Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Commun Biol ; 5(1): 1366, 2022 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-36513703

RESUMO

Cellular metabolism influences immune cell function, with mitochondrial fatty acid ß-oxidation and oxidative phosphorylation required for multiple immune cell phenotypes. Carnitine palmitoyltransferase 1a (Cpt1a) is considered the rate-limiting enzyme for mitochondrial metabolism of long-chain fatty acids, and Cpt1a deficiency is associated with infant mortality and infection risk. This study was undertaken to test the hypothesis that impairment in Cpt1a-dependent fatty acid oxidation results in increased susceptibility to infection. Screening the Cpt1a gene for common variants predicted to affect protein function revealed allele rs2229738_T, which was associated with pneumonia risk in a targeted human phenome association study. Pharmacologic inhibition of Cpt1a increases mortality and impairs control of the infection in a murine model of bacterial pneumonia. Susceptibility to pneumonia is associated with blunted neutrophilic responses in mice and humans that result from impaired neutrophil trafficking to the site of infection. Chemotaxis responsible for neutrophil trafficking requires Cpt1a-dependent mitochondrial fatty acid oxidation for amplification of chemoattractant signals. These findings identify Cpt1a as a potential host determinant of infection susceptibility and demonstrate a requirement for mitochondrial fatty acid oxidation in neutrophil biology.


Assuntos
Carnitina O-Palmitoiltransferase , Metabolismo dos Lipídeos , Neutrófilos , Animais , Humanos , Lactente , Camundongos , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Ácidos Graxos/metabolismo , Mitocôndrias/metabolismo , Neutrófilos/metabolismo
2.
Am J Physiol Renal Physiol ; 317(4): F922-F929, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31364379

RESUMO

Acute kidney injury is a common complication of severe sepsis and contributes to high mortality. The molecular mechanisms of acute kidney injury during sepsis are not fully understood. Because hemoproteins, including myoglobin and hemoglobin, are known to mediate kidney injury during rhabdomyolysis, we hypothesized that cell-free hemoglobin (CFH) would exacerbate acute kidney injury during sepsis. Sepsis was induced in mice by intraperitoneal injection of cecal slurry (CS). To mimic elevated levels of CFH observed during human sepsis, mice also received a retroorbital injection of CFH or dextrose control. Four groups of mice were analyzed: sham treated (sham), CFH alone, CS alone, and CS + CFH. The addition of CFH to CS reduced 48-h survival compared with CS alone (67% vs. 97%, P = 0.001) and increased the severity of illness. After 24 and 48 h, CS + CFH mice had a reduced glomerular filtration rate from baseline, whereas sham, CFH, and CS mice maintained baseline glomerular filtration rate. Biomarkers of acute kidney injury, neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1), were markedly elevated in CS+CFH compared with CS (8-fold for NGAL and 2.4-fold for KIM-1, P < 0.002 for each) after 48 h. Histological examination showed a trend toward increased tubular injury in CS + CFH-exposed kidneys compared with CS-exposed kidneys. However, there were similar levels of renal oxidative injury and apoptosis in the CS + CFH group compared with the CS group. Kidney levels of multiple proinflammatory cytokines were similar between CS and CS + CFH groups. Human renal tubule cells (HK-2) exposed to CFH demonstrated increased cytotoxicity. Together, these results show that CFH exacerbates acute kidney injury in a mouse model of experimental sepsis, potentially through increased renal tubular injury.


Assuntos
Injúria Renal Aguda/patologia , Hemoglobinas/toxicidade , Sepse/patologia , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/fisiopatologia , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sistema Livre de Células , Citocinas/metabolismo , Feminino , Taxa de Filtração Glomerular , Receptor Celular 1 do Vírus da Hepatite A/metabolismo , Túbulos Renais/metabolismo , Túbulos Renais/patologia , Lipocalina-2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sepse/complicações , Análise de Sobrevida
3.
JCI Insight ; 3(2)2018 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-29367464

RESUMO

Primary graft dysfunction (PGD) is acute lung injury within 72 hours of lung transplantation. We hypothesized that cell-free hemoglobin (CFH) contributes to PGD by increasing lung microvascular permeability and tested this in patients, ex vivo human lungs, and cultured human lung microvascular endothelial cells. In a nested case control study of 40 patients with severe PGD at 72 hours and 80 matched controls without PGD, elevated preoperative CFH was independently associated with increased PGD risk (odds ratio [OR] 2.75, 95%CI, 1.23-6.16, P = 0.014). The effect of CFH on PGD was magnified by reperfusion fraction of inspired oxygen (FiO2) ≥ 0.40 (OR 3.41, P = 0.031). Isolated perfused human lungs exposed to intravascular CFH (100 mg/dl) developed increased vascular permeability as measured by lung weight (CFH 14.4% vs. control 0.65%, P = 0.047) and extravasation of Evans blue-labeled albumin dye (EBD) into the airspace (P = 0.027). CFH (1 mg/dl) also increased paracellular permeability of human pulmonary microvascular endothelial cell monolayers (hPMVECs). Hyperoxia (FiO2 = 0.95) increased human lung and hPMVEC permeability compared with normoxia (FiO2 = 0.21). Treatment with acetaminophen (15 µg/ml), a specific hemoprotein reductant, prevented CFH-dependent permeability in human lungs (P = 0.046) and hPMVECs (P = 0.037). In summary, CFH may mediate PGD through oxidative effects on microvascular permeability, which are augmented by hyperoxia and abrogated by acetaminophen.


Assuntos
Hemoglobinas/imunologia , Hiperóxia/imunologia , Transplante de Pulmão/efeitos adversos , Disfunção Primária do Enxerto/imunologia , Acetaminofen/farmacologia , Aloenxertos/irrigação sanguínea , Aloenxertos/imunologia , Aloenxertos/patologia , Permeabilidade Capilar/efeitos dos fármacos , Permeabilidade Capilar/imunologia , Estudos de Casos e Controles , Linhagem Celular , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Feminino , Hemoglobinas/antagonistas & inibidores , Humanos , Hiperóxia/sangue , Hiperóxia/patologia , Pulmão/irrigação sanguínea , Pulmão/citologia , Pulmão/imunologia , Pulmão/patologia , Masculino , Microvasos/citologia , Microvasos/metabolismo , Pessoa de Meia-Idade , Estresse Oxidativo/imunologia , Disfunção Primária do Enxerto/sangue , Disfunção Primária do Enxerto/patologia
4.
Biochem Biophys Res Commun ; 495(1): 433-437, 2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-29129689

RESUMO

BACKGROUND: Increased endothelial permeability is central to shock and organ dysfunction in sepsis but therapeutics targeted to known mediators of increased endothelial permeability have been unsuccessful in patient studies. We previously reported that cell-free hemoglobin (CFH) is elevated in the majority of patients with sepsis and is associated with organ dysfunction, poor clinical outcomes and elevated markers of oxidant injury. Others have shown that Vitamin C (ascorbate) may have endothelial protective effects in sepsis. In this study, we tested the hypothesis that high levels of CFH, as seen in the circulation of patients with sepsis, disrupt endothelial barrier integrity. METHODS: Human umbilical vein endothelial cells (HUVEC) were grown to confluence and treated with CFH with or without ascorbate. Monolayer permeability was measured by Electric Cell-substrate Impedance Sensing (ECIS) or transfer of 14C-inulin. Viability was measured by trypan blue exclusion. Intracellular ascorbate was measured by HPLC. RESULTS: CFH increased permeability in a dose- and time-dependent manner with 1 mg/ml of CFH increasing inulin transfer by 50% without affecting cell viability. CFH (1 mg/ml) also caused a dramatic reduction in intracellular ascorbate in the same time frame (1.4 mM without CFH, 0.23 mM 18 h after 1 mg/ml CFH, p < 0.05). Pre-treatment of HUVECs with ascorbate attenuated CFH induced permeability. CONCLUSIONS: CFH increases endothelial permeability in part through depletion of intracellular ascorbate. Supplementation of ascorbate can attenuate increases in permeability mediated by CFH suggesting a possible therapeutic approach in sepsis.


Assuntos
Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Permeabilidade Capilar/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Hemoglobinas/metabolismo , Antioxidantes/metabolismo , Ácido Ascórbico/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Sepse/tratamento farmacológico , Sepse/metabolismo
5.
Ann Am Thorac Soc ; 14(Supplement_3): S251-S252, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28945473

RESUMO

RATIONALE: Cell-free hemoglobin (CFH) is a potent oxidant associated with poor clinical outcomes in a variety of clinical settings. Recent studies suggest that acetaminophen (APAP), a specific hemoprotein reductant, can abrogate CFH-mediated oxidative injury and organ dysfunction. Preoperative plasma CFH levels are independently associated with primary graft dysfunction (PGD) after lung transplant ( 1 ). OBJECTIVES: Our objectives were to determine whether CFH would increase lung vascular permeability in the isolated perfused human lung and whether APAP would limit these effects. METHODS: Human lungs declined for transplant were inflated and perfused with Dulbecco's modified Eagle medium/5% albumin at a pulmonary artery pressure of 8-12 mm Hg. After steady state was achieved, CFH (100 mg/dl) was added to the perfusate ± APAP (15 µg/ml). Lung permeability was measured by continuous monitoring of lung weight gain and by extravasation of Evans blue dye-labeled albumin from the vasculature into bronchoalveolar lavage. To test the mechanism of increased permeability, human pulmonary microvascular endothelial cells were exposed to CFH (0.5 mg/ml) ± APAP (160 µM) for 24 hours and permeability was assessed by electrical cell-substrate impedance sensing. MEASUREMENT AND MAIN RESULTS: In the isolated perfused human lung, CFH increased lung permeability over 2 hours compared with control lungs (12% vs. 2% weight gain from baseline, P = 0.03). Increased vascular permeability was confirmed by a 4.8-fold increase in Evans blue dye-labeled albumin in the airspace compared with control lungs. Pretreatment with APAP prevented lung weight gain (P = 0.06 vs. CFH). In human pulmonary microvascular endothelial cells, CFH increased monolayer permeability (P = 0.03 vs. control), and this was attenuated by APAP (P = 0.045 vs. CFH). CONCLUSIONS: Circulating CFH increases vascular permeability in the isolated perfused human lung and paracellular permeability in lung microvascular endothelial cells. These effects may explain the association of plasma CFH levels with PGD. The hemoprotein reductant APAP attenuates the effects of CFH and merits further exploration as a potential therapy for PGD prevention.


Assuntos
Acetaminofen/farmacologia , Permeabilidade Capilar/fisiologia , Hemoglobinas/metabolismo , Transplante de Pulmão/efeitos adversos , Pulmão/irrigação sanguínea , Disfunção Primária do Enxerto/prevenção & controle , Células Endoteliais/fisiologia , Humanos
6.
Shock ; 48(1): 54-60, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28125528

RESUMO

Although studies in rat cultured pulmonary artery endothelial cells, perfused lungs, and intact mice support the concept that oxidative mitochondrial (mt) DNA damage triggers acute lung injury (ALI), it has not yet been determined whether enhanced mtDNA repair forestalls development of ALI and its progression to multiple organ system failure (MOSF). Accordingly, here we examined the effect of a fusion protein construct targeting the DNA glycosylase, Ogg1, to mitochondria in a rat model intra-tracheal Pseudomonas aeruginosa (strain 103; PA103)-induced ALI and MOSF. Relative to controls, animals given PA103 displayed increases in lung vascular filtration coefficient accompanied by transient lung tissue oxidative mtDNA damage and variable changes in mtDNA copy number without evidence of nuclear DNA damage. The approximate 40% of animals surviving 24 h after bacterial administration exhibited multiple organ dysfunction, manifest as increased serum and tissue-specific indices of kidney and liver failure, along with depressed heart rate and blood pressure. While administration of mt-targeted Ogg1 to control animals was innocuous, the active fusion protein, but not a DNA repair-deficient mutant, prevented bacteria-induced increases in lung tissue oxidative mtDNA damage, failed to alter mtDNA copy number, and attenuated lung endothelial barrier degradation. These changes were associated with suppression of liver, kidney, and cardiovascular dysfunction and with decreased 24 h mortality. Collectively, the present findings indicate that oxidative mtDNA damage to lung tissue initiates PA103-induced ALI and MOSF in rats.


Assuntos
Lesão Pulmonar Aguda/genética , Dano ao DNA/genética , DNA Mitocondrial/genética , Insuficiência de Múltiplos Órgãos/genética , Lesão Pulmonar Aguda/microbiologia , Animais , DNA Glicosilases/genética , Masculino , Estresse Oxidativo/genética , Estresse Oxidativo/fisiologia , Pseudomonas aeruginosa/patogenicidade , Ratos , Ratos Sprague-Dawley , Traqueia/microbiologia
7.
Am J Physiol Lung Cell Mol Physiol ; 308(10): L1078-85, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25795724

RESUMO

Fragments of the mitochondrial genome released into the systemic circulation after mechanical trauma, termed mitochondrial DNA damage-associated molecular patterns (mtDNA DAMPs), are thought to mediate the systemic inflammatory response syndrome. The close association between circulating mtDNA DAMP levels and outcome in sepsis suggests that bacteria also might be a stimulus for mtDNA DAMP release. To test this hypothesis, we measured mtDNA DAMP abundance in medium perfusing isolated rat lungs challenged with an intratracheal instillation of 5 × 10(7) colony-forming units of Pseudomonas aeruginosa (strain 103; PA103). Intratracheal PA103 caused rapid accumulation of selected 200-bp sequences of the mitochondrial genome in rat lung perfusate accompanied by marked increases in both lung tissue oxidative mtDNA damage and in the vascular filtration coefficient (Kf). Increases in lung tissue mtDNA damage, perfusate mtDNA DAMP abundance, and Kf were blocked by addition to the perfusion medium of a fusion protein targeting the DNA repair enzyme Ogg1 to mitochondria. Intra-arterial injection of mtDNA DAMPs prepared from rat liver mimicked the effect of PA103 on both Kf and lung mtDNA integrity. Effects of mtDNA and PA103 on Kf were also attenuated by an oligodeoxynucleotide inhibitor of Toll-like receptor 9 (TLR-9) by mitochondria-targeted Ogg1 and by addition of DNase1 to the perfusion medium. Collectively, these findings are consistent with a model wherein PA103 causes oxidative mtDNA damage leading to a feed-forward cycle of mtDNA DAMP formation and TLR-9-dependent mtDNA damage that culminates in acute lung injury.


Assuntos
Lesão Pulmonar Aguda , Dano ao DNA , DNA Mitocondrial/metabolismo , Pulmão , Infecções por Pseudomonas , Pseudomonas aeruginosa , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , DNA Glicosilases/metabolismo , Desoxirribonuclease I/farmacologia , Pulmão/irrigação sanguínea , Pulmão/metabolismo , Pulmão/patologia , Masculino , Oligodesoxirribonucleotídeos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Perfusão , Infecções por Pseudomonas/metabolismo , Infecções por Pseudomonas/patologia , Ratos , Ratos Sprague-Dawley , Receptor Toll-Like 9/agonistas , Receptor Toll-Like 9/metabolismo
8.
Ann Surg ; 258(4): 591-6; discussion 596-8, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23979273

RESUMO

OBJECTIVE: Our objective was to execute a prospective cohort study to determine relationships between plasma mtDNA DAMP levels and the occurrence of systemic inflammatory response syndrome (SIRS), multiple organ dysfunction syndrome (MODS), and mortality. BACKGROUND: Mitochondrial DNA damage-associated molecular patterns (DAMPs) accumulate in the circulation after severe injury. Observations in animal models demonstrate that mtDNA DAMPs contribute to organ dysfunction; however, the link between plasma mtDNA DAMPs and outcome in severely injured human subjects has not been established. METHODS: DNA was isolated from plasma samples taken from severely injured patients at hospital days 0, 1, and 2. Real-time PCR was used to quantify selected ≈200 base pair sequences of mtDNA within the COX1, ND1, and ND6 genes, as well as from the D-Loop transcriptional regulatory region. MODS was defined as a Denver Multiple Organ Failure score of 4 or greater. RESULTS: MtDNA DAMPs were quantified as PCR threshold cycle number. Lower threshold cycles indicate increased mtDNA DAMP content. Patients with SIRS had significantly increased mtDNA DAMP levels in all 4 sequences examined (32.14 ± 0.90 vs 29.00 ± 1.15 for COX1, 31.90 ± 0.47 vs 30.16 ± 1.42 for ND1, 32.40 ± 0.61 vs 28.94 ± 1.13 for ND6, and 33.12 ± 0.83 vs 28.30 ± 1.14 for D-Loop). Patients who developed MODS also had elevated mtDNA DAMP levels compared with those who did not (32.57 ± 0.74 vs 27.12 ± 0.66 for COX1, 32.45 ± 0.65 vs 28.20 ± 0.73 for ND1, 32.52 ± 0.56 vs 27.60 ± 0.79 for ND6, and 32.85 ± 0.75 vs 27.86 ± 1.27 for D-Loop). Patients with above-median mtDNA DAMP levels had a significantly elevated relative risk for mortality. Four patients died secondary to severe MODS. CONCLUSIONS: These findings comprise the first observational evidence that plasma mtDNA DAMPs is associated with the evolution of SIRS, MODS, and mortality in severely injured human subjects.


Assuntos
DNA Mitocondrial/sangue , Insuficiência de Múltiplos Órgãos/diagnóstico , Síndrome de Resposta Inflamatória Sistêmica/diagnóstico , Ferimentos e Lesões/complicações , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Estudos de Coortes , Ciclo-Oxigenase 1/sangue , Ciclo-Oxigenase 1/genética , Feminino , Marcadores Genéticos , Humanos , Escala de Gravidade do Ferimento , Masculino , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos/sangue , Insuficiência de Múltiplos Órgãos/etiologia , Insuficiência de Múltiplos Órgãos/mortalidade , NADH Desidrogenase/sangue , NADH Desidrogenase/genética , Prognóstico , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo Real , Síndrome de Resposta Inflamatória Sistêmica/sangue , Síndrome de Resposta Inflamatória Sistêmica/etiologia , Síndrome de Resposta Inflamatória Sistêmica/mortalidade , Ferimentos e Lesões/sangue , Ferimentos e Lesões/mortalidade
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...