Interaction of PSD-95 with potassium channels visualized by fluorescence lifetime-based resonance energy transfer imaging

Resonance energy transfer (RET) has been extensively used to estimate the distance between two different fluorophores. This study demonstrates how protein-protein interactions can be visualized and quantified in living cells by time-correlated single-photon counting (TCSPC) imaging techniques that exploit the RET between appropriate fluorescent labels. We used this method to investigate the association of the potassium inward rectifier channel Kir2.1 and the neuronal PDZ protein PSD-95, which has been implicated in subcellular targeting and clustering of ion channels. Our data show that the two proteins not only colocalize within clusters but also interact with each other. Moreover, the data allow a spatially resolved quantification of this protein-protein interaction with respect to the relative number and the proximity between interacting molecules. Depending on the subcellular localization, a fraction of 20 to 60% of PSD-95 molecules interacted with Kir2.1 channels, approximating their fluorescent labels by less than 5 nm.     

Vitamin D food fortification in European countries: the underused potential to prevent cancer deaths

European Journal of Epidemiology - Background: Meta-analyses of randomized controlled trials have shown that vitamin D supplementation reduces cancer mortality by 13%. Vitamin D fortification of...     

Evidence for an inherent,ATP-stimulated uptake of calcium into chromaffin granules

Summary Partially purified chromaffin granules (granular fraction), crude mitochondria (mitochondrial fraction) and a microsomal fraction were prepared from bovine adrenals by differential centrifugation and characterized by their catecholamine content, succinate dehydrogenase and glucose-6-phosphatase activity. During isotonic incubation with 0.1 mM ⁴⁵Ca²⁺ all fractions showed an uptake of ⁴⁵Ca²⁺, which was stimulated by ATP. In addition, after incubation the granular fraction was further fractionated by sucrose density gradient centrifugation (2.0–1.3 M sucrose; 170,000g·60 min). In some of these experiments the granular fraction was incubated simultaneously with ⁴⁵Ca²⁺ and [³H](-)noradrenaline to compare the uptake of both. The rate of uptake of ⁴⁵Ca²⁺ into the 2.0 M sucrose fraction (characterized by the highest catecholamine content and the lowest succinate dehydrogenase activity) was doubled by ATP. The ATP-stimulated uptake of ⁴⁵Ca²⁺ into the 2.0 M sucrose fraction of chromaffin granules was inhibited by N-ethylmaleimide (NEM) (0.1 mM), 2,4-dinitrophenol (DNP) (0.1 mM), azide (1 mM), carbonyl cyanide mchlorophenylhydrazone (CCCP) (20M), atractyloside (50 M), ruthenium red (40 M) and amobarbital (1 mM). This inhibition pattern was different from that of the ATP-stimulated uptake of ⁴⁵Ca²⁺ into the microsomal fraction, but it was similar to that of the ATP-stimulated uptake of ⁴⁵Ca²⁺ into the mitochondrial fraction. However, the following differences are incompatible with the view that a mitochondrial contamination, with a highly active uptake, is responsible for the ATP-stimulated uptake of ⁴⁵Ca²⁺ into the 2.0 M sucrose fraction of chromaffin granules: a) The uptake of ⁴⁵Ca²⁺ into the mitochondrial fraction was insensitive to 1 mM amobarbital, whereas this agent inhibited the uptake of ⁴⁵Ca²⁺ into the 2.0 M sucrose fraction of chromaffin granules. b) Replacement of ATP by succinate stimulated the uptake of ⁴⁵Ca²⁺ into the mitochondrial fraction only. c) The dependence of the ATP-stimulated uptake of ⁴⁵Ca²⁺ on the concentration of ATP (0.1–5 mM) was determined: while uptake into the 2.0 M sucrose fraction of chromaffin granules exhibited saturation kinetics, that into the mitochondrial fraction was linearly related to the concentration of ATP. Interestingly, uptake of ⁴⁵Ca²⁺ into those fractions of chromaffin granules that are known to be contaminated with mitochondria (1.6 M sucrose fraction) exhibited bot a saturable and a nonsaturable component. d) The uptake of ⁴⁵Ca²⁺ into the mitochondrial fraction was more sensitive to 0.1 mM DNP than the uptake of ⁴⁵Ca²⁺ into the 2.0 M sucrose fraction of chromaffin granules.The comparison of the ATP-stimulated uptake of ⁴⁵Ca²⁺ with that of [³H](-)noradrenaline into the 2.0 M sucrose fraction of chromaffin granules revealed that amobarbital, N,N-dicyclohexylcarbodiimide (DCCD), DNP and increasing concentrations of ATP had the same influence on both uptake processes. From our results we conclude that an inherent, ATP-stimulated uptake of ⁴⁵Ca²⁺ exists in chromaffin granules. The effects of agents on the uptake of ⁴⁵Ca²⁺ and/or [³H](-)noradrenaline into chromaffin granules are discussed with regard to the granular uptake mechanisms.This work was supported by the Deutsche ForschungsgemeinschaftSome results were reported at the 18th Spring Meeting 1977 of the Deutsche Pharmakologische Gesellschaft (Burger and Häusler 1977)     

A Misinterpreted Case of Aorta Prosthesis Endocarditis: Remember The Phenomenon of Microbubbles

A 17‐year‐old male with a history of newly implanted mechanical valve at the aortic position, presented with fever, rigors, and painful cutaneous abscesses on his lower extremities and was suspected for infective endocarditis. Transthoracic echocardiography (TTE) showed a vegetation‐like structure following the movement of the mechanical heart valve (MHV), which eventually proved to be a product of degassing microbubbles (MB).     

Long-Term Risk of Infective Endocarditis After Transcatheter Aortic Valve Replacement

Background

Patients undergoing surgical aortic valve replacement (SAVR) are considered at high risk of infective endocarditis (IE). However, data on the risk of IE following transcatheter aortic valve replacement (TAVR) are sparse and limited by the lack of long-term follow-up as well as a direct comparison with patients undergoing SAVR.

Embolic protection devices for carotid artery stenting: better results than stenting without protection?

AIMS: Carotid artery stenting (CAS) for carotid artery stenoses has become an alternative to carotid endarterectomy. However, CAS itself can cause cerebral ischaemic events. Embolic protection devices (PD) promise to reduce the incidence of these events. METHODS AND RESULTS: From July 1996 to March 2003, 1483 patients from 26 hospitals were included in the prospective CAS Registry of the ALKK study group. A PD was used in 668 of 1483 patients (45%). The use of a PD has grown rapidly over the years and reached 100% in 2003. Patients treated with a PD had prior carotid artery dilatation more often (3.5% versus 1%, p < 0.001), a prior myocardial infarction (34% versus 27.4%, p = 0.007) and a history of arterial hypertension (89.9% versus 78.6%, p = 0.007) compared to patients treated without a PD. A thrombus was more often visible in patients treated under distal protection (16.5% versus 8%, p < 0.001). The use of a PD led to a 10-min longer intervention (45 min versus 35 min median, p < 0.001). Patients treated with a PD had a lower rate of ipsilateral stroke (1.7% versus 4.1%, p = 0.007) and a lower rate of all non-fatal strokes and all deaths (2.1% versus 4.9%, p = 0.004) during the hospital stay. This was confirmed by multiple logistic regression analysis (adjusted OR = 0.45, 95% CI: 0.23-0.91, p = 0.026). A similar reduction could be found for symptomatic as well as asymptomatic carotid artery stenoses. CONCLUSION: Since 1996 there has been a steady increase in the use of PDs for CAS, with a 100% use in 2003. The use of a PD may lower the rate of ipsilateral strokes during CAS.     

Fibrillin-1 genotype and risk of prevalent hypertension: A study in two independent populations

Abstract Objective. Mutations in the fibrillin-1 gene are the cause of Marfan syndrome. We wanted to investigate the relationship between a mutation in this gene and risk of prevalent hypertension. Methods. In a cross-sectional study, the effect of a G-A substitution in intron 27 in the fibrillin-1 gene (rs11856553) on risk of prevalent hypertension was studied in two large population-based studies: the Health 2006 study, consisting of 3193 women and men, age 18-69 years, and the MONICA10 study, consisting of 2408 women and men, age 41-72 years. In 1646 MONICA10 participants, blood pressure (BP) was also measured by 24-h ambulatory recordings. Results. Among the 3193 Health 2006 participants 23 had the G-A variant, and among the 2408 MONICA10 participants 18 had the G-A variant. In Health 2006, the odds ratio estimate (95% confidence intervals) for the G-A variant for risk of hypertension, defined as systolic (S) BP ≥?140 mmHg or diastolic (D) BP ≥?90 mmHg or on antihypertensive medicine, was 2.67 (1.14-6.18), p =?0.022. The corresponding figure for moderate to severe hypertension, defined as SBP ≥?160 mmHg or DBP ≥?100 mmHg, was 9.68 (4.24-22.12), p 相似文献