treatment decisions-i.e., serum ferritin-may not be the preferred peripheral indicator when attempting to promote brain iron uptake. The future direction of this line of investigation is to identify the factor(s) in the CSF that influence iron transport at the level of the BBB.


Assuntos
Barreira Hematoencefálica/metabolismo , Líquido Cefalorraquidiano/metabolismo , Eritropoese/fisiologia , Ferritinas/metabolismo , Hemoglobinas , Ferro/metabolismo , Transdução de Sinais/fisiologia , Transferrina/metabolismo , Animais , Bovinos , Células Cultivadas , Ferritinas/sangue , Ferritinas/líquido cefalorraquidiano , Humanos , Ferro/sangue , Ferro/líquido cefalorraquidiano , Síndrome das Pernas Inquietas/terapia , Transferrina/líquido cefalorraquidiano
14.
PLoS One ; 15(4): e0231057, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32240239

RESUMO

OBJECTIVES: Though elevated ferritin level and decreased lung function both predispose people to cardio-metabolic disease, few reports have investigated the association between them. Furthermore, it remains unclear whether the association reflects a change in iron stores or an epiphenomenon reflecting metabolic stress. Therefore, we looked for possible associations between ferritin, iron, and transferrin saturation (TSAT) and lung function to clarify the role of iron-related parameters in healthy men. METHODS: We conducted a cohort study of 42,927 healthy Korean men (mean age: 38.6 years). Percent predicted forced expiratory volume in one second (FEV1%) and forced vital capacity (FVC%) were categorized into quartiles. Adjusted odds ratios (aORs) and 95% confidence intervals (using the highest quartile as reference) were calculated for hyperferritinemia, high iron, and high TSAT after controlling for potential confounders. RESULTS: The median ferritin level was 199.8 (141.5-275.6) ng/mL. The prevalence of hyperferritinemia (defined as >300 ng/mL) was 19.3%. Subjects with hyperferritinemia had lower FEV1% and FVC% than those with normal ferritin level with a slight difference, but those were statistically significant (99.22% vs.99.61% for FEV1%, p = 0.015 and 98.43% vs. 98.87% for FVC, p = 0.001). However, FEV1/FVC ratio was not significantly different between groups (P = 0.797). Compared with the highest quartile, the aORs for hyperferritinemia across decreasing quartiles were 1.081 (1.005-1.163), 1.100 (1.007-1.200), and 1.140 (1.053-1.233) for FEV1% (p for trend = 0.007) and 1.094 (1.018-1.176), 1.101 (1.021-1.188), and 1.150 (1.056-1.252) for FVC% (p for trend = 0.001). However, neither FEV1% nor FVC% was associated with iron or TSAT. CONCLUSIONS: Hyperferritinemia was associated with decreased lung function in healthy Korean men, but iron and TSAT were not. Longitudinal follow-up studies are required to validate our findings.


Assuntos
Ferritinas/sangue , Ferro/sangue , Pulmão/fisiologia , Testes de Função Respiratória , Transferrina/metabolismo , Adulto , Grupo com Ancestrais do Continente Asiático , Biomarcadores/sangue , Estudos Transversais , Humanos , Modelos Logísticos , Masculino , Análise Multivariada , República da Coreia
15.
Proc Natl Acad Sci U S A ; 117(13): 7317-7325, 2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32188787

RESUMO

Iron sequestration is a recognized innate immune mechanism against invading pathogens mediated by iron-binding proteins called transferrins. Despite many studies on antimicrobial activity of transferrins in vitro, their specific in vivo functions are poorly understood. Here we use Drosophila melanogaster as an in vivo model to investigate the role of transferrins in host defense. We find that systemic infections with a variety of pathogens trigger a hypoferremic response in flies, namely, iron withdrawal from the hemolymph and accumulation in the fat body. Notably, this hypoferremia to infection requires Drosophila nuclear factor κB (NF-κB) immune pathways, Toll and Imd, revealing that these pathways also mediate nutritional immunity in flies. Next, we show that the iron transporter Tsf1 is induced by infections downstream of the Toll and Imd pathways and is necessary for iron relocation from the hemolymph to the fat body. Consistent with elevated iron levels in the hemolymph, Tsf1 mutants exhibited increased susceptibility to Pseudomonas bacteria and Mucorales fungi, which could be rescued by chemical chelation of iron. Furthermore, using siderophore-deficient Pseudomonas aeruginosa, we discover that the siderophore pyoverdine is necessary for pathogenesis in wild-type flies, but it becomes dispensable in Tsf1 mutants due to excessive iron present in the hemolymph of these flies. As such, our study reveals that, similar to mammals, Drosophila uses iron limitation as an immune defense mechanism mediated by conserved iron-transporting proteins transferrins. Our in vivo work, together with accumulating in vitro studies, supports the immune role of insect transferrins against infections via an iron withholding strategy.


Assuntos
Proteínas de Drosophila/metabolismo , Ferro/metabolismo , Transferrina/metabolismo , Animais , Proteínas de Drosophila/imunologia , Drosophila melanogaster , Hemolinfa/imunologia , Hemolinfa/metabolismo , Imunidade Inata , Ferro/imunologia , NF-kappa B/metabolismo , Pseudomonas aeruginosa/metabolismo , Sideróforos/metabolismo , Transferrina/imunologia
16.
Biochem Biophys Res Commun ; 525(3): 626-632, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32122653

RESUMO

BACKGROUND: When proliferating tumor cells expand to areas distant from vascular sites, poor diffusion of oxygen and nutrients occur, generating a restrictive hypoxic gradient in which susceptible tumor cells die. The heterogeneous population surviving hypoxia and metabolic starvation include de-differentiated cancer stem cells (CSC), capable of self-renewing tumor-initiating cells (TICs), or those that divide asymmetrically to produce non-tumor-initiating differentiated (NTI-D) cell progeny. Under such restrictive conditions, both populations slowly proliferate, entering quiescence or senescence, when exiting from cell cycle progression. This may drive chemoresistance and tumor recurrence, since most anti-cancer treatments target rapidly proliferating cells. PURPOSE: Since persistent or additional stress may increase NTI-D cells conversion to TICs, we investigated whether nutrient depletion or hypoxia influence expression of tyrosinase, a crucial enzyme for melanin synthesis, and B16 melanoma survival, when exposed to iron-dependent cell death oxidative stress produced by the Fenton reaction, resembling ferroptosis. RESULTS: -a) proliferating B16 melanoma with 10% serum-supplementation (10%S) normoxically express hypoxia inducible factor 1α (HIF1α) but lose tyrosinase, in contrast to those transiently exposed to (SF) serum-free medium, in which both HIF1α and tyrosinase are co-expressed; b) in contrast to the resistance to SNP toxicity in (SF) cells with higher tyrosinase expression, those in (10%S) are killed by iron from nitroprusside/ferricyanide (SNP) irrespective of exogenous H2O2, in a reaction antagonized by the anti-oxidant and MEK inhibitor UO126; c) Moreover, under transient serum depletion, SNP cooperates with hypoxia (1.5% oxygen), prolonging B16 melanoma (SF) survival; d) the hypoxia mimetic CoCl2 inhibits proliferation-associated cyclin A, irrespective of SNP, in (10%S) cells or in transiently serum-depleted (SF) cells. However, only in the latter cells, CoCl2 but not SNP, induce loss of HIF1α and apoptosis-associated PARP cleavage; e) longer term adaptation to survive serum depletion, generates (SS) cells resistant to SNP toxicity, which aerobically co-express HIF1α and tyrosinase. In SS B16 melanoma, exogenous non-toxic 100 µM H2O2 super-induces the ratio of tyrosinase to HIF1α. However, co-treatment of SS-B16 cells with SNP plus exogenous H2O2, partly increases PARP cleavage by reciprocally decreasing tyrosinase expression. SIGNIFICANCE: - These results suggest that a phenotypic plasticity in response to depletion of nutrients and/or oxygen, helps decide whether melanoma cells undergo either death by ferroptosis, or resistance to it, when challenged by the same exogenous oxidative stress (iron ± H2O2).


Assuntos
Ferroptose/efeitos dos fármacos , Melanoma Experimental/patologia , Nitroprussiato/farmacologia , Soro/metabolismo , Animais , Butadienos/farmacologia , Hipóxia Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cobalto/farmacologia , Meios de Cultura Livres de Soro , Ciclina A/metabolismo , Peróxido de Hidrogênio/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Camundongos , Monofenol Mono-Oxigenase/metabolismo , Nitrilos/farmacologia , Poli(ADP-Ribose) Polimerases/metabolismo , Transferrina/deficiência , Transferrina/metabolismo
17.
Int J Mol Sci ; 21(4)2020 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-32085672

RESUMO

Tubulinopathies are rare neurological disorders caused by alterations in tubulin structure and function, giving rise to a wide range of brain abnormalities involving neuronal proliferation, migration, differentiation and axon guidance. TUBB is one of the ten ß-tubulin encoding genes present in the human genome and is broadly expressed in the developing central nervous system and the skin. Mutations in TUBB are responsible for two distinct pathological conditions: the first is characterized by microcephaly and complex structural brain malformations and the second, also known as "circumferential skin creases Kunze type" (CSC-KT), is associated to neurological features, excess skin folding and growth retardation. We used a combination of immunocytochemical and cellular approaches to explore, on patients' derived fibroblasts, the functional consequences of two TUBB variants: the novel mutation (p.N52S), associated with basal ganglia and cerebellar dysgenesis, and the previously reported variant (p.M73T), linked to microcephaly, corpus callosum agenesis and CSC-KT skin phenotype. Our results demonstrate that these variants impair microtubule (MT) function and dynamics. Most importantly, our studies show an altered epidermal growth factor (EGF) and transferrin (Tf) intracellular vesicle trafficking in both patients' fibroblasts, suggesting a specific role of TUBB in MT-dependent vesicular transport.


Assuntos
Microtúbulos/metabolismo , Mutação/genética , Vesículas Transportadoras/metabolismo , Tubulina (Proteína)/genética , Sequência de Aminoácidos , Movimento Celular/efeitos dos fármacos , Criança , Análise Mutacional de DNA , Fator de Crescimento Epidérmico/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Imagem por Ressonância Magnética , Modelos Moleculares , Nocodazol/farmacologia , Fenótipo , Transporte Proteico , Transferrina/metabolismo , Tubulina (Proteína)/química
18.
Molecules ; 25(3)2020 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-32046362

RESUMO

A series of water-soluble copper(II) complexes based on 2,9-dimethyl-1,10-phenanthroline (dmphen) and mixed-ligands, containing PTA=O (1,3,5-triaza-7-phosphaadamantane-7-oxide) have been synthesized and fully characterized. Two types of complexes have been obtained, monocationic [Cu(NO3)(O-PTA=O)(dmphen)][PF6] (1), [Cu(Cl)(dmphen)2][PF6] (2), and neutral [Cu(NO3)2(dmphen)] (3). The solid-state structures of all complexes have been determined by single-crystal X-ray diffraction. Magnetic studies for the complex 1-3 indicated a very weak antiferromagnetic interaction between copper(II) ions in crystal lattice. Complexes were successfully evaluated for their cytotoxic activities on the normal human dermal fibroblast (NHDF) cell line and the antitumor activity using the human lung carcinoma (A549), epithelioid cervix carcinoma (HeLa), colon (LoVo), and breast adenocarcinoma (MCF-7) cell lines. Complexes 1 and 3 revealed lower toxicity to NHDF than A549 and HeLa cells, meanwhile compound 2 appeared to be more toxic to NHDF cell line in comparison to all cancer lines. Additionally, interactions between the complexes and human apo-transferrin (apo-Tf) using fluorescence and circular dichroism (CD) spectroscopy were also investigated. All compounds interacted with apo-transferrin, causing same changes of the protein conformation. Electrostatic interactions dominate in the 1/2 - apo- Tf systems and hydrophobic and ionic interactions in the case of 3.


Assuntos
Adamantano/química , Antineoplásicos/síntese química , Apoproteínas/química , Complexos de Coordenação/síntese química , Cobre/química , Fenantrolinas/química , Transferrina/química , Células A549 , Adamantano/análogos & derivados , Antineoplásicos/farmacologia , Apoproteínas/metabolismo , Cátions Bivalentes , Cátions Monovalentes , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/farmacologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Concentração Inibidora 50 , Cinética , Células MCF-7 , Óxidos/química , Ligação Proteica , Termodinâmica , Transferrina/metabolismo
19.
Int J Mol Sci ; 21(3)2020 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-32041196

RESUMO

Patients with hereditary hemochromatosis and non-transfusion-dependent hereditary anemia develop predominantly liver iron-overload. We present a unique method allowing quantification of liver iron retention in humans during first-pass of 59Fe-labeled iron through the portal system, using standard ferrokinetic techniques measuring red cell iron uptake after oral and intravenous 59Fe administration. We present data from patients with iron deficiency (ID; N = 47), hereditary hemochromatosis (HH; N = 121) and non-transfusion-dependent hereditary anemia (HA; N = 40). Mean mucosal iron uptake and mucosal iron transfer (±SD) were elevated in patients with HH (59 ± 18%, 80 ± 15% respectively), HA (65 ± 17%, 74 ± 18%) and ID (84 ± 14%, 94 ± 6%) compared to healthy controls (43 ± 19%, 64 ± 18%) (p < 0.05) resulting in increased iron retention after 14 days compared to healthy controls in all groups (p < 0.01). The fraction of retained iron utilized for red cell production was 0.37 ± 0.17 in untreated HA, 0.55 ± 0.20 in untreated HH and 0.99 ± 0.22 in ID (p < 0.01). Interestingly, compared to red blood cell iron utilization after oral iron administration, red blood cell iron utilization was higher after injection of transferrin-bound iron in HA and HH. Liver iron retention was considerably higher in HH and HA compared to ID. We hypothesize that albumin serves as a scavenger of absorbed Fe(II) for delivering albumin-bound Fe(III) to hepatocytes.


Assuntos
Anemia Hemolítica Congênita/tratamento farmacológico , Anemia Ferropriva/tratamento farmacológico , Hemocromatose/tratamento farmacológico , Radioisótopos de Ferro/administração & dosagem , Fígado/química , Administração Intravenosa , Administração Oral , Adolescente , Adulto , Idoso , Anemia Hemolítica Congênita/metabolismo , Anemia Ferropriva/metabolismo , Estudos de Casos e Controles , Feminino , Hemocromatose/metabolismo , Humanos , Radioisótopos de Ferro/farmacocinética , Masculino , Pessoa de Meia-Idade , Albumina Sérica Humana/metabolismo , Transferrina/metabolismo , Adulto Jovem
20.
Analyst ; 145(5): 1737-1748, 2020 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-31913371

RESUMO

Glycosylation influences the structure and functionality of glycoproteins, and is regulated by genetic and environmental factors. The types and abundance of glycans on glycoproteins can vary due to diseases such as cancer, inflammation, autoimmune and neurodegenerative disorders. Due to the crucial role glycans play in modulating protein function, glycosylation analysis could lead to the discovery of novel biomarkers and is of prime importance in controlling the quality of glycoprotein biopharmaceuticals. Here, we present a method for the identification and quantification of glycoforms directly on intact proteins, after immunoaffinity purification from biological fluids. The method was validated and applied to serum transferrin and the biopharmaceutical trastuzumab. The accuracy of the method, expressed as the relative error (RE), ranged from 2.1 (at high concentrations) to 7.9% (at low concentrations), and intra- and inter-day precision, expressed as relative standard deviation (RSD), was 3.2 and 8.2%, respectively. The sensitivity and linearity of the method were suitable for serum analysis and the LOQ was calculated to be 3.1 and 4.4 µg mL-1 for transferrin (TFN) and trastuzumab (TRA), respectively. Its application to transferrin from five healthy human serum samples yielded concentrations between 1.61 and 3.17 mg mL-1, which are in agreement with blood reference levels. In parallel, the structure of the identified glycans was determined by ion mobility spectrometry coupled with tandem mass spectrometry. No chromatographic separation was required and sample preparation was performed in a semi-automatic manner, facilitating the handling of up to 12 samples at a time. This method should be useful for clinical laboratories and for the quality control of large batches of biopharmaceuticals.


Assuntos
Espectrometria de Mobilidade Iônica/métodos , Polissacarídeos/análise , Espectrometria de Massas em Tandem/métodos , Transferrina/análise , Trastuzumab/sangue , Glicosilação , Humanos , Polissacarídeos/química , Polissacarídeos/metabolismo , Transferrina/química , Transferrina/metabolismo , Trastuzumab/química , Trastuzumab/metabolismo
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