Interleukin 10-coated nanoparticle systems compared for molecular imaging of atherosclerotic lesions.

Atherosclerosis (AS) is one of the leading causes of mortality in high-income countries. Early diagnosis of vulnerable atherosclerotic lesions is one of the biggest challenges currently facing cardiovascular medicine. The present study focuses on developing targeted nanoparticles (NPs) in order to improve the detection of vulnerable atherosclerotic-plaques. Various biomarkers involved in the pathogenesis of atherosclerotic-plaques have been identified and one of these promising candidates for diagnostic targeting is interleukin 10 (IL10). IL10 has been shown to be a key anti-inflammatory responding cytokine in the early stages of atherogenesis, and has already been used for therapeutic interventions in humans and mice. IL10, the targeting sequence, was coupled to two different types of NPs: protamine-oligonucleotide NPs (proticles) and sterically stabilized liposomes in order to address the question of whether the recognition and detection of atherosclerotic-lesions is primarily determined by the targeting sequence itself, or whether it depends on the NP carrier system to which the biomarker is coupled. Each IL10-targeted NP was assessed based on its sensitivity and selectivity toward characterizing atherosclerotic-plaque lesions using an apolipoprotein E-deficient mouse as the model of atherosclerosis. Aortas from apolipoprotein E-deficient mice fed a high fat diet, were stained with either fluorescence-labeled IL10 or IL10-coupled NPs. Ex vivo imaging was performed using confocal laser-scanning microscopy. We found that IL10-targeted proticles generated a stronger signal by accumulating at the surface of atherosclerotic-plaques, while IL10-targeted, sterically stabilized liposomes showed a staining pattern deeper in the plaque compared to the fluorescence-labeled IL10 alone. Our results point to a promising route for enhanced in vivo imaging using IL10-targeted NPs. NPs allow a higher payload of signal emitting molecules to be delivered to the atherosclerotic-plaques, thus improving signal detection. Importantly, this allows for the opportunity to visualize different areas within the plaque scenario, depending on the nature of the applied nanocarrier.

Rate of creatinine equilibration in whole blood.

The classic assumption that a large fraction of blood creatinine remains sequestered within erythrocytes when blood is dialyzed has been challenged by recent observations where approximately 60% of erythrocyte water appeared accessible to diffusive creatinine transport during a dialyzer transit. This discrepancy provided the motivation to revisit and reanalyze the equilibration of creatinine across the erythrocyte membrane in a series of in vitro studies with normal human blood under erythrocyte loading and unloading conditions at 37 degrees C. The time course of plasma creatinine concentrations measured by a kinetic picric acid assay was analyzed using a 2-compartment model. In 7 experiments, the equilibration constant was 0.052 +/- 0.013/min, corresponding to a mean half-life of 13.8 +/- 2.8 minutes, and comparable for erythrocyte loading and unloading. With these values and with mean dialyzer transit times in the range of 20 seconds the fraction of erythrocyte water accessible to diffusive clearance is in the range of 2%. These results are comparable to what has been measured with radiolabeled markers almost half a century ago. Therefore, when dialyzer outlet concentrations are sampled without equilibration the effective diffusion volume flow rate for creatinine is close to plasma water flow and does not include sizeable fractions of erythrocyte water flow.

Effect of hemolysis and free hemoglobin on optical hematocrit measurements in the extracorporeal circulation.

Clinically significant hemolysis is a rare but serious problem in dialysis. Because hemolysis affects red blood cell count and optical density of plasma it has been speculated whether techniques used for online blood volume monitoring would be useful to detect hemolysis. In this study the influence of free hemoglobin on hematocrit and relative blood volume changes measured by optical means (CritLine, HemaMetrics, Kaysville, UT) were examined using an in vitro model with bovine blood. Free hemoglobin solutions were added in steps to circulating whole blood at baseline hematocrits covering a range from 30% to 60% and at blood flows of approximately 200 and 400 ml/min. The free hemoglobin concentration reached was in the range of 2 to 3 g/dl. The presence of free hemoglobin led to a relative increase in hematocrit in the range of 0.3% per 0.1 g of free hemoglobin per dl (+3% dl/g). As an increase in hematocrit is interpreted as a decrease in blood volume, this change referred to an apparent decrease in relative blood volume in the same order of magnitude (-3% dl/g). Effects were more pronounced at low baseline hematocrit. Thus, although optical hematocrit readings are affected by the presence of free hemoglobin the changes at levels associated with clinical symptoms appear to be too small to be accurately detected in the in vivo situation where the hematocrit and the resulting optical signal is affected by various physiological processes and therefore much noisier.

Effects of beta2-glycoprotein-I on platelet aggregation in cord versus adult whole blood.

We present a peculiarity of the neonatal hemostatic system that might contribute to establish a procoagulant readiness in neonatal blood by sensitizing neonatal platelets for ADP stimulation. beta2-glycoprotein-I (beta2-GP-I) is a plasma constituent capable of suppressing ADP-induced platelet aggregation. We found significant lower levels of beta2-GP-I in cord vs. adult plasma (120 +/- 27 vs. 180 +/- 37 microg/mL, P<0.001). We demonstrate dose-dependent inhibition of ADP-induced platelet aggregation in cord whole blood (WB) in the presence of increasing amounts of beta2-GP-I, evaluated by means of WB aggregometry employing the impedance method. Particularly, raising the beta2-GP-I concentration in cord WB from neonatal level up to the respective adult value caused significant reduction of amplitude (from 9.5 +/- 2.7 to 2.8 +/- 0.9 Omega, P<0.001) and of slope (from 5.9 +/- 2.4 to 1.89 +/- 0.9 Omega/min, P<0.001), and a significant prolongation of the aggregation time (from 51.8 +/- 22.9 to 110.8 +/- 60.3 s, P<0.001). In conclusion, physiological low levels of beta2-GP-I in cord WB cause enhanced responsiveness of neonatal platelets to ADP stimulation. This mechanism might help to explain the clinically observed well-functioning hemostasis in neonates.