PROSPER delivery of universal preventive interventions with young adolescents: long-term effects on emerging adult substance misuse and associated risk behaviors.

BACKGROUND: Substance misuse and associated health-risking behaviors are prevalent in emerging adulthood. There is a knowledge gap concerning the post-high school effects of community-based delivery systems for universal preventive interventions implemented during young adolescence. This study reports effects of the PROSPER delivery system through age 19, 7.5 years past baseline. METHODS: A cohort sequential design included 28 public school districts randomly assigned to the PROSPER partnership delivery system or usual-programming conditions. PROSPER community teams implemented a family-focused intervention in 6th grade and a school-based intervention in 7th grade. Outcomes for the age 19, post-high school report included lifetime, current, and frequency of substance misuse, as well as antisocial and health-risking sexual behaviors. Intent-to-treat, multi-level analyses of covariance of point-in-time outcomes were conducted, along with analyses of risk-related moderation of intervention effects. RESULTS: Results showed emerging adults from PROSPER communities reported significantly lower substance misuse across a range of types of substances, with relative reduction rates of up to 41.0%. No significant findings were observed for associated antisocial and health-risking sexual behavior indices; or for lifetime rates of sexually transmitted infections. Risk-related moderation effects were non-significant, suggesting generally comparable outcomes across higher- and lower-risk subgroups of emerging adults. CONCLUSIONS: The PROSPER delivery system for brief universal preventive interventions has potential for public health impact by reducing long-term substance misuse, with positive results extending beyond high school.

Lipopolysaccharide is a potent monocyte/macrophage-specific stimulator of human immunodeficiency virus type 1 expression.

Lipopolysaccharide (LPS) potently stimulates human immunodeficiency virus type 1-long terminal repeat (HIV-1-LTR) CAT constructs transfected into monocyte/macrophage-like cell lines but not a T cell line. This effect appears to be mediated through the induction of nuclear factor kappa B (NF-kappa B). Electrophoretic mobility shift assays demonstrate that LPS induces a DNA binding activity indistinguishable from NF-kappa B in U937 and THP-1 cells. LPS is also shown to dramatically increase HIV-1 production from a chronically infected monocyte/macrophage-like cloned cell line, U1, which produces very low levels of HIV-1 at baseline. The stimulation of viral production from this cell line occurs only if these cells are treated with granulocyte/macrophage colony-stimulating factor (GM-CSF) before treatment with LPS. This stimulation of HIV-1 production is correlated with an increase in the level of HIV-1 RNA and and activation of NF-kappa B. LPS is not able to induce HIV-1 production in a cloned T cell line. The effect of LPS on HIV-1 replication occurs at picogram per milliliter concentrations and may be clinically significant in understanding the variability of the natural history of HIV-1 infection.

The human immunodeficiency virus-1 nef gene product: a positive factor for viral infection and replication in primary lymphocytes and macrophages.

Considerable controversy and uncertainty have surrounded the biological function of the Human Immunodeficiency Virus (HIV)-1 nef gene product. Initial studies suggested that this early, nonstructural viral protein functioned as a negative regulatory factor; thus, it was proposed to play a role in establishing or maintaining viral latency. In contrast, studies in Simian Immunodeficiency Virus (SIV)mac-infected rhesus monkeys have suggested that Nef is not a negative factor but rather plays a central role in promoting high-level viral replication and is required for viral pathogenesis in vivo. We sought to define a tissue culture system that would approximate the in vivo setting for virus infection in order to assess the role of HIV-1 Nef in viral replication. We show that infection of mitogen-activated peripheral blood mononuclear cells (PBMC) with Nef+ HIV results in enhanced replication as evidenced by earlier gag p24 expression when compared with infections performed with nef mutant viruses. Moreover, when unstimulated freshly isolated PBMC are infected with Nef+ and Nef- viruses and then subsequently activated with mitogen, the Nef-induced difference in viral replication kinetics is even more pronounced, with the Nef- viruses requiring much more time in culture for appreciable growth. A positive effect of Nef on viral replication was also observed in primary macrophages infected with a recombinant of YU-2, a patient-derived molecular clone with macrophage tropism. These positive effects of Nef on viral replication are dependent on the initial multiplicity of infection (MOI), in that infections of unstimulated PBMC at low MOI are most dependent upon intact nef for subsequent viral growth. We now provide evidence that the Nef+ HIV is more infectious than Nef- HIV from both a tissue culture infectious dose analysis, and a single-cell HIV infection assay. In the latter case, we demonstrate that infection with equivalent doses of HIV based on virion-associated gag p24 yields five- to sixfold more infected cells if Nef+ viral stocks were used. Furthermore, we find that the differential infectivity is not dependent on CD4 down-regulation as Nef+ virus produced from transfected COS cells lacking CD4 is also more infectious. However, normalization of PBMC infections to equivalent infectivity between that of the Nef+ and Nef- viruses continues to reveal delayed viral replication in the absence of Nef, suggesting that secondary viral spread in PBMC is also enhanced in Nef+ infections. We demonstrate this directly by showing a 13-15-fold increase in infectivity of PBMC-derived Nef+ HIC.(ABSTRACT TRUNCATED AT 400 WORDS)

Activation of virus replication after vaccination of HIV-1-infected individuals.

Little is known about the factors that govern the level of HIV-1 replication in infected individuals. Recent studies (using potent antiviral drugs) of the kinetics of HIV-1 replication in vivo have demonstrated that steady-state levels of viremia are sustained by continuous rounds of de novo infection and the associated rapid turnover of CD4+ T lymphocytes. However, no information is available concerning the biologic variables that determine the size of the pool of T cells that are susceptible to virus infection or the amount of virus produced from infected cells. Furthermore, it is not known whether all CD4+ T lymphocytes are equally susceptible to HIV-1 infection at a given time or whether the infection is focused on cells of a particular state of activation or antigenic specificity. Although HIV-1 replication in culture is known to be greatly facilitated by T cell activation, the ability of specific antigenic stimulation to augment HIV-1 replication in vivo has not been studied. We sought to determine whether vaccination of HIV-1-infected adults leads to activation of virus replication and the targeting of vaccine antigen-responsive T cells for virus infection and destruction. Should T cell activation resulting from exposure to environmental antigens prove to be an important determinant of the steady-state levels of HIV-1 replication in vivo and lead to the preferential loss of specific populations of CD4+ T lymphocytes, it would have significant implications for our understanding of and therapeutic strategies for HIV-1 disease. To begin to address these issues, HIV-1-infected individuals and uninfected controls were studied by measurement of immune responses to influenza antigens and quantitation of virion-associated plasma HIV-1 RNA levels at baseline and at intervals after immunization with the trivalent influenza vaccine. Influenza vaccination resulted in readily demonstrable but transient increases in plasma HIV-1 RNA levels, indicative of activation of viral replication, in HIV-1-infected individuals with preserved ability to immunologically respond to vaccine antigens. Activation of HIV-1 replication by vaccination was more often seen and of greater magnitude in individuals who displayed a T cell proliferative response to vaccine antigens at baseline and in those who mounted a significant serologic response after vaccination. The fold increase in viremia, as well as the rates of increase of HIV-1 in plasma after vaccination and rates of viral decline after peak viremia, were higher in individuals with higher CD4+ T cell counts.(ABSTRACT TRUNCATED AT 400 WORDS)

Engineering poliovirus as a vaccine vector for the expression of diverse antigens.

As a step toward developing poliovirus as a vaccine vector, poliovirus recombinants were constructed by fusing exogenous peptides (up to 400 amino acids) and an artificial cleavage site for viral protease 3Cpro to the amino terminus of the viral polyprotein. Viral replication proceeded normally. An extended polyprotein was produced in infected cells and proteolytically processed into the complete array of viral proteins plus the foreign peptide, which was excluded from mature virions. The recombinants retained exogenous sequences through successive rounds of replication in culture and in vivo. Infection of animals with recombinants elicited a humoral immune response to the foreign peptides.

Harmonization of strategies for the validation of quantitative analytical procedures: a SFSTP proposal part IV. Examples of application.

A harmonized approach for the validation of analytical methods based on accuracy profile was introduced by a SFSTP commission on the validation of analytical procedure. This fourth and last document aims at illustrating this methodology and the statistics used. Therefore the validation of real case methods are proposed such as methods for the quality control of drugs, for the quantitation of impurities in drug substances, for bioanalysis or for the determination of nutriments. Furthermore, different types of analytical methods are used in order to demonstrate the applicability of the proposed approach to a wide range of methods such as liquid chromatography (LC-UV, LC-MS), spectrophotometry or ELISA.

Factors associated with recent unsuppressed viral load in HIV-1-infected patients in care on first-line antiretroviral therapy in South Africa.

Unsuppressed viral load (VL) in patients on antiretroviral therapy (ART) occurs when treatment fails to suppress a person's VL and is associated with decreased survival and increased HIV transmission. The objective of this study was to evaluate factors associated with unsuppressed VL (VL > 400 copies/ml) in patients currently in care on first-line ART for ≥ 6 months attending South African public healthcare facilities. We analysed electronic medical records of ART patients with a VL result on record who started ART between January 2004 and April 2016 from 271 public health facilities. We present descriptive and multivariable logistic regression for unsuppressed VL at last visit using a priori variables. We included 244,370 patients (69% female) on first-line ART in April 2016 for ≥ 6 months. Median age at ART start was 33 years (7% were < 15 years old). Median duration on ART was 3.7 years. Adjusting for other variables, factors associated with having an unsuppressed VL at the most recent visit among patients in care included: (1) < 15 years old at ART start (adjusted odds ratio [aOR]=2.58; 95% CI = 2.37, 2.81) versus 15-49 years at ART start, (2) male gender (aOR = 1.29; 95% CI = 1.25, 1.35), (3) 6-12 months on ART versus longer (aOR = 1.34; 95% CI = 1.29, 1.40), (4) on tuberculosis (TB) treatment (aOR = 1.78; 95% CI = 1.48, 2.13), and (5) prior ART exposure versus none (aOR = 1.20; 95% CI = 1.08, 1.32). Approximately 85% of the ART cohort who were in care had achieved viral suppression, though men, youth/adolescents, patients with prior ART exposure, those with short duration of ART, and patients on TB treatment had increased odds of not achieving viral suppression. There is a need to develop and evaluate targeted interventions for ART patients in care who are at high risk of unsuppressed VL.

AIDS: decline and fall of immune surveillance?

Recent observations cast doubt on the view that cytotoxic T cells play a key role in keeping HIV-1 infection in check, and that it is the decline in this mechanism of immune surveillance that permits progression to AIDS.

Donor MHC and adhesion molecules in transplant arteriosclerosis.

Transplant-associated arteriosclerosis remains an obstacle to long-term graft survival. To determine the contribution to transplant arteriosclerosis of MHC and adhesion molecules from cells of the donor vasculature, we allografted carotid artery loops from six mutant mouse strains into immunocompetent CBA/CaJ recipients. The donor mice were deficient in either MHC I molecules or MHC II molecules, both MHC I and MHC II molecules, the adhesion molecule P-selectin, intercellular adhesion molecule (ICAM)-1, or both P-selectin and ICAM-1. Donor arteries in which ICAM-1, MHC II, or both MHC I and MHC II were absent showed reductions in neointima formation of 52%, 33%, and 38%, respectively, due primarily to a reduction in smooth muscle cell (SMC) accumulation. In P-selectin-deficient donor arteries, neointima formation did not differ from that in controls. In donor arteries lacking both P-selectin and ICAM-1, the size of the neointima was similar to that in those lacking ICAM-1 alone. In contrast, neointima formation increased by 52% in MHC I-deficient donor arteries. The number of CD4-positive T cells increased by 2.8-fold in MHC I-deficient arteries, and that of alpha-actin-positive SMCs by twofold. These observations indicate that ICAM-1 and MHC II molecules expressed in the donor vessel wall may promote transplant-associated arteriosclerosis. MHC I molecules expressed in the donor may have a protective effect.

Molecular insights into human immunodeficiency virus type 1 pathogenesis.

Infection with human immunodeficiency virus type 1 leads to a persistent but progressive cytopathic process that culminates in the near complete destruction of the CD4+ subset of T cells. The levels of human immunodeficiency virus type 1 replication and virus burden increase throughout the clinical course of disease reflecting a balance between the viral and cellular regulatory influences as well as the ability of the host immune system to eliminate infected T cells. Human immunodeficiency virus type 1 replication is dependent on the state of cellular activation and involves both inducible host cell derived transcription factors and at least three virus-derived gene products. Further study of the mechanism of action of these factors, particularly those encoded by the virus, may facilitate the future development of highly specific and effective therapies for human immunodeficiency virus type 1.

Obese adults with primary growth hormone resistance (Laron Syndrome) have normal endothelial function.

OBJECTIVE: Classic Laron Syndrome (LS) is a recessive disease of insulin-like growth factor I (IGF-I) deficiency and primary growth hormone insensitivity, clinically characterized by dwarfism and marked obesity. The aim of the current study was to investigate the impact of long-term IGF-I deficiency on flow-mediated dilation (FMD) in 11 non-IGF-I-treated LS adults with long-term IGF-I deficiency who on stress echocardiography were found to have reduced cardiac dimensions and output, but normal left ventricular (LV) ejection fraction at rest and LV contractile reserve following stress. DESIGN: Following an overnight fast we assessed percent improvement in endothelium-dependent FMD (%FMD) and endothelium-independent nitroglycerin (%NTG)-mediated vasodilation non-invasively in the brachial artery, using high resolution ultrasound in 11 non-treated adult patients with LS without known coronary artery disease, and compared them to 11 age- and sex-matched healthy controls. All subjects underwent symptom-limited exercise testing (Bruce protocol). RESULTS: LS patients had a significantly higher body mass index (29+/-6 vs. 25+/-2 kg/m(2), p=0.04), lower low-density lipoprotein cholesterol (142+/-28 vs. 176+/-12 mg/dl, p=0.03) and a smaller mean brachial artery diameter (4.63+/-0.72 vs. 5.70+/-1.06 mm, p=0.01) compared to controls. However, brachial artery %FMD and %NTG were not significantly different between the LS patients and controls (13.1+/-6.2% vs. 15.4+/-5.2%, p=0.28 and 22.3+/-6.0% vs. 18.9+/-6.2%, p=0.30; respectively). Cardiac performance, assessed by exercise duration time and metabolic equivalents (METs), was significantly greater in control subjects than in LS patients (10.3+/-2.0 vs. 6.0+/-1.4 min, p<0.01 and 10.2+/-2.0 vs. 7.2+/-1.4 METs, p<0.01; respectively). CONCLUSIONS: FMD was found to be within normal limits in non-IGF-I-treated adult patients with LS, despite congenital absence of IGF-I and obesity.

Harmonization of strategies for the validation of quantitative analytical procedures. A SFSTP proposal--part III.

In the first two documents [Ph. Hubert, J.J. Nguyen-Huu, B. Boulanger, E. Chapuzet, P. Chiap, N. Cohen, P.A. Compagnon, W. Dewé, M. Feinberg, M. Lallier, M. Laurentie, N. Mercier, G. Muzard, C. Nivet, L. Valat, J. Pharm. Biomed. Anal. 36 (2004) 579-586; Ph. Hubert, J.J. Nguyen-Huu, B. Boulanger, E. Chapuzet, P. Chiap, N. Cohen, P.A. Compagnon, W. Dewé, M. Feinberg, M. Lallier, M. Laurentie, N. Mercier, G. Muzard, C. Nivet, L. Valat, E. Rozet, J. Pharm. Biomed. Anal., in press], a recent SFSTP Commission on the validation of analytical procedure has introduced a harmonized approach for the validation of analytical procedures. In order to complete this guide, the statistical methodology allowing to correctly conclude about the validity of a procedure is proposed in this third part of the guide. Indeed all the steps to obtain the decision tool namely the accuracy profile are described and illustrated step by step by a numerical example. This tool, based on the concept of total error (bias+standard deviation) build with a beta-expectation tolerance interval, allows to easily take the right decision and simultaneously minimizing the risk of the future use of the analytical procedure.

Harmonization of strategies for the validation of quantitative analytical procedures. A SFSTP proposal--part II.

As reported in a previous paper, the main objective of the new commission of the Société Française des Sciences et Techniques Pharmaceutiques (SFSTP) was the harmonisation of approaches for the validation of quantitative analytical procedures. In a series of meetings, members of this Commission have first tried to review the objectives of analytical methods and the objectives of validation methods and to recommend the use of two-sided beta-expectation tolerance intervals for total error of validation samples (accuracy profile) in the acceptance/rejection of analytical method in validation phase. In the context of the harmonization, the other objectives were: (i) to propose a consensus on the norms usually recognized, while widely incorporating the ISO terminology; (ii) to recommend to validate the analytical procedure accordingly to the way it will be used in routine; (iii) to elaborate a rational, practical and statistically reliable strategy to assure the quality of the analytical results generated. This strategy has been formalised in a guide and the three latter objectives made by the Commission are summarised in the present paper which is the second part of summary report of the SFSTP commission. The SFSTP guide has been produced to help analysts to validate their analytical methods. It is the result of a consensus between professionals having expertise in analytical and/or statistical fields. The suggestions presented in this paper should therefore help the analyst to design and perform the minimum number validation experiments needed to obtain all the required information to establish and demonstrate the reliability of its analytical procedure.

Evolution of the uniquely adaptable lentiviral envelope in a natural reservoir host.

BACKGROUND: The ability of emerging pathogens to infect new species is likely related to the diversity of pathogen variants present in existing reservoirs and their degree of genomic plasticity, which determines their ability to adapt to new environments. Certain simian immunodeficiency viruses (SIVcpz, SIVsm) have demonstrated tremendous success in infecting new species, including humans, resulting in the HIV-1 and HIV-2 epidemics. Although SIV diversification has been studied on a population level, the essential substrates for cross-species transmission, namely SIV sequence diversity and the types and extent of viral diversification present in individual reservoir animals have not been elucidated. To characterize this intra-host SIV diversity, we performed sequence analyses of clonal viral envelope (env) V1V2 and gag p27 variants present in individual SIVsm-infected sooty mangabeys over time. RESULTS: SIVsm demonstrated extensive intra-animal V1V2 length variation and amino acid diversity (le38%), and continual variation in V1V2 N-linked glycosylation consensus sequence frequency and location. Positive selection was the predominant evolutionary force. Temporal sequence shifts suggested continual selection, likely due to evolving antibody responses. In contrast, gag p27 was predominantly under purifying selection. SIVsm V1V2 sequence diversification is at least as great as that in HIV-1 infected humans, indicating that extensive viral diversification in and of itself does not inevitably lead to AIDS. CONCLUSION: Positive diversifying selection in this natural reservoir host is the engine that has driven the evolution of the uniquely adaptable SIV/HIV envelope protein. These studies emphasize the importance of retroviral diversification within individual host reservoir animals as a critical substrate in facilitating cross-species transmission.

The HIV-1 nef gene acts as a positive viral infectivity factor.

The gene product of the nef gene of HIV-1 acts both to increase the infectivity of viral particles and to reduce the expression of the CD4 receptor molecule on the cell surface. These two functions of Nef may be related, in that downregulation of CD4 may promote the production of HIV that has greater infectivity.

Impact of atrioventricular compliance on pulmonary artery pressure in mitral stenosis: an exercise echocardiographic study.

BACKGROUND: The decay of the pressure gradient across a stenotic mitral valve is determined by the size of the orifice and net AV compliance (C(n)). We have observed a group of symptomatic patients, usually in sinus rhythm, characterized by pulmonary hypertension (particularly during exercise) despite a relatively large mitral valve area by pressure half-time. We speculated that this discrepancy was due to low atrial compliance causing both pulmonary hypertension and a steep decay of the transmitral pressure gradient despite significant stenosis. We therefore tested the hypothesis that C(n) is an important physiological determinant of pulmonary artery pressure at rest and during exercise in mitral stenosis. METHODS AND RESULTS: Twenty patients with mitral stenosis were examined by Doppler echocardiography. C(n), calculated from the ratio of effective mitral valve area (continuity equation) and the E-wave downslope, ranged from 1.7 to 8.1 mL/mm Hg. Systolic pulmonary artery pressure (PAP) increased from 43+/-12 mm Hg at rest to 71+/-23 mm Hg (range, 40 to 110 mm Hg) during exercise. There was a particularly close correlation between C(n) and exercise PAP (r=-0.85). Patients with a low compliance were more symptomatic (P<0.025). Catheter- and Doppler-derived values for C(n), determined in 10 cases, correlated well (r=0.79). CONCLUSIONS: C(n), which can be noninvasively assessed, is an important physiological determinant of PAP in mitral stenosis. Patients with low C(n) represent an important clinical entity, with symptoms corresponding to severe increases in PAP during stress echocardiography.

Effect of thrombolytic therapy on the evolution of significant mitral regurgitation in patients with a first inferior myocardial infarction.

OBJECTIVES: This study was designed to test the hypothesis that reperfusion therapy with thrombolysis will prevent the development of significant mitral regurgitation in patients with inferior myocardial infarction. BACKGROUND: The value of thrombolytic therapy in patients with inferior or posterior wall myocardial infarction has been controversial. We hypothesized that successful reperfusion therapy with intravenous thrombolysis may reduce the incidence and severity of postinfarction mitral regurgitation in this patient group. METHODS: We prospectively studied 104 patients with a first inferior myocardial infarction. Thrombolytic therapy was administered to 55 patients (treatment group) 3.2 +/- 2.1 h after the onset of symptoms. The other 49 patients formed the control group. Doppler echocardiographic color flow imaging was performed in all patients within 24 h, at 7 to 10 days and at 28 to 30 days after myocardial infarction. Significant mitral regurgitation was defined as moderate or severe (grade 2 or 3). RESULTS: No significant differences in baseline clinical characteristics were observed between the treatment and control groups. The overall incidence rates of significant mitral regurgitation at 24 h, 7 to 10 days and at 28 to 30 days were 10 (10%) of 104 patients, 18 (17%) of 104 patients and 11 (11%) of 100 patients, respectively. Multivariate analysis reveals the following independent predictors of the occurrence of significant mitral regurgitation: female gender (at 7 to 10 days, odds ratio 5.3, 90% confidence interval [CI] 1.8 to 15.5; at 28 to 30 days, odds ratio 3.7, 90% CI 1.1 to 12.7), heart failure (at 7 to 10 days, odds ratio 7.7, 90% CI 2.2 to 26.9) and transient complete atrioventricular block (at 24 h of myocardial infarction, odds ratio 5.8, 90% CI 1.2 to 27). Compared with the control group, the treatment group exhibited marked reduction in the incidence of significant mitral regurgitation at 24 h (16% vs. 4%; odds ratio 0.1, 90% CI 0.0 to 0.7); at 7 to 10 days (24% vs. 11%; odds ratio 0.3, 90% CI 0.1 to 0.9) and at 28 to 30 days (15% vs. 7%; odds ratio 0.4, 90% CI 0.1 to 1.6). Severe (grade 3) mitral regurgitation developed in five patients in the control group but in no patient in the treatment group. CONCLUSIONS: Thrombolytic therapy in the patients with a first inferior myocardial infarction was associated with a reduced incidence of significant mitral regurgitation. These results support the use of such therapy in patients with inferior myocardial infarction.

Left ventricular pseudohypertrophy in cardiac tamponade: an echocardiographic study in a canine model.

OBJECTIVES: This study was designed to establish whether left ventricular pseudohypertrophy in cardiac tamponade can be reproducibly induced in an experimental canine model and to investigate the mechanism of its production. BACKGROUND: Past experimental and clinical studies have shown reduction of ventricular volumes resulting from cardiac tamponade. Left ventricular pseudohypertrophy, a transient thickening of myocardial walls, was recently described as a new echocardiographic sign of cardiac tamponade. METHODS: Cardiac tamponade was induced in seven anesthetized open chest dogs with serial bolus injections of 50 ml each of 0.9% saline solution into the pericardial sac. Under hemodynamic monitoring, M-mode and two-dimensional echocardiographic measurements were performed from a right parasternal window at each stage of graded cardiac tamponade. RESULTS: There was a progressive increase of interventricular septal and posterior wall diastolic thickness. Mean wall thickness (interventricular septal thickness + posterior wall thickness divided by 2) was 9.8 +/- 1.3 mm at baseline, 14.3 +/- 0.9 mm at peak tamponade and 9.0 +/- 1.5 mm after fluid withdrawal (p < 0.0001). Mean wall thickness correlated directly with the severity of cardiac tamponade, as estimated from the level of right arterial pressures (r = 0.75 and p < 0.0001), and with the decrease of left ventricular cavity volume (r = -0.67 and p < 0.0001). Left ventricular mass did not change significantly. CONCLUSIONS: Left ventricular pseudohypertrophy is a constant manifestation of cardiac tamponade in a canine model. The degree of myocardial thickening correlates with the reduction of ventricular dimensions and with the severity of hemodynamic compromise, representing a constant facet of heart remodeling in cardiac tamponade.