Structure, sulfatide binding properties, and inhibition of platelet aggregation by a disabled-2 protein-derived peptide.

Disabled-2 (Dab2) targets membranes and triggers a wide range of biological events, including endocytosis and platelet aggregation. Dab2, through its phosphotyrosine-binding (PTB) domain, inhibits platelet aggregation by competing with fibrinogen for α(IIb)ß(3) integrin receptor binding. We have recently shown that the N-terminal region, including the PTB domain (N-PTB), drives Dab2 to the platelet membrane surface by binding to sulfatides through two sulfatide-binding motifs, modulating the extent of platelet aggregation. The three-dimensional structure of a Dab2-derived peptide encompassing the sulfatide-binding motifs has been determined in dodecylphosphocholine micelles using NMR spectroscopy. Dab2 sulfatide-binding motif contains two helices when embedded in micelles, reversibly binds to sulfatides with moderate affinity, lies parallel to the micelle surface, and when added to a platelet mixture, reduces the number and size of sulfatide-induced aggregates. Overall, our findings identify and structurally characterize a minimal region in Dab2 that modulates platelet homotypic interactions, all of which provide the foundation for rational design of a new generation of anti-aggregatory low-molecular mass molecules for therapeutic purposes.

Left ventricular vortex formation is unaffected by diastolic impairment.

Normal left ventricular (LV) filling occurs rapidly early in diastole caused by a progressive pressure gradient within the ventricle and with a low left atrial pressure. This normal diastolic function is altered in patients with heart failure. Such impairment of diastolic filling is manifested as an abrupt deceleration of the early filling wave velocity. Although variations within the early filling wave have been observed previously, the underlying hydrodynamic mechanisms are not well understood. Previously, it was proposed that the mitral annulus vortex ring formation time was the total duration of early diastolic filling and provided a measure of the efficiency of diastolic filling. However, we found that the favorable LV pressure difference driving early diastolic filling becomes zero simultaneously with the deceleration of the early filling wave propagation velocity and pinch-off of the LV vortex ring. Thus we calculated the vortex ring formation time using the duration of the early diastolic filling wave from its initiation to the time of the early filling wave propagation velocity deceleration when pinch-off occurs. This formation time does not vary with decreasing intraventricular pressure difference or with degree of diastolic dysfunction. Thus we conclude the vortex ring pinch-off occurs before the completion of early diastole, and its formation time remains invariant to changes of diastolic function.

Disabled-2 modulates homotypic and heterotypic platelet interactions by binding to sulfatides.

Disabled-2 (Dab2) inhibits platelet aggregation by competing with fibrinogen for binding to the α(IIb) ß(3) integrin receptor, an interaction that is modulated by Dab2 binding to sulfatides at the outer leaflet of the platelet plasma membrane. The disaggregatory function of Dab2 has been mapped to its N-terminus phosphotyrosine-binding (N-PTB) domain. Our data show that the surface levels of P-selectin, a platelet transmembrane protein known to bind sulfatides and promote cell-cell interactions, are reduced by Dab2 N-PTB, an event that is reversed in the presence of a mutant form of the protein that is deficient in sulfatide but not in integrin binding. Importantly, Dab2 N-PTB, but not its sulfatide binding-deficient form, was able to prevent sulfatide-induced platelet aggregation when tested under haemodynamic conditions in microfluidic devices at flow rates with shear stress levels corresponding to those found in vein microcirculation. Moreover, the regulatory role of Dab2 N-PTB extends to platelet-leucocyte adhesion and aggregation events, suggesting a multi-target role for Dab2 in haemostasis.

Assessment of left ventricular diastolic function using 4-dimensional phase-contrast cardiac magnetic resonance.

OBJECTIVES: Phase-contrast magnetic resonance imaging can potentially assess the dynamics of left ventricular (LV) early diastolic filling. METHODS: Fifteen participants underwent phase-contrast magnetic resonance imaging on a 1.5-T whole-body Avanto scanner (Siemens Healthcare, Erlangen, Germany). Left ventricular intracavitary velocities were measured in 3 orthogonal directions. Imaging parameters included a repetition time of 92.45 milliseconds, an echo time of 2.88 milliseconds, a flip angle of 30 degrees, and a velocity-encoding range of 100 to 150 cm/s. RESULTS: The color vector analysis provided a visual assessment of LV diastolic flow. In normal subjects, there was rapid organized early diastolic flow that extended from the mitral valve to the LV apex. In patients with LV diastolic dysfunction, organized high-velocity flow stopped in the mid-left ventricle. CONCLUSIONS: Four-dimensional phase-contrast cardiovascular magnetic resonance can differentiate between normal and abnormal diastolic flow propagation within the left ventricle.

Vortices formed on the mitral valve tips aid normal left ventricular filling.

For the left ventricle (LV) to function as an effective pump it must be able to fill from a low left atrial pressure. However, this ability is lost in patients with heart failure. We investigated LV filling by measuring the cardiac blood flow using 2D phase contrast magnetic resonance imaging and quantified the intraventricular pressure gradients and the strength and location of vortices. In normal subjects, blood flows towards the apex prior to the mitral valve opening, and the mitral annulus moves rapidly away after the valve opens, with both effects enhancing the vortex ring at the mitral valve tips. Instead of being a passive by-product of the process as was previously believed, this ring facilitates filling by reducing convective losses and enhancing the function of the LV as a suction pump. The virtual channel thus created by the vortices may help insure efficient mass transfer for the left atrium to the LV apex. Impairment of this mechanism contributes to diastolic dysfunction, with LV filling becoming dependent on left atrial pressure, which can lead to eventual heart failure. Better understanding of the mechanics of this progression may lead to more accurate diagnosis and treatment of this disease.

Loss of adrenergic augmentation of diastolic intra-LV pressure difference in patients with diastolic dysfunction: evaluation by color M-mode echocardiography.

OBJECTIVES: The aim of this study was to evaluate the hypothesis that the adrenergic response of the intraventricular pressure difference (IVPD) is reduced in patients with preserved ejection fraction (EF) and diastolic dysfunction (DD). BACKGROUND: In early diastole, there is a progressive IVPD extending from the left atrium (LA) to the left ventricular (LV) apex. In response to adrenergic stimulation, as occurs during exercise, the IVPD increases allowing rapid filling without an abnormal increase in LA pressure. Patients with heart failure with a reduced EF have impaired adrenergic augmentation of the IVPD. METHODS: We studied 166 consecutive patients undergoing dobutamine stress echocardiography who had no inducible ischemia and an EF ≥50%, of which 21 had normal diastolic function, 14 had impaired relaxation (grade 1), 80 had pseudonormal filling (grade 2), and 51 had restrictive filling (grade 3). Color M-mode Doppler (CMMD) images of mitral inflow were obtained at rest and during low (10 µg/kg/min) and peak (20 to 40 µg/kg/min) doses of dobutamine. The total IVPD from the LA to LV apex, LA to mid-LV, and mid-LV to the LV apex were calculated using the CMMD data to integrate the Euler equation. RESULTS: Total IVPD was not different between groups at rest. With dobutamine, the total IVPD increased by 2.20 ± 1.95 mm Hg in normal subjects and by only 0.73 ± 1.33 mm Hg, 1.84 ± 1.63 mm Hg, and 1.08 ± 1.57 mm Hg in patients with grades 1, 2, and 3 DD, respectively. This difference was due to a failure in augmentation of IVPD from the mid-LV to the LV apex, indicating reduced apical ventricular suction with DD, whereas the IVPD from the LA to the mid-LV responded similarly to dobutamine in normal subjects and those with DD. CONCLUSIONS: In patients with preserved EF, DD is associated with a reduced adrenergic augmentation of the IVPD from the mid-LV to the LV apex, reflecting less apical suction.

Evaluation of LV diastolic function from color M-mode echocardiography.

OBJECTIVES: this study evaluated early diastolic filling dynamics using a semiautomated objective analysis of filling velocities obtained from color M-mode echocardiography. BACKGROUND: diastolic function can be evaluated from color M-mode echocardiography by measuring the early diastolic flow propagation velocity (Vp) from the slope of a single linear approximation of an isovelocity contour. However, this method has limitations and may not accurately represent diastolic filling. METHODS: we used a semiautomated objective analysis of color M-mode echocardiograms from a development cohort of 125 patients with varying diastolic function to quantify left ventricular filling velocities. Early diastolic filling was not accurately described with a single propagation velocity; instead, the rapid initial filling velocity abruptly decelerated to a slower terminal velocity. Then, we evaluated a new measure of diastolic function in a separate group of 160 patients. RESULTS: compared with normal filling, diastolic dysfunction with restricted filling had a lower initial velocity (53 ± 21 cm/s vs. 87 ± 29 cm/s, p < 0.001), and the deceleration point occurred closer to the mitral annulus (2.4 ± 0.6 cm vs. 3.1 ± 0.7 cm, p < 0.05). The product of the initial velocity and the distance to the deceleration point from the mitral annulus, indicating the strength of the early filling (Vs), was progressively reduced with diastolic dysfunction. In a separate validation cohort of 160 patients, Vs better recognized diastolic dysfunction (classified by reduced diastolic intraventricular pressure gradient, elevated pulmonary capillary wedge pressure, or elevated B-type natriuretic peptide) than Vp did. CONCLUSIONS: early diastolic flow propagation occurs with an initial rapid velocity that abruptly decelerates to a terminal velocity. With diastolic dysfunction, the initial velocity is slower and the deceleration point occurs closer to the mitral annulus than with normal filling. A new parameter that combines these 2 effects (Vs) provides a more accurate assessment of diastolic function than the conventional propagation velocity.

In vitro comparison of the effect of stent configuration on wall shear stress using time-resolved particle image velocimetry.

Time resolved particle image velocimetry was used to measure wall shear stress (WSS) and oscillatory shear index (OSI) within a 3.0 mm diameter compliant vessel model implanted with an Abbott Vascular XIENCE V stent in five configurations: baseline, over-expanded, increased vessel diameter, two overlapped stents, and increased stent length. Flow through unstented vessels was also tested for comparison. Flow conditions featured a realistic coronary pressure-flow offset and reversal at average flow rates corresponding to resting (Re=160, f=70 bpm) and exercise conditions (Re=300, f=120 bpm). Comparisons revealed that the WSS was similar for all cases behind the first strut and downstream of the device, indicating that changes in configuration have little effect downstream. However, there were notable differences within each stent revealing reduced WSS values for all cases due to the stent-imposed expansion of the vessel wall (0.20-9.29 dynes/cm2 for Re=160 and d=3.0 mm). Over-expanding the stent with a second balloon affected the alignment of the stent geometry, and led to higher WSS at the inlet and lower values at mid-stent. The overlapped stents showed disturbed flow and a WSS deficit region downstream of the overlapped region. Analysis of the longer stent showed that the WSS within the vessel recovers with distance. An overall correlation was noted between decreased WSS values and elevated OSI. Results of this study are important because decreased WSS has been implicated in endothelial cell changes and increased restenosis, and clinical research has shown that a link exists between deployment configurations and negative patient outcomes.

In vitro, time-resolved PIV comparison of the effect of stent design on wall shear stress.

The effect of stent design on wall shear stress (WSS) and oscillatory shear index (OSI) was studied in vitro using time-resolved digital particle image velocimetry (DPIV). Four drug-eluting stents [XIENCE V (Abbott Vascular), TAXUS Liberté (Boston Scientific), Endeavor (Medtronic), and Cypher (J&J Cordis)] and a bare-metal stent [VISION (Abbott Vascular)] were implanted into compliant vessel models, and the flow was measured in physiologically accurate coronary conditions featuring reversal and realistic offsets between pressure and flowrate. DPIV measurements were made at three locations under two different flow rates (resting: Re = 160, f = 70 bpm and exercise: Re = 300, f = 120 bpm). It was observed that design substantially affected the WSS experienced at the vessel walls. Averaged values between struts ranged from 2.05 dynes/cm(2) (Cypher) to 8.52 dynes/cm(2) (XIENCE V) in resting conditions, and from 3.72 dynes/cm(2) (Cypher) to 14.66 dynes/cm(2) (VISION) for the exercise state. Within the stent, the WSS dropped and the OSI increased immediately distal to each strut. In addition, an inverse correlation between average WSS and OSI existed. Comparisons with recently published results from animal studies show strong correlation between the measured WSS and observed endothelial cell coverage. These results suggest the importance of stent design on the WSS experienced by endothelial cells in coronary arteries.