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Find video protocols related to scientific articles indexed in Pubmed.
HIV Nef promotes BLyS/BAFF expression by blood dendritic cells during the course of infection in humans.
J. Infect. Dis.
PUBLISHED: 11-08-2014
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Dendritic cells (DCs) modulate B-cell survival and differentiation, mainly through production of growth factors such as B lymphocyte stimulator (BLyS/BAFF). We have recently shown that in HIV-1-infected rapid and classic progressors B-cell dysregulations were associated with increased BLyS/BAFF expression in plasma and by blood myeloid DCs (mDCs), in contrast to aviremic slow progressors, also known as elite controllers. In previous work with HIV-transgenic mice, B-cell dysregulations were concomitant with altered mDCs and dependant on HIV negative factor (Nef). We now report that HIV-1-Nef is detected in plasma and BLyS/BAFF over-expressing blood mDCs of HIV-1-infected rapid and classic progressors, early on infection and despite successful therapy, whereas it is low to undetectable in aviremic slow progressors. In vitro, HIV-1-Nef drives monocyte-derived DCs towards BLyS/BAFF over-expression, through a process involving STAT1. Importantly, this is counteracted in presence of all-trans retinoic acid. Nef thus contributes to high BLyS/BAFF pro-inflammatory profiles in HIV-1-infected individuals.
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Short Communication: Anti-HIV-1 Envelope Immunoglobulin Gs in Blood and Cervicovaginal Samples of Beninese Commercial Sex Workers.
AIDS Res. Hum. Retroviruses
PUBLISHED: 10-30-2014
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Abstract Characterization of the immune correlates of protection against HIV infection is crucial for the development of preventive strategies. This study examined HIV-1 envelope (Env) glycoproteins, specifically immunoglobulin G (IgG), in systemic and mucosal compartments of female Beninese commercial sex workers (CSWs). Samples of 23 HIV-1-positive and 20 highly exposed HIV-1-seronegative (HESN) CSWs were studied. HIV-1 Env-specific IgG detection in sera and cervicovaginal lavages (CVLs) from the study population was done by cell-based ELISA. The HIV neutralizing activity was evaluated with a neutralization assay. The HIV-1-specific antibody-dependent cellular cytotoxicity (ADCC) response of the cohort was measured with a FACS-based assay evaluating the ADCC-mediated elimination of gp120-coated target cells. No anti-HIV-1 Env-specific IgG neutralizing or ADCC activities were detected in samples from HESN CSWs. Samples from HIV-1-infected CSWs presented ADCC activity in both sera and CVLs. Anti-Env IgG from sera and CVLs from HIV-1-infected CSWs preferentially recognized Env in its CD4-bound conformation. HIV-1-infected CSWs have ADCC-mediating IgG that preferentially recognizes Env in its CD4-bound conformation at the mucosal site.
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The HIV-1 gp120 CD4-bound conformation is preferentially targeted by ADCC-mediating antibodies in sera from HIV-1-infected individuals.
J. Virol.
PUBLISHED: 10-24-2014
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Recent studies have linked antibody Fc-mediated effector functions with protection or control of HIV-1 and SIV infections. Interestingly, the presence of antibodies with potent antibody-dependent cellular cytotoxicity (ADCC) activity in the Thai RV144 vaccine trial was suggested to correlate with a decreased HIV-1 acquisition risk. These antibodies were recently found to recognize HIV envelope (Env) epitopes exposed upon Env - CD4 interaction. CD4 downregulation by Nef and Vpu, as well as Vpu-mediated BST-2 antagonism, were reported to modulate exposure of those CD4-induced HIV-1 Env epitopes and therefore were proposed to play a role in reducing the susceptibility of infected cells to ADCC mediated by this class of antibodies. Here we report a high prevalence of antibodies recognizing CD4-induced HIV-1 Env epitopes in sera from HIV-1 infected individuals, which correlated with their ability to mediate ADCC responses against HIV-1 infected cells exposing these Env epitopes at the cell surface. Furthermore, our results indicate that Env variable regions V1, V2, V3 and V5 do not represent a major determinant for ADCC responses mediated by sera from HIV-1-infected individuals. Altogether, these findings suggest that HIV-1 tightly controls the exposure of certain Env epitopes at the surface of infected cells in order to prevent elimination by Fc-effector functions.
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Conformational Evaluation of HIV-1 Trimeric Envelope Glycoproteins Using a Cell-based ELISA Assay.
J Vis Exp
PUBLISHED: 10-07-2014
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HIV-1 envelope glycoproteins (Env) mediate viral entry into target cells and are essential to the infectious cycle. Understanding how those glycoproteins are able to fuel the fusion process through their conformational changes could lead to the design of better, more effective immunogens for vaccine strategies. Here we describe a cell-based ELISA assay that allows studying the recognition of trimeric HIV-1 Env by monoclonal antibodies. Following expression of HIV-1 trimeric Env at the surface of transfected cells, conformation specific anti-Env antibodies are incubated with the cells. A horseradish peroxidase-conjugated secondary antibody and a simple chemiluminescence reaction are then used to detect bound antibodies. This system is highly flexible and can detect Env conformational changes induced by soluble CD4 or cellular proteins. It requires minimal amount of material and no highly-specialized equipment or know-how. Thus, this technique can be established for medium to high throughput screening of antigens and antibodies, such as newly-isolated antibodies.
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Evolution of a novel pathway leading to dolutegravir resistance in a patient harbouring N155H and multiclass drug resistance.
J. Antimicrob. Chemother.
PUBLISHED: 10-05-2014
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Dolutegravir has been recently approved for treatment-naive and -experienced HIV-infected subjects, including integrase inhibitor (INI)-experienced patients. Dolutegravir is a second-generation INI that can overcome many prior raltegravir and elvitegravir failures. Here, we report the evolution of resistance to dolutegravir in a highly treatment-experienced patient harbouring the major N155H mutation consequent to raltegravir treatment failure.
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Potential of a spectroscopic measurement method using adding-doubling to retrieve the bulk optical properties of dense microalgal media.
Appl Spectrosc
PUBLISHED: 10-01-2014
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In the context of algal mass cultivation, current techniques used for the characterization of algal cells require time-consuming sample preparation and a large amount of costly, standard instrumentation. As the physical and chemical properties of the algal cells strongly affect their optical properties, the optical characterization is seen as a promising method to provide an early diagnosis in the context of mass cultivation monitoring. This article explores the potential of a spectroscopic measurement method coupled with the inversion of the radiative transfer theory for the retrieval of the bulk optical properties of dense algal samples. Total transmittance and total reflectance measurements were performed over the 380-1020 nm range on dense algal samples with a double integrating sphere setup. The bulk absorption and scattering coefficients were thus extracted over the 380-1020 nm range by inverting the radiative transfer theory using inverse-adding-doubling computations. The experimental results are presented and discussed; the configuration of the optical setup remains a critical point. The absorption coefficients obtained for the four samples of this study appear not to be more informative about pigment composition than would be classical methods in analytical spectroscopy; however, there is a real added value in measuring the reduced scattering coefficient, as it appears to be strongly correlated to the size distribution of the algal cells.
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Flow cytometry-based assay to study HIV-1 gp120 specific antibody-dependent cellular cytotoxicity responses.
J. Virol. Methods
PUBLISHED: 08-12-2014
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Increased attention on the role of Fc-mediated effector functions against HIV-1 has led to renewed interest into the role that antibody-dependent cellular cytotoxicity (ADCC) could play in controlling viral transmission and/or the rate of disease progression. While (51)Chromium release assays have traditionally been used to study ADCC responses against HIV-1, a number of alternative flow-cytometry-based assays were recently developed. In this study, an alternative flow-cytometry-based assay was established to allow non-radioactive measurement of ADCC-mediated elimination of HIV-1 gp120 envelope glycoprotein (Env)-coated target cells. This assay relies on staining target and effector cells with different dyes, which allows precise gating and permits the calculation of the number of surviving target cells by normalization to flow-cytometry particles. By using small concentrations of recombinant gp120 Env, suitable targets cells that recapitulate the ADCC response mediated against HIV-1-infected cells were generated. Finally, this method was applied successfully to screen human sera for ADCC activity directed against HIV-1 gp120 Env.
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A dual-labeled Annexin A5 is not suited for SPECT imaging of brain cell death in experimental murine stroke.
J. Cereb. Blood Flow Metab.
PUBLISHED: 04-02-2014
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Cell death is one of the pathophysiological hallmarks after stroke. Markers to image cell death pathways in vivo are highly desirable. We previously showed that fluorescently labeled Annexin A5 (AnxA5), which binds specifically to phosphatidylserine (PS) on dead/dying cells, can be used in experimental stroke for monitoring cell death with optical imaging. Here we investigated whether dual-labeled AnxA5 (technetium and fluorescence label) can be used for single-photon emission computed tomography (SPECT) of cell death in the same model. C57Bl6/N mice were subjected to 60-minute middle cerebral artery occlusion (MCAO) and underwent SPECT imaging at 24, 48, and 72 hours afterwards. They were injected intravenously with either PS-binding AnxA5 or the nonfunctional AnxA5 (negative control), labeled with 99mTc and Alexa Fluor 568, respectively. After SPECT imaging, brain sections were cut for autoradiography and fluorescence microscopy. Ethanol-induced cell death in the femur muscle was used as positive control. We detected dual-labeled AnxA5 in the model of ethanol-induced cell death in the femur muscle, but not after MCAO at any time point, either with SPECT or with ex vivo autoradiography or fluorescence microscopy. Dual-labeled AnxA5 appears to be unsuited for visualizing death of brain cells in this MCAO model.
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Determinants of human papillomavirus coinfections among Montreal university students: the influence of behavioral and biologic factors.
Cancer Epidemiol. Biomarkers Prev.
PUBLISHED: 02-25-2014
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Human papillomavirus (HPV) coinfections are common among HPV-infected individuals, but the significance and etiology of these infections remain unclear. Though current evidence suggests that women with coinfections have increased HPV exposure (i.e., more sexual partners), it is also hypothesized that these women may represent a subgroup with increased biologic susceptibility. This study sought to examine determinants of coinfections in a cohort of young women, examining both behavioral and biologic factors related to HPV acquisition over time.
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XTEN-annexin A5: XTEN allows complete expression of long-circulating protein-based imaging probes as recombinant alternative to PEGylation.
J. Nucl. Med.
PUBLISHED: 02-18-2014
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The coupling of polyethylene glycol (PEG) to proteins (PEGylation) has become a standard method to prolong blood circulation of imaging probes and other proteins, liposomes, and nanoparticles. However, concerns have arisen about the safety of PEG, especially with respect to its poor biodegradability and antibody formation, including new evidence about preformed anti-PEG antibodies in a quarter of healthy blood donors. Here, we apply a new hydrophilic polypeptide XTEN to extend the blood half-life of an imaging probe. As an example, we chose annexin A5 (AnxA5), a recombinant 35-kD protein extensively used for the in vitro and in vivo detection of apoptosis, that has a blood half-life of less than 7 min in mice, limiting its accumulation in target tissues and therefore limiting its utility as an imaging reagent.
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3D front face solid-phase fluorescence spectroscopy combined with Independent Components Analysis to characterize organic matter in model soils.
Talanta
PUBLISHED: 02-13-2014
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Soil organic matter (SOM) is a very complex and heterogeneous system which complicates its characterization. In fact, the methods classically used to characterize SOM are time- and solvent-consuming and insufficiently informative. The aim of this work is to study the potential of 3D solid-phase front face fluorescence (3D-SPFFF) spectroscopy to quickly provide a relevant and objective characterization of SOM as an alternative to the existing methods. Different soil models were prepared to simulate natural soil composition and were analyzed by 3D front-face fluorescence spectroscopy without prior preparation. The spectra were then treated using Independent Components Analysis. In this way, different organic molecules such as cellulose, proteins and amino acids used in the soil models were identified. The results of this study clearly indicate that 3D-SPFFF spectroscopy could be an easy, reliable and practical analytical method that could be an alternative to the classical methods in order to study SOM. The use of solid samples revealed some interactions that may occur in natural soils (self-quenching in the case of cellulose) and gave more accurate fluorescence signals for different components of the analyzed soil models. Independent Components Analysis (ICA) has demonstrated its power to extract the most informative signals and thus facilitate the interpretation of the complex 3D fluorescence data.
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Raman model development for the protein conformational state classification in different freeze-dried formulations.
Anal. Chim. Acta
PUBLISHED: 01-15-2014
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The aim of this work is to build a multivariate calibration (MVC) model from Raman spectra for the prediction of the protein conformational state class (i.e. native-like or non-native) in different freeze-dried pharmaceutical formulations of a model protein lactate dehydrogenase (LDH). As this model would be intended to facilitate and better understand formulation and process development, it should allow acceptable classification performance despite variations in formulation type and batch. Therefore, it was attempted to (1) find which factors interfere the Raman spectra, (2) understand them, and (3) make the MVC model robust for them. A variance analysis within the Raman spectral data space identified significant spectral background variations among certain formulation types and batches in the studied samples. Raw material (i.e. LDH) batch variability and the presence of a Maillard reaction in formulations were the main reasons for this. We demonstrate the successful use of both exhaustive calibration and external parameter orthogonalization (EPO) pre-processing for making the Raman classification model more robust for the expected spectral interferences.
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A new bifunctional chelator enables facile biocoupling and radiolabeling as the basis for a bioconjugation kit.
Chembiochem
PUBLISHED: 01-03-2014
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A new tridentate bifunctional chelator, N-(-2-picolyl)(-4-hydroxy)(-3-amino)benzoic acid (PHAB), was designed to efficiently coordinate the [(99m)Tc(CO)3](+) core and facilitate coupling reactions to biomolecules. The chelator can be procured in the form of the corresponding benzotriazole ester (PHAB-OBT), which can be stored and used as a bioconjugation kit. PHAB-OBT reacts with modified carbohydrates with high selectivity and efficiency in a single step in both aqueous and organic media. As is desirable for a kit, no complicated chemical bench work is required. Glycoconjugate postlabeling resulted in neutral radiolabeled glycans with high radiochemical yields. Prelabeling approaches were assessed by successive reaction of PHAB-OBT with the [(99m)Tc(CO)3](+) core and a modified galactose model. The radiolabeled galactose was obtained in 84% yield as defined by HPLC analysis. Biodistribution of the radioactive (99m)Tc-labeled chelator, as well as the glycoconjugates, were examined in mice. Noticeably different biodistribution patterns were observed that reflect trends in the uptake of carbohydrate analogues by various organs.
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A Significant Reduction in the Frequency of HIV-1 Drug Resistance in Québec from 2001 to 2011 Is Associated with a Decrease in the Monitored Viral Load.
PLoS ONE
PUBLISHED: 01-01-2014
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HIV drug resistance represents a major threat for effective treatment. We assessed the trends in the frequency of drug resistance mutations and the monitored viral load (VL) in treatment-naïve (TN) and treatment-experienced (TE) individuals infected with HIV-1 in Québec, Canada, between 2001 and 2011.
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IL-10 and lymphotoxin-? expression profiles within marginal zone-like B-cell populations are associated with control of HIV-1 disease progression.
PLoS ONE
PUBLISHED: 01-01-2014
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Understanding how the immune system facilitates or controls HIV-1 disease progression has important implications for the design of effective interventions. We report that although B-cell dysregulations associated with HIV-1 disease progression are accompanied by an overall decrease in the percentage of total blood B-cells, we observe an increase in relative frequencies of cells presenting characteristics of both transitional immature and first-line marginal zone (MZ) B-cell populations, we designated as precursor MZ-like B-cells. B-cells with similar attributes have been associated with IL-10 expression and "regulatory" potential. As such, the relative frequencies of precursor MZ-like B-cells expressing IL-10 are increased in the blood of viremic HIV-1-infected individuals when compared to HIV-negative subjects. Importantly, in aviremic HIV-1 Elite-Controllers (EC), we found unaltered relative percentages of precursor MZ-like B-cells which presented normal IL-10 expression patterns. Furthermore, EC had increased relative frequencies of blood MZ-like B-cells expressing LT-?. Thus in contrast to viremic HIV-1-infected individuals, EC present MZ-like B-cell populations which IL-10 and LT-? expression profiles may favour homeostasis of immune responses and lymphoid microenvironments.
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Clinical prediction and diagnosis of neurosyphilis in HIV-infected patients with early Syphilis.
J. Clin. Microbiol.
PUBLISHED: 10-02-2013
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The diagnosis of neurosyphilis (NS) is a challenge, especially in HIV-infected patients, and the criteria for deciding when to perform a lumbar puncture (LP) in HIV-infected patients with syphilis are controversial. We retrospectively reviewed demographic, clinical, and laboratory data from 122 cases of HIV-infected patients with documented early syphilis who underwent an LP to rule out NS, and we evaluated 3 laboratory-developed validated real-time PCR assays, the Treponema pallidum particle agglutination (TPPA) assay, the fluorescent treponemal antibody absorption (FTA-ABS) assay, and the line immunoassay INNO-LIA Syphilis, for the diagnosis of NS from cerebrospinal fluid (CSF) samples of these patients. NS was defined by a reactive CSF-VDRL test result and/or a CSF white blood cell (WBC) count of >20 cells/?l. Thirty of the 122 patients (24.6%) had early NS. Headache, visual symptoms, a CD4 cell count of <500 cells/?l, and viremia, as defined by an HIV-1 RNA count of ?50 copies/ml, were associated with NS in multivariate analysis (P = <0.001 for each factor). Blood serum rapid plasma reagin (RPR) titers were not associated with early NS (P = 0.575). For the diagnosis of NS, the PCR, FTA-ABS, TPPA, and INNO-LIA assays had sensitivities of 58%, 100%, 68%, and 100%, specificities of 67%, 12%, 49%, and 13%, and negative predictive values of 85%, 100%, 84%, and 100%, respectively. Visual disturbances, headache, uncontrolled HIV-1 viremia, and a CD4 cell count of <500 cells/?l were predictors of NS in HIV-infected patients with early syphilis, while blood serum RPR titers were not; therefore, RPR titers should not be used as the sole criterion for deciding whether to perform an LP in early syphilis. When applied to CSF samples, the INNO-LIA Syphilis assay easily helped rule out NS.
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Application of independent component analysis on Raman images of a pharmaceutical drug product: Pure spectra determination and spatial distribution of constituents.
J Pharm Biomed Anal
PUBLISHED: 09-12-2013
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Independent component analysis (ICA) was used as a blind source separation method on a Raman image of a pharmaceutical tablet. Calculations were performed without a priori knowledge concerning the formulation. The aim was to extract the pure signals from the initial data set in order to examine the distribution of actives and major excipients within the tablet. As a method based on the decomposition of a matrix of mixtures of several components, the number of independent component to choose is a critical step of the analysis. The ICA_by_blocks method, based on the calculation of several models using an increasing number of independent components on initial matrix blocks, was used. The calculated ICA signals were compared with the pure spectra of the formulation compounds. High correlations between the two active principal ingredient spectra and their corresponding calculated signals were observed giving a good overview of the distributions of these compounds within the tablet. Information from the major excipients (lactose and avicel) was found in several independent components but the ICA approach provides high level of information concerning their distribution within the tablet. However, the results could vary considerably by changing the number of independent components or the preprocessing method. Indeed, it was shown that under-decomposition of the matrix could lead to better signal quality (compared to the pure spectra) but in that case the contributions due to minor components or effects were not correctly identified and extracted. On the contrary, over-decomposition of the original dataset could provide information about low concentration compounds at the expense of some loss of signal interpretability for the other compounds.
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The association between human leukocyte antigen (HLA)-G polymorphisms and human papillomavirus (HPV) infection in Inuit women of northern Quebec.
Hum. Immunol.
PUBLISHED: 08-12-2013
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The human leukocyte antigen (HLA)-G molecules act as negative regulators of the immune response. We analyzed the associations between HLA G polymorphisms and human papillomavirus (HPV) infection and squamous intraepithelial lesions (SIL) in Inuit women from Nunavik, northern Quebec.
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Juvenile respiratory papillomatosis: risk factors for severity.
J. Med. Virol.
PUBLISHED: 03-11-2013
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Juvenile recurrent respiratory papillomatosis is caused mainly by human papillomavirus genotypes 6 or 11, acquired at birth or during pregnancy from an infected mother. Recurrent respiratory papillomatosis is characterized by recurring warts growing most commonly in the larynx. Multiple surgical procedures and the risk of airway obstruction contribute to the devastating impact of this disease. Some children will go into remission after a few surgeries whereas others will require repeated interventions over several years. Further understanding of the risk factors associated with severity may contribute to tailored treatments. A retrospective study of cases diagnosed between January 1995 and December 2008 was conducted to study determinants of severe forms of juvenile recurrent respiratory papillomatosis. Demographic and clinical variables were abstracted from childrens medical charts and mothers delivery charts. Viral factors (HPV genotyping and viral load) were studied from archived biopsies. Specific HLA class II alleles and killer-cell immunoglobulin-like receptors genes were tested from saliva samples. Logistic regression was performed to identify risk factors for severity. Overall, 31 pediatric cases of recurrent respiratory papillomatosis were identified. The only significant factor associated with severe forms of recurrent respiratory papillomatosis was the maternal history of condylomas during pregnancy (OR: 12.05 [P=0.05]). The analysis failed to identify risk factors that could be used clinically to identify recurrent respiratory papillomatosis cases likely to take a severe course. Although too early to determine, vaccination against the HPV types involved most commonly in recurrent respiratory papillomatosis may provide the best hope to prevent severe forms of this disease.
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Translational diffusion of probe molecules under high pressure: a study by fluorescence recovery after photobleaching technique.
Rev Sci Instrum
PUBLISHED: 03-08-2013
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We present fluorescence recovery measurements after photobleaching performed under high pressure in liquids that fill square-section fused silica micro-capillaries. These micro-capillaries withstand pressure up to 2500 bar for a wall thickness of about 140 ?m and fit easily on the microscope stage. This technique allows the translational diffusion coefficient of fluorescent molecules in liquids to be measured as a function of pressure. When the liquid sample is far from its glass transition the translational diffusive coefficient is in agreement with the Stokes-Einstein equation. As the glass transition is approached by further increasing the pressure, decoupling of the measured diffusion coefficient from the Stokes-Einstein relation is observed. These are the first measurements that combine the fluorescence recovery technique and high hydrostatic pressures. This experimental setup can also be used either with diamond or sapphire anvil cells in order to span a larger pressure range.
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Raman spectroscopy and multivariate analysis for the rapid discrimination between native-like and non-native states in freeze-dried protein formulations.
Eur J Pharm Biopharm
PUBLISHED: 01-10-2013
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This study investigates whether Raman spectroscopy combined with multivariate analysis (MVA) enables a rapid and direct differentiation between two classes of conformational states, i.e., native-like and non-native proteins, in freeze-dried formulations. A data set comprising of 99 spectra, both from native-like and various types of non-native freeze-dried protein formulations, was obtained by freeze-drying lactate dehydrogenase (LDH) as model protein under various conditions. Changes in the secondary structure in the solid freeze-dried proteins were determined through visual interpretation of the blank corrected second derivative amide I band in the ATR-FTIR spectra (further called FTIR spectra) and served as an independent reference to assign class labels. Exploratory analysis and supervised classification, using Principal Components Analysis (PCA) and Partial Least Squares - Linear Discriminant Analysis (PLS-LDA), respectively, revealed that Raman spectroscopy is with 95% accuracy able to correctly discriminate between native-like and non-native states in the tested freeze-dried LDH formulations. Backbone (i.e., amide III) and side chain sensitive spectral regions proved important for making the discrimination between both classes. As discrimination was not influenced by the spectral signals from the tested excipients, there was no need for blank corrections. The Raman model may allow direct and automated analysis of the investigated quality attribute, opening possibilities for a real time and in-line quality indication as a future step. However, the sensitivity of the method should be further investigated and where possible improved.
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The frequency of HLA alleles in a population of Inuit women of northern Quebec.
Int J Circumpolar Health
PUBLISHED: 01-01-2013
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Human leukocyte antigen (HLA) alleles code for proteins that are involved in the recognition of foreign antigens and activation of the immune system. The frequency of HLA alleles varies across different populations.
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vanA-containing Enterococcus faecium susceptible to vancomycin and teicoplanin because of major nucleotide deletions in Tn1546.
J. Antimicrob. Chemother.
PUBLISHED: 09-15-2011
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During the course of routine screening for vancomycin-resistant enterococci (VRE), we found six Enterococcus faecium isolates positive for vanA by PCR, but susceptible to vancomycin and teicoplanin by phenotypic testing. The aim of this study was to characterize the genetic composition of the Tn1546 vanA gene cluster of these isolates.
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Ciprofloxacin-resistant Shigella sonnei among men who have sex with men, Canada, 2010.
Emerging Infect. Dis.
PUBLISHED: 09-06-2011
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In 2010, we observed isolates with matching pulsed-field gel electrophoresis patterns from 13 cases of ciprofloxacin-resistant Shigella sonnei in Montréal. We report on the emergence of this resistance type and a study of resistance mechanisms. The investigation suggested local transmission among men who have sex with men associated with sex venues.
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Transmission clustering drives the onward spread of the HIV epidemic among men who have sex with men in Quebec.
J. Infect. Dis.
PUBLISHED: 09-02-2011
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Phylodynamic analysis and epidemiologic data identified 3 patterns of spread of primary human immunodeficiency virus type 1 infection (PHI) among men who have sex with men (2001-2009): 420 unique PHIs, 102 small clusters (2-4 PHIs per cluster, n = 280), and 46 large clusters (5-31 PHIs per cluster, n = 450). Large clusters disproportionately increased from 25.2% of PHIs in 2005 to 39.1% in 2009 (?(2) = 33.9, P < .001). Scalar expansion of large clusters over 11 months (interquartile range, 3.5-25.5 months) correlated with cluster membership size (r(2) = 0.174, F = 4.424, P = .047). PHI cohort data revealed variations in social networks and risk behaviors among the 3 groups, suggesting the need for tailored prevention measures.
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High level of soluble HLA-G in the female genital tract of Beninese commercial sex workers is associated with HIV-1 infection.
PLoS ONE
PUBLISHED: 06-24-2011
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Most HIV infections are transmitted across mucosal epithelium. Understanding the role of innate and specific mucosal immunity in susceptibility or protection against HIV infection, as well as the effect of HIV infection on mucosal immunity, are of fundamental importance. HLA-G is a powerful modulator of the immune response. The aim of this study was to investigate whether soluble HLA-G (sHLA-G) expression in the female genital tract is associated with HIV-1 infection.
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A comprehensive study of polymorphic sites along the HLA-G gene: implication for gene regulation and evolution.
Mol. Biol. Evol.
PUBLISHED: 05-28-2011
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HLA-G molecule plays an important role on immune response regulation and has been implicated on the inhibition of T and natural killer cell cytolytic function and inhibition of allogeneic T-cell proliferation. Due to its immune-modulator properties, the HLA-G gene expression has been associated with the outcome of allograft and of autoimmune, infectious, and malignant disorders. Several lines of evidence indicate that HLA-G polymorphisms at the 5-upstream regulatory region (5 URR) and 3-untranslated region (3 UTR) may influence the HLA-G expression levels. Because Brazilians represent one of the most heterogeneous populations in the world with the widest HLA-G coding region variability already detected among the studied populations, a high level of variability and haplotype diversity would be expected in Brazilians. On this basis, the 5 URR, coding, and 3 UTR variability were evaluated in a Brazilian series consisting of 100 healthy bone marrow donors, as well as the linkage disequilibrium pattern along the gene and the extended haplotypes encompassing several gene segment variations. The HLA-G locus seems to present six different HLA-G lineages showing functional variations mainly in nucleotides of the regulatory regions. Differences were observed at the 5 URR in positions that either coincide with or are close to transcription factor-binding sites and at the 3 UTR mainly in positions that have already been reported to influence HLA-G mRNA availability. We report several lines of evidence for balancing selection acting on the regulatory regions, which may indicate that these HLA-G lineages may be related to the differential HLA-G expression profiles.
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Effects of chronic renal failure on kidney drug transporters and cytochrome P450 in rats.
Drug Metab. Dispos.
PUBLISHED: 04-27-2011
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Chronic renal failure (CRF) leads to decreased drug renal clearance due to a reduction in the glomerular filtration rate. However, little is known about how renal failure affects renal metabolism and elimination of drugs. Because both depend on the activity of uptake and efflux by renal transporters as well as enzymes in tubular cells, the purpose of this study was to investigate the effects of CRF on the expression and activity of select renal drug transporters and cytochrome P450. Two groups of rats were studied: control and CRF (induced by 5/6 nephrectomy). Compared with control rats, we observed reductions in the expression of both protein and mRNA of Cyp1a, sodium-dependent phosphate transport protein 1, organic anion transporter (Oat)1, 2, and 3, OatK1/K2, organic anion-transporting polypeptide (Oatp)1 and 4c1, P-glycoprotein, and urate transporter 1, whereas an induction in the protein and mRNA expression of Mrp2, 3, and 4 and Oatp2 and 3 was observed. Cyp3a expression remained unchanged. Similar results were obtained by incubating a human proximal tubule cell line (human kidney-2) with sera from CRF rats, suggesting the presence of uremic modulators. Finally, the renal elimination of [(3)H]digoxin and [(14)C]benzylpenicillin was decreased in CRF rats, compared with controls, as shown by a 4- and 9-fold accumulation, respectively, of these drugs in kidneys of rats in CRF. Our results demonstrate that CRF affects the expression and activity of several kidney drug transporters leading to the intrarenal accumulation of drugs and reduced renal clearance that could, at least partially, explain the tubular toxicity of many drugs.
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Short communication: persistence of high blood levels of the chemokines CCL2, CCL19, and CCL20 during the course of HIV infection.
AIDS Res. Hum. Retroviruses
PUBLISHED: 01-23-2011
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Dendritic cells (DCs) are important mediators of the immune response against HIV and yet blood DC numbers fall substantially during HIV infection. Here we report that blood levels of the DC-tissue tropic chemokines monocyte chemotactic protein (MCP-1/CCL2), macrophage inflammatory proteins (MIP-3?/CCL20), and MIP-3?/CCL19 remained elevated throughout the course of HIV infection suggesting that the relatively low levels of blood circulating DCs may be due to active recruitment of these cells to peripheral sites to fight disease progression.
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Human leukocyte antigen (HLA)-E and HLA-G polymorphisms in human papillomavirus infection susceptibility and persistence.
Hum. Immunol.
PUBLISHED: 01-13-2011
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Human leukocyte antigen (HLA)-E and HLA-G molecules act as powerful modulators of innate and adaptive immune responses. The study examined whether HLA-E and/or HLA-G polymorphisms are associated with human papillomavirus (HPV) infection susceptibility and persistence in 636 female university students in Montreal. HLA-G*01:01:02 and HLA-G*01:01:08 alleles were associated with increased risk of HPV-16 (odds ratio (OR) = 2.10, 95% confidence interval (CI), 1.11-3.96) and any infections with HPV types from ? species 1, 8, 10, and 13 (OR = 2.72, 95% CI, 1.11-6.68). HLA-G*01:01:02 and HLA-G*01:03 alleles were associated with persistent HPV-16 (OR = 2.07, 95% CI, 1.16-3.68) and persistent infections with HPV types from ? species 2, 3, 4, and 15 (OR = 2.99, 95% CI, 1.12-8.00). HLA-E polymorphism was not associated with risk of acquisition or persistence of HPV infection. These results suggest that HLA-G molecules may play a role in mediating HPV infection risk.
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Discrimination of corn from monocotyledonous weeds with ultraviolet (UV) induced fluorescence.
Appl Spectrosc
PUBLISHED: 01-08-2011
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In production agriculture, savings in herbicides can be achieved if weeds can be discriminated from crop, allowing the targeting of weed control to weed-infested areas only. Previous studies demonstrated the potential of ultraviolet (UV) induced fluorescence to discriminate corn from weeds and recently, robust models have been obtained for the discrimination between monocots (including corn) and dicots. Here, we developed a new approach to achieve robust discrimination of monocot weeds from corn. To this end, four corn hybrids (Elite 60T05, Monsanto DKC 26-78, Pioneer 39Y85 (RR), and Syngenta N2555 (Bt, LL)) and four monocot weeds (Digitaria ischaemum (Schreb.) I, Echinochloa crus-galli (L.) Beauv., Panicum capillare (L.), and Setaria glauca (L.) Beauv.) were grown either in a greenhouse or in a growth cabinet and UV (327 nm) induced fluorescence spectra (400 to 755 nm) were measured under controlled or uncontrolled ambient light intensity and temperature. This resulted in three contrasting data sets suitable for testing the robustness of discrimination models. In the blue-green region (400 to 550 nm), the shape of the spectra did not contain any useful information for discrimination. Therefore, the integral of the blue-green region (415 to 455 nm) was used as a normalizing factor for the red fluorescence intensity (670 to 755 nm). The shape of the normalized red fluorescence spectra did not contribute to the discrimination and in the end, only the integral of the normalized red fluorescence intensity was left as a single discriminant variable. Applying a threshold on this variable minimizing the classification error resulted in calibration errors ranging from 14.2% to 15.8%, but this threshold varied largely between data sets. Therefore, to achieve robustness, a model calibration scheme was developed based on the collection of a calibration data set from 75 corn plants. From this set, a new threshold can be estimated as the 85% quantile on the cumulative frequency curve of the integral of the normalized red fluorescence. With this approach the classification error was nearly constant (16.0% to 18.5%), thereby indicating the potential of UV-induced fluorescence to reliably discriminate corn from monocot weeds.
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Subtype diversity associated with the development of HIV-1 resistance to integrase inhibitors.
J. Med. Virol.
PUBLISHED: 01-05-2011
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We used genotypic and phylogenetic analysis to determine integrase diversity among subtypes, and studied natural polymorphisms and mutations implicated in resistance to integrase inhibitors (INI) in treatment-naïve persons (n = 220) and -experienced individuals (n = 24). Phylogenetics revealed 7 and 10% inter-subtype diversity in the integrase and reverse transcriptase (RT)/protease regions, respectively. Integrase sequencing identified a novel A/B recombinant in which all viruses in a male-sex-male (MSM) transmission cluster (n = 12) appeared to possess subtype B in integrase and subtype A in the remainder of the pol region. Natural variations and signature polymorphisms were observed at codon positions 140, 148, 151, 157, and 160 among HIV subtypes. These variations predicted higher genetic barriers to G140S and G140C in subtypes C, CRF02_AG, and A/CRF01_AE, as well as higher genetic barriers toward acquisition of V151I in subtypes CRF02_AG and A/CRF01_AE. The E157Q and E160Q mutational motif was observed in 35% of INI-naïve patients harboring subtype C infections, indicating intra-subtype variations. Thirteen patients failed raltegravir (RAL)-containing regimens within 8 ± 1 months, in association with the major Q148K/R/H and G140A/S (n = 8/24) or N155H (n = 5/24) mutational pathways. Of note, the remaining patients on RAL regimens for 14 ± 3 months harbored no or only minor integrase mutations/polymorphisms (T66I, T97A, H114P, S119P, A124S, G163R, I203M, R263K). These results demonstrate the importance of understanding subtype variability in the development of resistance to INIs.
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Implications of the polymorphism of HLA-G on its function, regulation, evolution and disease association.
Cell. Mol. Life Sci.
PUBLISHED: 10-21-2010
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The HLA-G gene displays several peculiarities that are distinct from those of classical HLA class I genes. The unique structure of the HLA-G molecule permits a restricted peptide presentation and allows the modulation of the cells of the immune system. Although polymorphic sites may potentially influence all biological functions of HLA-G, those present at the promoter and 3 untranslated regions have been particularly studied in experimental and pathological conditions. The relatively low polymorphism observed in the MHC-G coding region both in humans and apes may represent a strong selective pressure for invariance, whereas, in regulatory regions several lines of evidence support the role of balancing selection. Since HLA-G has immunomodulatory properties, the understanding of gene regulation and the role of polymorphic sites on gene function may permit an individualized approach for the future use of HLA-G for therapeutic purposes.
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Household transmission of the 2009 pandemic A/H1N1 influenza virus: elevated laboratory?confirmed secondary attack rates and evidence of asymptomatic infections.
Clin. Infect. Dis.
PUBLISHED: 10-05-2010
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Characterizing household transmission of the 2009 pandemic A/H1N1 influenza virus (pH1N1) is critical for the design of effective public health measures to mitigate spread. Our objectives were to estimate the secondary attack rates (SARs), the proportion of asymptomatic infections, and risk factors for pH1N1 transmission within households on the basis of active clinical follow-up and laboratory-confirmed outcomes.
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High expression levels of B lymphocyte stimulator (BLyS) by dendritic cells correlate with HIV-related B-cell disease progression in humans.
Blood
PUBLISHED: 09-24-2010
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In view of assessing the possible contribution of dendritic cells (DCs) to HIV-related B-cell disorders, we have longitudinally measured B lymphocyte stimulator (BLyS) surface expression by myeloid DCs (mDCs) and concentrations of B-cell growth factors in the blood of subjects undergoing primary HIV infection with different rates of disease progression. We report that BLyS surface expression by mature mDCs and precursors as well as blood levels of BLyS, a proliferation-inducing ligand (APRIL), interleukin-6 (IL-6), and IL-10 increased above normal levels in both rapid and normal HIV progressors as quickly as in the acute phase of infection and persisting throughout the course of disease despite successful therapy. Consequently, hyperglobulinemia and high blood levels of circulating activated mature B cells and precursor/activated marginal zone (MZ)-like B cells were found throughout follow-up for both rapid and normal progressors. In contrast, mDC cell-surface expression of BLyS as well as blood levels of BLyS, immunoglobulin, activated mature B cells, and precursor/activated MZ-like B cells in aviremic slow progressors were similar to those observed in healthy donors. Interestingly, the levels of mature MZ B cells were significantly reduced in slow progressors. Our results suggest that DCs might modulate the outcome of the HIV-related B-cell disease progression through the expression of BLyS.
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Organic anion transporting polypeptide 1B1 (OATP1B1) and OATP1B3: genetic variability and haplotype analysis in white Canadians.
Drug Metab. Pharmacokinet.
PUBLISHED: 09-22-2010
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Organic anion transporting polypeptide 1B1 (OATP1B1) and OATP1B3 are human hepatocyte transporters that mediate the uptake of various endogenous and exogenous substances. Genetic variations in solute carrier transporter 1B1 (SLCO1B1) and SLCO1B3 genes, which encode OATP1B1 and OATP1B3 proteins, could affect the pharmacokinetics of drugs leading to interindividual differences in drug responses. The full extent of SLCO1B1 and SLCO1B3 polymorphisms in white Canadians was analyzed using DNA sequencing procedures. We identified 49 and 41 nucleotide sequence variants leading to 10 and 9 major haplotypes in SLCO1B1 and SLCO1B3 genes, respectively. We report several novel mutations within regulatory and coding regions that could affect gene transcription, translation and function. Comparison with other studies revealed that the distribution of SLCO1B1 and SLCO1B3 polymorphisms and haplotypes differs widely across populations. Data from this survey will ultimately contribute to the design of pharmacogenetic studies in the Canadian population.
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The surface molecular functionality of decellularized extracellular matrices.
Biomaterials
PUBLISHED: 08-17-2010
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Decellularization of tissues and organs is a successful platform technology for creating scaffolding materials for tissue engineering and regenerative medicine. It has been suggested that the success of these materials upon implantation is due to the molecular signals provided by the remaining scaffold extracellular matrix (ECM) components presented to probing cells in vivo as they repopulate the surface. For this study, decellularized matrices were created from esophagus, bladder, and small intestine harvested from adult male Fischer 344 rats. The three decellularized matrices (each originating from source tissues which included an epithelial lining on their luminal surfaces) were immunostained for collagen IV and laminin to determine basement membrane retention. Scanning electron micrographs of the surfaces were used to provide insight into the surface topography of each of the decellularized tissues. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) was used to generate high-resolution mass spectra for the surfaces of each scaffold. This surface-sensitive technique allows for detailed molecular analysis of the outermost 1-2 nm of a material and has been applied previously to thin protein films and secreted ECM proteins on poly(N-isopropyl acrylamide) (polyNIPAAM) surfaces. To extract trends from within the complex ToF-SIMS dataset, a multivariate analysis technique, principal component analysis (PCA), was employed. Using this method, a molecular fingerprint of each surface was created and separation was seen in the PCA scores between the decellularized esophagus and the decellularized small intestine samples. The PCA scores for the decellularized bladder sample fell between the previous two decellularized samples. Protein films of common extracellular matrix constituents (collagen IV, collagen I, laminin, and Matrigel) were also investigated. The PCA results from these protein films were used to develop qualitative hypotheses for the relationship of the key fragments identified from the PCA of the decellularized ECMs.
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Determinants of human papillomavirus infection among Inuit women of northern Quebec, Canada.
Sex Transm Dis
PUBLISHED: 05-18-2010
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We investigated risk factors for prevalent high-risk human papillomavirus (HR-HPV) in Inuit women from Quebec. Younger age and having 10 or more lifetime sexual partners were associated with HR-HPV. Findings suggest that for older women, markers of recent sexual activity are more predictive of HR-HPV status than markers of lifetime sexual history.
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Predicting tipranavir and darunavir resistance using genotypic, phenotypic, and virtual phenotypic resistance patterns: an independent cohort analysis of clinical isolates highly resistant to all other protease inhibitors.
Antimicrob. Agents Chemother.
PUBLISHED: 04-05-2010
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Genotypic interpretation systems (GISs) for darunavir and tipranavir susceptibility are rarely tested by the use of independent data sets. The virtual phenotype (the phenotype determined by Virco [the "Vircotype"]) was used to interpret all genotypes in Québec, Canada, and phenotypes were determined for isolates predicted to be resistant to all protease inhibitors other than darunavir and tipranavir. We used multivariate analyses to predict relative phenotypic susceptibility to darunavir and tipranavir. We compared the performance characteristics of the Agence Nationale de Recherche sur le Sida scoring algorithm, the Stanford HIV database scoring algorithm (with separate analyses of the discrete and numerical scores), the Vircotype, and the darunavir and tipranavir manufacturers scores for prediction of the phenotype. Of the 100 isolates whose phenotypes were determined, 89 and 72 were susceptible to darunavir and tipranavir, respectively. In multivariate analyses, the presence of I84V and V82T and the lack of L10F predicted that the isolates would be more susceptible to darunavir than tipranavir. The presence of I54L, V32I, and I47V predicted that the isolates would be more susceptible to tipranavir. All GISs except the system that provided the Stanford HIV database discrete score performed well in predicting the darunavir resistance phenotype (R(2) = 0.61 to 0.69); the R(2) value for the Stanford HIV database discrete scoring system was 0.38. Other than the system that provided the Vircotype (R(2) = 0.80), all GISs performed poorly in predicting the tipranavir resistance phenotype (R(2) = 0.00 to 0.31). In this independent cohort harboring highly protease inhibitor-resistant HIV isolates, reduced phenotypic susceptibility to darunavir and tipranavir was rare. Generally, GISs predict susceptibility to darunavir substantially better than they predict susceptibility to tipranavir.
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Performance of an agar dilution method and a Vitek 2 card for detection of inducible clindamycin resistance in Staphylococcus spp.
J. Clin. Microbiol.
PUBLISHED: 02-17-2010
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The D-zone test detects inducible clindamycin resistance in Staphylococcus spp. Two other methods not described by the Clinical and Laboratory Standards Institute (CLSI) are available to test for this resistance mechanism: an agar dilution method and new Vitek 2 cards. This study evaluated the performance of both methods in detecting inducible clindamycin resistance. Nonduplicate clinical strains of Staphylococcus spp. (111 Staphylococcus aureus and 52 coagulase-negative staphylococcus strains), intermediate or resistant to erythromycin but susceptible to clindamycin, were obtained from three hospitals in Montreal, Quebec, Canada. Molecular analysis to detect resistance genes was conducted on all strains. A Mueller-Hinton agar containing 1 mg of erythromycin and 0.5 mg of clindamycin/liter was used for the dilution method, and two inocula were tested: 10(4) and 10(5) CFU per spot. Plates were read at 24 and 48 h. The Vitek 2 AST-P580 card was used according to the manufacturers recommendations. The results were compared to those of the D-zone test. The D-zone test was positive in 134 of 163 (82%) strains. With the 10(4) CFU inoculum, the sensitivities were 84 and 99% at 24 and 48 h, respectively. The 10(5) CFU inoculum increased the sensitivities at 24 and 48 h to 91 and 100%, respectively. The specificity was 100% for the 10(4) CFU inoculum at 24 h and 97% for the other combinations. The sensitivity and specificity for the Vitek 2 card were 93 and 100%, respectively. The performance of both the agar dilution method and the Vitek 2 card was good, but these methods were not as sensitive as the D-zone test at 24 h.
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Improved discrimination between monocotyledonous and dicotyledonous plants for weed control based on the blue-green region of ultraviolet-induced fluorescence spectra.
Appl Spectrosc
PUBLISHED: 02-06-2010
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Precision weeding by spot spraying in real time requires sensors to discriminate between weeds and crop without contact. Among the optical based solutions, the ultraviolet (UV) induced fluorescence of the plants appears as a promising alternative. In a first paper, the feasibility of discriminating between corn hybrids, monocotyledonous, and dicotyledonous weeds was demonstrated on the basis of the complete spectra. Some considerations about the different sources of fluorescence oriented the focus to the blue-green fluorescence (BGF) part, ignoring the chlorophyll fluorescence that is inherently more variable in time. This paper investigates the potential of performing weed/crop discrimination on the basis of several large spectral bands in the BGF area. A partial least squares discriminant analysis (PLS-DA) was performed on a set of 1908 spectra of corn and weed plants over 3 years and various growing conditions. The discrimination between monocotyledonous and dicotyledonous plants based on the blue-green fluorescence yielded robust models (classification error between 1.3 and 4.6% for between-year validation). On the basis of the analysis of the PLS-DA model, two large bands were chosen in the blue-green fluorescence zone (400-425 nm and 425-490 nm). A linear discriminant analysis based on the signal from these two bands also provided very robust inter-year results (classification error from 1.5% to 5.2%). The same selection process was applied to discriminate between monocotyledonous weeds and maize but yielded no robust models (up to 50% inter-year error). Further work will be required to solve this problem and provide a complete UV fluorescence based sensor for weed-maize discrimination.
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Assessing yeast viability from cell size measurements?
J. Biotechnol.
PUBLISHED: 01-18-2010
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During microbial cell cultures, environmental conditions affect cell physiology and subsequently process efficiency. Physiological changes result in changing cell morphology, such as cell size variations. The aim of this work was to study cell size evolution of a Saccharomyces cerevisiae population exposed to various stresses during alcoholic batch fermentations, and to evaluate the potential use of cell size measurements to infer cell viability. During a reference culture, without perturbation, viability as assessed by propidium iodide staining (PI) remained 100% and mean cell diameter was found to be above 5microm. A rapid temperature shift from 33 to 43 degrees C at 50gl(-1) of ethanol resulted in an immediate arrest of growth and triggered a progressive loss of viability from 100% to 0% and a decrease of mean cell diameter from 5.2 to 3.7microm. Cell size distribution curves obtained with a cell counter showed an increasing subpopulation of significantly smaller cells. At single-cell level, combined microscopy size measurements and PI staining showed that this subpopulation was exclusively composed of dead cells. Similar results were obtained after acetic acid or furfural additions. Accordingly, a multivariate data analysis was achieved to estimate the ratio of dead cells from cell size distributions obtained using the cell counter.
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Improvement of direct calibration in spectroscopy.
Anal. Chim. Acta
PUBLISHED: 01-16-2010
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Several linear calibration methods have been proposed for predicting the concentration of a particular compound from a spectrum. Some methods are based on experimental data, such as Partial Least Square Regression. Other methods are based on expert data, e.g. Direct Calibration. This article proposes a new method, called Improved Direct Calibration, which uses expert and experimental information. It performs a projection onto the pure interest spectrum, after correcting it from influence factors. No calibration dataset is necessary to build this model. This method has been successfully applied to the quantification of ethanol in musts during fermentation, using near infrared spectrometry.
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Blood soluble human leukocyte antigen G levels are associated with human immunodeficiency virus type 1 infection in Beninese commercial sex workers.
Hum. Immunol.
PUBLISHED: 09-15-2009
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Human leukocyte antigen (HLA)-G is a powerful modulator of the immune response. The aim of this study was to investigate whether soluble HLA-G (sHLA-G) expression is associated with human immunodeficiency virus type 1 (HIV-1) infection. HIV-1-infected female commercial sex workers (CSWs) had significantly lower levels of plasma sHLA-G compared with those in both the HIV-1-uninfected CSW and the non-CSW groups. The presence of HLA-G*010101, HLA-G*010404 alleles, and the 3-untranslated region (3UTR) genetic variant at position 3,952 were all significantly associated with lower plasma sHLA-G levels in the HIV-1-infected CSWs, whereas the HLA-G 3UTR 14-bp sequence insertion was also associated with lower plasma sHLA-G levels in the overall population. When adjustment was made for all significant variables, the reduced expression of sHLA-G in the plasma remained significantly associated with HIV-1 infection and the HLA-G 3UTR 14-bp insertion homozygote genotype. This study demonstrates that low levels of plasma sHLA-G are associated with HIV-1 infection.
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Chemokine expression patterns in the systemic and genital tract compartments are associated with HIV-1 infection in women from Benin.
J. Clin. Immunol.
PUBLISHED: 09-03-2009
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Understanding the genital mucosal immunity and the factors involved in linking innate to adaptive immunity is crucial for the design of efficient preventive strategies against human immunodeficiency virus (HIV)-1.
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Surface characterization of extracellular matrix scaffolds.
Biomaterials
PUBLISHED: 08-13-2009
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Extracellular matrix (ECM) scaffolds prepared from different tissue sources or using different methods have been demonstrated to have distinctive effects upon cell adhesion patterns and the ability to support and maintain differentiated phenotypes. It is unknown whether the molecular composition or the ultrastructure of the ECM plays a greater role in determining the phenotype of the cells with which it comes into contact. However, when implanted, the topology and ligand landscape of the material will determine the host molecules that bind and the type and behavior of cells that mediate the host response. Therefore, a comprehensive understanding of surface characteristics is essential in the design of scaffolds for specific clinical applications. The surface characteristics of ECM scaffolds derived from porcine urinary bladder, small intestine, and liver as well as the effects of two commonly used methods of chemical cross-linking upon UBM were investigated. Electron microscopy and time of flight secondary ion mass spectroscopy were used to examine the surface characteristics of the scaffolds. The results show that ECM scaffolds have unique morphologic and structural properties which are dependant on the organ or tissue from which the scaffold is harvested. Furthermore, the results show that the surface characteristics of an ECM scaffold are changed through chemical cross-linking.
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Detection and typing of human papillomavirus nucleic acids in biological fluids.
Public Health Genomics
PUBLISHED: 08-11-2009
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Human papillomaviruses (HPV) are the etiologic agents of cancer of the uterine cervix and several other neoplasias. Detection of HPV infection will improve the sensitivity of primary and secondary screening of cervical cancer. The clinical indications for the use of HPV tests will have to consider the natural history of HPV infection and diseases, and the multiplicity of types involved. Signal amplification HPV DNA tests detect several high-risk HPV types, are standardized, commercially available and approved for clinical use. Nucleic acid amplification techniques are ideal methods for epidemiologic purposes since they minimize misclassification of HPV infection status and allow detection of infection with low viral burden. They are currently under evaluation for clinical use. PCR is the most widespread method for HPV typing, especially with the use of consensus primers and typing with reverse hybridization techniques. Novel promising HPV detection strategies are now proposed, such as HPV mRNA detection, and suspension or solid phase arrays. These novel techniques will have to be evaluated as stringently as actual assays in clinical studies. Although assays have been developed for the evaluation of viral load, viral integration and HPV polymorphism in molecular epidemiological studies, their role in clinical practice is not currently defined.
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Endogenous hepatocyte growth factor is a niche signal for subventricular zone neural stem cell amplification and self-renewal.
Stem Cells
PUBLISHED: 08-07-2009
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Neural stem cells persist in the adult mammalian brain, within the subventricular zone (SVZ). The endogenous mechanisms underpinning SVZ neural stem cell proliferation, self-renewal, and differentiation are not fully elucidated. In the present report, we describe a growth-stimulatory activity of liver explant-conditioned media on SVZ cell cultures and identify hepatocyte growth factor (HGF) as a major player in this effect. HGF exhibited a mitogenic activity on SVZ cell cultures in a mitogen-activated protein kinase (MAPK) (ERK1/2)-dependent manner as U0126, a specific MAPK inhibitor, blocked it. Combining a functional neurosphere forming assay with immunostaining for c-Met, along with markers of SVZ cells subtypes, demonstrated that HGF promotes the expansion of neural stem-like cells that form neurospheres and self-renew. Immunostaining, HGF enzyme-linked immunosorbent assay and Madin-Darby canine kidney cell scattering assay indicated that SVZ cell cultures produce and release HGF. SVZ cell-conditioned media induced proliferation on SVZ cell cultures, which was blocked by HGF-neutralizing antibodies, hence implying that endogenously produced HGF accounts for a major part in SVZ mitogenic activity. Brain sections immunostaining revealed that HGF is produced by nestin-expressing cells and c-Met is expressed within the SVZ by immature cells. HGF intracerebroventricular injection promoted SVZ cell proliferation and increased the ability of these cells exposed in vivo to HGF to form neurospheres in vitro, whereas intracerebroventricular injection of HGF-neutralizing antibodies decreased SVZ cell proliferation. The present study unravels a major role, both in vitro and in vivo, for endogenous HGF in SVZ neural stem cell growth and self-renewal.
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Potential impact of (68)Ga-DOTATOC PET/CT on stereotactic radiotherapy planning of meningiomas.
Eur. J. Nucl. Med. Mol. Imaging
PUBLISHED: 06-24-2009
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Since meningiomas show a high expression of somatostatin receptor subtype 2, PET with (68)Ga-DOTATOC was proposed as an additional imaging modality beside CT and MRI for planning radiotherapy. We investigated the input of (68)Ga-DOTATOC-PET/CT on the definition of the "gross tumour volume" (GTV) in meningiomas, in order to assess the potential value of this method.
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Functional genetic variants in DC-SIGNR are associated with mother-to-child transmission of HIV-1.
PLoS ONE
PUBLISHED: 05-26-2009
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Mother-to-child transmission (MTCT) is the main cause of HIV-1 infection in children worldwide. Given that the C-type lectin receptor, dendritic cell-specific ICAM-grabbing non-integrin-related (DC-SIGNR, also known as CD209L or liver/lymph node-specific ICAM-grabbing non-integrin (L-SIGN)), can interact with pathogens including HIV-1 and is expressed at the maternal-fetal interface, we hypothesized that it could influence MTCT of HIV-1.
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Persistence of high levels of blood soluble human leukocyte antigen-G is associated with rapid progression of HIV infection.
AIDS
PUBLISHED: 05-23-2009
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Human leukocyte antigen-G is an important suppressor of the immune response, and HIV can modulate its expression. Longitudinal monitoring of soluble human leukocyte antigen-G plasma levels in patients with primary HIV infection undergoing different rates of disease progression showed that levels were elevated in the early phases of infection and remained high throughout follow-up in rapid progressors who responded to antiretroviral therapy but were restored to normal levels in the chronic phase of infection in both untreated normal progressors and long-term nonprogressors.
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Synthetic Plasmodium-like hemozoin activates the immune response: a morphology - function study.
PLoS ONE
PUBLISHED: 04-23-2009
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Increasing evidence points to an important role for hemozoin (HZ), the malaria pigment, in the immunopathology related to this infection. However, there is no consensus as to whether HZ exerts its immunostimulatory activity in absence of other parasite or host components. Contamination of native HZ preparations and the lack of a unified protocol to produce crystals that mimic those of Plasmodium HZ (PHZ) are major technical limitants when performing functional studies with HZ. In fact, the most commonly used methods generate a heterogeneous nanocrystalline material. Thus, it is likely that such aggregates do not resemble to PHZ and differ in their inflammatory properties. To address this issue, the present study was designed to establish whether synthetic HZ (sHZ) crystals produced by different methods vary in their morphology and in their ability to activate immune responses. We report a new method of HZ synthesis (the precise aqueous acid-catalyzed method) that yields homogeneous sHZ crystals (Plasmodium-like HZ) which are very similar to PHZ in their size and physicochemical properties. Importantly, these crystals are devoid of protein and DNA contamination. Of interest, structure-function studies revealed that the size and shape of the synthetic crystals influences their ability to activate inflammatory responses (e.g. nitric oxide, chemokine and cytokine mRNA) in vitro and in vivo. In summary, our data confirm that sHZ possesses immunostimulatory properties and underline the importance of verifying by electron microscopy both the morphology and homogeneity of the synthetic crystals to ensure that they closely resemble those of the parasite. Periodic quality control experiments and unification of the method of HZ synthesis are key steps to unravel the role of HZ in malaria immunopathology.
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Low-risk human papillomavirus type 6 DNA load and integration in cervical samples from women with squamous intraepithelial lesions.
J. Clin. Virol.
PUBLISHED: 03-26-2009
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The association between human papillomavirus (HPV) viral load and high-grade squamous intraepithelial lesion (HSIL) of the uterine cervix has been demonstrated for high-risk HPV-16 but has not been investigated for low-risk HPV types.
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Modification of aminosilanized superparamagnetic nanoparticles: feasibility of multimodal detection using 3T MRI, small animal PET, and fluorescence imaging.
Mol Imaging Biol
PUBLISHED: 03-03-2009
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The aim of our study was to modify an aminosilane-coated superparamagnetic nanoparticle for cell labeling and subsequent multimodal imaging using magnetic resonance imaging (MRI), positron emission tomography (PET), and fluorescent imaging in vivo.
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HIV infection affects blood myeloid dendritic cells after successful therapy and despite nonprogressing clinical disease.
J. Infect. Dis.
PUBLISHED: 02-24-2009
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We assessed the longitudinal changes in blood myeloid dendritic cell (mDC) and plasmacytoid dendritic cell (pDC) populations in subjects with primary human immunodeficiency virus (HIV) infection undergoing different rates of disease progression. The relative level and degree of maturation of all cell populations decreased significantly in untreated individuals with acute infection. The most dramatic changes were observed in the rapid progressor group, correlating with their rate of clinical progression. Levels of mDCs remained lower than normal throughout follow-up for both rapid progressors who responded to antiretroviral therapy (ART) and untreated normal progressors. In contrast, mDC precursors were restored to normal levels during subsequent phases of infection in both rapid and normal progressors, and these levels were increased in long-term nonprogressors. pDC levels followed the pattern of CD4+ T cell fluctuations. These findings provide evidence for an ongoing process affecting mDCs after successful ART and despite nonprogressing clinical disease following HIV infection.
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Natural Immunity to HIV: a delicate balance between strength and control.
Clin. Dev. Immunol.
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Understanding how the mucosal immune system in the human female reproductive tract might prevent or facilitate HIV infection has important implications for the design of effective interventions. We and others have established cohorts of highly-exposed, HIV-seronegative individuals, such as HIV-uninfected commercial sex workers, who have remained HIV-negative after more than 5 years of active prostitution. Observations obtained in studies of such individuals, who represent a model of natural immunity to HIV, indicate that HIV resistance may be associated with the hosts capacity to preserve systemic integrity by constraining immune activity and controlling inflammatory conditions at the mucosal point of entry. This likely necessitates the orchestration of balanced, first-line and adaptive immune responses.
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Influence of SLCO1B3 genetic variations on tacrolimus pharmacokinetics in renal transplant recipients.
Drug Metab. Pharmacokinet.
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The immunosuppressive drug tacrolimus requires strict therapeutic monitoring due to its narrow therapeutic index and high interindividual variability. Organic anion transporting polypeptide 1B3 (OATP1B3) is a human hepatocyte transporter involved in the hepatobiliary elimination of diverse endogenous and exogenous substances. Genetic variations within the solute carrier (SLCO) 1B3 gene that encodes OATP1B3 may contribute to interindividual differences in tacrolimus disposition. The purpose of the present study is to investigate the association between SLCO1B3 polymorphisms and tacrolimus pharmacokinetics in renal transplant recipients. We found significant correlations between two linked coding nonsynomymous polymorphisms, T334G and G699A, and mean dose-adjusted tacrolimus trough blood concentrations during the first week post-transplantation (p = 0.04) and when the target dose (10-12 ng/ml) was obtained (p = 0.01). Patients carrying the homozygous mutant haplotype had 14.3-fold higher risk (95% confidence interval: 1.43-100; p = 0.02) of having blood tacrolimus concentrations above the median level, and thus being classified as poor OATP1B3 transporters, than carriers of one or two copies of the wild-type haplotype. This study shows, for the first time, that SLCO1B3 polymorphism is associated with tacrolimus exposure in the early post-transplant period.
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Brain perfusion SPECT in the mouse: normal pattern according to gender and age.
Neuroimage
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Regional cerebral blood flow (rCBF) is a useful surrogate marker of neuronal activity and a parameter of primary interest in the diagnosis of many diseases. The increasing use of mouse models spawns the demand for in vivo measurement of rCBF in the mouse. Small animal SPECT provides excellent spatial resolution at adequate sensitivity and is therefore a promising tool for imaging the mouse brain. This study evaluates the feasibility of mouse brain perfusion SPECT and assesses the regional pattern of normal Tc-99m-HMPAO uptake and the impact of age and gender. Whole-brain kinetics was compared between Tc-99m-HMPAO and Tc-99m-ECD using rapid dynamic planar scans in 10 mice. Assessment of the regional uptake pattern was restricted to the more suitable tracer, HMPAO. Two HMPAO SPECTs were performed in 18 juvenile mice aged 7.5 ± 1.5weeks, and in the same animals at young adulthood, 19.1 ± 4.0 weeks (nanoSPECT/CTplus, general purpose mouse apertures: 1.2kcps/MBq, 0.7mm FWHM). The 3-D MRI Digital Atlas Database of an adult C57BL/6J mouse brain was used for region-of-interest (ROI) analysis. SPECT images were stereotactically normalized using SPM8 and a custom made, left-right symmetric HMPAO template in atlas space. For testing lateral asymmetry, each SPECT was left-right flipped prior to stereotactical normalization. Flipped and unflipped SPECTs were compared by paired testing. Peak brain uptake was similar for ECD and HMPAO: 1.8 ± 0.2 and 2.1 ± 0.6 %ID (p=0.357). Washout after the peak was much faster for ECD than for HMPAO: 24 ± 7min vs. 4.6 ± 1.7h (p=0.001). The general linear model for repeated measures with gender as an intersubject factor revealed an increase in relative HMPAO uptake with age in the neocortex (p=0.018) and the hippocampus (p=0.012). A decrease was detected in the midbrain (p=0.025). Lateral asymmetry, with HMPAO uptake larger in the left hemisphere, was detected primarily in the neocortex, both at juvenile age (asymmetry index AI=2.7 ± 1.7%, p=0.000) and at young adult age (AI=2.4 ± 1.7%, p=0.000). Gender had no effect on asymmetry. Voxel-wise testing confirmed the ROI-based findings. In conclusion, high-resolution HMPAO SPECT is a promising technique for measuring rCBF in preclinical research. It indicates lateral asymmetry of rCBF in the mouse brain as well as age-related changes during late maturation. ECD is not suitable as tracer for brain SPECT in the mouse because of its fast clearance from tissue indicating an interspecies difference in esterase activity between mice and humans.
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Naturally-occurring genetic variants in human DC-SIGN increase HIV-1 capture, cell-transfer and risk of mother-to-child transmission.
PLoS ONE
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Mother-to-child transmission (MTCT) is the main cause of HIV-1 infection in children worldwide. Dendritic cell-specific ICAM-3 grabbing-nonintegrin (DC-SIGN, also known as CD209) is an HIV-1 receptor that enhances its transmission to T cells and is expressed on placental macrophages.
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Application of direct calibration in multivariate image analysis of heterogeneous materials.
Anal. Chim. Acta
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Many scientific instruments produce multivariate images characterized by three-way tables, an element of which represents the intensity value at a spatial location for a given spectral channel. A problem frequently encountered is to attempt estimating the contributions of some compounds at each location of these images. Usual regression methods of calibration, such as PLS, require having a matrix of calibration X (n×p) and the corresponding vector y of the dependent variable (n×1). X can be built up by sampling pixel-vectors in the images, but y is sometimes difficult to obtain, if the surface of the samples is formed by chemically heterogeneous regions. In this case, the quantitative analyses related to y may be difficult, if the pixels represent very small areas (for example on microscopic images) or very large ones (satellite images). This is for example the case when dealing with biological solid samples representing different tissues. Direct Calibration (DC), sometimes referred to as "spectral unmixing", do not require having such a calibration set. However, it is indeed needed to have both a matrix of "perturbing" pixel-vectors (noted K) and a vector of the "pure" component spectrum to be analyzed (p), which are more easily obtainable. For estimating the contribution, the unknown pixel vector x and the pure spectrum p are first projected orthogonally onto K giving the vectors x(?) onto p(?), respectively. The contribution is then estimated by a second projection of x(?) onto p(?). A method, based on principal component analysis, for determining the optimal dimensions of K is proposed. DC was applied on a collection of multivariate images of kernel of wheat to estimate the proportion of three tissues, namely out-layers, "waxy"endosperm and normal endosperm. The eventual results are presented as images of wheat kernels in false colors associated to the estimated proportions of the tissues. It is shown that DC is appropriate for estimating contributions in situations in which the more usual methods of calibration cannot be applied.
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Contribution of 68Ga-DOTATOC PET/CT to target volume delineation of skull base meningiomas treated with stereotactic radiation therapy.
Int. J. Radiat. Oncol. Biol. Phys.
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To investigate the potential impact of 68Ga-DOTATOC positron emission tomography (68Ga-DOTATOC-PET) in addition to magnetic resonance imaging (MRI) and computed tomography (CT) for retrospectively assessing the gross tumor volume (GTV) delineation of meningiomas of the skull base in patients treated with fractionated stereotactic radiation therapy (FSRT).
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Straightforward thiol-mediated protein labelling with DTPA: Synthesis of a highly active 111In-annexin A5-DTPA tracer.
EJNMMI Res
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Annexin A5 (anxA5) has been found useful for molecular imaging of apoptosis and other biological processes.
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Influence of dendritic cells on B-cell responses during HIV infection.
Clin. Dev. Immunol.
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Dendritic cells (DCs) modulate B-cell differentiation, activation, and survival mainly through production of growth factors such as B lymphocyte stimulator (BLyS/BAFF). DC populations have been reported to be affected in number, phenotype and function during HIV infection and such alterations may contribute to the dysregulation of the B-cell compartment. Herein, we reflect on the potential impact of DC on the pathogenesis of HIV-related B cell disorders, and how DC status may modulate the outcome of mucosal B cell responses against HIV, which are pivotal to the control of disease. A concept that could be extrapolated to the overall outcome of HIV disease, whereby control versus progression may reside in the hosts capacity to maintain DC homeostasis at mucosal sites, where DC populations present an inherent capacity of modulating the balance between tolerance and protection, and are amongst the earliest cell types to be exposed to the virus.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.