Citations

AbstractNatural killer (NK) cells are a subpopulation of lymphocytes that can mediate cytotoxicity of certain tumor cells, virus‐infected cells, and normal cells. In addition to their cytotoxic potential, NK cells secrete a variety of cytokines and chemokines that can modulate the function, growth, and differentiation of other immune cells. These different responses are initiated by the interaction of specific NK surface receptors with defined soluble or cell‐associated ligands. There are several different types of receptors on the NK cell surface including “triggering” receptors, adhesion molecules, cytokine receptors, and MHC‐recognizing killer‐cell inhibitory receptors. The functional response of an NK cell is the result of the integration of signals transduced by these different types of receptors. Some of these signaling pathways are similar to other lymphoid cells, but there are also unique features employed by NK cells. This overview focuses on receptor‐initiated signaling pathways that modulate NK functions.

AbstractCholecystokinin (CCK) and gastrin are structurally related peptide hormones synthesized and secreted by endocrine cells in the gastric antrum, the proximal small intestine, and the central and enteric nervous systems. A broad spectrum of cells express receptors and represent physiologic targets for these peptides. The major physiologic targets for gastrin in the oxyntic gastric mucosa, including the parietal cells and select enterochromaffin cells, while CCK has a broader range of targets, with most involved in nutrient assimilation and appetite control, and the regulation of anxiety. This unit describes binding assays for the determination of receptor number, ligand affinity, and inhibition constants for both CCKA and CCKB receptors, in addition to procedures detailing the preparation of tissues from various sources.

AbstractRadioligand binding assays can be used to characterize receptors in their natural environment (wild type) as well as those transfected into cell lines. They can be used to study receptor dynamics and localization, to identify novel chemical structures that interact with receptors, and to define ligand activity and selectivity. The technique thus serves as a valuable adjunct to other pharmacological and molecular biological approaches. This unit describes the major issues involved in developing a binding assay, such as selecting a radioligand, tissue preparation, optimization of assay parameters, and separation of bound from free ligand. Methods for analysis of the binding data are carefully considered, and tips for troubleshooting a binding assay are also provided.

AbstractThis unit provides a set of protocols for introducing recombinant genes into normal, injured, and atherosclerotic arteries. The protocols include animal preparation, surgical techniques, and delivery systems. Protocols describe gene delivery to normal, injured, and stented porcine iliofemoral arteries, employing a double balloon infusion catheter to deliver the vector. Another basic protocol describes gene delivery to atherosclerotic arteries using a hyperlipidemic double‐injury rabbit model, and requires surgical exposure of the artery and instillation of the gene vector via a catheter. Additional protocols describe gene delivery to normal and injured murine carotid and femoral arteries. An describes a percutaneous method for arterial gene delivery. These protocols may be adapted to deliver genes to either injured or noninjured atherosclerotic arteries.

AbstractThis unit describes three methods for the detection of caspase activation as cells undergo apoptosis. Simple and relatively quantitative enzymatic assays are provided using suitable substrates. Because the various low‐molecular‐weight substrates available for these assays are not selective, however, the assays do not accurately distinguish between various caspases. Immunoblotting is described for following the activation of specific caspases. When coupled with subcellular fractionation, this method can provide large amounts of temporal and spatial information about caspase activation. Finally, affinity labeling protocols are provided for detecting active caspases in whole‐cell lysates or subcellular fractions.

AbstractUnderstanding of transmembrane signaling during NK‐cell activation has greatly expanded during the past few years. The discovery and characterization of novel triggering and inhibitory receptors have revealed the complexity of these processes. This unit focuses on receptor‐initiated signaling pathways that modulate NK functions. Establishing the roles of different signaling pathways in NK cells is a crucial step in the design of therapeutic approaches for selective enhancement or suppression of NK‐cell activation.

Background— Preclinical systolic or diastolic dysfunction is associated with increased morbidity and mortality. We postulated that plasma brain natriuretic peptide (BNP) might serve as a biomarker for preclinical ventricular dysfunction (PCVD) but that the discriminatory values for BNP may vary with age and sex. Methods and Results— We measured BNP, systolic and diastolic ventricular function, and clinical parameters in 2042 randomly selected residents of Olmsted County, Minn, aged 45 years or older. For preclinical systolic dysfunction, the areas under the receiver operating characteristics curve were higher for those with more severe (0.82 to 0.92) than any (0.51 to 0.74) systolic dysfunction and were similar in men and women and in younger and older persons. For preclinical diastolic dysfunction, the areas under the receiver operating characteristics curve were higher for those with moderate-to-severe (0.74 to 0.79) than any (0.52 to 0.68) diastolic dysfunction and were similar regardless of age or sex. Optimal discriminatory values of BNP varied with age and sex. Considering the prevalence of preclinical systolic or diastolic dysfunction and the predictive characteristics observed, using BNP to screen for PCVD would necessitate echo in 10% to 40% of those screened, with most confirmatory echocardiograms being negative, and would miss 10% to 60% of those affected. Conclusions— BNP is a suboptimal screening test for PCVD in the population.

Background— In this study we evaluate the diastolic deformation of ischemic/reperfused myocardium and relate this deformation to tissue elastic properties. Methods and Results— Farm pigs were subjected to left anterior descending coronary artery occlusion followed by reperfusion to create either stunning (n=12) or transmural myocardial infarction (n=12). Ultrasound-derived radial strain rates (SR) and strain were measured in the ischemic and remote walls. Myocardial stiffness was estimated from diastolic pressure–wall thickness relationship obtained from preload alterations. At reperfusion, end-systolic strain (ε sys ) was significantly reduced in both stunned and infarcted walls compared with their remote walls (3±3% versus 26±2% and 1±0% versus 33±5%, respectively; P <0.0001) or baseline values. Diastolic passive deformation (ε A ) and rates of deformation during early (E SR ) and late (A SR ) diastole were comparable between stunned and remote walls (ε A : 7.3±1.6% versus 7.9±1.9%; E SR : −2.7±0.4 s −1 versus −2.6±0.5 s −1 ; A SR : −1.8±0.2 s −1 versus −1.9±0.3 s −1 ; P =NS for all) but were of significantly lower magnitude in infarcted walls versus remote walls (ε A : 1.1±0.2% versus 11.4±1.9%; E SR : −0.3±0.1 s −1 versus −2.4±0.4 s −1 ; A SR : −0.3±0.1 s −1 versus −2.5±0.4 s −1 ; P <0.0001 for all). Stiffness coefficient of exponential diastolic pressure–wall thickness relation was higher for infarcted ( P <0.05) but not for stunned walls ( P =NS) compared with their remote walls. Conclusions— Early after postischemic reperfusion and in the presence of severely reduced systolic deformation, diastolic passive deformation (and rates of deformation) can distinguish stiff, noncompliant, transmurally infarcted myocardial walls from those more compliant walls containing viable but stunned myocardium.

Background— Restenosis prediction from published studies is hampered by inadequate sample size and incomplete angiographic follow-up. The prediction of restenosis with the existing variables is poor. The aim of the present study was to include the clinical and angiographic variables commonly associated with angiographic restenosis and develop a prediction model for restenosis from the PRESTO database. Methods and Results— This study included 1312 patients with a single lesion enrolled in the angiographic substudy of the PRESTO trial. We constructed 2 risk scores. The first used preprocedural variables (female gender, vessel size [≤2.5 mm, 2.5 to 3 mm, 3 to 3.5 mm, 3.5 to 4 mm, >4 mm], lesion length >20 mm, diabetes, smoking status, type C lesion, any previous percutaneous coronary intervention [PCI], and unstable angina) derived from previous studies. Estimated restenosis rates and corresponding variability for each possible level of the resultant risk score were obtained via bootstrapping techniques. The area under the receiver-operator characteristic (ROC) curve was 0.63, indicating modest discriminatory ability to predict restenosis. The second approach constructed a multiple logistic regression model considering significant univariate clinical and angiographic predictors of restenosis identified from the PRESTO database (treated diabetes mellitus, nonsmoker, vessel size, lesion length, American College of Cardiology/American Heart Association type C lesion, ostial location, and previous PCI). The area under the ROC curve for this risk score was also 0.63. Conclusions— The preprocedural clinical and angiographic variables from available studies and from the PRESTO trial have only modest predictive ability for restenosis after PCI.

Abstract Retrospective studies have suggested that combination antibiotic therapy for severe bacteremic pneumococcal pneumonia may reduce mortality. We assessed this issue in a prospective, multicenter, international observational study of 844 adult patients with bacteremia due to Streptococcus pneumoniae. The effect of combination antibiotic therapy versus monotherapy on mortality was examined by univariate analyses and by logistic regression models. The 14-day mortality was not significantly different for the two groups. However, among critically ill patients, combination antibiotic therapy was associated with lower 14-day mortality (23.4 versus 55.3%, p = 0.0015). This improvement in survival was independent of country of origin, intensive care unit support, class of antibiotics, or in vitro activity of the antibiotics prescribed. Combination antibiotic therapy improved survival among critically ill patients with bacteremic pneumococcal illness.