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1.
J Am Chem Soc ; 143(18): 6981-6989, 2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-33905249

RESUMO

The function of proteins is linked to their conformations that can be resolved with several high-resolution methods. However, only a few methods can provide the temporal order of intermediates and conformational changes, with each having its limitations. Here, we combine pulsed electron-electron double resonance spectroscopy with a microsecond freeze-hyperquenching setup to achieve spatiotemporal resolution in the angstrom range and lower microsecond time scale. We show that the conformational change of the Cα-helix in the cyclic nucleotide-binding domain of the Mesorhizobium loti potassium channel occurs within about 150 µs and can be resolved with angstrom precision. Thus, this approach holds great promise for obtaining 4D landscapes of conformational changes in biomolecules.

2.
Nat Commun ; 11(1): 5682, 2020 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-33173168

RESUMO

The Rhizoclosmatium globosum genome encodes three rhodopsin-guanylyl cyclases (RGCs), which are predicted to facilitate visual orientation of the fungal zoospores. Here, we show that RGC1 and RGC2 function as light-activated cyclases only upon heterodimerization with RGC3 (NeoR). RGC1/2 utilize conventional green or blue-light-sensitive rhodopsins (λmax = 550 and 480 nm, respectively), with short-lived signaling states, responsible for light-activation of the enzyme. The bistable NeoR is photoswitchable between a near-infrared-sensitive (NIR, λmax = 690 nm) highly fluorescent state (QF = 0.2) and a UV-sensitive non-fluorescent state, thereby modulating the activity by NIR pre-illumination. No other rhodopsin has been reported so far to be functional as a heterooligomer, or as having such a long wavelength absorption or high fluorescence yield. Site-specific mutagenesis and hybrid quantum mechanics/molecular mechanics simulations support the idea that the unusual photochemical properties result from the rigidity of the retinal chromophore and a unique counterion triad composed of two glutamic and one aspartic acids. These findings substantially expand our understanding of the natural potential and limitations of spectral tuning in rhodopsin photoreceptors.


Assuntos
Quitridiomicetos/genética , Rodopsina , Biologia Computacional , Fluorescência , Corantes Fluorescentes/química , Corantes Fluorescentes/isolamento & purificação , Genes Fúngicos , Genoma Fúngico , Mutagênese Sítio-Dirigida , Processos Fotoquímicos , Células Fotorreceptoras/fisiologia , Rodopsina/biossíntese , Rodopsina/química , Rodopsina/genética
3.
Proc Natl Acad Sci U S A ; 117(24): 13783-13791, 2020 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-32467169

RESUMO

Proton (H+) channels are special: They select protons against other ions that are up to a millionfold more abundant. Only a few proton channels have been identified so far. Here, we identify a family of voltage-gated "pacemaker" channels, HCNL1, that are exquisitely selective for protons. HCNL1 activates during hyperpolarization and conducts protons into the cytosol. Surprisingly, protons permeate through the channel's voltage-sensing domain, whereas the pore domain is nonfunctional. Key to proton permeation is a methionine residue that interrupts the series of regularly spaced arginine residues in the S4 voltage sensor. HCNL1 forms a tetramer and thus contains four proton pores. Unlike classic HCN channels, HCNL1 is not gated by cyclic nucleotides. The channel is present in zebrafish sperm and carries a proton inward current that acidifies the cytosol. Our results suggest that protons rather than cyclic nucleotides serve as cellular messengers in zebrafish sperm. Through small modifications in two key functional domains, HCNL1 evolutionarily adapted to a low-Na+ freshwater environment to conserve sperm's ability to depolarize.


Assuntos
Peixe-Zebra/metabolismo , Sequência de Aminoácidos , Animais , Transporte Biológico , Masculino , Família Multigênica , Prótons , Espermatozoides/metabolismo , Peixe-Zebra/genética
4.
EMBO J ; 39(4): e102723, 2020 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-31880004

RESUMO

Cilia serve as cellular antennae that translate sensory information into physiological responses. In the sperm flagellum, a single chemoattractant molecule can trigger a Ca2+ rise that controls motility. The mechanisms underlying such ultra-sensitivity are ill-defined. Here, we determine by mass spectrometry the copy number of nineteen chemosensory signaling proteins in sperm flagella from the sea urchin Arbacia punctulata. Proteins are up to 1,000-fold more abundant than the free cellular messengers cAMP, cGMP, H+ , and Ca2+ . Opto-chemical techniques show that high protein concentrations kinetically compartmentalize the flagellum: Within milliseconds, cGMP is relayed from the receptor guanylate cyclase to a cGMP-gated channel that serves as a perfect chemo-electrical transducer. cGMP is rapidly hydrolyzed, possibly via "substrate channeling" from the channel to the phosphodiesterase PDE5. The channel/PDE5 tandem encodes cGMP turnover rates rather than concentrations. The rate-detection mechanism allows continuous stimulus sampling over a wide dynamic range. The textbook notion of signal amplification-few enzyme molecules process many messenger molecules-does not hold for sperm flagella. Instead, high protein concentrations ascertain messenger detection. Similar mechanisms may occur in other small compartments like primary cilia or dendritic spines.


Assuntos
Arbacia/fisiologia , Quimiotaxia , Proteômica , Transdução de Sinais , Animais , Arbacia/ultraestrutura , Cálcio/metabolismo , Cílios/fisiologia , Cílios/ultraestrutura , GMP Cíclico/metabolismo , Tomografia com Microscopia Eletrônica , Flagelos/fisiologia , Flagelos/ultraestrutura , Guanilato Ciclase/metabolismo , Masculino , Espectrometria de Massas , Espermatozoides/fisiologia , Espermatozoides/ultraestrutura
5.
Eur Heart J Cardiovasc Imaging ; 20(10): 1094-1101, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31327014

RESUMO

AIMS: To assess the survival benefit of aortic valve replacement (AVR) in patients with normal flow low gradient severe aortic stenosis (AS). METHODS AND RESULTS: A retrospective study of prospectively collected data of 276 patients (mean age 75 ± 15 years, 51% male) with normal transaortic flow [flow rate (FR) ≥200 mL/s or stroke volume index (SVi) ≥35 mL/m2] and severe AS (aortic valve area <1.0 cm2). The outcome measure was all-cause mortality. Of the 276 patients, 151 (55%) were medically treated, while 125 (45%) underwent an AVR. Over a mean follow-up of 3.2 ± 1.8 years (range 0-6.9 years), a total of 96 (34.8%) deaths occurred: 17 (13.6%) in AVR group vs. 79 (52.3%) in those medically treated, when transaortic flow was defined by FR (P < 0.001). When transaortic flow was defined by SVi, a total of 79 (31.3%) deaths occurred: 18 (15.1%) in AVR group vs. 61 (45.9%) in medically treated (P < 0.001). In a propensity-matched multivariable Cox regression analysis adjusting for age, gender, body surface area, smoking, hypertension, diabetes mellitus, atrial fibrillation, peripheral vascular disease, chronic kidney disease, left ventricular ejection fraction, left ventricular mass, and mean aortic gradient, not having AVR was associated with a 6.3-fold higher hazard ratio (HR) of all-cause mortality [HR 6.28, 95% confidence interval (CI) 3.34-13.16; P < 0.001] when flow was defined by FR. In the SVi-guided model, it was 3.83-fold (HR 3.83, 95% CI 2.30-6.37; P < 0.001). CONCLUSION: In patients with normal flow low gradient severe AS, AVR was associated with a significantly improved survival compared with those who received standard medical treatment.


Assuntos
Estenose da Valva Aórtica/diagnóstico por imagem , Estenose da Valva Aórtica/mortalidade , Estenose da Valva Aórtica/cirurgia , Ecocardiografia Doppler , Implante de Prótese de Valva Cardíaca/métodos , Idoso , Velocidade do Fluxo Sanguíneo , Comorbidade , Feminino , Humanos , Londres , Masculino , Pontuação de Propensão , Estudos Retrospectivos , Fatores de Risco , Índice de Gravidade de Doença , Taxa de Sobrevida
6.
Diabetes Care ; 42(7): 1225-1233, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31036547

RESUMO

OBJECTIVE: Altered plasma amino acid levels have been implicated as markers of risk for incident type 2 diabetes; however, amino acids are also related to established diabetes risk factors. Therefore, potential for confounding and the impact from competing risks require evaluation. RESEARCH DESIGN AND METHODS: We prospectively followed 2,519 individuals with coronary artery disease but without diabetes. Mixed Gaussian modeling identified potential for confounding. Confounding, defined as a change in effect estimate (≥10%), was investigated by comparing amino acid-incident diabetes risk in a Cox model containing age and sex with that in models adjusted for potential confounders (BMI, estimated glomerular filtration rate, HDL cholesterol, triacylglycerol, C-reactive protein), which were further adjusted for plasma glucose, competing risks, and multiple comparisons (false discovery rate = 0.05, Benjamini-Hochberg method). Finally, component-wise likelihood-based boosting analysis including amino acids and confounders was performed and adjusted for competing risks in order to identify an optimal submodel for predicting incident diabetes. RESULTS: The mean age of the source population was 61.9 years; 72% were men. During a median follow-up of 10.3 years, 267 incident cases of diabetes were identified. In age- and sex-adjusted models, several amino acids, including the branched-chain amino acids, significantly predicted incident diabetes. Adjustment for confounders, however, attenuated associations. Further adjustment for glucose and multiple comparisons rendered only arginine significant (hazard ratio/1 SD 1.21 [95% CI 1.07-1.37]). The optimal submodel included arginine and asparagine. CONCLUSIONS: Adjustment for relevant clinical factors attenuated the amino acid-incident diabetes risk. Although these findings do not preclude the potential pathogenic role of other amino acids, they suggest that plasma arginine is independently associated with incident diabetes. Both arginine and asparagine were identified in an optimal model for predicting new-onset type 2 diabetes.


Assuntos
Aminoácidos/sangue , Doença da Artéria Coronariana/sangue , Doença da Artéria Coronariana/epidemiologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/epidemiologia , Idoso , Biomarcadores/sangue , Proteína C-Reativa/análise , Proteína C-Reativa/metabolismo , Estudos de Coortes , Doença da Artéria Coronariana/complicações , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/diagnóstico , Angiopatias Diabéticas/sangue , Angiopatias Diabéticas/epidemiologia , Feminino , Seguimentos , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Prognóstico , Fatores de Risco
7.
Methods Cell Biol ; 151: 487-517, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948028

RESUMO

Sperm from sea urchins are attracted by chemical cues released by the egg-a mechanism called chemotaxis. We describe here the signaling pathway and molecular components endowing sperm with single-molecule sensitivity. Chemotactic signaling and behavioral responses occur on a timescale of a few milliseconds to seconds. We describe the techniques and chemical tools used to resolve the signaling events in time. The techniques include rapid-mixing devices, rapid stroboscopic microscopy, and photolysis of caged second messengers and chemoattractants.


Assuntos
Óptica e Fotônica/métodos , Motilidade Espermática/genética , Espermatozoides/crescimento & desenvolvimento , Estroboscopia/métodos , Animais , Fatores Quimiotáticos/química , Cinética , Masculino , Ouriços-do-Mar/crescimento & desenvolvimento , Espermatozoides/ultraestrutura
8.
J Am Heart Assoc ; 7(22): e010735, 2018 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-30571488

RESUMO

Background Exaggerated blood pressure response during exercise predicts future hypertension and cardiovascular events in general population and different patients groups. However, its clinical and prognostic implications in patients with aortic stenosis have not been previously evaluated. Methods and Results We retrospectively studied 301 patients with moderate to severe asymptomatic aortic stenosis (aged 65±12 years) who underwent echocardiography and a modified Bruce exercise treadmill test. An exaggerated blood pressure response was defined as peak systolic blood pressure ≥190 mm Hg. An abnormal blood pressure response (either blunted or exaggerated) was found in 58% of patients and abnormal left ventricular geometry in 82%. There was no difference in the rates of abnormal blood pressure responses between patients with moderate and severe aortic stenosis ([exaggerated blood pressure response: 21% versus 22%, P=0.876] and [blunted blood pressure response: 35% versus 40%, P=0.647]). Patients with exaggerated blood pressure response (21%) were more likely to be older, have hypertension, higher pretest systolic blood pressure, left ventricular ejection fraction and mass, and increased arterial stiffness (all P<0.05). In a multivariate logistic regression analysis, an exaggerated blood pressure response was associated with higher pulse pressure/stroke volume index (odds ratio 2.45, 95% confidence interval 1.02-6.00, P=0.037) and left ventricular mass (odds ratio 2.04, 95% confidence interval 1.23-3.38, P=0.012) independent of age, hypertension, aortic annulus and left atrium diameter, and left ventricular ejection fraction. Conclusions In those with aortic stenosis, exaggerated blood pressure was strongly related to higher resting blood pressure values, left ventricular mass, and increased arterial stiffness independent of hypertension.


Assuntos
Estenose da Valva Aórtica/fisiopatologia , Pressão Sanguínea/fisiologia , Teste de Esforço , Função Ventricular Esquerda/fisiologia , Doença Aguda , Idoso , Estenose da Valva Aórtica/diagnóstico , Estenose da Valva Aórtica/diagnóstico por imagem , Doenças Assintomáticas , Ecocardiografia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
9.
Nat Commun ; 9(1): 4611, 2018 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-30397200

RESUMO

Optogenetics enables manipulation of biological processes with light at high spatio-temporal resolution to control the behavior of cells, networks, or even whole animals. In contrast to the performance of excitatory rhodopsins, the effectiveness of inhibitory optogenetic tools is still insufficient. Here we report a two-component optical silencer system comprising photoactivated adenylyl cyclases (PACs) and the small cyclic nucleotide-gated potassium channel SthK. Activation of this 'PAC-K' silencer by brief pulses of low-intensity blue light causes robust and reversible silencing of cardiomyocyte excitation and neuronal firing. In vivo expression of PAC-K in mouse and zebrafish neurons is well tolerated, where blue light inhibits neuronal activity and blocks motor responses. In combination with red-light absorbing channelrhodopsins, the distinct action spectra of PACs allow independent bimodal control of neuronal activity. PAC-K represents a reliable optogenetic silencer with intrinsic amplification for sustained potassium-mediated hyperpolarization, conferring high operational light sensitivity to the cells of interest.


Assuntos
Optogenética/métodos , Canais de Potássio/genética , Canais de Potássio/metabolismo , Canais de Potássio/efeitos da radiação , Elementos Silenciadores Transcricionais , Adenilil Ciclases/genética , Adenilil Ciclases/metabolismo , Adenilil Ciclases/efeitos da radiação , Animais , Animais Geneticamente Modificados , Channelrhodopsins/efeitos da radiação , Expressão Gênica/genética , Expressão Gênica/efeitos da radiação , Células HEK293 , Humanos , Luz , Camundongos , Modelos Animais , Miócitos Cardíacos/metabolismo , Neurônios/metabolismo , Neurônios/efeitos da radiação , Rodopsina/farmacologia , Peixe-Zebra
10.
Int J Cardiol ; 267: 100-106, 2018 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-29957250

RESUMO

BACKGROUND: Plasma trimethylamine-N-oxide (TMAO) is associated with cardiovascular disease; however specific relationships with cardiac arrhythmias are unknown. We evaluated the association between plasma TMAO and incident atrial fibrillation (AF). METHODS: Risk associations were explored among 3797 patients with suspected stable angina in the Western Norway Coronary Angiography Cohort (WECAC) and verified in 3143 elderly participants in the community-based Hordaland Health Study (HUSK). Information on endpoints was obtained from nationwide registries. RESULTS: Median follow-up was 7.3 and 10.8 years in the WECAC and HUSK cohorts, respectively, and 412 (10.9%) and 484 (15.4%) subjects were registered with incident AF. The age and gender adjusted HRs were 1.16, 95% CI 1.05-1.28 and 1.10, 95% CI 1.004-1.19 per 1 SD increase in log-transformed plasma TMAO. Adjusting for hypertension, BMI, smoking, diabetes, or intake of total choline, a TMAO precursor, did not materially influence the risk associations. Among patients in WECAC, further extensive adjustment for other AF risk factors yielded similar results. Adding TMAO to traditional AF risk factors (age, gender, hypertension, BMI, smoking and diabetes) yielded a continuous net reclassification improvement of 0.108, 95% CI 0.015-0.202 and 0.139, 95% CI 0.042-0.235. CONCLUSIONS: Plasma TMAO was associated with and improved reclassification of incident AF in two independent Norwegian cohorts with long-term follow-up. The relationship was independent of traditional AF risk factors, as well as of dietary choline intake. Our findings motivate further studies to explore endogenous metabolic factors influencing the relationship between TMAO and cardiovascular disease.


Assuntos
Fibrilação Atrial/sangue , Metilaminas , Idoso , Fibrilação Atrial/diagnóstico , Fibrilação Atrial/epidemiologia , Colina/sangue , Feminino , Seguimentos , Humanos , Incidência , Masculino , Metilaminas/sangue , Metilaminas/metabolismo , Pessoa de Meia-Idade , Noruega/epidemiologia , Prognóstico , Fatores de Risco
11.
Heart ; 104(22): 1836-1842, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29654094

RESUMO

OBJECTIVE: To assess the safety and tolerability of treadmill exercise testing and the association of revealed symptoms with outcome in apparently asymptomatic patients with moderate to severe aortic stenosis (AS). METHODS: A retrospective cohort study of 316 patients (age 65±12 years, 67% men) with moderate and severe AS who underwent echocardiography and modified Bruce exercise treadmill tests (ETTs) at a specialist valve clinic. The outcome measures were aortic valve replacement (AVR), all-cause mortality or a composite of AVR and all-cause mortality. RESULTS: At baseline, there were 210 (66%) patients with moderate and 106 (34%) with severe AS. There were 264 (83%) events. 234 (74%) patients reached an indication for AVR, 145 (69%) with moderate and 88 (83%) with severe AS (p<0.05). Of the 30 (9%) deaths recoded during follow-up, 20 (67%) were cardiovascular related. In total, 797 exercise tests (mean 2.5±2.1 per patient) were performed. No serious adverse events were observed. The prevalence of revealed symptoms at baseline ETT was 29% (n=91) and was significantly higher in severe AS compared with moderate AS (38%vs23%, p=0.008). Symptoms were revealed in 18%-59% of patients during serial ETT conducted over a follow-up period of 34.9 (SD 35.1) months. The event-free survival at 24 months with revealed symptoms was 46%±4% and without revealed symptoms was 70%±4%. CONCLUSIONS: ETT in patients with moderate or severe AS is safe and tolerable. Serial exercise testing is useful to reveal symptoms not volunteered on the history and adds incremental prognostic information to baseline testing.


Assuntos
Estenose da Valva Aórtica/diagnóstico , Valva Aórtica/fisiopatologia , Teste de Esforço , Idoso , Idoso de 80 Anos ou mais , Valva Aórtica/cirurgia , Estenose da Valva Aórtica/mortalidade , Estenose da Valva Aórtica/fisiopatologia , Estenose da Valva Aórtica/cirurgia , Doenças Assintomáticas , Ecocardiografia , Eletrocardiografia , Teste de Esforço/efeitos adversos , Feminino , Implante de Prótese de Valva Cardíaca , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Intervalo Livre de Progressão , Estudos Retrospectivos , Fatores de Risco , Índice de Gravidade de Doença , Fatores de Tempo , Resultado do Tratamento
12.
Atherosclerosis ; 272: 175-181, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29621698

RESUMO

BACKGROUND AND AIMS: Methionine (Met) is an essential amino acid involved in methylation reactions and lipid metabolism. A Met-deficient diet may cause hepatic lipid accumulation, which is considered an independent risk factor for atherosclerosis. However, the prospective relationship between circulating Met and incident acute myocardial infarction (AMI) is unknown. METHODS: We studied the associations of plasma Met and incident AMI in 4156 patients (77% men; median age 62 years) with stable angina pectoris, among whom the majority received lipid lowering therapy with statins. Risk associations were estimated using Cox-regression analyses. RESULTS: Plasma Met was negatively related to age, serum levels of total cholesterol, low-density lipoprotein cholesterol (LDL-C) and apolipoprotein (apo) B at baseline (all p≤0.05). During a median follow-up of 7.5 years, 534 (12.8%) patients experienced an AMI. There was no overall association between plasma Met and incident AMI; however, plasma Met was inversely associated with risk among patients with high as compared to low levels of serum LDL-C or apo B 100 (multivariate adjusted HRs per SD [95% CI] 0.84 [0.73-0.96] and 0.83[0.73-0.95], respectively; p-interaction ≤0.02). Trends towards an inverse risk relationship were also observed among those younger than 62 years and patients without diabetes or hypertension. CONCLUSIONS: Low plasma Met was associated with increased risk of AMI in patients with high circulating levels of atherogenic lipids, but also in subgroups with presumably lower cardiovascular risk. The determinants of Met status and their relation with residual cardiovascular risk in patients with coronary heart disease should be further investigated.


Assuntos
Lipídeos/sangue , Metionina/sangue , Infarto do Miocárdio/sangue , Infarto do Miocárdio/diagnóstico , Fatores Etários , Idoso , Angina Estável/sangue , Doenças Cardiovasculares/epidemiologia , LDL-Colesterol/sangue , Angiografia Coronária , Doença das Coronárias/sangue , Diabetes Mellitus/sangue , Feminino , Seguimentos , Humanos , Hipertensão/sangue , Masculino , Pessoa de Meia-Idade , Modelos de Riscos Proporcionais , Estudos Prospectivos , Medição de Risco , Fatores de Risco
13.
Thromb Haemost ; 118(4): 778-790, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29458232

RESUMO

Systemic fibrinogen and neopterin are related to inflammation. We investigated the prognostic utility and possible interactions of these biomarkers in stable coronary artery disease (SCAD) patients undergoing coronary angiography. We included 3,545 patients with suspected stable angina with a median follow-up of 7.3 and 10.2 years for incident acute myocardial infarction (AMI) and all-cause mortality, respectively. Prospective associations were explored by Cox regression. Potential effect modifications were investigated according to strata of fibrinogen, neopterin or high-sensitivity troponin T (hsTnT) below and above the median, as well as gender and smoking habits. During follow-up, 543 patients experienced an AMI and 769 patients died. In a multivariable model, the hazard ratios (HRs; 95% confidence interval [CI]) per 1 SD increase for fibrinogen in relation to these endpoints were 1.30 (1.20, 1.42; p < 0.001) and 1.22 (1.13, 1.31; p < 0.001), respectively. For neopterin, the HRs (95% CI) were 1.31 (1.23, 1.40; p < 0.001) and 1.24 (1.15, 1.34; p < 0.001), respectively. No significant interaction between fibrinogen and neopterin was observed. The prognostic utility of neopterin for incident AMI was improved in patients with an hsTnT above the median, for total mortality in non-smokers, and for both total mortality and AMI in females. In conclusion, both fibrinogen and neopterin were associated with future AMI and total mortality, but had low discriminatory impact. No interaction was observed between these two biomarkers. The prognostic utility of neopterin was improved in patients with hsTnT levels above the median, and in females and non-smokers.


Assuntos
Doença da Artéria Coronariana/sangue , Fibrinogênio/análise , Infarto do Miocárdio/sangue , Neopterina/sangue , Idoso , Biomarcadores/sangue , Estudos de Coortes , Doença da Artéria Coronariana/complicações , Doença da Artéria Coronariana/mortalidade , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/complicações , Infarto do Miocárdio/mortalidade , Prognóstico , Modelos de Riscos Proporcionais , Estudos Prospectivos , Risco , Fumar , Troponina T/sangue
15.
J Am Heart Assoc ; 6(11)2017 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-29097387

RESUMO

BACKGROUND: Plasma total homocysteine (tHcy) is related to plasma neopterin, an indicator of interferon-γ-mediated immune activation, and both biomarkers positively predict cardiovascular risk. We examined whether the association between tHcy and subsequent risk of acute myocardial infarction (AMI) was modified by systemic concentrations of neopterin and C-reactive protein among patients with coronary heart disease. METHODS AND RESULTS: By Cox modeling, we explored the association between tHcy and risk of AMI in 4164 patients with suspected stable angina pectoris. Subgroup analyses were performed according to median levels of neopterin and C-reactive protein. A replication study was performed among 3749 patients with AMI at baseline. Median follow-up was 7.3 and 8.3 years among patients with stable angina pectoris and AMI, respectively. tHcy and neopterin correlated in both cohorts (rs=0.34 and rs=0.30 among stable angina pectoris and AMI patients, respectively, both P<0.001). tHcy predicted AMI in both cohorts, independent of B-vitamin treatment. However, significant risk associations were confined to patients with plasma neopterin above the median (hazard ratios [95% confidence interval] per 1-SD increment of log-transformed tHcy 1.38 [1.26-1.50] and 1.18 [1.10-1.26] among stable angina pectoris and AMI patients, respectively) (Pint<0.005 in both cohorts). Further, adding information on the interaction between tHcy and neopterin improved model discrimination and reclassification. tHcy and C-reactive protein were weakly related, and no effect modification was found by C-reactive protein. CONCLUSIONS: Among patients with coronary heart disease, tHcy predicted risk of AMI only in subjects with concomitantly elevated plasma neopterin. Our results motivate further research on the relationship between homocysteine metabolism, cellular immune activation, and atherothrombosis.


Assuntos
Angina Estável/sangue , Homocisteína/sangue , Hiper-Homocisteinemia/sangue , Mediadores da Inflamação/sangue , Infarto do Miocárdio/sangue , Neopterina/sangue , Idoso , Angina Estável/diagnóstico por imagem , Angina Estável/epidemiologia , Biomarcadores/sangue , Proteína C-Reativa/análise , Angiografia Coronária , Feminino , Humanos , Hiper-Homocisteinemia/diagnóstico , Hiper-Homocisteinemia/epidemiologia , Estimativa de Kaplan-Meier , Modelos Lineares , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio/epidemiologia , Noruega , Valor Preditivo dos Testes , Prognóstico , Modelos de Riscos Proporcionais , Estudos Prospectivos , Ensaios Clínicos Controlados Aleatórios como Assunto , Medição de Risco , Fatores de Risco , Regulação para Cima
16.
J Cardiovasc Pharmacol Ther ; 22(2): 179-188, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27390144

RESUMO

AIM: Insulin and glucose may have opposite effects when used to reduce ischemia-reperfusion injury. When insulin is administered alone, feeding state determines tolerance and further induces metabolic and hormonal changes. Higher insulin doses are needed for similar activation of cardioprotective Akt signaling in the fed compared to the fasted pig heart. Thus, the aim of the study was to investigate the effects of 2 prespecified insulin doses on infarct size, apoptosis, metabolism, and cardiac function in a clinically relevant, randomized large animal model using conventional percutaneous catheter intervention techniques and including different fasting states. METHODS AND RESULTS: Twenty-seven female pigs were subjected to 40-minute ischemia and 120-minute reperfusion. Pharmacological postconditioning with intracoronary infusions administered over 3 × 30 seconds was performed at immediate reperfusion. Animals were randomly assigned to 3 groups-preexperimental fasting and intracoronary saline ( controls), preexperimental fasting and 0.1U of insulin ( fasted Ins0.1U), and preexperimental feeding and 1.0U of insulin ( fed Ins1.0U). A significant reduction in infarct size was demonstrated in the fed Ins1.0U group ( P = .047) but not in the fasted Ins0.1U group ( P = .531) compared to controls (infarct size normalized to area at risk ± standard deviation: controls 70.2% ± 12.9%, fasted Ins0.1U 65.0% ± 9.4%, and fed Ins1.0U 54.4% ± 7.3%). Infarct limitation was associated with more uncleaved caspase-3 in the area of risk and the infarcted area, lower circulating free fatty acids, and less increase in heart rate during reperfusion. Fed animals had higher levels of glucose, carnitine, potassium, and normetanephrine and higher heart rate at baseline compared to controls. CONCLUSION: Insulin postconditioning reduced infarct size in the in vivo pig heart, but the beneficial effects were restricted to the highest dose, which is limited by side effects and can only be given to nonfasted animals. The finding challenges successful general use of insulin in the treatment of reperfusion injury in clinical acute myocardial infarction.

17.
Eur Heart J Cardiovasc Imaging ; 18(4): 404-412, 2017 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-27491438

RESUMO

Aims: In aortic valve stenosis (AS), having a small aortic root may influence both the assessment of AS severity and the treatment strategy. The aim was to test the prognostic implications of having a small aortic root in AS within a large prospective study. Methods and results: We used data from 4.3-year follow-up of 1560 patients with asymptomatic, initially mostly moderate AS enrolled in the Simvastatin and Ezetimibe in Aortic Stenosis study. A small aortic root was defined as inner aortic sinotubular junction diameter indexed for body height <1.4 cm/m in women and <1.5 cm/m in men. A small aortic root was found in 270 patients (17.3%) at baseline. Having a small aortic root was associated with larger aortic root wall thickness, higher pressure recovery, lower systemic arterial compliance, left ventricular mass index, and female sex in a multivariable logistic regression analysis (all P < 0.05). In the Cox regression analysis, having a small aortic root at baseline was associated with higher hazard rates of ischaemic cardiovascular events (n = 268; HR 1.55, 95% CI 1.16-2.06), non-haemorrhagic stroke (n = 55; HR 1.88, 95% CI 1.04-3.41), and cardiovascular death (n = 81; HR 2.08, 95% CI 1.28-3.39) (all P < 0.05) after adjusting for confounders, including randomized study treatment, sex, hypertension, AS severity, and aortic valve replacement. Conclusion: In AS patients without known cardiovascular disease or diabetes, having a small aortic root was associated with increased ischaemic cardiovascular events and mortality. The results suggest a relation between the presence of a small aortic root and that of subclinical atherosclerosis. Clinical trial registration: ClinicalTrials.gov identifier: NCT00092677.


Assuntos
Estenose da Valva Aórtica/diagnóstico por imagem , Estenose da Valva Aórtica/tratamento farmacológico , Valva Aórtica/diagnóstico por imagem , Ezetimiba/uso terapêutico , Sinvastatina/uso terapêutico , Idoso , Valva Aórtica/efeitos dos fármacos , Estenose da Valva Aórtica/mortalidade , Estenose da Valva Aórtica/fisiopatologia , Ecocardiografia , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Modelos de Riscos Proporcionais , Estudos Prospectivos , Medição de Risco , Índice de Gravidade de Doença , Taxa de Sobrevida , Fatores de Tempo , Resultado do Tratamento
18.
Biophys J ; 111(8): 1668-1678, 2016 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-27760354

RESUMO

Ligand-protein binding processes are essential in biological systems. A well-studied system is the binding of cyclic adenosine monophosphate to the cyclic nucleotide binding domain of the bacterial potassium channel MloK1. Strikingly, the measured on-rate for cyclic adenosine monophosphate binding is two orders of magnitude slower than a simple Smoluchowski diffusion model would suggest. To resolve this discrepancy and to characterize the ligand-binding path in structural and energetic terms, we calculated 1100 ligand-binding molecular dynamics trajectories and tested two scenarios: In the first scenario, the ligand transiently binds to the protein surface and then diffuses along the surface into the binding site. In the second scenario, only ligands that reach the protein surface in the vicinity of the binding site proceed into the binding site. Here, a binding funnel, which increasingly confines the translational as well as the rotational degrees of freedom, determines the binding pathways and limits the on-rate. From the simulations, we identified five surface binding states and calculated the rates between these surface binding states, the binding site, and the bulk. We find that the transient binding of the ligands to the surface binding states does not affect the on-rate, such that this effect alone cannot explain the observed low on-rate. Rather, by quantifying the translational and rotational degrees of freedom and by calculating the binding committor, our simulations confirmed the existence of a binding funnel as the main bottleneck. Direct binding via the binding funnel dominates the binding kinetics, and only ∼10% of all ligands proceed via the surface into the binding site. The simulations further predict an on-rate between 15 and 40µs-1(mol/l)-1, which agrees with the measured on-rate.


Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , AMP Cíclico/metabolismo , Simulação de Dinâmica Molecular , Canais de Potássio/química , Canais de Potássio/metabolismo , Ligação Proteica , Domínios Proteicos , Rotação
19.
Methods Mol Biol ; 1408: 93-105, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26965118

RESUMO

As a transformative approach in neuroscience and cell biology, optogenetics grants control over manifold cellular events with unprecedented spatiotemporal definition, reversibility, and noninvasiveness. Sensory photoreceptors serve as genetically encoded, light-regulated actuators and hence embody the cornerstone of optogenetics. To expand the scope of optogenetics, ever more naturally occurring photoreceptors are being characterized, and synthetic photoreceptors with customized, light-regulated function are being engineered. Perturbational control over intracellular cyclic-nucleotide-monophosphate (cNMP) levels is achieved via sensory photoreceptors that catalyze the making and breaking of these second messengers in response to light. To facilitate discovery, engineering and quantitative characterization of such light-regulated cNMP actuators, we have developed an efficient fluorometric assay. Both the formation and the hydrolysis of cNMPs are accompanied by proton release which can be quantified with the fluorescent pH indicator 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF). This assay equally applies to nucleotide cyclases, e.g., blue-light-activated bPAC, and to cNMP phosphodiesterases, e.g., red-light-activated LAPD. Key benefits include potential for parallelization and automation, as well as suitability for both purified enzymes and crude cell lysates. The BCECF assay hence stands to accelerate discovery and characterization of light-regulated actuators of cNMP metabolism.


Assuntos
AMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Fluorometria/métodos , Luz , Adenilil Ciclases/metabolismo , Animais , AMP Cíclico/análise , GMP Cíclico/análise , Ensaios Enzimáticos/métodos , Escherichia coli/metabolismo , Fluoresceínas/química , Guanilato Ciclase/metabolismo , Humanos , Hidrólise , Optogenética/métodos , Diester Fosfórico Hidrolases/metabolismo
20.
Clin Chem ; 62(5): 755-65, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26980210

RESUMO

BACKGROUND: Several compounds in the choline oxidation pathway are associated with insulin resistance and prevalent diabetes; however, prospective data are scarce.We explored the relationships between systemic and urinary choline-related metabolites and incident type 2 diabetes in an observational prospective study among Norwegian patients. METHODS: We explored risk associations by logistic regression among 3621 nondiabetic individuals with suspected stable angina pectoris, of whom 3242 provided urine samples. Reclassification of patients was investigated according to continuous net reclassification improvement (NRI >0). RESULTS: After median (25th to 75th percentile) follow-up of 7.5 (6.4-8.7) years, 233 patients (6.4%) were registered with incident type 2 diabetes. In models adjusted for age, sex, and fasting status, plasma betaine was inversely related to new-onset disease [odds ratio (OR) per 1 SD, 0.72; 95% CI, 0.62-0.83; P < 0.00001], whereas positive associations were observed for urine betaine (1.25; 1.09-1.43; P = 0.001), dimethylglycine (1.22; 1.06-1.40; P = 0.007), and sarcosine (1.30; 1.13-1.49; P < 0.001). The associations were maintained in a multivariable model adjusting for body mass index, hemoglobin A1c, urine albumin-to-creatinine ratio, estimated glomerular filtration rate, C-reactive protein, HDL cholesterol, and medications. Plasma betaine and urine sarcosine, the indices most strongly related to incident type 2 diabetes, improved reclassification [NRI >0 (95% CI) 0.33 (0.19-0.47) and 0.16 (0.01-0.31), respectively] and showed good within-person reproducibility. CONCLUSIONS: Systemic and urinary concentrations of several choline metabolites were associated with risk of incident type 2 diabetes, and relevant biomarkers may improve risk prediction.


Assuntos
Colina/metabolismo , Colina/urina , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/urina , Idoso , Diabetes Mellitus Tipo 2/diagnóstico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos
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