HIV-1 persistence in the central nervous system: viral and host determinants during antiretroviral therapy.

Despite remarkable therapeutic advances in the past two decades, the elimination of human immunodeficiency virus type 1 (HIV-1) from latent reservoirs constitutes a major barrier to eradication and preventing neurological disease associated with HIV/AIDS. Invasion of the central nervous system (CNS) by HIV-1 occurs early in infection, leading to viral infection and productive persistence in brain macrophage-like cells (BMCs) including resident microglia and infiltrating macrophages. HIV-1 persistence in the brain and chronic neuroinflammation occur despite effective treatment with antiretroviral therapy (ART). This review examines the evidence from clinical studies, in vivo and in vitro models for HIV-1 CNS persistence, as well as therapeutic considerations in targeting latent CNS reservoirs.

De Novo Belatacept in a Human Immunodeficiency Virus-Positive Kidney Transplant Recipient.

Benefits of belatacept-based immunosuppressive regimens in human immunodeficiency virus (HIV)-positive renal transplant recipients include avoidance of drug interactions between calcineurin inhibitors and highly active antiretroviral agents and decreased likelihood or severity of nonimmune toxicities such as new-onset diabetes after transplant, hyperlipidemia and hypertension. We report a successful case of de novo belatacept at >18 mo from transplant in an HIV-positive black man aged 50 years who received his first transplant from a living related kidney donor. To our knowledge, this case is the first reported of belatacept use in an HIV-positive renal transplant recipient.

Cramér-Rao Lower Bound for Point Based Image Registration With Heteroscedastic Error Model for Application in Single Molecule Microscopy.

The Cramér-Rao lower bound for the estimation of the affine transformation parameters in a multivariate heteroscedastic errors-in-variables model is derived. The model is suitable for feature-based image registration in which both sets of control points are localized with errors whose covariance matrices vary from point to point. With focus given to the registration of fluorescence microscopy images, the Cramér-Rao lower bound for the estimation of a feature's position (e.g., of a single molecule) in a registered image is also derived. In the particular case where all covariance matrices for the localization errors are scalar multiples of a common positive definite matrix (e.g., the identity matrix), as can be assumed in fluorescence microscopy, then simplified expressions for the Cramér-Rao lower bound are given. Under certain simplifying assumptions these expressions are shown to match asymptotic distributions for a previously presented set of estimators. Theoretical results are verified with simulations and experimental data.

Analysis of Point Based Image Registration Errors With Applications in Single Molecule Microscopy.

We present an asymptotic treatment of errors involved in point-based image registration where control point (CP) localization is subject to heteroscedastic noise; a suitable model for image registration in fluorescence microscopy. Assuming an affine transform, CPs are used to solve a multivariate regression problem. With measurement errors existing for both sets of CPs this is an errors-in-variable problem and linear least squares is inappropriate; the correct method being generalized least squares. To allow for point dependent errors the equivalence of a generalized maximum likelihood and heteroscedastic generalized least squares model is achieved allowing previously published asymptotic results to be extended to image registration. For a particularly useful model of heteroscedastic noise where covariance matrices are scalar multiples of a known matrix (including the case where covariance matrices are multiples of the identity) we provide closed form solutions to estimators and derive their distribution. We consider the target registration error (TRE) and define a new measure called the localization registration error (LRE) believed to be useful, especially in microscopy registration experiments. Assuming Gaussianity of the CP localization errors, it is shown that the asymptotic distribution for the TRE and LRE are themselves Gaussian and the parameterized distributions are derived. Results are successfully applied to registration in single molecule microscopy to derive the key dependence of the TRE and LRE variance on the number of CPs and their associated photon counts. Simulations show asymptotic results are robust for low CP numbers and non-Gaussianity. The method presented here is shown to outperform GLS on real imaging data.

IMAGE REGISTRATION ERROR ANALYSIS WITH APPLICATIONS IN SINGLE MOLECULE MICROSCOPY.

This paper is concerned with assessing localization errors emanating from the image registration of two monochromatic fluorescence microscopy images. Assuming an affine transform exists between images, registration in this setting typically involves using control points to solve a multivariate linear regression problem; however with measurement errors existing in both sets of variables the use of linear least squares is inappropriate. It is shown that image registration is an errors-in-variable problem and as such the correct method is to use generalized least squares. Traditionally this requires the measurement errors to be independent and identically distributed (iid); an assumption that is rarely satisfied in practical situations. An extension of the multivariate generalized least squares estimator that allows non-iid noise is applied. The distributional properties of the estimators are used to derive localization errors emanating from the image registration process in terms of photon counts and experimental parameters.

Measurement Errors in Fluorescence Microscopy Image Registration.

Image registration is an important processing step in fluorescence microscopy, for example in tracking or super-resolution methods. Precision localization of single fluorescent molecules from a quantum limited photon detection process, subject to Gaussian readout noise, is key to the use of single molecule microscopy. It is therefore important to know the effect that registration has on the accuracy of localizing a single molecule. Here we demonstrate a suitable approach to image registration that accounts for point-wise errors in localizing the control points typically used in fluorescence microscopy. This allows expressions for the localization errors caused by the registration process to be derived, showing dependence on the number of control points and their associated photon counts.

Effect of operator and institutional volume on clinical outcomes after percutaneous coronary interventions performed in Canada and the United States: a brief report from the Enhanced Suppression of the Platelet glycoprotein IIb/IIIa Receptor with Integrilin Therapy (ESPRIT) study.

BACKGROUND: The Enhanced Suppression of the Platelet glycoprotein IIb/IIIa Receptor with Integrilin Therapy (ESPRIT) trial compared the use of eptifibatide with placebo in 2064 coronary intervention patients. It was previously reported that Canadian patients had reduced rates of 30-day and one-year death, myocardial infarction (MI) or target vessel revascularization (TVR) compared with patients in the United States (US). OBJECTIVE: To examine whether operator or institutional volume differences explain the regional variation in clinical outcome. METHODS AND RESULTS: Each site received an operator and institutional volume survey. Fifty-seven sites (62%) returned complete data on 1338 patients. In this smaller cohort, Canadian patients had reduced rates of 30-day and one-year death, MI or TVR compared with US patients (6.3% versus 10.3% and 14.9% versus 20.1%, respectively; P<0.05 for both comparisons). Among 176 physicians with a median of 13 years experience, the median operator volume was 200 cases per year. Operators with fewer than 100 cases per year had higher rates of 30-day death, MI or TVR (13.2% versus 8.7%; P=0.18) and large MI (7.7% versus 3.3%; P=0.06) than those with 100 or more cases per year. The median institutional volume was 1064 cases per year. Canadian and US centres had similar operator and institutional volumes. By multivariate modelling, operator volume was not predictive of adverse clinical events. However, the rates of 30-day and one-year death, MI or TVR fell by 3% for every 100 patients treated by the institution (OR 0.97; P=0.058 and P=0.002, respectively). Enrollment in Canada was associated with improved outcomes at 30 days (OR 0.50; P=0.001) and one year (OR 0.66; P=0.001) despite inclusion of volume variables in the models. CONCLUSIONS: In the ESPRIT study, institutional volume was associated with a modest reduction in risk of death, MI or TVR over short- and long-term follow-up periods. The Canadian and US investigators and institutions selected in ESPRIT had similar annual procedural volumes. Therefore, volume variables did not explain the differential risk of clinical events observed for patients enrolled in the two countries.

First experience with direct, selective factor Xa inhibition in patients with non-ST-elevation acute coronary syndromes: results of the XaNADU-ACS Trial.

BACKGROUND: Unfractionated heparin is widely used in patients with non-ST-elevation acute coronary syndromes but has important limitations. Anticoagulants with predictable kinetics and anticoagulant effects, better efficacy, and greater safety are needed. OBJECTIVE: To investigate the efficacy and safety of a direct, selective factor Xa inhibitor, DX-9065a (Daiichi Pharmaceuticals LTD, Inc.) compared with heparin, in patients with non-ST-elevation acute coronary syndromes. PATIENTS AND METHODS: Patients (n = 402) from the USA, Canada, and Japan were randomized to blinded, weight-adjusted heparin, low-dose DX-9065a, or high-dose DX-9065a. RESULTS: The primary efficacy endpoint of death, myocardial infarction, urgent revascularization, or ischemia on continuous ST-segment monitoring occurred in 33.6%, 34.3%, and 31.3% of patients assigned to heparin, low-dose DX-9065a, and high-dose DX-9065a (P = 0.91 for heparin vs. combined DX-9065a). The composite of death, myocardial infarction, or urgent revascularization occurred in 19.5%, 19.3%, and 11.9% (P = 0.125 for heparin vs. high-dose DX-9065a) of patients; major or minor bleeding occurred in 7.7%, 4.2%, and 7.0% of patients; and major bleeding in 3.3%, 0.8%, and 0.9% of patients. Higher concentrations of DX-9065a were associated with a lower likelihood of ischemic events (P = 0.03) and a non-significant tendency toward a higher likelihood of major bleeding (P = 0.32). CONCLUSIONS: In this small phase II trial, there was a non-significant tendency toward a reduction in ischemic events and bleeding with DX-9065a compared with heparin in patients with acute coronary syndromes. The absence of an effect on ST-monitor ischemia warrants further investigation. These data provide the rationale for adequately powered studies of DX-9065a in acute coronary syndromes or percutaneous intervention.

Initial experience with factor-Xa inhibition in percutaneous coronary intervention: the XaNADU-PCI Pilot.

BACKGROUND: Direct factor (F)Xa inhibition is an attractive method to limit thrombotic complications during percutaneous coronary intervention (PCI). OBJECTIVES: To investigate drug levels achieved, effect on coagulation markers, and preliminary efficacy and safety of several doses of DX-9065a, an intravenous, small molecule, direct, reversible FXa inhibitor during PCI. PATIENTS AND METHODS: Patients undergoing elective, native-vessel PCI (n = 175) were randomized 4 : 1 to open-label DX-9065a or heparin in one of four sequential stages. DX-9065a regimens in stages I-III were designed to achieve concentrations of > 100 ng mL-1, > 75 ng mL-1, and > 150 ng mL-1. Stage IV used the stage III regimen but included patients recently given heparin. RESULTS: At 15 min median (minimum) DX-9065a plasma levels were 192 (176), 122 (117), 334 (221), and 429 (231) ng mL-1 in stages I-IV, respectively. Median whole-blood international normalized ratios (INRs) were 2.6 (interquartile range 2.5, 2.7), 1.9 (1.8, 2.0), 3.2 (3.0, 4.1), and 3.8 (3.4, 4.6), and anti-FXa levels were 0.36 (0.32, 0.38), 0.33 (0.26, 0.39), 0.45 (0.41, 0.51), and 0.62 (0.52, 0.65) U mL-1, respectively. Stage II enrollment was stopped (n = 7) after one serious thrombotic event. Ischemic and bleeding events were rare and, in this small population, showed no clear relation to DX-9065a dose. CONCLUSIONS: Elective PCI is feasible using a direct FXa inhibitor for anticoagulation. Predictable plasma drug levels can be rapidly obtained with double-bolus and infusion DX-9065a dosing. Monitoring of DX-9065a may be possible using whole-blood INR. Direct FXa inhibition is a novel and potentially promising approach to anticoagulation during PCI that deserves further study.

Severe thrombocytopenia possibly related to readministration of eptifibatide.

Thrombocytopenia following eptifibatide therapy is a rare event, and acute severe thrombocytopenia following readministration has not been reported. We describe a case of acute severe thrombocytopenia following reexposure to eptifibatide during percutaneous coronary intervention. We continue to monitor platelet counts at 2-4 hr and 16-24 hr following administration of any glycoprotein IIb/IIIa inhibitor. Cathet Cardiovasc Intervent 2001;54:63-67.

Predictors of improvement in left ventricular function after percutaneous revascularization of occluded coronary arteries: a report from the Total Occlusion Study of Canada (TOSCA).

BACKGROUND: The Total Occlusion Study of Canada (TOSCA) is a multicenter, randomized trial evaluating the effect of stenting with > =1 heparin-coated stent on long-term patency after percutaneous coronary intervention by balloon angioplasty of occluded coronary arteries. The purpose of the current study was to compare the effect of stenting and balloon angioplasty on global left ventricular ejection fraction (LVEF) and regional wall motion and to examine what clinical and angiographic factors may have an effect on left ventricular function in this setting. METHODS AND RESULTS: Analysis at the core angiographic laboratory of paired baseline and follow-up left ventricular angiograms, as well as target vessel patency, was possible in 244 of 410 cases. An improvement in LVEF was observed in the entire group (59.4% +/- 11% to 61.0% +/- 11%, P =.003). The LVEF change was +1.84 +/- 7.54 in the stent group (P =.009) and 1.28 +/- 8.16 in the percutaneous transluminal coronary angioplasty group (P =.085). There was no significant intergroup difference. Patients with duration of occlusion < or =6 weeks had an improvement in LVEF (+2.98 +/- 8.68, P =.0006), whereas those with an occlusion duration of > 6 weeks had no improvement (+0.48 +/- 7.01, P not significant). Multivariate analysis revealed baseline LVEF <60%, duration of occlusion < or =6 weeks, and Canadian Cardiology Society angina class I or II to be independent predictors of improvement in LVEF. CONCLUSIONS: The restoration of coronary patency of nonacute occluded coronary arteries is associated with a small but significant improvement in regional and global left ventricular function, especially in patients with recent occlusions and depressed left ventricular function. In spite of significant effect on long-term patency, stenting of nonacute coronary occlusions does not result in significantly better left ventricular function compared with balloon angioplasty in this setting.

Comparison of two platelet glycoprotein IIb/IIIa inhibitors, tirofiban and abciximab, for the prevention of ischemic events with percutaneous coronary revascularization.

BACKGROUND: In the setting of percutaneous coronary revascularization, agents in the class known as platelet glycoprotein IIb/IIIa inhibitors have significantly reduced the incidence of death or nonfatal myocardial infarction at 30 days. We assessed whether there are differences in safety or efficacy between two such inhibitors, tirofiban and abciximab. METHODS: Using a double-blind, double-dummy design at 149 hospitals in 18 countries, we randomly assigned patients to receive either tirofiban or abciximab before undergoing percutaneous coronary revascularization with the intent to perform stenting. The primary end point was a composite of death, nonfatal myocardial infarction, or urgent target-vessel revascularization at 30 days. The trial was designed and statistically powered to demonstrate the noninferiority of tirofiban as compared with abciximab. RESULTS: The primary end point occurred more frequently among the 2398 patients in the tirofiban group than among the 2411 patients in the abciximab group (7.6 percent vs. 6.0 percent; hazard ratio, 1.26; one-sided 95 percent confidence interval of 1.51, demonstrating lack of equivalence, and two-sided 95 percent confidence interval of 1.01 to 1.57, demonstrating the superiority of abciximab over tirofiban; P=0.038). The magnitude and the direction of the effect were similar for each component of the composite end point (hazard ratio for death, 1.21; hazard ratio for myocardial infarction, 1.27; and hazard ratio for urgent target-vessel revascularization, 1.26), and the difference in the incidence of myocardial infarction between the tirofiban group and the abciximab group was significant (6.9 percent and 5.4 percent, respectively; P=0.04). The relative benefit of abciximab was consistent regardless of age, sex, the presence or absence of diabetes, or the presence or absence of pretreatment with clopidogrel. There were no significant differences in the rates of major bleeding complications or transfusions, but tirofiban was associated with a lower rate of minor bleeding episodes and thrombocytopenia. CONCLUSIONS: Although the trial was intended to assess the noninferiority of tirofiban as compared with abciximab, the findings demonstrated that tirofiban offered less protection from major ischemic events than did abciximab.

RNA trafficking signals in human immunodeficiency virus type 1.

Intracellular trafficking of retroviral RNAs is a potential mechanism to target viral gene expression to specific regions of infected cells. Here we show that the human immunodeficiency virus type 1 (HIV-1) genome contains two sequences similar to the hnRNP A2 response element (A2RE), a cis-acting RNA trafficking sequence that binds to the trans-acting trafficking factor, hnRNP A2, and mediates a specific RNA trafficking pathway characterized extensively in oligodendrocytes. The two HIV-1 sequences, designated A2RE-1, within the major homology region of the gag gene, and A2RE-2, in a region of overlap between the vpr and tat genes, both bind to hnRNP A2 in vitro and are necessary and sufficient for RNA transport in oligodendrocytes in vivo. A single base change (A8G) in either sequence reduces hnRNP A2 binding and, in the case of A2RE-2, inhibits RNA transport. A2RE-mediated RNA transport is microtubule and hnRNP A2 dependent. Differentially labelled gag and vpr RNAs, containing A2RE-1 and A2RE-2, respectively, coassemble into the same RNA trafficking granules and are cotransported to the periphery of the cell. tat RNA, although it contains A2RE-2, is not transported as efficiently as vpr RNA. An A2RE/hnRNP A2-mediated trafficking pathway for HIV RNA is proposed, and the role of RNA trafficking in targeting HIV gene expression is discussed.

In vitro and in vivo comparison of three different intravascular ultrasound catheter designs.

Intravascular ultrasound (IVUS) is an invasive imaging modality, which provides detailed two-dimensional images of blood vessels. There are currently two different types of IVUS catheters available, namely, the phased-array and the mechanical designs. The operating ultrasound frequency of these catheters ranges from 20 to 40 MHz. This study sought to evaluate the image quality, accuracy of diameter and pullback length measurements, and catheter handling characteristics of three different IVUS catheters currently available for clinical use using both in vitro phantom models and in patients undergoing percutaneous coronary intervention (PCI). In gelatin phantom models, image quality assessed on a semiquantitative scale was significantly different between the three IVUS catheters (P = 0.01) with the 40-MHz catheter providing the best images. Accuracy of lumen diameter measurements, when compared to optical microscopy, were similar between the three IVUS catheter designs (all R(2) = 0.99). There were no significant differences in accuracy of pullback length measurements in vitro between the three designs. However, there were differences in the performance of the three IVUS catheters when used for preinterventional imaging in patients undergoing PCI. Both mechanical IVUS catheters were associated with lower procedural, fluoroscopy, and lesion crossing times compared to the phased-array catheter (all P < 0.05). There were no significant differences between the catheters during postinterventional IVUS imaging. There were also small but potentially important differences with regards to clinical events and complications associated with the use of the different IVUS catheters during the PCI procedures, reflecting differences in catheter design.

Bleeding risks with abciximab after full-dose thrombolysis in rescue or urgent angioplasty for acute myocardial infarction.

BACKGROUND: The bleeding risk associated with platelet glycoprotein IIb/IIIa inhibition in patients undergoing percutaneous transluminal coronary angioplasty (PTCA) after full-dose thrombolysis for acute myocardial infarction (AMI) is unclear. We examined the risk and predictors of bleeding complications in patients with AMI who received abciximab during rescue or urgent PTCA after full-dose thrombolytic therapy. METHODS: A multicenter retrospective cohort of 147 consecutive patients who underwent PTCA within 48 hours after full-dose thrombolysis for AMI was studied. Bleeding events (major, minor, nuisance) from the onset of AMI to discharge were compared between those who received abciximab (n = 57) and those who did not (n = 90). RESULTS: Baseline clinical characteristics were similar between the two groups. Despite lower doses of procedural heparin, the incidence of non-coronary artery bypass graft-related major and minor bleeding was higher in the abciximab group than in controls (63% vs 39%, P =.004). Although the risk of major bleeding was 4-fold with abciximab (12% vs 3%, P =.04), only one intracranial and one fatal bleeding event occurred. Multivariable regression identified abciximab therapy as the most powerful independent predictor of combined major and minor bleeding, with a hazard risk ratio of 1.9 (P =.04). CONCLUSIONS: In the setting of rescue or urgent PTCA within 48 hours after full-dose thrombolytic therapy after AMI, major and particularly minor bleeding were frequently encountered. The adjunctive use of abciximab increased these bleeding risks by approximately 2-fold.

Human Jurkat lymphocytes clones differ in their capacity to support productive human immunodeficiency virus type 1 multiplication.

CD4-positive human lymphocytic cell lines are essential tools for the study of human immunodeficiency virus (HIV) replication. Jurkat cells are among cells more frequently used for this purpose. In the current study, various cell clones or cells stocks derived from this cell line were shown to vary substantially in their response to viral infection and in their ability to support productive virus multiplication. The formation of syncytia, the effect of Vpu on viral export, and especially the specific infectivity of the viruses released, can vary significantly among independent cell stocks. This suggests that the choice of an adequate cell clone or cell line could be critical while evaluating specific properties of the virus and further stresses the limitations of tissue culture models. However, these observations also raise the possibility of exploiting these differences among cells to study specific aspects of host-cell interactions contributing to viral multiplication.

Y stenting of a bifurcation stenosis using a new radiopaque stent.

A case of Y stenting using radiopaque stents for a bifurcation stenosis is described. Angiography and intravascular ultrasound demonstrate that in spite of optimal stent placement there is an area of the lesion that is not fully covered by the stents. This may predispose to subacute thrombosis and restenosis. This case illustrates a potential deficiency of this type of bifurcation stenting.

Influence of human immunodeficiency virus type 1 envelope glycoprotein YXXL endocytosis/polarization signal on viral accessory protein functions.

INTRODUCTION: A tyrosine-based targeting signal in the intracytoplasmic domain of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein is required for basolateral targeting of viral budding in polarized epithelial cells, concentration of viral assembly at one pole of infected lymphocytes, and rapid endocytosis of the glycoprotein from the plasma membrane. In HIV-1, the process of viral assembly and budding is complex and involves the participation of viral accessory proteins. STUDY DESIGN/METHODS: In this study, we examined whether the functions of the Nef, Vif, and Vpu proteins can influence or be influenced by the presence of envelope targeting signal in epithelial cells or lymphocytes. A series of proviral DNAs combining a mutation that inactivates the targeting signal with independent mutations in the three accessory proteins was constructed for this purpose. RESULTS: It was found that none of these three accessory proteins affected the basolateral release of the virus in polarized MDCK cells. Reciprocally, a mutation abolishing targeting of the viral envelope glycoprotein did not affect the phenotype conferred by the accessory proteins. Interestingly, the mutation abolishing targeting increased viral infectivity only in the presence of the Vpu protein. This phenotype was found to be associated with the release-enhancing function of Vpu and with an increased incorporation of viral envelope glycoprotein in virions. CONCLUSIONS: These data clearly show that accessory protein functions, and more specifically Vpu modulation of viral infectivity, can be affected by variations in the viral envelope glycoprotein.

Human immunodeficiency virus type 1 Vpr-mediated G(2) cell cycle arrest: Vpr interferes with cell cycle signaling cascades by interacting with the B subunit of serine/threonine protein phosphatase 2A.

The Vpr protein of primate lentiviruses arrests cell cycling at the G(2)/M phase through an inactivation of cyclin B-p34(cdc2) and its upstream regulator cdc25. We provide here biochemical and functional evidence demonstrating that human immunodeficiency virus type 1 (HIV-1) Vpr mediates G(2) arrest by forming a complex with protein phosphatase 2A (PP2A), an upstream regulator of cdc25. Vpr associates with PP2A through a specific interaction with the B55 regulatory subunit. This interaction is necessary but not sufficient for G(2) arrest. Interestingly, we found that Vpr association with B55-containing PP2A targets the enzymatic complex to the nucleus and, importantly, enhances the recruitment and dephosphorylation of the cdc25 substrate. Our data suggest that Vpr mediates G(2) arrest by enhancing the nuclear import of PP2A and by positively modulating its catalytic activity towards active phosphorylated nuclear cdc25.