Donor variability in HIV binding to peripheral blood mononuclear cells

<p>Abstract</p> <p>Background</p> <p>HIV infection of cells varies greatly between individuals, with multiple steps in the replication cycle potentially contributing to the variability. Although entry and post-entry variability of HIV infection levels in cells has been demonstrated, variability in HIV binding has not been examined. In this study, we examined variability of HIV binding to peripheral blood mononuclear cells (PBMC) from different donors.</p> <p>Results</p> <p>HIV binding to PBMC varied up to 3.9-fold between individuals and was independent of CD4. Replication of HIV in donor PBMC required CD4 and paralleled virus binding trends of donor PBMC. To assess the stability of virus binding phenotypes over time, HIV was bound to donors with low- and high-binding phenotypes. The binding phenotypes were maintained when tested weekly over a 4-week period for 3 of 4 donors, while one high-binding donor decreased to lower binding on the 4th week. The low- and high-binding phenotypes were also preserved across different HIV strains. Experiments performed to determine if there was an association between HIV binding levels and specific cell subset levels within PBMC showed no correlation, suggesting that HIV binds to multiple cell subsets.</p> <p>Conclusion</p> <p>These results show that differences exist in HIV binding to donor PBMC. Our data also show that HIV binding to donor PBMC is CD4-independent and can change over time, suggesting that virus binding variability is due to differences in the expression of changeable cell-surface host factors. Taken together, this study highlights the impact of cell-surface factors in HIV binding to, and infection of, PBMC which likely represents an important step in HIV infection <it>in vivo</it>.</p

Evaluation of transduction efficiency in macrophage colony-stimulating factor differentiated human macrophages using HIV-1 based lentiviral vectors

<p>Abstract</p> <p>Background</p> <p>Monocyte-derived macrophages contribute to atherosclerotic plaque formation. Therefore, manipulating macrophage function could have significant therapeutic value. The objective of this study was to determine transduction efficiency of two HIV-based lentiviral vector configurations as delivery systems for the transduction of primary human blood monocyte-derived macrophages.</p> <p>Results</p> <p>Human blood monocytes were transduced using two VSV-G pseudotyped HIV-1 based lentiviral vectors containing EGFP expression driven by either native HIV-LTR (VRX494) or EF1α promoters (VRX1090). Lentiviral vectors were added to cultured macrophages at different times and multiplicities of infection (MOI). Transduction efficiency was assessed using fluorescence microscopy and flow cytometry. Macrophages transduced between 2 and 120 hours after culturing showed the highest transduction efficiency at 2-hours transduction time. Subsequently, cells were transduced 2 hours after culturing at various vector concentrations (MOIs of 5, 10, 25 and 50) to determine the amount of lentiviral vector particles required to maximally transduce human monocyte-derived macrophages. On day 7, all transduced cultures showed EGFP-positive cells by microscopy. Flow cytometric analysis showed with all MOIs a peak shift corresponding to the presence of EGFP-positive cells. For VRX494, transduction efficiency was maximal at an MOI of 25 to 50 and ranged between 58 and 67%. For VRX1090, transduction efficiency was maximal at an MOI of 10 and ranged between 80 and 90%. Thus, transductions performed with VRX1090 showed a higher number of EGFP-positive cells than VRX494.</p> <p>Conclusions</p> <p>This report shows that VSV-G pseudotyped HIV-based lentiviral vectors can efficiently transduce human blood monocyte-derived macrophages early during differentiation using low particle numbers that do not interfere with differentiation of monocytes into macrophages.</p

Increased glucose transporter-1 expression on intermediate monocytes from HIV-infected women with subclinical cardiovascular disease

People living with HIV (PLWH) have chronic immune activation and increased cardiovascular disease (CVD) risk. Activation of monocytes and T lymphocytes causes up-regulation of glucose transporter-1 (GLUT1) for efficient function. PLWH have an increased percentage of GLUT1-expressing monocytes and T lymphocytes, but it is unclear if these cells are associated with CVD. We evaluated expression of GLUT1 and CD38 on monocyte and T lymphocyte populations from HIV-infected women with subclinical CVD

Genome sequencing reveals Zika virus diversity and spread in the Americas

Although the recent Zika virus (ZIKV) epidemic in the Americas and its link to birth defects have attracted a great deal of attention, much remains unknown about ZIKV disease epidemiology and ZIKV evolution, in part owing to a lack of genomic data. Here we address this gap in knowledge by using multiple sequencing approaches to generate 110 ZIKV genomes from clinical and mosquito samples from 10 countries and territories, greatly expanding the observed viral genetic diversity from this outbreak. We analysed the timing and patterns of introductions into distinct geographic regions; our phylogenetic evidence suggests rapid expansion of the outbreak in Brazil and multiple introductions of outbreak strains into Puerto Rico, Honduras, Colombia, other Caribbean islands, and the continental United States. We find that ZIKV circulated undetected in multiple regions for many months before the first locally transmitted cases were confirmed, highlighting the importance of surveillance of viral infections. We identify mutations with possible functional implications for ZIKV biology and pathogenesis, as well as those that might be relevant to the effectiveness of diagnostic tests

Chikungunya Virus Infection and Acute Elevation of Serum Prostate-Specific Antigen

A man with prostate cancer on a regime of active surveillance had a laboratory-confirmed acute Chikungunya virus infection. The patient experienced a sudden increase in serum Prostate-Specific Antigen (PSA) during the acute illness that caused him anxiety and confounded interpretation of the PSA test. Six weeks after the onset of Chikungunya Fever symptoms, the elevated serum PSA returned to baseline. The association of Chikungunya Fever and elevated serum PSA may result in misinterpretation of the PSA test, triggering unnecessary prostate biopsy or other management errors

Fluorescent pegylated nanoparticles demonstrate fluid-phase pinocytosis by macrophages in mouse atherosclerotic lesions

The uptake of lipoproteins by macrophages is a critical step in the development of atherosclerotic lesions. Cultured monocyte-derived macrophages take up large amounts of native LDL by receptor-independent fluid-phase pinocytosis, either constitutively or in response to specific activating stimuli, depending on the macrophage phenotype. We therefore sought to determine whether fluid-phase pinocytosis occurs in vivo in macrophages in atherosclerotic lesions. We demonstrated that fluorescent pegylated nanoparticles similar in size to LDL (specifically nontargeted Qtracker quantum dot and AngioSPARK nanoparticles) can serve as models of LDL uptake by fluid-phase pinocytosis in cultured human monocyte-derived macrophages and mouse bone marrow-derived macrophages. Using fluorescence microscopy, we showed that atherosclerosis-prone Apoe-knockout mice injected with these nanoparticles displayed massive accumulation of the nanoparticles within CD68 + macrophages, including lipid-containing foam cells, in atherosclerotic lesions in the aortic arch. Similar results were obtained when atherosclerotic mouse aortas were cultured with nanoparticles in vitro. These results show that macrophages within atherosclerotic lesions can take up LDL-sized nanoparticles by fluid-phase pinocytosis and indicate that fluid-phase pinocytosis of LDL is a mechanism for macrophage foam cell formation in vivo

Activation by Inflammatory Stimuli Increases Neutrophil Binding of Human Immunodeficiency Virus Type 1 and Subsequent Infection of Lymphocytes

Resting neutrophils bind human immunodeficiency virus type 1 (HIV-1) and efficiently transfer infection to lymphocytes. The present study shows that a brief activation by inflammatory stimuli increases the neutrophil binding levels of both R5 and X4 isolates of HIV-1 at least twofold. The binding occurs independently of CD4, gp120, and incubation temperature and is observed with HIV-1 propagated either in lymphocytes or in HEK293 cells. Significantly, HIV-1 bound to the activated neutrophils accelerates the infection of activated lymphocytes compared to free HIV-1 or to HIV-1 bound to resting neutrophils. It is proposed that these events may contribute to the increased risk of HIV-1 transmission at sites of mucosal infection

Murine bone marrow-derived macrophages differentiated with GM-CSF become foam cells by PI3K?-dependent fluid-phase pinocytosis of native LDL

Accumulation of cholesterol by macrophage uptake of LDL is a key event in the formation of atherosclerotic plaques. Previous research has shown that granulocyte-macrophage colony-stimulating factor (GM-CSF) is present in atherosclerotic plaques and promotes aortic lipid accumulation. However, it has not been determined whether murine GM-CSF-differentiated macrophages take up LDL to become foam cells. GM-CSF-differentiated macrophages from LDL receptor-null mice were incubated with LDL, resulting in massive macrophage cholesterol accumulation. Incubation of LDL receptor-null or wild-type macrophages with increasing concentrations of ¹²?I-LDL showed nonsaturable macrophage LDL uptake that was linearly related to the amount of LDL added, indicating that LDL uptake was mediated by fluid-phase pinocytosis. Previous studies suggest that phosphoinositide 3-kinases (PI3K) mediate macrophage fluid-phase pinocytosis, although the isoform mediating this process has not been determined. Because PI3K? is known to promote aortic lipid accumulation, we investigated its role in mediating macrophage fluid-phase pinocytosis of LDL. Wild-type macrophages incubated with LDL and the PI3K? inhibitor AS605240 or PI3K?-null macrophages incubated with LDL showed an ?50% reduction in LDL uptake and cholesterol accumulation compared with wild-type macrophages incubated with LDL only. These results show that GM-CSF-differentiated murine macrophages become foam cells by fluid-phase pinocytosis of LDL and identify PI3K? as contributing to this process