Maraviroc (UK-427,857), a Potent, Orally Bioavailable, and Selective Small-Molecule Inhibitor of Chemokine Receptor CCR5 with Broad-Spectrum Anti-Human Immunodeficiency Virus Type 1 Activity

PubMed Central

Dorr, Patrick; Westby, Mike; Dobbs, Susan; Griffin, Paul; Irvine, Becky; Macartney, Malcolm; Mori, Julie; Rickett, Graham; Smith-Burchnell, Caroline; Napier, Carolyn; Webster, Rob; Armour, Duncan; Price, David; Stammen, Blanda; Wood, Anthony; Perros, Manos

2005-01-01

Maraviroc (UK-427,857) is a selective CCR5 antagonist with potent anti-human immunodeficiency virus type 1 (HIV-1) activity and favorable pharmacological properties. Maraviroc is the product of a medicinal chemistry effort initiated following identification of an imidazopyridine CCR5 ligand from a high-throughput screen of the Pfizer compound file. Maraviroc demonstrated potent antiviral activity against all CCR5-tropic HIV-1 viruses tested, including 43 primary isolates from various clades and diverse geographic origin (geometric mean 90% inhibitory concentration of 2.0 nM). Maraviroc was active against 200 clinically derived HIV-1 envelope-recombinant pseudoviruses, 100 of which were derived from viruses resistant to existing drug classes. There was little difference in the sensitivity of the 200 viruses to maraviroc, as illustrated by the biological cutoff in this assay (= geometric mean plus two standard deviations [SD] of 1.7-fold). The mechanism of action of maraviroc was established using cell-based assays, where it blocked binding of viral envelope, gp120, to CCR5 to prevent the membrane fusion events necessary for viral entry. Maraviroc did not affect CCR5 cell surface levels or associated intracellular signaling, confirming it as a functional antagonist of CCR5. Maraviroc has no detectable in vitro cytotoxicity and is highly selective for CCR5, as confirmed against a wide range of receptors and enzymes, including the hERG ion channel (50% inhibitory concentration, >10 μM), indicating potential for an excellent clinical safety profile. Studies in preclinical in vitro and in vivo models predicted maraviroc to have human pharmacokinetics consistent with once- or twice-daily dosing following oral administration. Clinical trials are ongoing to further investigate the potential of using maraviroc for the treatment of HIV-1 infection and AIDS. PMID:16251317

Functional impact of HIV coreceptor-binding site mutations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Biscone, Mark J.; Miamidian, John L.; Muchiri, John M.

2006-07-20

The bridging sheet region of the gp120 subunit of the HIV-1 Env protein interacts with the major virus coreceptors, CCR5 and CXCR4. We examined the impact of mutations in and adjacent to the bridging sheet region of an X4 tropic HIV-1 on membrane fusion and entry inhibitor susceptibility. When the V3-loop of this Env was changed so that CCR5 was used, the effects of these same mutations on CCR5 use were assayed as well. We found that coreceptor-binding site mutations had greater effects on CXCR4-mediated fusion and infection than when CCR5 was used as a coreceptor, perhaps related to differencesmore » in coreceptor affinity. The mutations also reduced use of the alternative coreceptors CCR3 and CCR8 to varying degrees, indicating that the bridging sheet region is important for the efficient utilization of both major and minor HIV coreceptors. As seen before with a primary R5 virus strain, bridging sheet mutations increased susceptibility to the CCR5 inhibitor TAK-779, which correlated with CCR5 binding efficiency. Bridging sheet mutations also conferred increased susceptibility to the CXCR4 ligand AMD-3100 in the context of the X4 tropic Env. However, these mutations had little effect on the rate of membrane fusion and little effect on susceptibility to enfuvirtide, a membrane fusion inhibitor whose activity is dependent in part on the rate of Env-mediated membrane fusion. Thus, mutations that reduce coreceptor binding and enhance susceptibility to coreceptor inhibitors can affect fusion and enfuvirtide susceptibility in an Env context-dependent manner.« less

Preclinical discovery and development of maraviroc for the treatment of HIV.

PubMed

Veljkovic, Nevena; Vucicevic, Jelica; Tassini, Sabrina; Glisic, Sanja; Veljkovic, Veljko; Radi, Marco

2015-06-01

Maraviroc is a first-in-class antiretroviral (ARV) drug acting on a host cell target (CCR5), which blocks the entry of the HIV virus into the cell. Maraviroc is currently indicated for combination ARV treatment in adults infected only with CCR5-tropic HIV-1. This drug discovery case history focuses on the key studies that led to the discovery and approval of maraviroc, as well as on post-launch clinical reports. The article is based on the data reported in published preclinical and clinical studies, conference posters and on drug package data. The profound understanding of HIV's entry mechanisms has provided a strong biological rationale for targeting the chemokine receptor CCR5. The CCR5-antagonist mariviroc, with its unique mode of action and excellent safety profile, is an important therapeutic option for HIV patients. In general, the authors believe that targeting host factors is a useful approach for combating new and re-emerging transmissible diseases, as well as pathogens that easily become resistant to common antiviral drugs. Maraviroc, offering a potent and safe cellular receptor-mediated pharmacological response to HIV, has paved the way for the development of a new generation of host-targeting antivirals.

Activity of the HIV-1 Attachment Inhibitor BMS-626529, the Active Component of the Prodrug BMS-663068, against CD4-Independent Viruses and HIV-1 Envelopes Resistant to Other Entry Inhibitors

PubMed Central

Li, Zhufang; Zhou, Nannan; Sun, Yongnian; Ray, Neelanjana; Lataillade, Max; Hanna, George J.

2013-01-01

BMS-626529 is a novel small-molecule HIV-1 attachment inhibitor active against both CCR5- and CXCR4-tropic viruses. BMS-626529 functions by preventing gp120 from binding to CD4. A prodrug of this compound, BMS-663068, is currently in clinical development. As a theoretical resistance pathway to BMS-663068 could be the development of a CD4-independent phenotype, we examined the activity of BMS-626529 against CD4-independent viruses and investigated whether resistance to BMS-626529 could be associated with a CD4-independent phenotype. Finally, we evaluated whether cross-resistance exists between BMS-626529 and other HIV-1 entry inhibitors. Two laboratory-derived envelopes with a CD4-independent phenotype (one CXCR4 tropic and one CCR5 tropic), five envelopes from clinical isolates with preexisting BMS-626529 resistance, and several site-specific mutant BMS-626529-resistant envelopes were examined for their dependence on CD4 for infectivity or susceptibility to BMS-626529. Viruses resistant to other entry inhibitors (enfuvirtide, maraviroc, and ibalizumab) were also examined for susceptibility to BMS-626529. Both CD4-independent laboratory isolates retained sensitivity to BMS-626529 in CD4− cells, while HIV-1 envelopes from viruses resistant to BMS-626529 exhibited no evidence of a CD4-independent phenotype. BMS-626529 also exhibited inhibitory activity against ibalizumab- and enfuvirtide-resistant envelopes. While there appeared to be some association between maraviroc resistance and reduced susceptibility to BMS-626529, an absolute correlation cannot be presumed, since some CCR5-tropic maraviroc-resistant envelopes remained sensitive to BMS-626529. Clinical use of the prodrug BMS-663068 is unlikely to promote resistance via generation of CD4-independent virus. No cross-resistance between BMS-626529 and other HIV entry inhibitors was observed, which could allow for sequential or concurrent use with different classes of entry inhibitors. PMID:23774428

Response to vicriviroc in treatment-experienced subjects, as determined by an enhanced-sensitivity coreceptor tropism assay: reanalysis of AIDS clinical trials group A5211.

PubMed

Su, Zhaohui; Gulick, Roy M; Krambrink, Amy; Coakley, Eoin; Hughes, Michael D; Han, Dong; Flexner, Charles; Wilkin, Timothy J; Skolnik, Paul R; Greaves, Wayne L; Kuritzkes, Daniel R; Reeves, Jacqueline D

2009-12-01

The enhanced-sensitivity Trofile assay (Monogram Biosciences) was used to retest coreceptor use at both study screening and study entry for 118 treatment-experienced subjects in AIDS Clinical Trials Group A5211 who had CCR5-tropic (R5) virus detected by the original Trofile assay at study screening. Among 90 recipients of vicriviroc, a significantly (P< .001) greater mean reduction in HIV-1 RNA was observed in 72 subjects with R5 virus versus 15 subjects reclassified as having dual/mixed-tropic viruses at screening: -1.11 versus -0.09 log(10) copies/mL at day 14 and -1.91 versus -0.57 log(10) copies/mL at week 24, respectively. Results suggest that the enhanced-sensitivity assay is a better screening tool for determining patient eligibility for CCR5 antagonist therapy.

Platelet factor 4 (CXCL4) facilitates human macrophage infection with HIV-1 and potentiates virus replication.

PubMed

Schwartzkopff, Franziska; Grimm, Tobias A; Lankford, Carla S R; Fields, Karen; Wang, Jiun; Brandt, Ernst; Clouse, Kathleen A

2009-12-01

Platelet factor 4 (CXCL4), a member of the CXC chemokine subfamily released in high amounts by activated platelets, has been identified as a monocyte survival factor that induces monocyte differentiation into macrophages. Although CXCL4 has been shown to have biological effects unique to chemokines, nothing is known about the role of CXCL4-derived human macrophages or CXCL4 in human immunodeficiency virus (HIV) disease. In this study, CXCL4-derived macrophages are compared with macrophage-colony stimulating factor (M-CSF)-derived macrophages for their ability to support HIV-1 replication. We show that CXCL4-derived macrophages can be infected with macrophage-tropic HIV-1 that uses either CC-chemokine receptor 5 (CCR5) or CXC-chemokine receptor 4 (CXCR4) as a co-receptor for viral entry. We also find that M-CSF and the chemokines, monocyte chemoattractant protein 1 (MCP-1; CCL2) and macrophage-inflammatory-protein-1-alpha (MIP-1alpha; CCL3) are produced upon R5- and X4-tropic HIV-1 replication in both M-CSF- and CXCL4-derived human macrophages. In addition, CXCL4 added to M-CSF-derived macrophages after virus adsorption and maintained throughout the infection enhances HIV-1 replication. We thus propose a novel role for CXCL4 in HIV disease.

[Maraviroc: clinical trials results].

PubMed

Chidiac, C; Katlama, C; Yeni, P

2008-03-01

Just over a decade after identification of chemokine receptors CCR5 and CXCR4 as coreceptors for HIV, maraviroc (Celsentri), the first CCR5 antagonist, has recently obtained its Marketing Authorization in the United States and Europe, for treatment of treatment-experienced adult patients infected with only CCR5-tropic HIV-1 detectable. CCR5 antagonists, after fusion inhibitor enfuvirtide available since 2003, also belong to entry inhibitors. These molecules, unlike previous antiretrovirals, do not target the virus but its target cell by blocking viral penetration. Maraviroc has shown its clinical efficacy in patients failing other antiretroviral classes. Its safety profile was similar to placebo in two large phase III trials. However, careful assessment of both hepatic and immunologic safety of this new therapeutic class is needed. Viral tropism testing has to be investigated before using maraviroc in the clinic, because CCR5 antagonists are not active against CXCR4 viruses. For the moment indicated for the treatment-experienced patient population, maraviroc could in the future benefit to other types of patients, depending on ongoing trials results.

Maraviroc: a coreceptor CCR5 antagonist for management of HIV infection.

PubMed

Yost, Raymond; Pasquale, Timothy R; Sahloff, Eric G

2009-04-15

The mechanism of action, pharmacology, pharmacokinetics, clinical efficacy, drug interactions, adverse effects, dosage and administration, cost, and role in therapy of maraviroc are reviewed. Maraviroc is the first CCR5 coreceptor antagonist to receive marketing approval from the Food and Drug Administration (FDA) for the treatment of CCR5-tropic human immunodeficiency virus (HIV) infection as part of an optimized antiretroviral regimen in treatment-experienced patients. As 50% or more of treatment-experienced patients may be infected with CXCR4-tropic virus, a tropism assay should be performed before initiating maraviroc therapy. The majority of evidence supporting maraviroc's use comes from two studies of HIV-infected, treatment-experienced patients. Pooled analysis from these two studies revealed that twice-daily maraviroc decreased HIV-1 RNA by 1.84 log copies/mL, compared with 0.78 log copy/mL with placebo. Forty-six percent of subjects attained an HIV-1 RNA concentration of <50 copies/mL, compared with only 17% with placebo. In a trial of treatment-naive HIV-infected individuals, maraviroc failed to show noninferiority to efavirenz. Maraviroc is metabolized by cytochrome P-450 isoenzyme 3A4 and is subject to interactions with inhibitors and inducers of that isoenzyme, such as the protease inhibitors (except tipranavir), efavirenz, and rifampin. Resistance has been reported with maraviroc, but specific mechanisms are still poorly understood. The most common adverse effects reported with maraviroc were diarrhea, nausea, fatigue, and headache. Available data support the use of maraviroc, the first CCR5 antagonist to receive FDA marketing approval, as part of an optimized antiretroviral regimen in treatment-experienced patients infected with CCR5-tropic HIV.

HIV Type 1 from a Patient with Baseline Resistance to CCR5 Antagonists Uses Drug-Bound Receptor for Entry

PubMed Central

Tilton, John C.; Amrine-Madsen, Heather; Miamidian, John L.; Kitrinos, Kathryn M.; Pfaff, Jennifer; Demarest, James F.; Ray, Neelanjana; Jeffrey, Jerry L.; Labranche, Celia C.

2010-01-01

Abstract CCR5 antagonists are a new class of antiretroviral drugs that block viral entry by disrupting interactions between the viral envelope (Env) glycoprotein and coreceptor. During the CCR100136 (EPIC) Phase IIb study of the CCR5 antagonist aplaviroc (APL) in treatment-naive individuals, a patient was identified who harbored virus strains that exhibited partial resistance to APL at the time of virologic failure. Retrospectively, it was found that APL resistance was present at baseline as well. To investigate the mechanism of APL resistance in this patient, we cloned HIV-1 env genes from plasma obtained at baseline and after virologic failure. Approximately 85% of cloned Envs were functional, and all exhibited partial resistance to APL. All Envs were R5-tropic, were partially resistant to other CCR5 antagonists including maraviroc on cells with high CCR5 expression, but remained sensitive to the fusion inhibitor enfuvirtide. Competition studies with natural CCR5 ligands revealed that the mechanism of drug resistance entailed the use of the drug-bound conformation of CCR5 by the Env proteins obtained from this individual. The degree of drug resistance varied between Env clones, and also varied depending on the cell line used or the donor from whom the primary T cells were obtained. Thus, both virus and host factors contribute to CCR5 antagonist resistance. This study shows that R5 HIV-1 strains resistant to CCR5 inhibitors can arise in patients, confirming a mechanism of resistance previously characterized in vitro. In addition, some patients can harbor CCR5 antagonist-resistant viruses prior to treatment, which may have implications for the clinical use of this new class of antiretrovirals. PMID:20055594

In vivo evolution of HIV-1 co-receptor usage and sensitivity to chemokine-mediated suppression.

PubMed

Scarlatti, G; Tresoldi, E; Björndal, A; Fredriksson, R; Colognesi, C; Deng, H K; Malnati, M S; Plebani, A; Siccardi, A G; Littman, D R; Fenyö, E M; Lusso, P

1997-11-01

Following the identification of the C-C chemokines RANTES, MIP-1alpha and MIP-1beta as major human immunodeficiency virus (HIV)-suppressive factors produced by CD8+ T cells, several chemokine receptors were found to serve as membrane co-receptors for primate immunodeficiency lentiretroviruses. The two most widely used co-receptors thus far recognized, CCR5 and CXCR4, are expressed by both activated T lymphocytes and mononuclear phagocytes. CCR5, a specific RANTES, MIP-1alpha and MIP-1 receptor, is used preferentially by non-MT2-tropic HIV-1 and HIV-2 strains and by simian immunodeficiency virus (SIV), whereas CXCR4, a receptor for the C-X-C chemokine SDF-1, is used by MT2-tropic HIV-1 and HIV-2, but not by SIV. Other receptors with a more restricted cellular distribution, such as CCR2b, CCR3 and STRL33, can also function as co-receptors for selected viral isolates. The third variable region (V3) of the gp120 envelope glycoprotein of HIV-1 has been fingered as a critical determinant of the co-receptor choice. Here, we document a consistent pattern of evolution of viral co-receptor usage and sensitivity to chemokine-mediated suppression in a longitudinal follow-up of children with progressive HIV-1 infection. Viral isolates obtained during the asymptomatic stages generally used only CCR5 as a co-receptor and were inhibited by RANTES, MIP-1alpha and MIP-1beta, but not by SDF-1. By contrast, the majority of the isolates derived after the progression of the disease were resistant to C-C chemokines, having acquired the ability to use CXCR4 and, in some cases, CCR3, while gradually losing CCR5 usage. Surprisingly, most of these isolates were also insensitive to SDF-1, even when used in combination with RANTES. An early acquisition of CXCR4 usage predicted a poor prognosis. In children who progressed to AIDS without a shift to CXCR4 usage, all the sequential isolates were CCR5-dependent but showed a reduced sensitivity to C-C chemokines. Discrete changes in the V3 domain of gp120 were associated with the loss of sensitivity to C-C chemokines and the shift in co-receptor usage. These results suggest an adaptive evolution of HIV-1 in vivo, leading to escape from the control of the antiviral C-C chemokines.

Exclusive and persistent use of the entry coreceptor CXCR4 by human immunodeficiency virus type 1 from a subject homozygous for CCR5 delta32.

PubMed

Michael, N L; Nelson, J A; KewalRamani, V N; Chang, G; O'Brien, S J; Mascola, J R; Volsky, B; Louder, M; White, G C; Littman, D R; Swanstrom, R; O'Brien, T R

1998-07-01

Individuals who are homozygous for the 32-bp deletion in the gene coding for the chemokine receptor and major human immunodeficiency virus type 1 (HIV-1) coreceptor CCR5 (CCR5 -/-) lack functional cell surface CCR5 molecules and are relatively resistant to HIV-1 infection. HIV-1 infection in CCR5 -/- individuals, although rare, has been increasingly documented. We now report that the viral quasispecies from one such individual throughout disease is homogenous, T cell line tropic, and phenotypically syncytium inducing (SI); exclusively uses CXCR4; and replicates well in CCR5 -/- primary T cells. The recently discovered coreceptors BOB and Bonzo are not used. Although early and persistent SI variants have been described in longitudinal studies, this is the first demonstration of exclusive and persistent CXCR4 usage. With the caveat that the earliest viruses available from this subject were from approximately 4 years following primary infection, these data suggest that HIV-1 infection can be mediated and persistently maintained by viruses which exclusively utilize CXCR4. The lack of evolution toward the available minor coreceptors in this subject underscores the dominant biological roles of the major coreceptors CCR5 and CXCR4. This and two similar subjects (R. Biti, R. Ffrench, J. Young, B. Bennetts, G. Stewart, and T. Liang, Nat. Med. 3:252-253, 1997; I. Theodoreu, L. Meyer, M. Magierowska, C. Katlama, and C. Rouzioux, Lancet 349:1219-1220, 1997) showed relatively rapid CD4+ T-cell declines despite average or low initial viral RNA load. Since viruses which use CXCR4 exclusively cannot infect macrophages, these data have implications for the relative infection of the T-cell compartment versus the macrophage compartment in vivo and for the development of CCR5-based therapeutics.

Maraviroc (Celsentri) for multidrug-resistant human immunodeficiency virus (HIV)-1.

PubMed

Ndegwa, S

2007-12-01

(1) Maraviroc belongs to a new class of antiretroviral drugs designed to block entry of HIV-1 into CD4+ T-cells via the CCR5 coreceptor. It is indicated for combination therapy in treatment-experienced adults infected with CCR5-tropic HIV-1 that is resistant to multiple antiretroviral agents. (2) Results from two randomized controlled trials (RCTs) indicate that in treatment experienced patients, maraviroc, combined with optimized background therapy (OBT), significantly decreases the level of HIV-1 RNA in the blood (viral load) when compared with OBT alone. The number of patients achieving undetectable viral loads and CD4+ cell count increases were also significantly higher in those receiving maraviroc. (3) Most patients experiencing treatment failure with maraviroc exhibit tropism changes from CCR5-tropic to CXCR4-using virus, but there is no evidence of disease progression. (4) Adverse effects reported with maraviroc include cough, fever, upper respiratory tract infections, rash, muscle and joint pain, abdominal pain, and postural hypotension (dizziness). No significant increases in cardiovascular events, hepatotoxicity, infections or malignancies have been reported with short-term maraviroc therapy. Several post-marketing studies will assess maraviroc's long-term safety for immune function, liver function, malignancy, cardiac events, and risks associated with changes in tropism. (5) Results from an ongoing trial in treatment naive patients suggest that maraviroc may not be superior in terms of viral suppression to standard therapy, but may significantly increase the number of CD4+ T-cells.

Prognostic value of a CCR5 defective allele in pediatric HIV-1 infection.

PubMed Central

Romiti, M. L.; Colognesi, C.; Cancrini, C.; Mas, A.; Berrino, M.; Salvatori, F.; Orlandi, P.; Jansson, M.; Palomba, E.; Plebani, A.; Bertran, J. M.; Hernandez, M.; de Martino, M.; Amoroso, A.; Tovo, P. A.; Rossi, P.; Espanol, T.; Scarlatti, G.

2000-01-01

BACKGROUND: A deletion of 32 base pairs in the CCR5 gene (delta32 CCR5) has been linked to resistance to HIV-1 infection in exposed adults and to the delay of disease progression in infected adults. MATERIALS AND METHODS: To determine the role of delta32 CCR5 in disease progression of HIV-1 infected children born to seropositive mothers, we studied a polymerase chain reaction in 301 HIV-1 infected, 262 HIV-1 exposed-uninfected and 47 HIV-1 unexposed-uninfected children of Spanish and Italian origin. Infected children were further divided into two groups according to their rate of HIV-1 disease progression: rapid progressors who developed severe clinical and/or immunological conditions within the second year of life, and delayed progressors with any other evolution of disease. Among the latter were the long-term, non-progressors (LTNP) who presented with mild or no symptoms of HIV-1 infection above 8 years of age. Viral phenotype was studied for 45 delayed progressors. RESULTS: No correlation was found between delta32 CCR5 and mother-to-child transmission of HIV-1. However, the frequency of the deletion was substantially higher in LTNP, compared with delayed (p = 0.019) and rapid progressors (p = 0.0003). In children carrying the delta32 CCRS mutation, the presence of MT-2 tropic virus isolate was associated with a severe immune suppression (p = 0.028); whereas, the presence of MT-2 negative viruses correlated with LTNP (p = 0.010). CONCLUSIONS: Given the rapidity and simplicity of the assay, the delta32 CCR5 mutation may be a useful predictive marker to identify children with delayed disease progression who, consequently, may not require immediate antiretroviral treatment. PMID:10803406

Prognostic value of a CCR5 defective allele in pediatric HIV-1 infection.

PubMed

Romiti, M L; Colognesi, C; Cancrini, C; Mas, A; Berrino, M; Salvatori, F; Orlandi, P; Jansson, M; Palomba, E; Plebani, A; Bertran, J M; Hernandez, M; de Martino, M; Amoroso, A; Tovo, P A; Rossi, P; Espanol, T; Scarlatti, G

2000-01-01

A deletion of 32 base pairs in the CCR5 gene (delta32 CCR5) has been linked to resistance to HIV-1 infection in exposed adults and to the delay of disease progression in infected adults. To determine the role of delta32 CCR5 in disease progression of HIV-1 infected children born to seropositive mothers, we studied a polymerase chain reaction in 301 HIV-1 infected, 262 HIV-1 exposed-uninfected and 47 HIV-1 unexposed-uninfected children of Spanish and Italian origin. Infected children were further divided into two groups according to their rate of HIV-1 disease progression: rapid progressors who developed severe clinical and/or immunological conditions within the second year of life, and delayed progressors with any other evolution of disease. Among the latter were the long-term, non-progressors (LTNP) who presented with mild or no symptoms of HIV-1 infection above 8 years of age. Viral phenotype was studied for 45 delayed progressors. No correlation was found between delta32 CCR5 and mother-to-child transmission of HIV-1. However, the frequency of the deletion was substantially higher in LTNP, compared with delayed (p = 0.019) and rapid progressors (p = 0.0003). In children carrying the delta32 CCRS mutation, the presence of MT-2 tropic virus isolate was associated with a severe immune suppression (p = 0.028); whereas, the presence of MT-2 negative viruses correlated with LTNP (p = 0.010). Given the rapidity and simplicity of the assay, the delta32 CCR5 mutation may be a useful predictive marker to identify children with delayed disease progression who, consequently, may not require immediate antiretroviral treatment.

Reanalysis of Coreceptor Tropism in HIV-1–Infected Adults Using a Phenotypic Assay with Enhanced Sensitivity

PubMed Central

Goetz, Mathew Bidwell; Leduc, Robert; Skowron, Gail; Su, Zhaohui; Chan, Ellen S.; Heera, Jayyant; Chapman, Doug; Spritzler, John; Reeves, Jacqueline D.; Gulick, Roy M.; Coakley, Eoin

2011-01-01

The enhanced-sensitivity Trofile assay (TF-ES; Monogram Biosciences) was used to retest coreceptor tropism samples from 4 different cohorts of HIV-1–infected patients. Nine percent to 26% of patients with CCR5-tropic virus by the original Trofile assay had CXCR4-using virus by TF-ES. Lower CD4 cell counts were associated with CXCR4-using virus in all cohorts. PMID:21427401

High-Sequence Diversity and Rapid Virus Turnover Contribute to Higher Rates of Coreceptor Switching in Treatment-Experienced Subjects with HIV-1 Viremia.

PubMed

Nedellec, Rebecca; Herbeck, Joshua T; Hunt, Peter W; Deeks, Steven G; Mullins, James I; Anton, Elizabeth D; Reeves, Jacqueline D; Mosier, Donald E

2017-03-01

Coreceptor switching from CCR5 to CXCR4 is common during chronic HIV-1 infection, but is even more common in individuals who have failed antiretroviral therapy (ART). Prior studies have suggested rapid mutation and/or recombination of HIV-1 envelope (env) genes during coreceptor switching. We compared the functional and genotypic changes in env of viruses from viremic subjects who had failed ART just before and after coreceptor switching and compared those to viruses from matched subjects without coreceptor switching. Analysis of multiple unique functional env clones from each subject revealed extensive diversity at both sample time points and rapid diversification of sequences during the 4-month interval in viruses from both 9 subjects with coreceptor switching and 15 control subjects. Only two subjects had envs with evidence of recombination. Three findings distinguished env clones from subjects with coreceptor switching from controls: (1) lower entry efficiency via CCR5; (2) longer V1/V2 regions; and (3), lower nadir CD4 T cell counts during prior years of infection. Most of these subjects harbored virus with lower replicative capacity associated with protease (PR) and/or reverse transcriptase inhibitor resistance mutations, and the extensive diversification tended to lead either to improved entry efficiency via CCR5 or the gain of entry function via CXCR4. These results suggest that R5X4 or X4 variants emerge from a diverse, low-fitness landscape shaped by chronic infection, multiple ART resistance mutations, the availability of target cells, and reduced entry efficiency via CCR5.

Lymphotropic Virions Affect Chemokine Receptor-Mediated Neural Signaling and Apoptosis: Implications for Human Immunodeficiency Virus Type 1-Associated Dementia

PubMed Central

Zheng, Jialin; Ghorpade, Anuja; Niemann, Douglas; Cotter, Robin L.; Thylin, Michael R.; Epstein, Leon; Swartz, Jennifer M.; Shepard, Robin B.; Liu, Xiaojuan; Nukuna, Adeline; Gendelman, Howard E.

1999-01-01

Chemokine receptors pivotal for human immunodeficiency virus type 1 (HIV-1) infection in lymphocytes and macrophages (CCR3, CCR5, and CXCR4) are expressed on neural cells (microglia, astrocytes, and/or neurons). It is these cells which are damaged during progressive HIV-1 infection of the central nervous system. We theorize that viral coreceptors could effect neural cell damage during HIV-1-associated dementia (HAD) without simultaneously affecting viral replication. To these ends, we studied the ability of diverse viral strains to affect intracellular signaling and apoptosis of neurons, astrocytes, and monocyte-derived macrophages. Inhibition of cyclic AMP, activation of inositol 1,4,5-trisphosphate, and apoptosis were induced by diverse HIV-1 strains, principally in neurons. Virions from T-cell-tropic (T-tropic) strains (MN, IIIB, and Lai) produced the most significant alterations in signaling of neurons and astrocytes. The HIV-1 envelope glycoprotein, gp120, induced markedly less neural damage than purified virions. Macrophage-tropic (M-tropic) strains (ADA, JR-FL, Bal, MS-CSF, and DJV) produced the least neural damage, while 89.6, a dual-tropic HIV-1 strain, elicited intermediate neural cell damage. All T-tropic strain-mediated neuronal impairments were blocked by the CXCR4 antibody, 12G5. In contrast, the M-tropic strains were only partially blocked by 12G5. CXCR4-mediated neuronal apoptosis was confirmed in pure populations of rat cerebellar granule neurons and was blocked by HA1004, an inhibitor of calcium/calmodulin-dependent protein kinase II, protein kinase A, and protein kinase C. Taken together, these results suggest that progeny HIV-1 virions can influence neuronal signal transduction and apoptosis. This process occurs, in part, through CXCR4 and is independent of CD4 binding. T-tropic viruses that traffic in and out of the brain during progressive HIV-1 disease may play an important role in HAD neuropathogenesis. PMID:10482576

CD4 T cell decline following HIV seroconversion in individuals with and without CXCR4-tropic virus.

PubMed

Ghosn, Jade; Bayan, Tatiana; Meixenberger, Karolin; Tran, Laurent; Frange, Pierre; d'Arminio Monforte, Antonella; Zangerle, Robert; de Mendoza, Carmen; Krastinova, Evguenia; Porter, Kholoud; Meyer, Laurence; Chaix, Marie-Laure

2017-10-01

The natural clinical and immunological courses following HIV seroconversion with CXCR4-tropic or dual-mixed (X4/DM) viruses are controversial. We compared spontaneous immunological outcome in patients harbouring an X4/DM virus at the time of seroconversion with those harbouring a CCR5-tropic (R5) virus. Data were included from patients participating in CASCADE, a large cohort collaboration of HIV seroconverters, with ≥2 years of follow-up since seroconversion. The HIV envelope gene was sequenced from frozen plasma samples collected at enrolment, and HIV tropism was determined using Geno2Pheno (false-positive rate 10%). The spontaneous CD4 T cell evolution was compared by modelling CD4 kinetics using linear mixed-effects models with random intercept and random slope. A total of 1387 patients were eligible. Median time between seroconversion and enrolment was 1 month (range 0-3). At enrolment, 202 of 1387 (15%) harboured an X4/DM-tropic virus. CD4 decrease slopes were not significantly different according to HIV-1 tropism during the first 30 months after seroconversion. No marked change in these results was found after adjusting for age, year of seroconversion and baseline HIV viral load. Time to antiretroviral treatment initiation was not statistically different between patients harbouring an R5 (20.76 months) and those harbouring an X4/DM-tropic virus (22.86 months, logrank test P = 0.32). Conclusions: In this large cohort collaboration, 15% of the patients harboured an X4/DM virus close to HIV seroconversion. Patients harbouring X4/DM-tropic viruses close to seroconversion did not have an increased risk of disease progression, estimated by the decline in CD4 T cell count or time to combined ART initiation. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

High-Sequence Diversity and Rapid Virus Turnover Contribute to Higher Rates of Coreceptor Switching in Treatment-Experienced Subjects with HIV-1 Viremia

PubMed Central

Nedellec, Rebecca; Herbeck, Joshua T.; Hunt, Peter W.; Deeks, Steven G.; Mullins, James I.; Anton, Elizabeth D.; Reeves, Jacqueline D.

2017-01-01

Abstract Coreceptor switching from CCR5 to CXCR4 is common during chronic HIV-1 infection, but is even more common in individuals who have failed antiretroviral therapy (ART). Prior studies have suggested rapid mutation and/or recombination of HIV-1 envelope (env) genes during coreceptor switching. We compared the functional and genotypic changes in env of viruses from viremic subjects who had failed ART just before and after coreceptor switching and compared those to viruses from matched subjects without coreceptor switching. Analysis of multiple unique functional env clones from each subject revealed extensive diversity at both sample time points and rapid diversification of sequences during the 4-month interval in viruses from both 9 subjects with coreceptor switching and 15 control subjects. Only two subjects had envs with evidence of recombination. Three findings distinguished env clones from subjects with coreceptor switching from controls: (1) lower entry efficiency via CCR5; (2) longer V1/V2 regions; and (3), lower nadir CD4 T cell counts during prior years of infection. Most of these subjects harbored virus with lower replicative capacity associated with protease (PR) and/or reverse transcriptase inhibitor resistance mutations, and the extensive diversification tended to lead either to improved entry efficiency via CCR5 or the gain of entry function via CXCR4. These results suggest that R5X4 or X4 variants emerge from a diverse, low-fitness landscape shaped by chronic infection, multiple ART resistance mutations, the availability of target cells, and reduced entry efficiency via CCR5. PMID:27604829

Characterization of emergent HIV resistance in treatment-naive subjects enrolled in a vicriviroc phase 2 trial.

PubMed

McNicholas, Paul; Wei, Yi; Whitcomb, Jeannette; Greaves, Wayne; Black, Todd A; Tremblay, Cecile L; Strizki, Julie M

2010-05-15

Vicriviroc is a C-C motif chemokine receptor 5 (CCR5) antagonist that is in clinical development for the treatment of human immunodeficiency virus type 1 (HIV-1) infection. This study explored the molecular basis for the development of phenotypically resistant virus. HIV-1 RNA from treatment-naive subjects who experienced virological failure in a phase 2 dose-finding trial was evaluated for coreceptor usage and susceptibility. For viruses that exhibited reduced susceptibility to vicriviroc, envelope clones were phenotypically and genotypically characterized. Twenty-six vicriviroc-treated subjects experienced virological failure; for 24 the virus remained CCR5-tropic, and 2 had dual/X4 virus. Reduced susceptibility to vicriviroc, manifested as decreases in the maximum percent inhibition value (no increase in median inhibitory concentration), was detected in 4 of the 26 subjects who experienced virological failure. Clonal analysis of envelopes in samples from these 4 subjects revealed multiple sequence changes in gp160, principally within the variable domain 1/variable domain 2, variable domain 3, and variable domain 4 loops. However, no consistent pattern of mutations was observed across subjects. In this study, only a small proportion of treatment failures were associated with tropism changes or reduced susceptibility to vicriviroc. Genotypic analysis of cloned env sequences revealed no specific mutational pattern associated with reduced susceptibility to vicriviroc, although numerous changes were observed in the variable domain 3 loop and in other regions of gp160.

Reanalysis of coreceptor tropism in HIV-1-infected adults using a phenotypic assay with enhanced sensitivity.

PubMed

Wilkin, Timothy J; Goetz, Mathew Bidwell; Leduc, Robert; Skowron, Gail; Su, Zhaohui; Chan, Ellen S; Heera, Jayyant; Chapman, Doug; Spritzler, John; Reeves, Jacqueline D; Gulick, Roy M; Coakley, Eoin

2011-04-01

The enhanced-sensitivity Trofile assay (TF-ES; Monogram Biosciences) was used to retest coreceptor tropism samples from 4 different cohorts of HIV-1-infected patients. Nine percent to 26% of patients with CCR5-tropic virus by the original Trofile assay had CXCR4-using virus by TF-ES. Lower CD4 cell counts were associated with CXCR4-using virus in all cohorts. © The Author 2011. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved.

Preferential susceptibility of Th9 and Th2 CD4+ T cells to X4-tropic HIV-1 infection.

PubMed

Orlova-Fink, Nina; Chowdhury, Fatema Z; Sun, Xiaoming; Harrington, Sean; Rosenberg, Eric S; Yu, Xu G; Lichterfeld, Mathias

2017-10-23

The functional polarization of CD4 T cells determines their antimicrobial effector profile, but may also impact the susceptibility to infection with HIV-1. Here, we analyzed the susceptibility of CD4 T cells with different functional polarization to infection with X4 and R5-tropic HIV-1. CD4 T cells with a Th1, Th2, Th17, and Th9 polarization were subjected to in-vitro infection assays with X4, R5, or vesicular stomatitis virus-G protein-pseudotyped HIV-1. In addition, we sorted differentially polarized CD4 T-cell subsets from individuals treated with antiretroviral therapy and analyzed the tropism of viral env sequences. Th9-polarized CD4 T cells and, to a lesser extent, Th2-polarized CD4 T cells expressed higher surface levels of CXCR4, and are more permissive to X4-tropic infection in vitro. In contrast, Th1 and Th17 CD4 T cells exhibited stronger surface expression of CCR5, and were more susceptible to infection with R5-tropic viruses. Correspondingly, the distribution of X4-tropic viral sequences in antiretroviral therapy-treated HIV-1-infected patients was biased toward Th9/Th2 cells, whereas R5-tropic sequences were more frequently observed in Th17 cells. CD4 T-cell polarization is associated with a distinct susceptibility to X4 and R5-tropic HIV-1 infection.

CXCR4-using HIV variants in a cohort of Black men who have sex with men: HIV Prevention Trials Network 061.

PubMed

Chen, Iris; Huang, Wei; Connor, Matthew B; Frantzell, Arne; Cummings, Vanessa; Beauchamp, Geetha G; Griffith, Sam; Fields, Sheldon D; Scott, Hyman M; Shoptaw, Steven; Del Rio, Carlos; Magnus, Manya; Mannheimer, Sharon; Tieu, Hong-Van; Wheeler, Darrell P; Mayer, Kenneth H; Koblin, Beryl A; Eshleman, Susan H

2016-07-01

To evaluate factors associated with HIV tropism among Black men who have sex with men (MSM) in the United States enrolled in a clinical study (HIV Prevention Trials Network 061). HIV tropism was analyzed using a phenotypic assay (Trofile assay, Monogram Biosciences). Samples were analyzed from 43 men who were HIV infected at enrollment and reported either exclusive insertive intercourse or exclusive receptive intercourse; samples were also analyzed from 20 men who were HIV uninfected at enrollment and seroconverted during the study. Clonal analysis of individual viral variants was performed for seroconverters who had dual/mixed (DM) viruses. DM viruses were detected in samples from 11 (26%) of the 43 HIV-infected men analyzed at the enrollment visit; HIV tropism did not differ between those reporting exclusive insertive vs receptive intercourse. DM viruses were also detected in five (25%) of the 20 seroconverters. DM viruses were associated with lower CD4 cell counts. Seroconverters with DM viruses had dual-tropic viruses only or mixed populations of CCR5- and dual-tropic viruses. DM viruses were frequently detected among Black MSM in this study, including seroconverters. Further studies are needed to understand factors driving transmission and selection of CXCR4- and dual-tropic viruses among Black MSM.

Inhibitory function of adapter-related protein complex 2 alpha 1 subunit in the process of nuclear translocation of human immunodeficiency virus type 1 genome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kitagawa, Yukiko; Department of Oral and Maxillofacial Surgery II, Osaka University, Osaka 565-0871; Kameoka, Masanori

2008-03-30

The transfection of human cells with siRNA against adapter-related protein complex 2 alpha 1 subunit (AP2{alpha}) was revealed to significantly up-regulate the replication of human immunodeficiency virus type 1 (HIV-1). This effect was confirmed by cell infection with vesicular stomatitis virus G protein-pseudotyped HIV-1 as well as CXCR4-tropic and CCR5-tropic HIV-1. Viral adsorption, viral entry and reverse transcription processes were not affected by cell transfection with siRNA against AP2{alpha}. In contrast, viral nuclear translocation as well as the integration process was significantly up-regulated in cells transfected with siRNA against AP2{alpha}. Confocal fluorescence microscopy revealed that a subpopulation of AP2{alpha} wasmore » not only localized in the cytoplasm but was also partly co-localized with lamin B, importin {beta} and Nup153, implying that AP2{alpha} negatively regulates HIV-1 replication in the process of nuclear translocation of viral DNA in the cytoplasm or the perinuclear region. We propose that AP2{alpha} may be a novel target for disrupting HIV-1 replication in the early stage of the viral life cycle.« less

CXCR4-using HIV variants in a cohort of Black men who have sex with men: HIV Prevention Trials Network 061

PubMed Central

Chen, Iris; Huang, Wei; Connor, Matthew B.; Frantzell, Arne; Cummings, Vanessa; Beauchamp, Geetha G.; Griffith, Sam; Fields, Sheldon D.; Scott, Hyman M.; Shoptaw, Steven; del Rio, Carlos; Magnus, Manya; Mannheimer, Sharon; Tieu, Hong-Van; Wheeler, Darrell P.; Mayer, Kenneth H.; Koblin, Beryl A.; Eshleman, Susan H.

2016-01-01

Objective To evaluate factors associated with HIV tropism among Black men who have sex with men (MSM) in the United States enrolled in a clinical study (HIV Prevention Trials Network 061). Methods HIV tropism was analyzed using a phenotypic assay (Trofile assay, Monogram Biosciences). Samples were analyzed from 43 men who were HIV infected at enrollment and reported either exclusive insertive intercourse or exclusive receptive intercourse; samples were also analyzed from 20 men who were HIV uninfected at enrollment and seroconverted during the study. Clonal analysis of individual viral variants was performed for seroconverters who had dual/mixed viruses. Results Dual/mixed viruses were detected in samples from 11 (26%) of the 43 HIV-infected men analyzed at the enrollment visit; HIV tropism did not differ between those reporting exclusive insertive vs. receptive intercourse. Dual/mixed viruses were also detected in five (25%) of the 20 seroconverters. Dual/mixed viruses were associated with lower CD4 cell counts. Seroconverters with dual/mixed viruses had dual-tropic viruses only or mixed populations of CCR5− and dual-tropic viruses. Conclusions Dual/mixed viruses were frequently detected among Black MSM in this study, including seroconverters. Further studies are needed to understand factors driving transmission and selection of CXCR4− and dual-tropic viruses among Black MSM. PMID:27300696

Simian Immunodeficiency Virus SIVagm Efficiently Utilizes Non-CCR5 Entry Pathways in African Green Monkey Lymphocytes: Potential Role for GPR15 and CXCR6 as Viral Coreceptors.

PubMed

Riddick, Nadeene E; Wu, Fan; Matsuda, Kenta; Whitted, Sonya; Ourmanov, Ilnour; Goldstein, Simoy; Goeken, Robert M; Plishka, Ronald J; Buckler-White, Alicia; Brenchley, Jason M; Hirsch, Vanessa M

2015-12-09

African green monkeys (AGM) are natural hosts of simian immunodeficiency virus (SIV), and infection in these animals is generally nonpathogenic, whereas infection of nonnatural hosts, such as rhesus macaques (RM), is commonly pathogenic. CCR5 has been described as the primary entry coreceptor for SIV in vivo, while human-derived CXCR6 and GPR15 also appear to be used in vitro. However, sooty mangabeys that are genetically deficient in CCR5 due to an out-of-frame deletion are infectible with SIVsmm, indicating that SIVsmm can use alternative coreceptors in vivo. In this study, we examined the CCR5 dependence of SIV strains derived from vervet AGM (SIVagmVer) and the ability of AGM-derived GPR15 and CXCR6 to serve as potential entry coreceptors. We found that SIVagmVer replicated efficiently in AGM and RM peripheral blood mononuclear cells (PBMC) in the presence of the CCR5 antagonist maraviroc, despite the fact that maraviroc was capable of blocking the CCR5-tropic strains SIVmac239, SIVsmE543-3, and simian-human immunodeficiency virus SHIV-AD8 in RM PBMC. We also found that AGM CXCR6 and AGM GPR15, to a lesser extent, supported entry of pseudotype viruses bearing SIVagm envelopes, including SIVagm transmitted/founder envelopes. Lastly, we found that CCR5, GPR15, and CXCR6 mRNAs were detected in AGM and RM memory CD4(+) T cells. These results suggest that GPR15 and CXCR6 are expressed on AGM CD4(+) T cells and are potential alternative coreceptors for SIVagm use in vivo. These data suggest that the use of non-CCR5 entry pathways may be a common feature of SIV replication in natural host species, with the potential to contribute to nonpathogenicity in these animals. African green monkeys (AGM) are natural hosts of SIV, and infection in these animals generally does not cause AIDS, whereas SIV-infected rhesus macaques (RM) typically develop AIDS. Although it has been reported that SIV generally uses CD4 and CCR5 to enter target cells in vivo, other molecules, such as GPR15 and CXCR6, also function as SIV coreceptors in vitro. In this study, we investigated whether SIV from vervet AGM can use non-CCR5 entry pathways, as has been observed in sooty mangabeys. We found that SIVagmVer efficiently replicated in AGM and RM peripheral blood mononuclear cells in the presence of the CCR5 antagonist maraviroc, suggesting that non-CCR5 entry pathways can support SIVagm entry. We found that AGM-derived GPR15 and CXCR6 support SIVagmVer entry in vitro and may serve as entry coreceptors for SIVagm in vivo, since their mRNAs were detected in AGM memory CD4(+) T cells, the preferred target cells of SIV. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Simian Immunodeficiency Virus SIVagm Efficiently Utilizes Non-CCR5 Entry Pathways in African Green Monkey Lymphocytes: Potential Role for GPR15 and CXCR6 as Viral Coreceptors

PubMed Central

Riddick, Nadeene E.; Wu, Fan; Matsuda, Kenta; Whitted, Sonya; Ourmanov, Ilnour; Goldstein, Simoy; Goeken, Robert M.; Plishka, Ronald J.; Buckler-White, Alicia; Brenchley, Jason M.

2015-01-01

ABSTRACT African green monkeys (AGM) are natural hosts of simian immunodeficiency virus (SIV), and infection in these animals is generally nonpathogenic, whereas infection of nonnatural hosts, such as rhesus macaques (RM), is commonly pathogenic. CCR5 has been described as the primary entry coreceptor for SIV in vivo, while human-derived CXCR6 and GPR15 also appear to be used in vitro. However, sooty mangabeys that are genetically deficient in CCR5 due to an out-of-frame deletion are infectible with SIVsmm, indicating that SIVsmm can use alternative coreceptors in vivo. In this study, we examined the CCR5 dependence of SIV strains derived from vervet AGM (SIVagmVer) and the ability of AGM-derived GPR15 and CXCR6 to serve as potential entry coreceptors. We found that SIVagmVer replicated efficiently in AGM and RM peripheral blood mononuclear cells (PBMC) in the presence of the CCR5 antagonist maraviroc, despite the fact that maraviroc was capable of blocking the CCR5-tropic strains SIVmac239, SIVsmE543-3, and simian-human immunodeficiency virus SHIV-AD8 in RM PBMC. We also found that AGM CXCR6 and AGM GPR15, to a lesser extent, supported entry of pseudotype viruses bearing SIVagm envelopes, including SIVagm transmitted/founder envelopes. Lastly, we found that CCR5, GPR15, and CXCR6 mRNAs were detected in AGM and RM memory CD4+ T cells. These results suggest that GPR15 and CXCR6 are expressed on AGM CD4+ T cells and are potential alternative coreceptors for SIVagm use in vivo. These data suggest that the use of non-CCR5 entry pathways may be a common feature of SIV replication in natural host species, with the potential to contribute to nonpathogenicity in these animals. IMPORTANCE African green monkeys (AGM) are natural hosts of SIV, and infection in these animals generally does not cause AIDS, whereas SIV-infected rhesus macaques (RM) typically develop AIDS. Although it has been reported that SIV generally uses CD4 and CCR5 to enter target cells in vivo, other molecules, such as GPR15 and CXCR6, also function as SIV coreceptors in vitro. In this study, we investigated whether SIV from vervet AGM can use non-CCR5 entry pathways, as has been observed in sooty mangabeys. We found that SIVagmVer efficiently replicated in AGM and RM peripheral blood mononuclear cells in the presence of the CCR5 antagonist maraviroc, suggesting that non-CCR5 entry pathways can support SIVagm entry. We found that AGM-derived GPR15 and CXCR6 support SIVagmVer entry in vitro and may serve as entry coreceptors for SIVagm in vivo, since their mRNAs were detected in AGM memory CD4+ T cells, the preferred target cells of SIV. PMID:26656714

The Effects of the Recombinant CCR5 T4 Lysozyme Fusion Protein on HIV-1 Infection.

PubMed

Jin, Qingwen; Chen, Hong; Wang, Xingxia; Zhao, Liandong; Xu, Qingchen; Wang, Huijuan; Li, Guanyu; Yang, Xiaofan; Ma, Hongming; Wu, Haoquan; Ji, Xiaohui

2015-01-01

Insertion of T4 lysozyme (T4L) into the GPCR successfully enhanced GPCR protein stability and solubilization. However, the biological functions of the recombinant GPCR protein have not been analyzed. We engineered the CCR5-T4L mutant and expressed and purified the soluble recombinant protein using an E.coli expression system. The antiviral effects of this recombinant protein in THP-1 cell lines, primary human macrophages, and PBMCs from different donors were investigated. We also explored the possible mechanisms underlying the observed antiviral effects. Our data showed the biphasic inhibitory and promotion effects of different concentrations of soluble recombinant CCR5-T4L protein on R5 tropic human immunodeficiency virus-1 (HIV-1) infection in THP-1 cell lines, human macrophages, and PBMCs from clinical isolates. We demonstrated that soluble recombinant CCR5-T4L acts as a HIV-1 co-receptor, interacts with wild type CCR5, down-regulates the surface CCR5 expression in human macrophages, and interacts with CCL5 to inhibit macrophage migration. Using binding assays, we further determined that recombinant CCR5-T4L and [125I]-CCL5 compete for the same binding site on wild type CCR5. Our results suggest that recombinant CCR5-T4L protein marginally promotes HIV-1 infection at low concentrations and markedly inhibits infection at higher concentrations. This recombinant protein may be helpful in the future development of anti-HIV-1 therapeutic agents.

Abnormal immune response of CCR5-deficient mice to ocular infection with herpes simplex virus type 1

PubMed Central

Carr, Daniel J.J.; Ash, John; Lane, Thomas E.; Kuziel, William A.

2006-01-01

Summary Ocular herpes simplex virus type 1 (HSV-1) infection elicits a strong inflammatory response that is associated with production of the β chemokines CCL3 and CCL5, which share a common receptor, CCR5. To gain insight into the role of these molecules in ocular immune responses, we infected the corneas of WT and CCR5-deficient (CCR5-/-) mice with HSV-1 and measured inflammatory parameters. In the absence of CCR5, the early infiltration of neutrophils into the cornea was diminished. Associated with this aberrant leukocyte recruitment, neutrophils in CCR5-/- mice were restricted to the stroma whereas in wild type mice these cells trafficked to the stroma and epithelial layers of the infected cornea. Virus titers and cytokine/chemokine levels in the infected tissue of these mice were similar for the first 5 days after infection. However, by day 7 post-infection, the CCR5-/- mice showed a significant elevation in the chemokines CCL2, CCL5, CXCL9, and CXCL10 in the trigeminal ganglion and brain stem as well as a significant increase in viral burden. The increase in chemokine expression was associated with an increase in the infiltration of CD4 and/or CD8 T cells into the trigeminal ganglion and brain stem of CCR5-/- mice. Surprisingly, even though infected CCR5-/- mice were less efficient at controlling the progression of virus replication, there was no difference in mortality. These results suggest that, although CCR5 plays a role in regulating leukocyte trafficking and control of virus burden, compensatory mechanisms are involved in preventing mortality following HSV-1 infection. PMID:16476970

Different HIV-1 env frames: gp120 and ASP (antisense protein) biosynthesis, and theirs co-variation tropic amino acid signatures in X4- and R5-viruses.

PubMed

Dimonte, Salvatore

2017-01-01

Antisense protein (ASP) is the new actor of viral life of Human Immunodeficiency Virus type 1 (HIV-1) although proposed above 20 years ago. The asp ORF is into complementary strand of the gp120/gp41 junction of env gene. The ASP biological role remains little known. Knowing the Env markers of viral tropism, a dataset of sequences (660 strains) was used to analyze the hypothetical ASP involvement in CCR5 (R5) and/or CXCR4 (X4) co-receptor interaction. Preliminarily, prevalence of ASP and gp120 V3 mutations was performed; following association among mutations were elaborate. The classical V3 tropic-signatures were confirmed, and 36 R5- and 22 X4-tropic ASP mutations were found. Moreover, by analyzing the ASP sequences, 36 out of 179 amino acid positions significantly associated with different co-receptor usage were found. Several statistically significant associations between gp120 V3 and ASP mutations were observed. The dendrogram showed the existence of a cluster associated with R5-usage and a large cluster associated with X4-usage. These results show that gp120 V3 and specific amino acid changes in ASP are associated together with CXCR4 and/or CCR5-usage. These findings implement previous observations on unclear ASP functions. J. Med. Virol. 89:112-122, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Dengue virus requires the CC-chemokine receptor CCR5 for replication and infection development.

PubMed

Marques, Rafael E; Guabiraba, Rodrigo; Del Sarto, Juliana L; Rocha, Rebeca F; Queiroz, Ana Luiza; Cisalpino, Daniel; Marques, Pedro E; Pacca, Carolina C; Fagundes, Caio T; Menezes, Gustavo B; Nogueira, Maurício L; Souza, Danielle G; Teixeira, Mauro M

2015-08-01

Dengue is a mosquito-borne disease that affects millions of people worldwide yearly. Currently, there is no vaccine or specific treatment available. Further investigation on dengue pathogenesis is required to better understand the disease and to identify potential therapeutic targets. The chemokine system has been implicated in dengue pathogenesis, although the specific role of chemokines and their receptors remains elusive. Here we describe the role of the CC-chemokine receptor CCR5 in Dengue virus (DENV-2) infection. In vitro experiments showed that CCR5 is a host factor required for DENV-2 replication in human and mouse macrophages. DENV-2 infection induces the expression of CCR5 ligands. Incubation with an antagonist prevents CCR5 activation and reduces DENV-2 positive-stranded (+) RNA inside macrophages. Using an immunocompetent mouse model of DENV-2 infection we found that CCR5(-/-) mice were resistant to lethal infection, presenting at least 100-fold reduction of viral load in target organs and significant reduction in disease severity. This phenotype was reproduced in wild-type mice treated with CCR5-blocking compounds. Therefore, CCR5 is a host factor required for DENV-2 replication and disease development. Targeting CCR5 might represent a therapeutic strategy for dengue fever. These data bring new insights on the association between viral infections and the chemokine receptor CCR5. © 2015 John Wiley & Sons Ltd.

A Maraviroc-Resistant HIV-1 with Narrow Cross-Resistance to Other CCR5 Antagonists Depends on both N-Terminal and Extracellular Loop Domains of Drug-Bound CCR5▿

PubMed Central

Tilton, John C.; Wilen, Craig B.; Didigu, Chukwuka A.; Sinha, Rohini; Harrison, Jessamina E.; Agrawal-Gamse, Caroline; Henning, Elizabeth A.; Bushman, Frederick D.; Martin, Jeffrey N.; Deeks, Steven G.; Doms, Robert W.

2010-01-01

CCR5 antagonists inhibit HIV entry by binding to a coreceptor and inducing changes in the extracellular loops (ECLs) of CCR5. In this study, we analyzed viruses from 11 treatment-experienced patients who experienced virologic failure on treatment regimens containing the CCR5 antagonist maraviroc (MVC). Viruses from one patient developed high-level resistance to MVC during the course of treatment. Although resistance to one CCR5 antagonist is often associated with broad cross-resistance to other agents, these viruses remained sensitive to most other CCR5 antagonists, including vicriviroc and aplaviroc. MVC resistance was dependent upon mutations within the V3 loop of the viral envelope (Env) protein and was modulated by additional mutations in the V4 loop. Deep sequencing of pretreatment plasma viral RNA indicated that resistance appears to have occurred by evolution of drug-bound CCR5 use, despite the presence of viral sequences predictive of CXCR4 use. Envs obtained from this patient before and during MVC treatment were able to infect cells expressing very low CCR5 levels, indicating highly efficient use of a coreceptor. In contrast to previous reports in which CCR5 antagonist-resistant viruses interact predominantly with the N terminus of CCR5, these MVC-resistant Envs were also dependent upon the drug-modified ECLs of CCR5 for entry. Our results suggest a model of CCR5 cross-resistance whereby viruses that predominantly utilize the N terminus are broadly cross-resistant to multiple CCR5 antagonists, whereas viruses that require both the N terminus and antagonist-specific ECL changes demonstrate a narrow cross-resistance profile. PMID:20702642

Glycosylphosphatidylinositol-Anchored Anti-HIV scFv Efficiently Protects CD4 T Cells from HIV-1 Infection and Deletion in hu-PBL Mice

PubMed Central

Ye, Chaobaihui; Wang, Weiming; Cheng, Liang; Li, Guangming; Wen, Michael; Wang, Qi; Zhang, Qing; Li, Dan

2016-01-01

ABSTRACT Despite success in viral inhibition and CD4 T cell recovery by highly active antiretroviral treatment (HAART), HIV-1 is still not curable due to the persistence of the HIV-1 reservoir during treatment. One patient with acute myeloid leukemia who received allogeneic hematopoietic stem cell transplantation from a homozygous CCR5 Δ32 donor has had no detectable viremia for 9 years after HAART cessation. This case has inspired a field of HIV-1 cure research focusing on engineering HIV-1 resistance in permissive cells. Here, we employed a glycosylphosphatidylinositol (GPI)-scFv X5 approach to confer resistance of human primary CD4 T cells to HIV-1. We showed that primary CD4 T cells expressing GPI-scFv X5 were resistant to CCR5 (R5)-, CXCR4 (X4)-, and dual-tropic HIV-1 and had a survival advantage compared to control cells ex vivo. In a hu-PBL mouse study, GPI-scFv X5-transduced CD4 T cells were selected in peripheral blood and lymphoid tissues upon HIV-1 infection. Finally, GPI-scFv X5-transduced CD4 T cells, after being cotransfused with HIV-infected cells, showed significantly reduced viral loads and viral RNA copy numbers relative to CD4 cells in hu-PBL mice compared to mice with GPI-scFv AB65-transduced CD4 T cells. We conclude that GPI-scFv X5-modified CD4 T cells could potentially be used as a genetic intervention against both R5- and X4-tropic HIV-1 infections. IMPORTANCE Blocking of HIV-1 entry is one of most promising approaches for therapy. Genetic disruption of the HIV-1 coreceptor CCR5 by nucleases in T cells is under 2 clinical trials and leads to reduced viremia in patients. However, the emergence of viruses using the CXCR4 coreceptor is a concern for therapies applying single-coreceptor disruption. Here, we report that HIV-1-permissive CD4 T cells engineered with GPI-scFv X5 are resistant to R5-, X4-, or dual-tropic virus infection ex vivo. In a preclinical study using hu-PBL mice, we show that CD4 T cells were protected and that GPI-scFv X5-transduced cells were selected in HIV-1-infected animals. Moreover, we show that GPI-scFv X5-transduced CD4 T cells exerted a negative effect on virus replication in vivo. We conclude that GPI-scFv X5-modified CD4 T cells could potentially be used as a genetic intervention against both R5- and X4-tropic HIV-1 infections. PMID:27881659

Loss of a single N-linked glycan allows CD4-independent human immunodeficiency virus type 1 infection by altering the position of the gp120 V1/V2 variable loops.

PubMed

Kolchinsky, P; Kiprilov, E; Bartley, P; Rubinstein, R; Sodroski, J

2001-04-01

The gp120 envelope glycoprotein of primary human immunodeficiency virus type 1 (HIV-1) promotes virus entry by sequentially binding CD4 and the CCR5 chemokine receptor on the target cell. Previously, we adapted a primary HIV-1 isolate, ADA, to replicate in CD4-negative canine cells expressing human CCR5. The gp120 changes responsible for CD4-independent replication were limited to the V2 loop-V1/V2 stem. Here we show that elimination of a single glycosylation site at asparagine 197 in the V1/V2 stem is sufficient for CD4-independent gp120 binding to CCR5 and for HIV-1 entry into CD4-negative cells expressing CCR5. Deletion of the V1/V2 loops also allowed CD4-independent viral entry and gp120 binding to CCR5. The binding of the wild-type ADA gp120 to CCR5 was less dependent upon CD4 at 4 degrees C than at 37 degrees C. In the absence of the V1/V2 loops, neither removal of the N-linked carbohydrate at asparagine 197 nor lowering of the temperature increased the CD4-independent phenotypes. A CCR5-binding conformation of gp120, achieved by CD4 interaction or by modification of temperature, glycosylation, or variable loops, was preferentially recognized by the monoclonal antibody 48d. These results suggest that the CCR5-binding region of gp120 is occluded by the V1/V2 variable loops, the position of which can be modulated by temperature, CD4 binding, or an N-linked glycan in the V1/V2 stem.

Both Cerebral and Hematopoietic Deficiencies in CCR2 Result in Uncontrolled Herpes Simplex Virus Infection of the Central Nervous System in Mice.

PubMed

Menasria, Rafik; Canivet, Coraline; Piret, Jocelyne; Gosselin, Jean; Boivin, Guy

2016-01-01

CCR2 is a chemokine receptor expressed on the surface of blood leukocytes, particularly «Ly6Chi» inflammatory monocytes and microglia. Signaling through this receptor is thought to influence the immune activity of microglia as well as monocytes egress from the bone marrow (BM) and their trafficking into the central nervous system (CNS) in several neurological diseases. During experimental herpes simplex virus 1 (HSV-1) encephalitis (HSE), CCR2 deficiency has been reported to exacerbate the outcome of the disease. However, the precise contribution of CCR2 expressed in cells of the CNS or peripheral monocytes in the protection against HSE remains unclear. To dissect the differential role of CCR2 during HSE, chimeric mice with receptor deficiency in the brain or blood cells were generated by transplanting wild-type (WT) C57BL/6 or CCR2-/- BM-derived cells in CCR2-/- (WT→CCR2-/-) and WT (CCR2-/-→WT) mice, respectively. Our results indicate that following intranasal infection with 1.2x106 plaque forming units of HSV-1, CCR2 deficiency in hematopoietic cells and, to a lesser extent, in CNS exacerbates the outcome of HSE. Mortality rates of CCR2-/- (71.4%) and CCR2-/-→WT (57.1%) mice were significantly higher than that of WT (15.3%; P<0.01 and P<0.05, respectively) but the difference did not reach statistical significance for WT→CCR2-/- animals (42.8%; P = 0.16). Both peripheral and CNS deficiencies in CCR2 resulted in increased infectious viral titers and wider dissemination of HSV antigens in the brain as well as an overproduction of inflammatory cytokines and chemokines including IL-1β, IL-6, CCL2, CCL3 and CCL5. Furthermore, CCR2 deficiency in the hematopoietic system altered monocytes egress from the BM and their recruitment to the CNS, which may contribute to the failure in HSV-1 containment. Collectively, these data suggest that CCR2 expressed on cells of CNS and especially on peripheral monocytes is important for the control of HSV-1 replication and inflammatory environment during experimental HSE.

Both Cerebral and Hematopoietic Deficiencies in CCR2 Result in Uncontrolled Herpes Simplex Virus Infection of the Central Nervous System in Mice

PubMed Central

Menasria, Rafik; Canivet, Coraline; Piret, Jocelyne; Gosselin, Jean; Boivin, Guy

2016-01-01

CCR2 is a chemokine receptor expressed on the surface of blood leukocytes, particularly «Ly6Chi» inflammatory monocytes and microglia. Signaling through this receptor is thought to influence the immune activity of microglia as well as monocytes egress from the bone marrow (BM) and their trafficking into the central nervous system (CNS) in several neurological diseases. During experimental herpes simplex virus 1 (HSV-1) encephalitis (HSE), CCR2 deficiency has been reported to exacerbate the outcome of the disease. However, the precise contribution of CCR2 expressed in cells of the CNS or peripheral monocytes in the protection against HSE remains unclear. To dissect the differential role of CCR2 during HSE, chimeric mice with receptor deficiency in the brain or blood cells were generated by transplanting wild-type (WT) C57BL/6 or CCR2-/- BM-derived cells in CCR2-/- (WT→CCR2-/-) and WT (CCR2-/-→WT) mice, respectively. Our results indicate that following intranasal infection with 1.2x106 plaque forming units of HSV-1, CCR2 deficiency in hematopoietic cells and, to a lesser extent, in CNS exacerbates the outcome of HSE. Mortality rates of CCR2-/- (71.4%) and CCR2-/-→WT (57.1%) mice were significantly higher than that of WT (15.3%; P<0.01 and P<0.05, respectively) but the difference did not reach statistical significance for WT→CCR2-/- animals (42.8%; P = 0.16). Both peripheral and CNS deficiencies in CCR2 resulted in increased infectious viral titers and wider dissemination of HSV antigens in the brain as well as an overproduction of inflammatory cytokines and chemokines including IL-1β, IL-6, CCL2, CCL3 and CCL5. Furthermore, CCR2 deficiency in the hematopoietic system altered monocytes egress from the BM and their recruitment to the CNS, which may contribute to the failure in HSV-1 containment. Collectively, these data suggest that CCR2 expressed on cells of CNS and especially on peripheral monocytes is important for the control of HSV-1 replication and inflammatory environment during experimental HSE. PMID:27930721

Rapid and Simple Phenotypic Assay for Drug Susceptibility of Human Immunodeficiency Virus Type 1 Using CCR5-Expressing HeLa/CD4+ Cell Clone 1-10 (MAGIC-5)

PubMed Central

Hachiya, Atsuko; Aizawa-Matsuoka, Saori; Tanaka, Mari; Takahashi, Yukiko; Ida, Setsuko; Gatanaga, Hiroyuki; Hirabayashi, Yoshihiro; Kojima, Asato; Tatsumi, Masashi; Oka, Shinichi

2001-01-01

We describe a rapid and simple novel phenotypic assay for drug susceptibility of human immunodeficiency virus type-1 (HIV-1) using a CCR5-expressing HeLa/CD4+ cell clone 1-10 (MAGIC-5). MAGIC-5 cells produced large amounts of HIV-1 in culture supernatants, which enabled us to perform the phenotypic resistance assay. Determination of HIV-1 susceptibility to various protease inhibitors (PI) and nucleoside reverse transcriptase inhibitors was completed within 15 days in T-cell-tropic (X4) and macrophage-tropic (R5) viruses using fresh plasma samples containing at least 104 copies/ml. The nucleotide sequence of the envelope V3 region of HIV-1 in plasma was almost identical to that of the virus isolated by MAGIC-5 cells, suggesting a lack of selection bias in our assay. The assay variability was confined to within five-fold in all drugs examined. Accordingly, we used a 10-fold increase in the 50% inhibitory concentration as the cutoff value for viral resistance in the present assay. HIV-1 resistant to lamivudine, which was not detected by conventional genotypic assays, was isolated. In HIV-1 with PI-associated primary amino acid substitutions, our assay showed that drug resistance profiles correlated well with previously reported genotypic-assay data. Furthermore, our assay provided comprehensive results regarding PI resistance in the presence of multiple mutations. The novel assay successfully quantified the level of resistance of clinical HIV-1 isolates to a battery of anti-HIV drugs, indicating its clinical usefulness, particularly in patients who failed to respond to antiretroviral chemotherapy. PMID:11158746

Anionic Carbosilane Dendrimers Destabilize the GP120-CD4 Complex Blocking HIV-1 Entry and Cell to Cell Fusion.

PubMed

Guerrero-Beltran, Carlos; Rodriguez-Izquierdo, Ignacio; Serramia, Ma Jesus; Araya-Durán, Ingrid; Márquez-Miranda, Valeria; Gomez, Rafael; de la Mata, Francisco Javier; Leal, Manuel; González-Nilo, Fernando; Muñoz-Fernández, M Angeles

2018-05-16

Cell-to-cell transmission is the most effective pathway for the spread of human immunodeficiency virus (HIV-1). Infected cells expose virus-encoded fusion proteins on their surface as a consequence of HIV-1 replicative cycle that interacts with noninfected cells through CD4 receptor and CXCR4 coreceptor leading to the formation of giant multinucleated cells known as syncytia. Our group previously described the potent activity of dendrimers against CCR5-tropic viruses. Nevertheless, the study of G1-S4, G2-S16, and G3-S16 dendrimers in the context of X4-HIV-1 tropic cell-cell fusion referred to syncytium formation remains still unknown. These dendrimers showed a suitable biocompatibility in all cell lines studied and our results demonstrated that anionic carbosilane dendrimers G1-S4, G2-S16, and G3-S16 significantly inhibit the X4-HIV-1 infection, as well as syncytia formation, in a dose dependent manner. We also demonstrated that G2-S16 and G1-S4 significantly reduced syncytia formation in HIV-1 Env-mediated cell-to-cell fusion model. Molecular modeling and in silico models showed that G2-S16 dendrimer interfered with gp120-CD4 complex and demonstrated its potential use for a treatment.

Generation and Characterization of HIV-1 Transmitted and Founder Virus Consensus Sequence from Intravenous Drug Users in Xinjiang, China.

PubMed

Li, Fan; Ma, Liying; Feng, Yi; Hu, Jing; Ni, Na; Ruan, Yuhua; Shao, Yiming

2017-06-01

HIV-1 transmission in intravenous drug users (IDUs) has been characterized by high genetic multiplicity and suggests a greater challenge for HIV-1 infection blocking. We investigated a total of 749 sequences of full-length gp160 gene obtained by single genome sequencing (SGS) from 22 HIV-1 early infected IDUs in Xinjiang province, northwest China, and generated a transmitted and founder virus (T/F virus) consensus sequence (IDU.CON). The T/F virus was classified as subtype CRF07_BC and predicted to be CCR5-tropic virus. The variable region (V1, V2, and V4 loop) of IDU.CON showed length variation compared with the heterosexual T/F virus consensus sequence (HSX.CON) and homosexual T/F virus consensus sequence (MSM.CON). A total of 26 N-linked glycosylation sites were discovered in the IDU.CON sequence, which is less than that of MSM.CON and HSX.CON. Characterization of T/F virus from IDUs highlights the genetic make-up and complexity of virus near the moment of transmission or in early infection preceding systemic dissemination and is important toward the development of an effective HIV-1 preventive methods, including vaccines.

Identification of dual-tropic HIV-1 using evolved neural networks.

PubMed

Fogel, Gary B; Lamers, Susanna L; Liu, Enoch S; Salemi, Marco; McGrath, Michael S

2015-11-01

Blocking the binding of the envelope HIV-1 protein to immune cells is a popular concept for development of anti-HIV therapeutics. R5 HIV-1 binds CCR5, X4 HIV-1 binds CXCR4, and dual-tropic HIV-1 can bind either coreceptor for cellular entry. R5 viruses are associated with early infection and over time can evolve to X4 viruses that are associated with immune failure. Dual-tropic HIV-1 is less studied; however, it represents functional antigenic intermediates during the transition of R5 to X4 viruses. Viral tropism is linked partly to the HIV-1 envelope V3 domain, where the amino acid sequence helps dictate the receptor a particular virus will target; however, using V3 sequence information to identify dual-tropic HIV-1 isolates has remained difficult. Our goal in this study was to elucidate features of dual-tropic HIV-1 isolates that assist in the biological understanding of dual-tropism and develop an approach for their detection. Over 1559 HIV-1 subtype B sequences with known tropisms were analyzed. Each sequence was represented by 73 structural, biochemical and regional features. These features were provided to an evolved neural network classifier and evaluated using balanced and unbalanced data sets. The study resolved R5X4 viruses from R5 with an accuracy of 81.8% and from X4 with an accuracy of 78.8%. The approach also identified a set of V3 features (hydrophobicity, structural and polarity) that are associated with tropism transitions. The ability to distinguish R5X4 isolates will improve computational tropism decisions for R5 vs. X4 and assist in HIV-1 research and drug development efforts. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

[The expression of periphery blood leucocyte CCR3 and CCR5 in the children with Epstein-Barr virus associated infectious mononucleosis].

PubMed

Qi, Tie-xiong; Gao, Guo-hua; Liu, Shi-hua

2010-10-01

To explore the expression of periphery blood leucocyte CCR3 and CCR5 and to comprehend T helper cell in the Children with Epstein-Barr virus associated infectious mononucleosis. We defined the children according to the diagnosis criterion through Paul-Bunnell test inspecting the children's periphery blood unusual lymphocyte and detecting their anti-EBV-CA-IgM, anti-EBV-CA-IgG and anti-EBV-NA-IgG by ELISA and counted the ratio of CCR3 + and CCR5 + cells in lymphocytes with flow cytometry. The ratio of unusual lymphocyte in IM was higher than that of the healthy control group (P < 0.05). The ratio of CCR3 + cells in IM group was higher than that of the healthy control group (P < 0.05). The ratio of CCR5 + cells in IM group was significantly lower than that of the healthy control group. CCR3 + had direct interrelation with fever continued time and the ratio of unusual lymphocyte. There was a negative interrelation between CCR5 and fever continued time (P < 0.05). Children infectious of IM expressed higher level of CCR3 + and lower level of CCR5 + and there was a tendency of Th2 polarization with over production of T helper cell divide imbalance. CCR3 + and CCR5 + may be important targets to judge the degree of seriousness of IM.

Molecular Gymnastics: Mechanisms of HIV-1 Resistance to CCR5 Antagonists and Impact on Virus Phenotypes.

PubMed

Roche, Michael; Borm, Katharina; Flynn, Jacqueline K; Lewin, Sharon R; Churchill, Melissa J; Gorry, Paul R

2016-01-01

Human immunodeficiency virus type 1 (HIV-1) enters host cells through the binding of its envelope glycoproteins (Env) to the host cell receptor CD4 and then subsequent binding to a chemokine coreceptor, either CCR5 or CXCR4. CCR5 antagonists are a relatively recent class addition to the armamentarium of anti-HIV-1 drugs. These compounds act by binding to a hydrophobic pocket formed by the transmembrane helices of CCR5 and altering the conformation of the extracellular domains, such that they are no longer recognized by Env. Maraviroc is the first drug within this class to be licenced for use in HIV-1 therapy regimens. HIV resistance to CCR5 antagonists occurs either through outgrowth of pre-existing CXCR4-using viruses, or through acquisition of the ability of CCR5-using HIV-1 to use the antagonist bound form of CCR5. In the latter scenario, the mechanism underlying resistance is through complex alterations in the way that resistant Envs engage CCR5. These significant changes are unlikely to occur without consequence to the viral entry phenotype and may also open up new avenues to target CCR5 antagonist resistant viruses. This review discusses the mechanism of action of CCR5 antagonists, how HIV resistance to CCR5 antagonists occurs, and the subsequent effects on Env function.

Functional Mimetics of the HIV-1 CCR5 Co-Receptor Displayed on the Surface of Magnetic Liposomes.

PubMed

Kuzmina, Alona; Vaknin, Karin; Gdalevsky, Garik; Vyazmensky, Maria; Marks, Robert S; Taube, Ran; Engel, Stanislav

2015-01-01

Chemokine G protein coupled receptors, principally CCR5 or CXCR4, function as co-receptors for HIV-1 entry into CD4+ T cells. Initial binding of the viral envelope glycoprotein (Env) gp120 subunit to the host CD4 receptor induces a cascade of structural conformational changes that lead to the formation of a high-affinity co-receptor-binding site on gp120. Interaction between gp120 and the co-receptor leads to the exposure of epitopes on the viral gp41 that mediates fusion between viral and cell membranes. Soluble CD4 (sCD4) mimetics can act as an activation-based inhibitor of HIV-1 entry in vitro, as it induces similar structural changes in gp120, leading to increased virus infectivity in the short term but to virus Env inactivation in the long term. Despite promising clinical implications, sCD4 displays low efficiency in vivo, and in multiple HIV strains, it does not inhibit viral infection. This has been attributed to the slow kinetics of the sCD4-induced HIV Env inactivation and to the failure to obtain sufficient sCD4 mimetic levels in the serum. Here we present uniquely structured CCR5 co-receptor mimetics. We hypothesized that such mimetics will enhance sCD4-induced HIV Env inactivation and inhibition of HIV entry. Co-receptor mimetics were derived from CCR5 gp120-binding epitopes and functionalized with a palmitoyl group, which mediated their display on the surface of lipid-coated magnetic beads. CCR5-peptidoliposome mimetics bound to soluble gp120 and inhibited HIV-1 infectivity in a sCD4-dependent manner. We concluded that CCR5-peptidoliposomes increase the efficiency of sCD4 to inhibit HIV infection by acting as bait for sCD4-primed virus, catalyzing the premature discharge of its fusion potential.

Functional Mimetics of the HIV-1 CCR5 Co-Receptor Displayed on the Surface of Magnetic Liposomes

PubMed Central

Kuzmina, Alona; Vaknin, Karin; Gdalevsky, Garik; Vyazmensky, Maria; Marks, Robert S.; Taube, Ran

2015-01-01

Chemokine G protein coupled receptors, principally CCR5 or CXCR4, function as co-receptors for HIV-1 entry into CD4+ T cells. Initial binding of the viral envelope glycoprotein (Env) gp120 subunit to the host CD4 receptor induces a cascade of structural conformational changes that lead to the formation of a high-affinity co-receptor-binding site on gp120. Interaction between gp120 and the co-receptor leads to the exposure of epitopes on the viral gp41 that mediates fusion between viral and cell membranes. Soluble CD4 (sCD4) mimetics can act as an activation-based inhibitor of HIV-1 entry in vitro, as it induces similar structural changes in gp120, leading to increased virus infectivity in the short term but to virus Env inactivation in the long term. Despite promising clinical implications, sCD4 displays low efficiency in vivo, and in multiple HIV strains, it does not inhibit viral infection. This has been attributed to the slow kinetics of the sCD4-induced HIV Env inactivation and to the failure to obtain sufficient sCD4 mimetic levels in the serum. Here we present uniquely structured CCR5 co-receptor mimetics. We hypothesized that such mimetics will enhance sCD4-induced HIV Env inactivation and inhibition of HIV entry. Co-receptor mimetics were derived from CCR5 gp120-binding epitopes and functionalized with a palmitoyl group, which mediated their display on the surface of lipid-coated magnetic beads. CCR5-peptidoliposome mimetics bound to soluble gp120 and inhibited HIV-1 infectivity in a sCD4-dependent manner. We concluded that CCR5-peptidoliposomes increase the efficiency of sCD4 to inhibit HIV infection by acting as bait for sCD4-primed virus, catalyzing the premature discharge of its fusion potential. PMID:26629902

HIV coreceptor phenotyping in the clinical setting.

PubMed

Low, Andrew J; Swenson, Luke C; Harrigan, P Richard

2008-01-01

The introduction of CCR5 antagonists increases the options available for constructing antiretroviral regimens. However, this option is coupled with the caveat that patients should be tested for HIV coreceptor tropism prior to initiating CCR5 antagonist-based therapy. Failure to screen for CXCR4 usage increases the risk of using an ineffective drug, thus reducing the likelihood of viral suppression and increasing their risk for developing antiretroviral resistance. This review discusses current and future methods of determining HIV tropism, with a focus on their utility in the clinical setting for screening purposes. Some of these methods include recombinant phenotypic tests, such as the Monogram Trofile assay, as well as genotype-based predictors, heteroduplex tracking assays, and flow cytometry based methods. Currently, the best evidence supports the use of phenotypic methods, although other methods of screening for HIV coreceptor usage prior to the administration of CCR5 antagonists may reduce costs and increase turnaround time over phenotypic methods. The presence of low levels of X4 virus is a challenge to all assay methods, resulting in reduced sensitivity in clinical, patient-derived samples when compared to clonally derived samples. Gaining a better understanding of the output of these assays and correlating them with clinical progression and therapy response will provide some indication on how both genotype-based, and phenotypic assays for determining HIV coreceptor usage can be improved. In addition, leveraging new technologies capable of detecting low-level minority species may provide the most significant advances in ensuring that individuals with low levels of dual/mixed tropic virus are not inadvertently prescribed CCR5 antagonists.

HTLV-1 induces a Th1-like state in CD4+CCR4+ T cells

PubMed Central

Araya, Natsumi; Sato, Tomoo; Ando, Hitoshi; Tomaru, Utano; Yoshida, Mari; Coler-Reilly, Ariella; Yagishita, Naoko; Yamauchi, Junji; Hasegawa, Atsuhiko; Kannagi, Mari; Hasegawa, Yasuhiro; Takahashi, Katsunori; Kunitomo, Yasuo; Tanaka, Yuetsu; Nakajima, Toshihiro; Nishioka, Kusuki; Utsunomiya, Atae; Jacobson, Steven; Yamano, Yoshihisa

2014-01-01

Human T-lymphotropic virus type 1 (HTLV-1) is linked to multiple diseases, including the neuroinflammatory disease HTLV-1–associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T cell leukemia/lymphoma. Evidence suggests that HTLV-1, via the viral protein Tax, exploits CD4+ T cell plasticity and induces transcriptional changes in infected T cells that cause suppressive CD4+CD25+CCR4+ Tregs to lose expression of the transcription factor FOXP3 and produce IFN-γ, thus promoting inflammation. We hypothesized that transformation of HTLV-1–infected CCR4+ T cells into Th1-like cells plays a key role in the pathogenesis of HAM/TSP. Here, using patient cells and cell lines, we demonstrated that Tax, in cooperation with specificity protein 1 (Sp1), boosts expression of the Th1 master regulator T box transcription factor (T-bet) and consequently promotes production of IFN-γ. Evaluation of CSF and spinal cord lesions of HAM/TSP patients revealed the presence of abundant CD4+CCR4+ T cells that coexpressed the Th1 marker CXCR3 and produced T-bet and IFN-γ. Finally, treatment of isolated PBMCs and CNS cells from HAM/TSP patients with an antibody that targets CCR4+ T cells and induces cytotoxicity in these cells reduced both viral load and IFN-γ production, which suggests that targeting CCR4+ T cells may be a viable treatment option for HAM/TSP. PMID:24960164

Biological characterization of HIV type 1 envelope V3 regions from mothers and infants associated with perinatal transmission.

PubMed

Matala, E; Hahn, T; Yedavalli, V R; Ahmad, N

2001-12-10

Our previous study has shown that the human immunodeficiency virus type 1 (HIV-1) envelope V3 region minor genotypes of infected mothers were transmitted to their infants and predominated initially as a homogeneous virus population in the infants (Ahmad N, Baroudy BM, Baker RC, et al.: J Virol 1995;69:1001-1012). Here we have characterized the biological properties, including cellular tropism, replication efficiency, cytopathic effects, and coreceptor utilization, of these V3 region isolates from mothers and infants. Nineteen V3 region sequences from three mother-infant pairs, including the minor variants of mothers and the major variants of infants as characterized in our previous study, were reciprocally inserted into an HIV-1 infectious molecular clone, pNL4-3, and chimeric viruses were generated by DNA transfections into HeLa cells. Equal amounts of chimeric viruses were then used to infect T lymphocyte cell lines (A3.01 and MT-2), primary blood lymphocytes (PBLs), primary monocyte-derived macrophages (MDMs), and coreceptor cell lines. We found that the V3 region chimeras failed to replicate in T lymphocyte cell lines but replicated in MDMs and PBLs, albeit at reduced levels compared with R5 laboratory HIV-1 strains. In addition, the V3 region chimeras were able to infect the HOS-CD4(+)CCR5(+) cell line, suggesting CCR5 coreceptor utilization. Moreover, the V3 region chimeras were unable to induce syncytia in MT-2 cells, indicative of non-syncytium-inducing (NSI) phenotypes. In conclusion, the HIV-1 minor genotypes of infected mothers with macrophage-tropic and NSI or R5 phenotypes are transmitted to their infants and are initially maintained with the same properties.

Drug Resistance and Viral Tropism in HIV-1 Subtype C-Infected Patients in KwaZulu-Natal, South Africa: Implications for Future Treatment Options

PubMed Central

Singh, Ashika; Sunpath, Henry; Green, Taryn N.; Padayachi, Nagavelli; Hiramen, Keshni; Lie, Yolanda; Anton, Elizabeth D.; Murphy, Richard; Reeves, Jacqueline D.; Kuritzkes, Daniel R.; Ndung’u, Thumbi

2011-01-01

Background Drug resistance poses a significant challenge for the successful application of highly active antiretroviral therapy (HAART) globally. Furthermore, emergence of HIV-1 isolates that preferentially utilize CXCR4 as a coreceptor for cell entry, either as a consequence of natural viral evolution or HAART use may compromise the efficacy of CCR5 antagonists as alternative antiviral therapy. Methods We sequenced the pol gene of viruses from 45 individuals failing at least six months of HAART in Durban, South Africa to determine the prevalence and patterns of drug resistance mutations. Coreceptor usage profiles of these viruses and those from 45 HAART-naive individuals were analyzed using phenotypic and genotypic approaches. Results Ninety-five percent of HAART-failing patients had at least one drug resistance mutation. Thymidine analog mutations (TAMs) were present in 55% of patients with 9% of individuals possessing mutations indicative of the TAM1 pathway, 44% had TAM2 while 7% had mutations common to both pathways. Sixty percent of HAART-failing subjects had X4/dual//mixed-tropic viruses compared to 30% of HAART-naïve subjects (p<0.02). Genetic coreceptor usage prediction algorithms correlated with phenotypic results with 60% of samples from HAART-failing subjects predicted to possess CXCR4-using (X4/dual/mixed viruses) versus 15% of HAART-naïve patients. Conclusions The high proportion of TAMs and X4/dual/mixed HIV-1 viruses among patients failing therapy highlight the need for intensified monitoring of patients taking HAART and the problem of diminished drug options (including CCR5 antagonists) for patients failing therapy in resource-poor settings. PMID:21709569

Cell surface expression of CCR5 and other host factors influence the inhibition of HIV-1 infection of human lymphocytes by CCR5 ligands

PubMed Central

Ketas, Thomas J.; Kuhmann, Shawn E.; Palmer, Ashley; Zurita, Juan; He, Weijing; Ahuja, Sunil K.; Klasse, Per Johan; Moore, John P.

2007-01-01

Several CCR5 ligands, including small molecules and monoclonal antibodies (MAbs), are being developed as therapies for infection with strains of human immunodeficiency virus type 1 (HIV-1) that use CCR5 for entry (R5 viruses). The efficacy of such therapies could be influenced by inter-individual differences in host factors, such as CCR5 expression levels. To study this, we used peripheral blood mononuclear cells (PBMCs) from humans and rhesus macaques. The half-maximal inhibitory concentrations (IC50) of the small-molecule CCR5 ligands CMPD167, UK427,857 and SCH-D, and of the PRO 140 MAb, differ by >2 logs in a donor-dependent manner. We studied this variation by using flow cytometry to measure CCR5 expression on PBMCs from six of the human donors: the IC50 values of both SCH-D and PRO 140 correlated with CCR5 expression (R2 = 0.64 and 0.99, respectively). We also determined the efficacy of the CCR5 ligands against HIV-1 infection of HeLa-derived cell lines that express CD4 at the same level but vary 2-fold in CCR5 expression (JC.48 and JC.53 cells). The moderately greater CCR5 expression on the JC.53 than the JC.48 cells was associated with proportionately higher median IC50 values for all four CCR5 ligands but not for a soluble CD4-based inhibitor or a non-nucleoside reverse transcriptase inhibitor. We conclude that differences in CCR5 expression on human PBMCs, which can be affected by CCL3L1 gene dose, may influence the antiviral potency of CCR5 ligands in vitro, but other host factors are also likely to be involved. These host factors may affect the clinical activity of CCR5 inhibitors, including their use as topical microbicides to prevent HIV-1 transmission. PMID:17428518

Chemokine Receptor CCR5 Antagonist Maraviroc: Medicinal Chemistry and Clinical Applications

PubMed Central

Xu, Guoyan G.; Guo, Jia; Wu, Yuntao

2015-01-01

The human immunodeficiency virus (HIV) causes acquired immumodeficiency syndrome (AIDS), one of the worst global pandemic. The virus infects human CD4 T cells and macrophages, and causes CD4 depletion. HIV enters target cells through the binding of the viral envelope glycoprotein to CD4 and the chemokine coreceptor, CXCR4 or CCR5. In particular, the CCR5-utilizing viruses predominate in the blood during the disease course. CCR5 is expressed on the surface of various immune cells including macrophages, monocytes, microglia, dendric cells, and active memory CD4 T cells. In the human population, the CCR5 genomic mutation, CCR5Δ32, is associated with relative resistance to HIV. These findings paved the way for the discovery and development of CCR5 inhibitors to block HIV transmission and replication. Maraviroc, discovered as a CCR5 antagonist, is the only CCR5 inhibitor that has been approved by both US FDA and the European Medicines Agency (EMA) for treating HIV/AIDS patients. In this review, we summarize the medicinal chemistry and clinical studies of Maraviroc. PMID:25159165

De novo design of peptide immunogens that mimic the coiled coil region of human T-cell leukemia virus type-1 glycoprotein 21 transmembrane subunit for induction of native protein reactive neutralizing antibodies.

PubMed

Sundaram, Roshni; Lynch, Marcus P; Rawale, Sharad V; Sun, Yiping; Kazanji, Mirdad; Kaumaya, Pravin T P

2004-06-04

Peptide vaccines able to induce high affinity and protective neutralizing antibodies must rely in part on the design of antigenic epitopes that mimic the three-dimensional structure of the corresponding region in the native protein. We describe the design, structural characterization, immunogenicity, and neutralizing potential of antibodies elicited by conformational peptides derived from the human T-cell leukemia virus type 1 (HTLV-1) gp21 envelope glycoprotein spanning residues 347-374. We used a novel template design and a unique synthetic approach to construct two peptides (WCCR2T and CCR2T) that would each assemble into a triple helical coiled coil conformation mimicking the gp21 crystal structure. The peptide B-cell epitopes were grafted onto the epsilon side chains of three lysyl residues on a template backbone construct consisting of the sequence acetyl-XGKGKGKGCONH2 (where X represents the tetanus toxoid promiscuous T cell epitope (TT) sequence 580-599). Leucine substitutions were introduced at the a and d positions of the CCR2T sequence to maximize helical character and stability as shown by circular dichroism and guanidinium hydrochloride studies. Serum from an HTLV-1-infected patient was able to recognize the selected epitopes by enzyme-linked immunosorbent assay (ELISA). Mice immunized with the wild-type sequence (WCCR2T) and the mutant sequence (CCR2T) elicited high antibody titers that were capable of recognizing the native protein as shown by flow cytometry and whole virus ELISA. Sera and purified antibodies from immunized mice were able to reduce the formation of syncytia induced by the envelope glycoprotein of HTLV-1, suggesting that antibodies directed against the coiled coil region of gp21 are capable of disrupting cell-cell fusion. Our results indicate that these peptides represent potential candidates for use in a peptide vaccine against HTLV-1.

Dual R3R5 tropism characterizes cerebrospinal fluid HIV-1 isolates from individuals with high cerebrospinal fluid viral load.

PubMed

Karlsson, Ulf; Antonsson, Liselotte; Ljungberg, Bengt; Medstrand, Patrik; Esbjörnsson, Joakim; Jansson, Marianne; Gisslen, Magnus

2012-09-10

To study the use of major and alternative coreceptors by HIV-1 isolates obtained from paired plasma and cerebrospinal fluid (CSF) samples. Paired plasma and CSF isolates from HIV-1-infected individuals with varying clinical, virologic, and immunologic parameters were assessed for the ability to infect indicator cells expressing a panel of coreceptors with documented expression in the central nervous system (CNS). HIV-1 isolates obtained from plasma and CSF in 28 individuals with varying viral load, CD4 T-cell counts, and with or without AIDS-defining disease were analyzed for the ability to infect NP2.CD4 cells stably expressing a panel of HIV coreceptors (CCR5, CXCR4, CCR3, CXCR6, GPR1, APJ, ChemR23, RDC-1 or BLT1). All isolates from both plasma and CSF utilized CCR5 and/or CXCR4. However, the ability to use both CCR3 and CCR5 (R3R5) was more pronounced in CSF isolates and correlated with high CSF viral load and low CD4 T-cell count. Notably, four out of five CSF isolates of subtype C origin exhibited CXCR6 use, which coincided with high CSF viral load despite preserved CD4 T-cell counts. The use of other alternative coreceptors was less pronounced. Dual-tropic R3R5 HIV-1 isolates in CSF coincide with high CSF viral load and low CD4 T-cell counts. Frequent CXCR6 use by CSF-derived subtype C isolates indicates that subtype-specific differences in coreceptor use may exist that will not be acknowledged when assessing plasma virus isolates. The findings may also bare relevance for HIV-1 replication within the CNS, and consequently, for the neuropathogenesis of AIDS.

Decreased HIV Type 1 Transcription in CCR5-Δ32 Heterozygotes During Suppressive Antiretroviral Therapy

PubMed Central

Wang, Charlene; Abdel-Mohsen, Mohamed; Strain, Matthew C.; Lada, Steven M.; Yukl, Steven; Cockerham, Leslie R.; Pilcher, Christopher D.; Hecht, Frederick M.; Sinclair, Elizabeth; Liegler, Teri; Richman, Douglas D.; Deeks, Steven G.; Pillai, Satish K.

2014-01-01

Individuals who are heterozygous for the CCR5-Δ32 mutation provide a natural model to examine the effects of reduced CCR5 expression on human immunodeficiency virus (HIV) persistence. We evaluated the HIV reservoir in 18 CCR5-Δ32 heterozygotes and 54 CCR5 wild-type individuals during suppressive antiretroviral therapy. Cell-associated HIV RNA levels (P = .035), RNA to DNA transcriptional ratios (P = .013), and frequency of detectable HIV 2–long terminal repeat circular DNA (P = .013) were significantly lower in CD4+ T cells from CCR5-Δ32 heterozygotes. Cell-associated HIV RNA was significantly correlated with CCR5 surface expression on CD4+ T cells (r2 = 0.136; P = .002). Our findings suggest that curative strategies should further explore manipulation of CCR5. PMID:24935955

CCR2-64I polymorphism is not associated with altered CCR5 expression or coreceptor function.

PubMed

Mariani, R; Wong, S; Mulder, L C; Wilkinson, D A; Reinhart, A L; LaRosa, G; Nibbs, R; O'Brien, T R; Michael, N L; Connor, R I; Macdonald, M; Busch, M; Koup, R A; Landau, N R

1999-03-01

A polymorphism in the gene encoding CCR2 is associated with a delay in progression to AIDS in human immunodeficiency virus (HIV)-infected individuals. The polymorphism, CCR2-64I, changes valine 64 of CCR2 to isoleucine. However, it is not clear whether the effect on AIDS progression results from the amino acid change or whether the polymorphism marks a genetically linked, yet unidentified mutation that mediates the effect. Because the gene encoding CCR5, the major coreceptor for HIV type 1 primary isolates, lies 15 kb 3' to CCR2, linked mutations in the CCR5 promoter or other regulatory sequences could explain the association of CCR2-64I with slowed AIDS pathogenesis. Here, we show that CCR2-64I is efficiently expressed on the cell surface but does not have dominant negative activity on CCR5 coreceptor function. A panel of peripheral blood mononuclear cells (PBMC) from uninfected donors representing the various CCR5/CCR2 genotypes was assembled. Activated primary CD4(+) T cells of CCR2 64I/64I donors expressed cell surface CCR5 at levels comparable to those of CCR2 +/+ donors. A slight reduction in CCR5 expression was noted, although this was not statistically significant. CCR5 and CCR2 mRNA levels were nearly identical for each of the donor PBMC, regardless of genotype. Cell surface CCR5 and CCR2 levels were more variable than mRNA transcript levels, suggesting that an alternative mechanism may influence CCR5 cell surface levels. CCR2-64I is linked to the CCR5 promoter polymorphisms 208G, 303A, 627C, and 676A; however, in transfected promoter reporter constructs, these did not affect transcriptional activity. Taken together, these findings suggest that CCR2-64I does not act by influencing CCR5 transcription or mRNA levels.

Vaccination with live attenuated simian immunodeficiency virus causes dynamic changes in intestinal CD4+CCR5+ T cells

PubMed Central

2011-01-01

Background Vaccination with live attenuated SIV can protect against detectable infection with wild-type virus. We have investigated whether target cell depletion contributes to the protection observed. Following vaccination with live attenuated SIV the frequency of intestinal CD4+CCR5+ T cells, an early target of wild-type SIV infection and destruction, was determined at days 3, 7, 10, 21 and 125 post inoculation. Results In naive controls, modest frequencies of intestinal CD4+CCR5+ T cells were predominantly found within the LPL TTrM-1 and IEL TTrM-2 subsets. At day 3, LPL and IEL CD4+CCR5+ TEM cells were dramatically increased whilst less differentiated subsets were greatly reduced, consistent with activation-induced maturation. CCR5 expression remained high at day 7, although there was a shift in subset balance from CD4+CCR5+ TEM to less differentiated TTrM-2 cells. This increase in intestinal CD4+CCR5+ T cells preceded the peak of SIV RNA plasma loads measured at day 10. Greater than 65.9% depletion of intestinal CD4+CCR5+ T cells followed at day 10, but overall CD4+ T cell homeostasis was maintained by increased CD4+CCR5- T cells. At days 21 and 125, high numbers of intestinal CD4+CCR5- naive TN cells were detected concurrent with greatly increased CD4+CCR5+ LPL TTrM-2 and IEL TEM cells at day 125, yet SIV RNA plasma loads remained low. Conclusions This increase in intestinal CD4+CCR5+ T cells, following vaccination with live attenuated SIV, does not correlate with target cell depletion as a mechanism of protection. Instead, increased intestinal CD4+CCR5+ T cells may correlate with or contribute to the protection conferred by vaccination with live attenuated SIV. PMID:21291552

CCR5 Gene Disruption via Lentiviral Vectors Expressing Cas9 and Single Guided RNA Renders Cells Resistant to HIV-1 Infection

PubMed Central

Liu, Jingjing; Zhang, Di; Kimata, Jason T.; Zhou, Paul

2014-01-01

CCR5, a coreceptor for HIV-1 entry, is a major target for drug and genetic intervention against HIV-1. Genetic intervention strategies have knocked down CCR5 expression levels by shRNA or disrupted the CCR5 gene using zinc finger nucleases (ZFN) or Transcription activator-like effector nuclease (TALEN). In the present study, we silenced CCR5 via CRISPR associated protein 9 (Cas9) and single guided RNAs (sgRNAs). We constructed lentiviral vectors expressing Cas9 and CCR5 sgRNAs. We show that a single round transduction of lentiviral vectors expressing Cas9 and CCR5 sgRNAs into HIV-1 susceptible human CD4+ cells yields high frequencies of CCR5 gene disruption. CCR5 gene-disrupted cells are not only resistant to R5-tropic HIV-1, including transmitted/founder (T/F) HIV-1 isolates, but also have selective advantage over CCR5 gene-undisrupted cells during R5-tropic HIV-1 infection. Importantly, using T7 endonuclease I assay we did not detect genome mutations at potential off-target sites that are highly homologous to these CCR5 sgRNAs in stably transduced cells even at 84 days post transduction. Thus we conclude that silencing of CCR5 via Cas9 and CCR5-specific sgRNAs could be a viable alternative strategy for engineering resistance against HIV-1. PMID:25541967

Evolution of the Human Immunodeficiency Virus Type 1 Envelope during Infection Reveals Molecular Corollaries of Specificity for Coreceptor Utilization and AIDS Pathogenesis

PubMed Central

Hu, Qin-xue; Barry, Ashley Perkins; Wang, Zi-xuan; Connolly, Shanon M.; Peiper, Stephen C.; Greenberg, Michael L.

2000-01-01

The evolution of human immunodeficiency virus type 1 infection is associated with a shift in the target cell population, driven by variability in coreceptor utilization resulting from diversity in env. To elucidate the potential consequences of these changes for Env-mediated fusion over the course of AIDS, we examined the biological properties of serial viral isolates and determined coreceptor utilization by the products of env cloned from two individuals, followed from the detection of seroconversion throughout the course of their infection. One had a typical course, and the other had an accelerated progression. Early isolates were non-syncytium inducing, and the corresponding Env exclusively utilized CCR5, whereas Env from late phases of infection showed restricted utilization of CXCR4 in both patients. Env from subject SC24, who had a standard progression, demonstrated multitropism, manifested by utilization of CCR3, CXCR4, and CCR5 in the intervening period. In contrast, Env from patient SC51, who experienced early conversion to the syncytium-inducing phenotype, developed dualtropic coreceptor utilization of CCR5 and CXCR4. Genetic analysis of env from each isolate revealed that those with an X4 phenotype formed a distinct subcluster within each subject. Analysis of chimeras constructed from R5 and multispecific env from patient SC24 demonstrated that while the V3 domain played a dominant role in determining coreceptor utilization, sequences in the V4–V5 region also contributed to the latter phenotype. Immunoprecipitation experiments confirmed that the hybrid Env proteins were expressed at similar levels. These experiments demonstrate that progression from the R5 to X4 phenotype may occur through a multi- or dual-tropic intermediate and that multiple domains contribute to this process. PMID:11090186

CXCR6 and CCR5 localize T lymphocyte subsets in nasopharyngeal carcinoma.

PubMed

Parsonage, Greg; Machado, Lee Richard; Hui, Jan Wai-Ying; McLarnon, Andrew; Schmaler, Tilo; Balasothy, Meenarani; To, Ka-Fai; Vlantis, Alexander C; van Hasselt, Charles A; Lo, Kwok-Wai; Wong, Wai-Lap; Hui, Edwin Pun; Chan, Anthony Tak Cheung; Lee, Steven P

2012-03-01

The substantial T lymphocyte infiltrate found in cases of nasopharyngeal carcinoma (NPC) has been implicated in the promotion of both tumor growth and immune escape. Conversely, because malignant NPC cells harbor the Epstein-Barr virus, this tumor is a candidate for virus-specific T cell-based therapies. Preventing the accumulation of tumor-promoting T cells or enhancing the recruitment of tumor-specific cytotoxic T cells offers therapeutic potential. However, the mechanisms involved in T cell recruitment to this tumor are poorly understood. Comparing memory T cell subsets that have naturally infiltrated NPC tissue with their counterparts from matched blood revealed enrichment of CD8(+), CD4(+), and regulatory T cells expressing the chemokine receptor CXCR6 in tumor tissue. CD8(+) and (nonregulatory) CD4(+) T cells also were more frequently CCR5(+) in tumor than in blood. Ex vivo studies demonstrated that both receptors were functional. CXCL16 and CCL4, unique chemokine ligands for CXCR6 and CCR5, respectively, were expressed by the malignant cells in tumor tissue from the majority of NPC cases, as was another CCR5 ligand, CCL5. The strongest expression of CXCL16 was found on tumor-infiltrating cells. CCL4 was detected on the tumor vasculature in a majority of cases. These findings suggest that CXCR6 and CCR5 play important roles in T cell recruitment and/or retention in NPC and have implications for the pathogenesis and treatment of this tumor. Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Simian immunodeficiency virus SIVmac239 infection and simian human immunodeficiency virus SHIV89.6P infection result in progression to AIDS in cynomolgus macaques of Asian origin.

PubMed

Okamura, Tomotaka; Tsujimura, Yusuke; Soma, Shogo; Takahashi, Ichiro; Matsuo, Kazuhiro; Yasutomi, Yasuhiro

2016-12-01

Simian immunodeficiency virus (SIV) infection models in cynomolgus macaques are important for analysis of the pathogenesis of immunodeficiency virus and for studies on the efficacy of new vaccine candidates. However, very little is known about the pathogenesis of SIV or simian human immunodeficiency virus (SHIV) in cynomolgus macaques from different Asian countries. In the present study, we analysed the infectivity and pathogenicity of CCR5-tropic SIVmac and those of dual-tropic SHIV89.6P inoculated into cynomolgus macaques in Indonesian, Malaysian or Philippine origin. The plasma viral loads in macaques infected with either SIVmac239 or SHIV89.6P were maintained at high levels. CD4+ T cell levels in macaques infected with SIVmac239 gradually decreased. All of the macaques infected with SHIV89.6P showed greatly reduced CD4+ T-cell numbers within 6 weeks of infection. Eight of the 11 macaques infected with SIVmac239 were killed due to AIDS symptoms after 2-4.5 years, while four of the five macaques infected with SHIV89.6P were killed due to AIDS symptoms after 1-3.5 years. We also analysed cynomolgus macaques infected intrarectally with repeated low, medium or high doses of SIVmac239, SIVmac251 or SHIV89.6P. Infection was confirmed by quantitative RT-PCR at more than 5000, 300 and 500 TCID50 for SIVmac239, SIVmac251 and SHIV89.6P, respectively. The present study indicates that cynomolgus macaques of Asian origin are highly susceptible to SIVmac and SHIV infection by both intravenous and mucosal routes. These models will be useful for studies on virus pathogenesis, vaccination and therapeutics against human immunodeficiency virus/AIDS.

CCL2, but not its receptor, is essential to restrict immune privileged central nervous system-invasion of Japanese encephalitis virus via regulating accumulation of CD11b(+) Ly-6C(hi) monocytes.

PubMed

Kim, Jin Hyoung; Patil, Ajit Mahadev; Choi, Jin Young; Kim, Seong Bum; Uyangaa, Erdenebileg; Hossain, Ferdaus Mohd Altaf; Park, Sang-Youel; Lee, John Hwa; Kim, Koanhoi; Eo, Seong Kug

2016-10-01

Japanese encephalitis virus (JEV) is a re-emerging zoonotic flavivirus that poses an increasing threat to global health and welfare due to rapid changes in climate and demography. Although the CCR2-CCL2 axis plays an important role in trafficking CD11b(+) Ly-6C(hi) monocytes to regulate immunopathological diseases, little is known about their role in monocyte trafficking during viral encephalitis caused by JEV infection. Here, we explored the role of CCR2 and its ligand CCL2 in JE caused by JEV infection using CCR2- and CCL2-ablated murine models. Somewhat surprisingly, the ablation of CCR2 and CCL2 resulted in starkly contrasting susceptibility to JE. CCR2 ablation induced enhanced resistance to JE, whereas CCL2 ablation highly increased susceptibility to JE. This contrasting regulation of JE progression by CCR2 and CCL2 was coupled to central nervous system (CNS) infiltration of Ly-6C(hi) monocytes and Ly-6G(hi) granulocytes. There was also enhanced expression of CC and CXC chemokines in the CNS of CCL2-ablated mice, which appeared to induce CNS infiltration of these cell populations. However, our data revealed that contrasting regulation of JE in CCR2- and CCL2-ablated mice was unlikely to be mediated by innate natural killer and adaptive T-cell responses. Furthermore, CCL2 produced by haematopoietic stem cell-derived leucocytes played a dominant role in CNS accumulation of Ly-6C(hi) monocytes in infected bone marrow chimeric models, thereby exacerbating JE progression. Collectively, our data indicate that CCL2 plays an essential role in conferring protection against JE caused by JEV infection. In addition, blockage of CCR2, but not CCL2, will aid in the development of strategies for prophylactics and therapeutics of JE. © 2016 John Wiley & Sons Ltd.

Antigen-driven C–C Chemokine-mediated HIV-1 Suppression by CD4+ T Cells from Exposed Uninfected Individuals Expressing the Wild-type CCR-5 Allele

PubMed Central

Furci, Lucinda; Scarlatti, Gabriella; Burastero, Samuele; Tambussi, Giuseppe; Colognesi, Claudia; Quillent, Caroline; Longhi, Renato; Loverro, Patrizia; Borgonovo, Barbara; Gaffi, Davide; Carrow, Emily; Malnati, Mauro; Lusso, Paolo; Siccardi, Antonio G.; Lazzarin, Adriano; Beretta, Alberto

1997-01-01

Despite repeated exposure to HIV-1, certain individuals remain persistently uninfected. Such exposed uninfected (EU) people show evidence of HIV-1–specific T cell immunity and, in rare cases, selective resistance to infection by macrophage-tropic strains of HIV-1. The latter has been associated with a 32–base pair deletion in the C–C chemokine receptor gene CCR-5, the major coreceptor of macrophage-tropic strains of HIV-1. We have undertaken an analysis of the HIV-specific T cell responses in 12 EU individuals who were either homozygous for the wild-type CCR-5 allele or heterozygous for the deletion allele (CCR-5Δ32). We have found evidence of an oligoclonal T cell response mediated by helper T cells specific for a conserved region of the HIV-1 envelope. These cells produce very high levels of C–C chemokines when stimulated by the specific antigen and suppress selectively the replication of macrophage-tropic, but not T cell–tropic, strains of HIV-1. These chemokine-producing helper cells may be part of a protective immune response that could be potentially exploited for vaccine development. PMID:9236198

CCR5 Signal Transduction in Macrophages by Human Immunodeficiency Virus and Simian Immunodeficiency Virus Envelopes

PubMed Central

Arthos, James; Rubbert, Andrea; Rabin, Ronald L.; Cicala, Claudia; Machado, Elizabeth; Wildt, Kathryne; Hanbach, Meredith; Steenbeke, Tavis D.; Swofford, Ruth; Farber, Joshua M.; Fauci, Anthony S.

2000-01-01

The capacity of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelopes to transduce signals through chemokine coreceptors on macrophages was examined by measuring the ability of recombinant envelope proteins to mobilize intracellular calcium stores. Both HIV and SIV envelopes mobilized calcium via interactions with CCR5. The kinetics of these responses were similar to those observed when macrophages were treated with MIP-1β. Distinct differences in the capacity of envelopes to mediate calcium mobilization were observed. Envelopes derived from viruses capable of replicating in macrophages mobilized relatively high levels of calcium, while envelopes derived from viruses incapable of replicating in macrophages mobilized relatively low levels of calcium. The failure to efficiently mobilize calcium was not restricted to envelopes derived from CXCR4-utilizing isolates but also included envelopes derived from CCR5-utilizing isolates that fail to replicate in macrophages. We characterized one CCR5-utilizing isolate, 92MW959, which entered macrophages but failed to replicate. A recombinant envelope derived from this virus mobilized low levels of calcium. When macrophages were inoculated with 92MW959 in the presence of MIP-1α, viral replication was observed, indicating that a CC chemokine-mediated signal provided the necessary stimulus to allow the virus to complete its replication cycle. Although the role that envelope-CCR5 signal transduction plays in viral replication is not yet understood, it has been suggested that envelope-mediated signals facilitate early postfusion events in viral replication. The data presented here are consistent with this hypothesis and suggest that the differential capacity of viral envelopes to signal through CCR5 may influence their ability to replicate in macrophages. PMID:10864653

CCR5 signal transduction in macrophages by human immunodeficiency virus and simian immunodeficiency virus envelopes.

PubMed

Arthos, J; Rubbert, A; Rabin, R L; Cicala, C; Machado, E; Wildt, K; Hanbach, M; Steenbeke, T D; Swofford, R; Farber, J M; Fauci, A S

2000-07-01

The capacity of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelopes to transduce signals through chemokine coreceptors on macrophages was examined by measuring the ability of recombinant envelope proteins to mobilize intracellular calcium stores. Both HIV and SIV envelopes mobilized calcium via interactions with CCR5. The kinetics of these responses were similar to those observed when macrophages were treated with MIP-1beta. Distinct differences in the capacity of envelopes to mediate calcium mobilization were observed. Envelopes derived from viruses capable of replicating in macrophages mobilized relatively high levels of calcium, while envelopes derived from viruses incapable of replicating in macrophages mobilized relatively low levels of calcium. The failure to efficiently mobilize calcium was not restricted to envelopes derived from CXCR4-utilizing isolates but also included envelopes derived from CCR5-utilizing isolates that fail to replicate in macrophages. We characterized one CCR5-utilizing isolate, 92MW959, which entered macrophages but failed to replicate. A recombinant envelope derived from this virus mobilized low levels of calcium. When macrophages were inoculated with 92MW959 in the presence of MIP-1alpha, viral replication was observed, indicating that a CC chemokine-mediated signal provided the necessary stimulus to allow the virus to complete its replication cycle. Although the role that envelope-CCR5 signal transduction plays in viral replication is not yet understood, it has been suggested that envelope-mediated signals facilitate early postfusion events in viral replication. The data presented here are consistent with this hypothesis and suggest that the differential capacity of viral envelopes to signal through CCR5 may influence their ability to replicate in macrophages.

HIV-1 Clinical Isolates Resistant to CCR5 Antagonists Exhibit Delayed Entry Kinetics That Are Corrected in the Presence of Drug

PubMed Central

Putcharoen, Opass; Lee, Sun Hee; Henrich, Timothy J.; Hu, Zixin; Vanichanan, Jakapat; Coakley, Eoin; Greaves, Wayne; Gulick, Roy M.; Kuritzkes, Daniel R.

2012-01-01

HIV CCR5 antagonists select for env gene mutations that enable virus entry via drug-bound coreceptor. To investigate the mechanisms responsible for viral adaptation to drug-bound coreceptor-mediated entry, we studied viral isolates from three participants who developed CCR5 antagonist resistance during treatment with vicriviroc (VCV), an investigational small-molecule CCR5 antagonist. VCV-sensitive and -resistant viruses were isolated from one HIV subtype C- and two subtype B-infected participants; VCV-resistant isolates had mutations in the V3 loop of gp120 and were cross-resistant to TAK-779, an investigational antagonist, and maraviroc (MVC). All three resistant isolates contained a 306P mutation but had variable mutations elsewhere in the V3 stem. We used a virus-cell β-lactamase (BlaM) fusion assay to determine the entry kinetics of recombinant viruses that incorporated full-length VCV-sensitive and -resistant envelopes. VCV-resistant isolates exhibited delayed entry rates in the absence of drug, relative to pretherapy VCV-sensitive isolates. The addition of drug corrected these delays. These findings were generalizable across target cell types with a range of CD4 and CCR5 surface densities and were observed when either population-derived or clonal envelopes were used to construct recombinant viruses. V3 loop mutations alone were sufficient to restore virus entry in the presence of drug, and the accumulation of V3 mutations during VCV therapy led to progressively higher rates of viral entry. We propose that the restoration of pre-CCR5 antagonist therapy HIV entry kinetics drives the selection of V3 loop mutations and may represent a common mechanism that underlies the emergence of CCR5 antagonist resistance. PMID:22090117

HIV type 1 chemokine receptor usage in mother-to-child transmission.

PubMed

Salvatori, F; Scarlatti, G

2001-07-01

To investigate the role of the HIV-1 phenotype in mother-to-child HIV-1 transmission, we evaluated coreceptor usage and replication kinetics in chemokine receptor-expressing U87MG.CD4 cells of primary isolates from 32 HIV-1-infected mothers of Italian origin, none under preventive antiretroviral therapy, and from their infected infants. Five of 15 mothers of infected children and 2 of 17 mothers of uninfected children harbored viruses able to use CXCR4 as coreceptor. However, all isolates used CCR5, alone or in association with CXCR4. The replicative capacity in coreceptor-expressing cells of the viral isolates did not differ between the two groups of mothers. All mothers with an R5 virus transmitted a virus with the same coreceptor usage, whereas those four with a multitropic virus transmitted such a virus in one case. Although the presence of a mixed viral population was documented in the mothers, we did not observe transmission solely of X4 viruses. Interestingly, the only child infected with a multitropic virus carried a defective CCR5 allele. Analysis of the env V3 region of the provirus from this child revealed infection with multiple viral variants with a predominance of R5-type over X4-type sequences. These findings show that CCR5 usage of a viral isolate is not a discriminating risk factor for vertical transmission. Furthermore, X4 viruses can be transmitted to the newborn, although less frequently. In particular, we document the transmission of multiple viral variants with different coreceptor usage in a Delta32 CCR5 heterozygous child, and demonstrate that the heterozygous genotype per se does not contribute to the restriction of R5-type virus spread.

In vivo emergence of vicriviroc resistance in a human immunodeficiency virus type 1 subtype C-infected subject.

PubMed

Tsibris, Athe M N; Sagar, Manish; Gulick, Roy M; Su, Zhaohui; Hughes, Michael; Greaves, Wayne; Subramanian, Mani; Flexner, Charles; Giguel, Françoise; Leopold, Kay E; Coakley, Eoin; Kuritzkes, Daniel R

2008-08-01

Little is known about the in vivo development of resistance to human immunodeficiency virus type 1 (HIV-1) CCR5 antagonists. We studied 29 subjects with virologic failure from a phase IIb study of the CCR5 antagonist vicriviroc (VCV) and identified one individual with HIV-1 subtype C who developed VCV resistance. Studies with chimeric envelopes demonstrated that changes within the V3 loop were sufficient to confer VCV resistance. Resistant virus showed VCV-enhanced replication, cross-resistance to another CCR5 antagonist, TAK779, and increased sensitivity to aminooxypentane-RANTES and the CCR5 monoclonal antibody HGS004. Pretreatment V3 loop sequences reemerged following VCV discontinuation, implying that VCV resistance has associated fitness costs.

Induction of Simian AIDS in Infant Rhesus Macaques Infected with CCR5- or CXCR4-Utilizing Simian-Human Immunodeficiency Viruses Is Associated with Distinct Lesions of the Thymus

PubMed Central

Reyes, R. A.; Canfield, Don R.; Esser, Ursula; Adamson, Lourdes A.; Brown, Charles R.; Cheng-Mayer, Cecilia; Gardner, Murray B.; Harouse, Janet M.; Luciw, Paul A.

2004-01-01

Newborn rhesus macaques were infected with two chimeric simian-human immunodeficiency virus (SHIV) strains which contain unique human immunodeficiency virus type 1 (HIV-1) env genes and exhibit distinct phenotypes. Infection with either the CCR5-specific SHIVSF162P3 or the CXCR4-utilizing SHIVSF33A resulted in clinical manifestations consistent with simian AIDS. Most prominent in this study was the detection of severe thymic involution in all SHIVSF33A-infected infants, which is very similar to HIV-1-induced thymic dysfunction in children who exhibit a rapid pattern of disease progression. In contrast, SHIVSF162P3 induced only a minor disruption in thymic morphology. Consistent with the distribution of the coreceptors CXCR4 and CCR5 within the thymus, the expression of SHIVSF162P3 was restricted to the thymic medulla, whereas SHIVSF33A was preferentially detected in the cortex. This dichotomy of tissue tropism is similar to the differential tropism of HIV-1 isolates observed in the reconstituted human thymus in SCID-hu mice. Accordingly, our results show that the SHIV-monkey model can be used for the molecular dissection of cell and tissue tropisms controlled by the HIV-1 env gene and for the analysis of mechanisms of viral immunopathogenesis in AIDS. Furthermore, these findings could help explain the rapid progression of disease observed in some HIV-1-infected children. PMID:14747577

Analysis of HIV tropism in Ugandan infants

PubMed Central

Church, Jessica D.; Huang, Wei; Mwatha, Anthony; Musoke, Philippa; Jackson, J. Brooks; Bagenda, Danstan; Omer, Saad B.; Donnell, Deborah; Nakabiito, Clemensia; Eure, Chineta; Guay, Laura A.; Taylor, Allan; Bakaki, Paul M.; Matovu, Flavia; McConnell, Michelle; Fowler, Mary Glenn; Eshleman, Susan H.

2010-01-01

HIV-infected infants may have CXCR4-using (X4-tropic) HIV, CCR5-using (R5-tropic) HIV, or a mixture of R5-tropic and X4-tropic HIV (dual/mixed, DM HIV). The level of infectivity for R5 virus (R5-RLU) varies among HIV-infected infants. HIV tropism and R5-RLU were measured in samples from HIV-infected Ugandan infants using a commercial assay. DM HIV was detected in 7/72 (9.7%) infants at the time of HIV diagnosis (birth or 6–8 weeks of age, 4/15 (26.7%) with subtype D, 3/57 (5.3 %) with other subtypes, P=0.013). A transition from R5-tropic to DM HIV was observed in only two (6.7%) of 30 infants over 6–12 months. Six (85.7%) of seven infants with DM HIV died, compared to 21/67 (31.3%) infants with R5-tropic HIV (p=0.09). Higher R5-RLU at 6–8 weeks was not associated with decreased survival. Infants with in utero infection had a higher median R5-RLU than infants who were HIV-uninfected at birth (p=0.025). PMID:21073438

HIV Transmission

PubMed Central

Shaw, George M.; Hunter, Eric

2012-01-01

HIV-1 is transmitted by sexual contact across mucosal surfaces, by maternal-infant exposure, and by percutaneous inoculation. For reasons that are still incompletely understood, CCR5-tropic viruses (R5 viruses) are preferentially transmitted by all routes. Transmission is followed by an orderly appearance of viral and host markers of infection in the blood plasma. In the acute phase of infection, HIV-1 replicates exponentially and diversifies randomly, allowing for an unambiguous molecular identification of transmitted/founder virus genomes and a precise characterization of the population bottleneck to virus transmission. Sexual transmission of HIV-1 most often results in productive clinical infection arising from a single virus, highlighting the extreme bottleneck and inherent inefficiency in virus transmission. It remains to be determined if HIV-1 transmission is largely a stochastic process whereby any reasonably fit R5 virus can be transmitted or if there are features of transmitted/founder viruses that facilitate their transmission in a biologically meaningful way. Human tissue explant models of HIV-1 infection and animal models of SIV/SHIV/HIV-1 transmission, coupled with new challenge virus strains that more closely reflect transmitted/founder viruses, have the potential to elucidate fundamental mechanisms in HIV-1 transmission relevant to vaccine design and other prevention strategies. PMID:23043157

Influence of the CCR-5/MIP-1 α Axis in the Pathogenesis of Rocio Virus Encephalitis in a Mouse Model

PubMed Central

Chávez, Juliana H.; França, Rafael F. O.; Oliveira, Carlo J. F.; de Aquino, Maria T. P.; Farias, Kleber J. S.; Machado, Paula R. L.; de Oliveira, Thelma F. M.; Yokosawa, Jonny; Soares, Edson G.; da Silva, João S.; da Fonseca, Benedito A. L.; Figueiredo, Luiz T. M.

2013-01-01

Rocio virus (ROCV) caused an outbreak of human encephalitis during the 1970s in Brazil and its immunopathogenesis remains poorly understood. CC-chemokine receptor 5 (CCR5) is a chemokine receptor that binds to macrophage inflammatory protein (MIP-1 α). Both molecules are associated with inflammatory cells migration during infections. In this study, we demonstrated the importance of the CCR5 and MIP-1 α, in the outcome of viral encephalitis of ROCV-infected mice. CCR5 and MIP-1 α knockout mice survived longer than wild-type (WT) ROCV-infected animals. In addition, knockout mice had reduced inflammation in the brain. Assessment of brain viral load showed mice virus detection five days post-infection in wild-type and CCR5−/− mice, while MIP-1 α−/− mice had lower viral loads seven days post-infection. Knockout mice required a higher lethal dose than wild-type mice as well. The CCR5/MIP-1 α axis may contribute to migration of infected cells to the brain and consequently affect the pathogenesis during ROCV infection. PMID:24080631

Resistance to human immunodeficiency virus type 1 (HIV-1) generated by lentivirus vector-mediated delivery of the CCR5Δ32 gene despite detectable expression of the HIV-1 co-receptors

PubMed Central

Jin, Qingwen; Marsh, Jon; Cornetta, Kenneth; Alkhatib, Ghalib

2009-01-01

It has previously been demonstrated that there are two distinct mechanisms for genetic resistance to human immunodeficiency virus type 1 (HIV-1) conferred by the CCR5Δ32 gene: the loss of wild-type CCR5 surface expression and the generation of CCR5Δ32 protein, which interacts with CXCR4. To analyse the protective effects of long-term expression of the CCR5Δ32 protein, recombinant lentiviral vectors were used to deliver the CCR5Δ32 gene into human cell lines and primary peripheral blood mononuclear cells that had been immortalized by human T-cell leukemia virus type 1. Blasticidin S-resistant cell lines expressing the lentivirus-encoded CCR5Δ32 showed a significant reduction in HIV-1 Env-mediated fusion assays. It was shown that CD4+ T lymphocytes expressing the lentivirus-encoded CCR5Δ32 gene were highly resistant to infection by a primary but not by a laboratory-adapted X4 strain, suggesting different infectivity requirements. In contrast to previous studies that analysed the CCR5Δ32 protective effects in a transient expression system, this study showed that long-term expression of CCR5Δ32 conferred resistance to HIV-1 despite cell-surface expression of the HIV co-receptors. The results suggest an additional unknown mechanism for generating the CCR5Δ32 resistance phenotype and support the hypothesis that the CCR5Δ32 protein acts as an HIV-suppressive factor by altering the stoichiometry of the molecules involved in HIV-1 entry. The lentiviral-CCR5Δ32 vectors offer a method of generating HIV-resistant cells by delivery of the CCR5Δ32 gene that may be useful for stem cell- or T-cell-based gene therapy for HIV-1 infection. PMID:18796731

MCP-1 and CCR2 Contribute to Non-Lymphocyte-Mediated Brain Disease Induced by Fr98 Polytropic Retrovirus Infection in Mice: Role for Astrocytes in Retroviral Neuropathogenesis

PubMed Central

Peterson, Karin E.; Errett, John S.; Wei, Tao; Dimcheff, Derek E.; Ransohoff, Richard; Kuziel, William A.; Evans, Leonard; Chesebro, Bruce

2004-01-01

Virus infection of the central nervous system (CNS) often results in chemokine upregulation. Although often associated with lymphocyte recruitment, increased chemokine expression is also associated with non-lymphocyte-mediated CNS disease. In these instances, the effect of chemokine upregulation on neurological disease is unclear. In vitro, several chemokines including monocyte chemotactic protein 1 (MCP-1) protect neurons from apoptosis. Therefore, in vivo, chemokine upregulation may be a protective host response to CNS damage. Alternatively, chemokines may contribute to pathogenesis by stimulating intrinsic brain cells or recruiting macrophages to the brain. To investigate these possibilities, we studied a neurovirulent retrovirus, Fr98, that induces severe non-lymphocyte-mediated neurological disease and causes the upregulation of several chemokines that bind to chemokine receptors CCR2 and CCR5. Knockout mice deficient in CCR2 had reduced susceptibility to Fr98 pathogenesis, with significantly fewer mice developing clinical disease than did wild-type controls. In contrast, no reduction in Fr98-induced disease was observed in CCR5 knockout mice. Thus, signaling through CCR2, but not CCR5, plays an important role in Fr98-mediated pathogenesis. Three ligands for CCR2 (MCP-1, MCP-3, and MCP-5) were upregulated during Fr98 infection of the brain. Antibody-blocking experiments demonstrated that MCP-1 was important for retrovirus-induced neurological disease. In situ hybridization analysis revealed that MCP-1 was expressed by glial fibrillary acidic protein-positive astrocytes. Thus, astrocytes, previously not thought to play an effector role in the disease process were found to contribute to pathogenesis through the production of MCP-1. This study also demonstrates that chemokines can mediate pathogenesis in the CNS in the absence of lymphocytic infiltrate and gives credence to the hypothesis that chemokine upregulation is a mechanism by which retroviruses such as human immunodeficiency virus induce neurological damage. PMID:15163738

M- and T-tropic HIVs Promote Apoptosis in Rat Neurons

PubMed Central

Bachis, Alessia; Biggio, Francesca; Major, Eugene O.; Mocchetti, Italo

2009-01-01

Neuronal loss, reactive astrocytes, and other abnormalities are seen in the brain of individuals with acquired immune deficiency syndrome-associated Dementia Complex (ADC). Human immunodeficiency virus-1 (HIV-1) is believed to be the main agent causing ADC. However, little is known about the molecular and cellular mechanisms of HIV-1 neurotoxicity considering that HIV-1 does not infect post-mitotic neurons and that viral load does not necessarily correlate with ADC. Various viral proteins, such as the envelope protein gp120 and the transcription activator Tat, have been shown to induce neuronal apoptosis through direct and indirect mechanisms both in vitro and in vivo. Progeny HIV-1 virions can also cause neuronal death. However, it has not been fully established yet whether HIV-1 promotes neuronal apoptosis by a direct mechanism. To explore the neurotoxic effect of HIV-1, we exposed rat cerebellar granule cells and cortical neurons in culture to two different strains of HIV-1, IIIB and BaL, T- and M-tropic strains that utilize CXCR4 and CCR5 coreceptors, respectively, to infect cells. We observed that both viruses elicit a time-dependent apoptotic cell death in these cultures without inducing a productive infection as determined by the absence of the core protein of HIV-1, p24, in cell lysates. Instead, neurons were gp 120 positive, suggesting that the envelope protein is shed by the virus and then subsequently internalized by neurons. The CXCR4 receptor antagonist AMD3100 or the CCR5 receptor inhibitor D-Ala-peptide T-amide blocked HIV IIIB and HIV Bal neurotoxicity, respectively. In contrast, the N-methyl-D-aspartate receptor blocker MK801 failed to protect neurons from HIV-mediated apoptosis, suggesting that HIV-1 neurotoxicity can be initiated by the viral protein gp 120 binding to neuronal chemokine receptors. PMID:19034668

Microbiota induces tonic CCL2 systemic levels that control pDC trafficking in steady state.

PubMed

Swiecki, M; Miller, H L; Sesti-Costa, R; Cella, M; Gilfillan, S; Colonna, M

2017-07-01

Plasmacytoid dendritic cells (pDCs) detect viruses initiating antiviral type I interferon responses. The microbiota is known to shape immune responses, but whether it influences pDC homeostasis and/or function is poorly understood. By comparing pDCs in germ-free and specific pathogen-free mice, we found that the microbiota supports homeostatic trafficking by eliciting constitutive levels of the chemokine CCL2 that engages CCR2. Mononuclear phagocytes were required for tonic CCL2 levels. CCL2 was particularly important for trafficking of a CCR2 hi subset of pDCs that produced proinflammatory cytokines and was prone to apoptosis. We further demonstrated that CCR2 was also essential for pDC migration during inflammation. Wild-type (WT):Ccr2 -/- mixed bone marrow chimeras revealed that CCR2 promotes pDC migration in a cell-intrinsic manner. Overall, we identify a novel role for the microbiota in shaping immunity, which includes induction of CCL2 levels that control homeostatic trafficking of pDCs.

Phenotype Variation in Human Immunodeficiency virus Type 1 Transmission and Disease Progression

PubMed Central

Cavarelli, Mariangela; Scarlatti, Gabriella

2009-01-01

Human immunodeficiency virus type I (HIV-1) infects target cells through interaction with the CD4 molecule and chemokine receptors, mainly CCR5 and CXCR4. Viral isolates can be phenotypically classified based on the co-receptor they utilize to infect target cells. Thus, R5 and X4 virus use respectively CCR5 and CXCR4, whereas R5X4 virus can use either CCR5 or CXCR4. This review describes the central role played by co-receptor expression and usage for HIV-1 cell tropism, transmission and pathogenesis. We discuss various hypotheses proposed to explain the preferential transmission of R5 viruses and the mechanisms driving the change of HIV-1 co-receptor usage in the course of infection. Recent insights in the intrinsic variability of R5 viruses and their role in influencing disease progression in both adults and children are also discussed. PMID:19893208

Phenotype variation in human immunodeficiency virus type 1 transmission and disease progression.

PubMed

Cavarelli, Mariangela; Scarlatti, Gabriella

2009-01-01

Human immunodeficiency virus type I (HIV-1) infects target cells through interaction with the CD4 molecule and chemokine receptors, mainly CCR5 and CXCR4. Viral isolates can be phenotypically classified based on the co-receptor they utilize to infect target cells. Thus, R5 and X4 virus use respectively CCR5 and CXCR4, whereas R5X4 virus can use either CCR5 or CXCR4. This review describes the central role played by co-receptor expression and usage for HIV-1 cell tropism, transmission and pathogenesis. We discuss various hypotheses proposed to explain the preferential transmission of R5 viruses and the mechanisms driving the change of HIV-1 co-receptor usage in the course of infection. Recent insights in the intrinsic variability of R5 viruses and their role in influencing disease progression in both adults and children are also discussed.

A Novel CCR5 Mutation Common in Sooty Mangabeys Reveals SIVsmm Infection of CCR5-Null Natural Hosts and Efficient Alternative Coreceptor Use In Vivo

PubMed Central

Riddick, Nadeene E.; Hermann, Emilia A.; Loftin, Lamorris M.; Elliott, Sarah T.; Wey, Winston C.; Cervasi, Barbara; Taaffe, Jessica; Engram, Jessica C.; Li, Bing; Else, James G.; Li, Yingying; Hahn, Beatrice H.; Derdeyn, Cynthia A.; Sodora, Donald L.; Apetrei, Cristian; Paiardini, Mirko; Silvestri, Guido; Collman, Ronald G.

2010-01-01

In contrast to HIV infection in humans and SIV in macaques, SIV infection of natural hosts including sooty mangabeys (SM) is non-pathogenic despite robust virus replication. We identified a novel SM CCR5 allele containing a two base pair deletion (Δ2) encoding a truncated molecule that is not expressed on the cell surface and does not support SIV entry in vitro. The allele was present at a 26% frequency in a large SM colony, along with 3% for a CCR5Δ24 deletion allele that also abrogates surface expression. Overall, 8% of animals were homozygous for defective CCR5 alleles and 41% were heterozygous. The mutant allele was also present in wild SM in West Africa. CD8+ and CD4+ T cells displayed a gradient of CCR5 expression across genotype groups, which was highly significant for CD8+ cells. Remarkably, the prevalence of natural SIVsmm infection was not significantly different in animals lacking functional CCR5 compared to heterozygous and homozygous wild-type animals. Furthermore, animals lacking functional CCR5 had robust plasma viral loads, which were only modestly lower than wild-type animals. SIVsmm primary isolates infected both homozygous mutant and wild-type PBMC in a CCR5-independent manner in vitro, and Envs from both CCR5-null and wild-type infected animals used CXCR6, GPR15 and GPR1 in addition to CCR5 in transfected cells. These data clearly indicate that SIVsmm relies on CCR5-independent entry pathways in SM that are homozygous for defective CCR5 alleles and, while the extent of alternative coreceptor use in SM with CCR5 wild type alleles is uncertain, strongly suggest that SIVsmm tropism and host cell targeting in vivo is defined by the distribution and use of alternative entry pathways in addition to CCR5. SIVsmm entry through alternative pathways in vivo raises the possibility of novel CCR5-negative target cells that may be more expendable than CCR5+ cells and enable the virus to replicate efficiently without causing disease in the face of extremely restricted CCR5 expression seen in SM and several other natural host species. PMID:20865163

A novel CCR5 mutation common in sooty mangabeys reveals SIVsmm infection of CCR5-null natural hosts and efficient alternative coreceptor use in vivo.

PubMed

Riddick, Nadeene E; Hermann, Emilia A; Loftin, Lamorris M; Elliott, Sarah T; Wey, Winston C; Cervasi, Barbara; Taaffe, Jessica; Engram, Jessica C; Li, Bing; Else, James G; Li, Yingying; Hahn, Beatrice H; Derdeyn, Cynthia A; Sodora, Donald L; Apetrei, Cristian; Paiardini, Mirko; Silvestri, Guido; Collman, Ronald G

2010-08-26

In contrast to HIV infection in humans and SIV in macaques, SIV infection of natural hosts including sooty mangabeys (SM) is non-pathogenic despite robust virus replication. We identified a novel SM CCR5 allele containing a two base pair deletion (Δ2) encoding a truncated molecule that is not expressed on the cell surface and does not support SIV entry in vitro. The allele was present at a 26% frequency in a large SM colony, along with 3% for a CCR5Δ24 deletion allele that also abrogates surface expression. Overall, 8% of animals were homozygous for defective CCR5 alleles and 41% were heterozygous. The mutant allele was also present in wild SM in West Africa. CD8+ and CD4+ T cells displayed a gradient of CCR5 expression across genotype groups, which was highly significant for CD8+ cells. Remarkably, the prevalence of natural SIVsmm infection was not significantly different in animals lacking functional CCR5 compared to heterozygous and homozygous wild-type animals. Furthermore, animals lacking functional CCR5 had robust plasma viral loads, which were only modestly lower than wild-type animals. SIVsmm primary isolates infected both homozygous mutant and wild-type PBMC in a CCR5-independent manner in vitro, and Envs from both CCR5-null and wild-type infected animals used CXCR6, GPR15 and GPR1 in addition to CCR5 in transfected cells. These data clearly indicate that SIVsmm relies on CCR5-independent entry pathways in SM that are homozygous for defective CCR5 alleles and, while the extent of alternative coreceptor use in SM with CCR5 wild type alleles is uncertain, strongly suggest that SIVsmm tropism and host cell targeting in vivo is defined by the distribution and use of alternative entry pathways in addition to CCR5. SIVsmm entry through alternative pathways in vivo raises the possibility of novel CCR5-negative target cells that may be more expendable than CCR5+ cells and enable the virus to replicate efficiently without causing disease in the face of extremely restricted CCR5 expression seen in SM and several other natural host species.

Polymorphisms of CCL3L1/CCR5 genes and recurrence of hepatitis B in liver transplant recipients.

PubMed

Li, Hong; Xie, Hai-Yang; Zhou, Lin; Wang, Wei-Lin; Liang, Ting-Bo; Zhang, Min; Zheng, Shu-Sen

2011-12-01

The genetic diversity of chemokines and chemokine receptors has been associated with the outcome of hepatitis B virus infection. The aim of this study was to evaluate whether the copy number variation in the CCL3L1 gene and the polymorphisms of CCR5Δ32 and CCR5-2459A→G (rs1799987) are associated with recurrent hepatitis B in liver transplantation for hepatitis B virus infection-related end-stage liver disease. A total of 185 transplant recipients were enrolled in this study. The genomic DNA was extracted from whole blood, the copy number of the CCL3L1 gene was determined by a quantitative real-time PCR based assay, CCR5Δ32 was detected by a sizing PCR method, and a single-nucleotide polymorphism in CCR5-2459 was detected by restriction fragment length polymorphism PCR. No CCR5Δ32 mutation was detected in any of the individuals from China. Neither copy number variation nor polymorphism in CCR5-2459 was associated with post-transplant re-infection with hepatitis B virus. However, patients with fewer copies (<4) of the CCL3L1 gene compared with the population median in combination with the CCR5G allele had a significantly higher risk for recurrent hepatitis B (odds ratio=1.93, 95% CI: 1.00-3.69; P=0.047). Patients possessing the compound decreased functional genotype of both CCL3L1 and CCR5 genes might be more likely to have recurrence of hepatitis B after transplantation.

Concordant utilization of macrophage entry coreceptors by related variants within an HIV type 1 primary isolate viral swarm.

PubMed

Singh, A; Yi, Y; Isaacs, S N; Kolson, D L; Collman, R G

2001-07-01

There is considerable diversity among HIV-1 strains in terms of their ability to use entry coreceptors on macrophages, especially CXCR4, but it is not known whether virus-specific differences exist among related members of a viral swarm. Defining how entry coreceptors on primary target cells are utilized by the spectrum of HIV-1 variants that emerge in vivo is important for understanding the relationship between coreceptor selectivity and pathogenesis. HIV-1 89.6(PI) is a dual-tropic primary isolate, and the prototype 89.6-cloned R5X4 Env uses both CXCR4 and CCR5 on macrophages. We generated a panel of env clones from the 89.6(PI) quasispecies and found a mixture of R5, R5X4, and X4 variants on the basis of fusion and infection of coreceptor-transfected cell lines. Here we address the use of macrophage coreceptors by these related Envs by analyzing fusion and infection of primary monocyte-derived macrophages mediated specifically through each coreceptor. All R5X4 Envs utilized both CXCR4 and CCR5 on macrophages, while R5 variants used CCR5 only. One variant characterized in cell lines as X4 used both CXCR4 and CCR5 on macrophages. No Env variant fused with macrophages through alternative coreceptor pathways. Thus, there was heterogeneity in coreceptor use among the related Env variants, but use of each coreceptor specifically in macrophages was consistent among members of the viral swarm. Coreceptor use in transfected cells generally predicted use in primary macrophages, although for some Envs macrophages may be a more sensitive indicator of CCR5 use than transfected cell lines.

Engineering hematopoietic stem cells toward a functional cure of human immunodeficiency virus infection.

PubMed

Wang, Jianbin; Holmes, Michael C

2016-11-01

The battle with human immunodeficiency virus (HIV) has been ongoing for more than 30 years, and although progress has been made, there are still significant challenges remaining. A few unique features render HIV to be one of the toughest viruses to conquer in the modern medicine era, such as the ability to target the host immune system, persist by integrating into the host genome and adapt to a hostile environment such as a single anti-HIV medication by continuously evolving. The finding of combination anti-retroviral therapy (cART) about 2 decades ago has transformed the treatment options for HIV-infected patients and significantly improved patient outcomes. However, finding an HIV cure has proven to be extremely challenging with the only known exception being the so-called "Berlin patient," whose immune system was replaced by stem cell transplants from a donor missing one of HIV's key co-receptors (CCR5). The broad application of this approach is limited by the requirement of an HLA-matched donor who is also homozygous for the rare CCR5 delta32 deletion. On the other hand, the Berlin patient provided the proof of concept of a potential cure for HIV using HIV-resistant hematopoietic stem cells (HSCs), revitalizing the hope to find an HIV cure that is broadly applicable. Here we will review strategies and recent attempts to engineer HIV-resistant HSCs as a path to an HIV cure. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Identification of full-length transmitted/founder viruses and their progeny in primary HIV-1 infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Korber, Bette; Hraber, Peter; Giorgi, Elena

2009-01-01

Identification of transmitted/founder virus genomes and their progeny by is a novel strategy for probing the molecular basis of HIV-1 transmission and for evaluating the genetic imprint of viral and host factors that act to constrain or facilitate virus replication. Here, we show in a cohort of twelve acutely infected subjects (9 clade B; 3 clade C), that complete genomic sequences of transmitted/founder viruses could be inferred using single genome amplification of plasma viral RNA, direct amplicon sequencing, and a model of random virus evolution. This allowed for the precise identification, chemical synthesis, molecular cloning, and biological analysis of thosemore » viruses actually responsible for productive clinical infection and for a comprehensive mapping of sequential viral genomes and proteomes for mutations that are necessary or incidental to the establishment of HIV-1 persistence. Transmitted/founder viruses were CD4 and CCR5 tropic, replicated preferentially in activated primary T-Iymphocytes but not monocyte-derived macrophages, and were effectively shielded from most heterologous or broadly neutralizing antibodies. By 3 months of infection, the evolving viral quasispecies in three subjects showed mutational fixation at only 2-5 discreet genomic loci. By 6-12 months, mutational fixation was evident at 18-27 genomic loci. Some, but not all, of these mutations were attributable to virus escape from cytotoxic Tlymphocytes or neutralizing antibodies, suggesting that other viral or host factors may influence early HIV -1 fitness.« less

Single-Cell Tracking Reveals a Role for Pre-Existing CCR5+ Memory Th1 Cells in the Control of Rhinovirus-A39 After Experimental Challenge in Humans.

PubMed

Muehling, Lyndsey M; Turner, Ronald B; Brown, Kenneth B; Wright, Paul W; Patrie, James T; Lahtinen, Sampo J; Lehtinen, Markus J; Kwok, William W; Woodfolk, Judith A

2018-01-17

Little is known about T cells that respond to human rhinovirus in vivo, due to timing of infection, viral diversity, and complex T-cell specificities. We tracked circulating CD4+ T cells with identical epitope specificities that responded to intranasal challenge with rhinovirus (RV)-A39, and we assessed T-cell signatures in the nose. Cells were monitored using a mixture of 2 capsid-specific major histocompatibility complex II tetramers over a 7-week period, before and after RV-A39 challenge, in 16 human leukocyte antigen-DR4+ subjects who participated in a trial of Bifidobacterium lactis (Bl-04) supplementation. Pre-existing tetramer+ T cells were linked to delayed viral shedding, enriched for activated CCR5+ Th1 effectors, and included a minor interleukin-21+ T follicular helper cell subset. After RV challenge, expansion and activation of virus-specific CCR5+ Th1 effectors was restricted to subjects who had a rise in neutralizing antibodies, and tetramer-negative CCR5+ effector memory types were comodulated. In the nose, CXCR3-CCR5+ T cells present during acute infection were activated effector memory type, whereas CXCR3+ cells were central memory type, and cognate chemokine ligands were elevated over baseline. Probiotic had no T-cell effects. We conclude that virus-specific CCR5+ effector memory CD4+ T cells primed by previous exposure to related viruses contribute to the control of rhinovirus. © The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Induction of human immunodeficiency virus neutralizing antibodies using fusion complexes.

PubMed

Zipeto, Donato; Matucci, Andrea; Ripamonti, Chiara; Scarlatti, Gabriella; Rossolillo, Paola; Turci, Marco; Sartoris, Silvia; Tridente, Giuseppe; Bertazzoni, Umberto

2006-05-01

Human immunodeficiency virus-1 (HIV-1) infects cells by membrane fusion that is mediated by the envelope proteins gp120/gp41 and the cellular receptors CD4 and CCR5. During this process, some conserved viral epitopes are temporarily exposed and may induce a neutralizing antibody response when fixed in the fusogenic conformation. These transient structures are conserved and may be effective antigens for use in an anti-HIV-1 vaccine. In this study we tested different conditions of preparation of fusion complexes inducing neutralizing antibodies against both R5 and X4 tropic HIV-1 strains. Cell lines expressing HIV-1 gp120/gp41 and CD4-CCR5 were prepared and conditions for producing fusion complexes were tested. Complexes produced at different temperature and fixative combinations were used to immunize mice. Results indicated that (a) fusion complexes prepared at either 21 degrees C, 30 degrees C or 37 degrees C were immunogenic and induced neutralizing antibodies against both R5 and X4 HIV-1 heterologous isolates; (b) after extensive purification of antibodies there was no cytotoxic effect; (c) complexes prepared at 37 degrees C were more immunogenic and induced higher titers of neutralizing antibodies than complexes prepared at either 21 degrees C or 30 degrees C; (d) the fixative used did not affect the titer of neutralizing antibodies except for glutaraldehyde which was ineffective; (e) the neutralizing activity was retained after CD4-CCR5 antibody removal. The production of higher titers of neutralizing antibody with fusion complexes prepared at 37 degrees C, as compared to lower temperatures, may be related to the induction of antibodies against many different conformation intermediates that subsequently act synergistically at different steps in the fusion process.

Chemokine Receptor Ccr7 Restricts Fatal West Nile Virus Encephalitis.

PubMed

Bardina, Susana V; Brown, Julia A; Michlmayr, Daniela; Hoffman, Kevin W; Sum, Janet; Pletnev, Alexander G; Lira, Sergio A; Lim, Jean K

2017-05-15

West Nile virus (WNV) is a mosquito-transmitted flavivirus that can cause debilitating encephalitis. To delineate the mechanisms behind this pathology, we studied Ccr7-deficient mice, which afforded us the capacity to study infection in mice with disrupted peripheral cellular trafficking events. The loss of Ccr7 resulted in an immediate pan-leukocytosis that remained elevated throughout the infection. This leukocytosis resulted in a significant enhancement of leukocyte accumulation within the central nervous system (CNS). Despite an excess of virus-specific T cells in the CNS, Ccr7-deficient mice had significantly higher CNS viral loads and mortality rates than wild-type animals. Mechanistically, the elevated trafficking of infected myeloid cells into the brain in Ccr7-deficient mice resulted in increased levels of WNV in the CNS, thereby effectively contributing to neuroinflammation and lowering viral clearance. Combined, our experiments suggest that during WNV infection, Ccr7 is a gatekeeper for nonspecific viral transference to the brain. IMPORTANCE In this study, we show that Ccr7 is required for the sufficient migration of dendritic cells and T cells into the draining lymph node immediately following infection and for the restriction of leukocyte migration into the brain. Further, the severe loss of dendritic cells in the draining lymph node had no impact on viral replication in this organ, suggesting that WNV may migrate from the skin into the lymph node through another mechanism. Most importantly, we found that the loss of Ccr7 results in a significant leukocytosis, leading to hypercellularity within the CNS, where monocytes/macrophages contribute to CNS viremia, neuroinflammation, and increased mortality. Together, our data point to Ccr7 as a critical host defense restriction factor limiting neuroinflammation during acute viral infection. Copyright © 2017 American Society for Microbiology.

Determination of Coreceptor Usage of Human Immunodeficiency Virus Type 1 from Patient Plasma Samples by Using a Recombinant Phenotypic Assay

PubMed Central

Trouplin, Virginie; Salvatori, Francesca; Cappello, Fanny; Obry, Veronique; Brelot, Anne; Heveker, Nikolaus; Alizon, Marc; Scarlatti, Gabriella; Clavel, François; Mammano, Fabrizio

2001-01-01

We developed a recombinant virus technique to determine the coreceptor usage of human immunodeficiency virus type 1 (HIV-1) from plasma samples, the source expected to represent the most actively replicating virus population in infected subjects. This method is not subject to selective bias associated with virus isolation in culture, a step required for conventional tropism determination procedures. The addition of a simple subcloning step allowed semiquantitative evaluation of virus populations with a different coreceptor (CCR5 or CXCR4) usage specificity present in each plasma sample. This procedure detected mixtures of CCR5- and CXCR4-exclusive virus populations as well as dualtropic viral variants, in variable proportions. Sequence analysis of dualtropic clones indicated that changes in the V3 loop are necessary for the use of CXCR4 as a coreceptor, but the overall context of the V1-V3 region is important to preserve the capacity to use CCR5. This convenient technique can greatly assist the study of virus evolution and compartmentalization in infected individuals. PMID:11119595

Maraviroc Observational Study: The Impact of Expanded Resistance Testing and Clinical Considerations for Antiretroviral Regimen Selection in Treatment-Experienced Patients

PubMed Central

Wilkins, Sara-Anne; Tamhane, Ashutosh; Nevin, Christa R.; Mugavero, Michael J.; Raper, James L.; Napolitano, Laura A.; Saag, Michael S.

2013-01-01

Abstract Maraviroc (MVC) use has trailed that of other post-2006 antiretroviral therapy (ART) options for treatment-experienced patients. We explored the impact of free tropism testing on MVC utilization in our cohort and explored barriers to MVC utilization. The Maraviroc Outcomes Study (MOS) is an investigator-initiated industry-sponsored trial where consecutive ART-experienced patients receiving routine care with viral loads ≥1,000 copies/ml, and whose provider requested resistance testing and received standardized resistance testing (SRT; phenotype, genotype, coreceptor/tropism). Sociodemographic, clinical, and ART characteristics of those receiving SRT were compared to a historical cohort (HC). Subsequently, providers were surveyed regarding factors influencing selection of salvage ART therapy. The HC (n=165) had resistance testing 7/08–9/09, while prospective SRT (n=83) patients were enrolled 9/09–8/10. In the HC, 92% had genotypes, 2% had tropism assays, and 62% (n=102) changed ART after resistance testing (raltegravir 37%, etravirine 25%, darunavir 24%, MVC 1%). In the SRT cohort, 57% (n=48) changed regimens after standardized resistance testing (darunavir 48%, raltegravir 40%, and etravirine 19%). CCR5-tropic virus was identified in 43% of the SRT group, and MVC was used in 10% [or 20% of R5 tropic patients who underwent a subsequent regimen change (n=25)], a statistically significant (p=0.01) increase in utilization. The factors most strongly influencing utilization were unique patient circumstances (60%), clinical experience (55%), and potential side effects (40%). The addition of routine tropism testing to genotypic/phenotypic testing was associated with increased MVC utilization, raising the possibility that tropism testing may present a barrier to MVC use; however, additional barriers exist, and merit further evaluation. PMID:22881368

Novel AgoshRNA molecules for silencing of the CCR5 co-receptor for HIV-1 infection

PubMed Central

Herrera-Carrillo, Elena

2017-01-01

Allogeneic transplantation of blood stem cells from a CCR5-Δ32 homozygous donor to an HIV-infected individual, the “Berlin patient”, led to a cure. Since then there has been a search for approaches that mimic this intervention in a gene therapy setting. RNA interference (RNAi) has evolved as a powerful tool to regulate gene expression in a sequence-specific manner and can be used to inactivate the CCR5 mRNA. Short hairpin RNA (shRNA) molecules can impair CCR5 expression, but these molecules may cause unintended side effects and they will not be processed in cells that lack Dicer, such as monocytes. Dicer-independent RNAi pathways have opened opportunities for new AgoshRNA designs that rely exclusively on Ago2 for maturation. Furthermore, AgoshRNA processing yields a single active guide RNA, thus reducing off-target effects. In this study, we tested different AgoshRNA designs against CCR5. We selected AgoshRNAs that potently downregulated CCR5 expression on human T cells and peripheral blood mononuclear cells (PBMC) and that had no apparent adverse effect on T cell development as assessed in a competitive cell growth assay. CCR5 knockdown significantly protected T cells from CCR5 tropic HIV-1 infection. PMID:28542329

Protection of rhesus macaques from vaginal infection by vaginally delivered Maraviroc, an inhibitor of HIV-1 entry via the CCR5 co-receptor

PubMed Central

Veazey, Ronald S.; Ketas, Thomas J.; Dufour, Jason; Moroney-Rasmussen, Terri; Green, Linda C.; Klasse, P.J.; Moore, John P.

2010-01-01

An effective vaginal microbicide could reduce human immunodeficiency virus type 1 (HIV-1) transmission to women. Among microbicide candidates in clinical development is Maraviroc (MVC), a small molecule drug that binds the CCR5 co-receptor and impedes HIV-1 entry into cells. Delivered systemically, MVC reduces viral load in HIV-1-infected people, but its ability to prevent transmission is untested. We have now evaluated MVC as a vaginal microbicide, using a stringent model involving challenge of rhesus macaques with a high-dose of a CCR5-using virus, SHIV-162P3. Gel-formulated, prescription-grade MVC provided dose-dependent protection, half-maximally at 0.5 mM (0.25 mg/ml). The duration of protection was transient; the longer the delay between MVC application and virus challenge, the less protection (T1/2 ~ 4 h). As expected, MVC neither protected against challenge with a CXCR4-using virus, SHIV-KU1, nor exacerbated post-infection viremia. These findings validate MVC development as a vaginal microbicide for women, and should guide clinical programs. PMID:20629537

Co-Introduced Functional CCR2 Potentiates In Vivo Anti-Lung Cancer Functionality Mediated by T Cells Double Gene-Modified to Express WT1-Specific T-Cell Receptor

PubMed Central

Asai, Hiroaki; Fujiwara, Hiroshi; An, Jun; Ochi, Toshiki; Miyazaki, Yukihiro; Nagai, Kozo; Okamoto, Sachiko; Mineno, Junichi; Kuzushima, Kiyotaka; Shiku, Hiroshi; Inoue, Hirofumi; Yasukawa, Masaki

2013-01-01

Background and Purpose Although gene-modification of T cells to express tumor-related antigen-specific T-cell receptor (TCR) or chimeric antigen receptor (CAR) has clinically proved promise, there still remains room to improve the clinical efficacy of re-directed T-cell based antitumor adoptive therapy. In order to achieve more objective clinical responses using ex vivo-expanded tumor-responsive T cells, the infused T cells need to show adequate localized infiltration into the tumor. Methodology/Principal Findings Human lung cancer cells variously express a tumor antigen, Wilms' Tumor gene product 1 (WT1), and an inflammatory chemokine, CCL2. However, CCR2, the relevant receptor for CCL2, is rarely expressed on activated T-lymphocytes. A HLA-A2402+ human lung cancer cell line, LK79, which expresses high amounts of both CCL2 and WT1 mRNA, was employed as a target. Normal CD8+ T cells were retrovirally gene-modified to express both CCR2 and HLA-A*2402-restricted and WT1235–243 nonapeptide-specific TCR as an effector. Anti-tumor functionality mediated by these effector cells against LK79 cells was assessed both in vitro and in vivo. Finally the impact of CCL2 on WT1 epitope-responsive TCR signaling mediated by the effector cells was studied. Introduced CCR2 was functionally validated using gene-modified Jurkat cells and human CD3+ T cells both in vitro and in vivo. Double gene-modified CD3+ T cells successfully demonstrated both CCL2-tropic tumor trafficking and cytocidal reactivity against LK79 cells in vitro and in vivo. CCL2 augmented the WT1 epitope-responsive TCR signaling shown by relevant luciferase production in double gene-modified Jurkat/MA cells to express luciferase and WT1-specific TCR, and CCL2 also dose-dependently augmented WT1 epitope-responsive IFN-γ production and CD107a expression mediated by these double gene-modifiedCD3+ T cells. Conclusion/Significance Introduction of the CCL2/CCR2 axis successfully potentiated in vivo anti-lung cancer reactivity mediated by CD8+ T cells double gene-modified to express WT1-specific TCR and CCR2 not only via CCL2-tropic tumor trafficking, but also CCL2-enhanced WT1-responsiveness. PMID:23441216

CCR5 Knockout Prevents Neuronal Injury and Behavioral Impairment Induced in a Transgenic Mouse Model by a CXCR4-using HIV-1 Glycoprotein 1201

PubMed Central

Maung, Ricky; Hoefer, Melanie M.; Sanchez, Ana B.; Sejbuk, Natalia E.; Medders, Kathryn E.; Desai, Maya K.; Catalan, Irene C.; Dowling, Cari C.; de Rozieres, Cyrus M.; Garden, Gwenn A.; Russo, Rossella; Roberts, Amanda J.; Williams, Roy; Kaul, Marcus

2014-01-01

The innate immune system has been implicated in several neurodegenerative diseases, including human immunodeficiency virus (HIV)-1 associated dementia. Here we show that genetic ablation of CCR5 prevents microglial activation and neuronal damage in a transgenic model of HIV-associated brain injury induced by a CXCR4-utilizing viral envelope gp120. The CCR5 knockout (KO) also rescues spatial learning and memory in gp120-transgenic (tg) mice. However, the CCR5KO does not abrogate astrocytosis, indicating it can occur independently from neuronal injury and behavioral impairment. To further characterize the neuroprotective effect of CCR5-deficiency we performed a genome –wide gene expression analysis of brains from HIVgp120tg mice expressing or lacking CCR5 and non-transgenic controls. Comparison with a human brain microarray study reveals that brains of HIVgp120tg mice and HIV patients with neurocognitive impairment share numerous differentially regulated genes. Furthermore, brains of CCR5 wild-type (WT) and CCR5KO gp120tg mice express markers of an innate immune response. One of the most significantly up-regulated factors is the acute phase protein lipocalin-2 (LCN2). Using cerebrocortical cell cultures, we find that LCN2 is neurotoxic in a CCR5-dependent fashion while inhibition of CCR5 alone is not sufficient to abrogate neurotoxicity of a CXCR4-utilizing gp120. However, the combination of pharmacological CCR5 blockade and LCN2 protects neurons from toxicity of a CXCR4-utilizing gp120 thus recapitulating the finding in CCR5-deficient gp120tg mouse brain. Altogether, our study provides evidence for an indirect pathological role of CCR5 and a novel protective effect of LCN2 in combination with inhibition of CCR5 in HIV-associated brain injury. PMID:25031461

Protection of macaques from vaginal SHIV challenge by an orally delivered CCR5 inhibitor.

PubMed

Veazey, Ronald S; Springer, Martin S; Marx, Preston A; Dufour, Jason; Klasse, Per Johan; Moore, John P

2005-12-01

Pre-exposure oral prophylaxis with antiviral drugs is a potential method for preventing transmission of human immunodeficiency virus type 1 (HIV-1). We show that oral delivery of CMPD167, a small molecule that binds to the CCR5 coreceptor, for 10-14 d can protect a substantial proportion of macaques from vaginal infection with a CCR5-using virus (SHIV-162P3). The macaques that became infected despite receiving CMPD167 had reduced plasma viremia levels during the earliest stages of infection.

Induction of HIV-blocking anti-CCR5 IgA in Peyers's patches without histopathological alterations.

PubMed

Pastori, Claudia; Diomede, Lorenzo; Venuti, Assunta; Fisher, Gregory; Jarvik, Jonathan; Bomsel, Morgane; Sanvito, Francesca; Lopalco, Lucia

2014-04-01

The chemokine receptor CCR5 is essential for HIV infection and is thus a potential target for vaccine development. However, because CCR5 is a host protein, generation of anti-CCR5 antibodies requires the breaking of immune tolerance and thus carries the risk of autoimmune responses. In this study, performed in mice, we compared 3 different immunogens representing surface domains of murine CCR5, 4 different adjuvants, and 13 different immunization protocols, with the goal of eliciting HIV-blocking activity without inducing autoimmune dysfunction. In all cases the CCR5 sequences were presented as fusions to the Flock House virus (FHV) capsid precursor protein. We found that systemic immunization and mucosal boosting elicited CCR5-specific antibodies and achieved consistent priming in Peyer's patches, where most cells showed a phenotype corresponding to activated B cells and secreted high levels of IgA, representing up to one-third of the total HIV-blocking activity. Histopathological analysis revealed mild to moderate chronic inflammation in some tissues but failed in reporting signs of autoimmune dysfunction associated with immunizations. Antisera against immunogens representing the N terminus and extracellular loops 1 and 2 (Nter1 and ECL1 and ECL2) of CCR5 were generated. All showed specific anti-HIV activity, which was stronger in the anti-ECL1 and -ECL2 sera than in the anti-Nter sera. ECL1 and ECL2 antisera induced nearly complete long-lasting CCR5 downregulation of the receptor, and especially, their IgG-depleted fractions prevented HIV infection in neutralization and transcytosis assays. In conclusion, the ECL1 and ECL2 domains could offer a promising path to achieve significant anti-HIV activity in vivo. The study was the first to adopt a systematic strategy to compare the immunogenicities of all extracellular domains of the CCR5 molecule and to set optimal conditions leading to generation of specific antibodies in the mouse model. There were several relevant findings, which could be translated into human trials. (i) Prime (systemic) and boost (mucosal) immunization is the best protocol to induce anti-self antibodies with the expected properties. (ii) Aluminum is the best adjuvant in mice and thus can be easily used in nonhuman primates (NHP) and humans. (iii) The Flock House virus (FHV) system represents a valid delivery system, as the structure is well known and is not pathogenic for humans, and it is possible to introduce constrained regions able to elicit antibodies that recognize conformational epitopes. (iv) The best CCR5 vaccine candidate should include either extracellular loop 1 or 2 (ECL1 or ECL2), but not N terminus domains.

Cellular and molecular identity of tumor-associated macrophages in glioblastoma

PubMed Central

Chen, Zhihong; Feng, Xi; Herting, Cameron J.; Garcia, Virginia Alvarez; Nie, Kai; Pong, Winnie W.; Rasmussen, Rikke; Dwivedi, Bhakti; Seby, Sandra; Wolf, Susanne A.; Gutmann, David H.; Hambardzumyan, Dolores

2017-01-01

In glioblastoma (GBM), tumor-associated macrophages (TAM) represent up to one half of the cells of the tumor mass, including both infiltrating macrophages and resident brain microglia. In an effort to delineate the temporal and spatial dynamics of TAM composition during gliomagenesis, we employed two genetically engineered mouse models where oncogenic drivers and fluorescent reporters were expressed coordinately under the control of the monocyte/microglia-selective Cx3cr1 or Ccr2 promoters, respectively. Using this approach, we demonstrated that CX3CR1LoCCR2Hi monocytes were recruited to the glioblastoma, where they transitioned to CX3CR1HiCCR2Lo macrophages and CX3CR1HiCCR2− microglia-like cells. Infiltrating macrophages/monocytes constituted ~85% of the total TAM population, with resident microglia accounting for the ~15% remaining. Bone marrow-derived infiltrating macrophages/monocytes were recruited to the tumor early during GBM initiation, where they localized preferentially to perivascular areas. In contrast, resident microglia were localized mainly to peritumoral regions. RNA-sequencing analyses revealed differential gene expression patterns unique to infiltrating and resident cells, suggesting unique functions for each TAM population. Notably, limiting monocyte infiltration via Ccl2 genetic ablation prolonged the survival of tumor-bearing mice. Our findings illuminate the unique composition and functions of infiltrating and resident myeloid cells in GBM, establishing a rationale to target infiltrating cells in this neoplasm. PMID:28235764

Dualtropic CXCR6/CCR5 Simian Immunodeficiency Virus (SIV) Infection of Sooty Mangabey Primary Lymphocytes: Distinct Coreceptor Use in Natural versus Pathogenic Hosts of SIV.

PubMed

Elliott, Sarah T C; Wetzel, Katherine S; Francella, Nicholas; Bryan, Steven; Romero, Dino C; Riddick, Nadeene E; Shaheen, Farida; Vanderford, Thomas; Derdeyn, Cynthia A; Silvestri, Guido; Paiardini, Mirko; Collman, Ronald G

2015-09-01

Natural-host sooty mangabeys (SM) infected with simian immunodeficiency virus (SIV) exhibit high viral loads but do not develop disease, whereas infection of rhesus macaques (RM) causes CD4(+) T cell loss and AIDS. Several mechanisms have been proposed to explain these divergent outcomes, including differences in cell targeting, which have been linked to low expression of the canonical SIV entry receptor CCR5 on CD4(+) T cells of SM and other natural hosts. We previously showed that infection and high-level viremia occur even in a subset of SM that genetically lack functional CCR5, which indicates that alternative entry coreceptors are used by SIV in vivo in these animals. We also showed that SM CXCR6 is a robust coreceptor for SIVsmm in vitro. Here we identify CXCR6 as a principal entry pathway for SIV in SM primary lymphocytes. We show that ex vivo SIV infection of lymphocytes from CCR5 wild-type SM is mediated by both CXCR6 and CCR5. In contrast, infection of RM lymphocytes is fully dependent on CCR5. These data raise the possibility that CXCR6-directed tropism in CCR5-low natural hosts may alter CD4(+) T cell subset targeting compared with that in nonnatural hosts, enabling SIV to maintain high-level replication without leading to widespread CD4(+) T cell loss. Natural hosts of SIV, such as sooty mangabeys, sustain high viral loads but do not develop disease, while nonnatural hosts, like rhesus macaques, develop AIDS. Understanding this difference may help elucidate mechanisms of pathogenesis. Natural hosts have very low levels of the SIV entry coreceptor CCR5, suggesting that restricted entry may limit infection of certain target cells, although it is unclear how the virus replicates so robustly. Here we show that in sooty mangabey lymphocytes, infection is mediated by the alternative entry coreceptor CXCR6, as well as CCR5. In rhesus macaque lymphocytes, however, infection occurs entirely through CCR5. The use of CXCR6 for entry, combined with very low CCR5 levels, may redirect the virus to different cell targets in natural hosts. It is possible that differential targeting may favor infection of nonessential cells and limit infection of critical cells in natural hosts, thus contributing to benign outcome of infection. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Dualtropic CXCR6/CCR5 Simian Immunodeficiency Virus (SIV) Infection of Sooty Mangabey Primary Lymphocytes: Distinct Coreceptor Use in Natural versus Pathogenic Hosts of SIV

PubMed Central

Elliott, Sarah T. C.; Wetzel, Katherine S.; Francella, Nicholas; Bryan, Steven; Romero, Dino C.; Riddick, Nadeene E.; Shaheen, Farida; Vanderford, Thomas; Derdeyn, Cynthia A.; Silvestri, Guido; Paiardini, Mirko

2015-01-01

ABSTRACT Natural-host sooty mangabeys (SM) infected with simian immunodeficiency virus (SIV) exhibit high viral loads but do not develop disease, whereas infection of rhesus macaques (RM) causes CD4+ T cell loss and AIDS. Several mechanisms have been proposed to explain these divergent outcomes, including differences in cell targeting, which have been linked to low expression of the canonical SIV entry receptor CCR5 on CD4+ T cells of SM and other natural hosts. We previously showed that infection and high-level viremia occur even in a subset of SM that genetically lack functional CCR5, which indicates that alternative entry coreceptors are used by SIV in vivo in these animals. We also showed that SM CXCR6 is a robust coreceptor for SIVsmm in vitro. Here we identify CXCR6 as a principal entry pathway for SIV in SM primary lymphocytes. We show that ex vivo SIV infection of lymphocytes from CCR5 wild-type SM is mediated by both CXCR6 and CCR5. In contrast, infection of RM lymphocytes is fully dependent on CCR5. These data raise the possibility that CXCR6-directed tropism in CCR5-low natural hosts may alter CD4+ T cell subset targeting compared with that in nonnatural hosts, enabling SIV to maintain high-level replication without leading to widespread CD4+ T cell loss. IMPORTANCE Natural hosts of SIV, such as sooty mangabeys, sustain high viral loads but do not develop disease, while nonnatural hosts, like rhesus macaques, develop AIDS. Understanding this difference may help elucidate mechanisms of pathogenesis. Natural hosts have very low levels of the SIV entry coreceptor CCR5, suggesting that restricted entry may limit infection of certain target cells, although it is unclear how the virus replicates so robustly. Here we show that in sooty mangabey lymphocytes, infection is mediated by the alternative entry coreceptor CXCR6, as well as CCR5. In rhesus macaque lymphocytes, however, infection occurs entirely through CCR5. The use of CXCR6 for entry, combined with very low CCR5 levels, may redirect the virus to different cell targets in natural hosts. It is possible that differential targeting may favor infection of nonessential cells and limit infection of critical cells in natural hosts, thus contributing to benign outcome of infection. PMID:26109719

Multiple-cohort genetic association study reveals CXCR6 as a new chemokine receptor involved in long-term nonprogression to AIDS.

PubMed

Limou, Sophie; Coulonges, Cédric; Herbeck, Joshua T; van Manen, Daniëlle; An, Ping; Le Clerc, Sigrid; Delaneau, Olivier; Diop, Gora; Taing, Lieng; Montes, Matthieu; van't Wout, Angélique B; Gottlieb, Geoffrey S; Therwath, Amu; Rouzioux, Christine; Delfraissy, Jean-François; Lelièvre, Jean-Daniel; Lévy, Yves; Hercberg, Serge; Dina, Christian; Phair, John; Donfield, Sharyne; Goedert, James J; Buchbinder, Susan; Estaquier, Jérôme; Schächter, François; Gut, Ivo; Froguel, Philippe; Mullins, James I; Schuitemaker, Hanneke; Winkler, Cheryl; Zagury, Jean-François

2010-09-15

The compilation of previous genomewide association studies of AIDS shows a major polymorphism in the HCP5 gene associated with both control of the viral load and long-term nonprogression (LTNP) to AIDS. To look for genetic variants that affect LTNP without necessary control of the viral load, we reanalyzed the genomewide data of the unique LTNP Genomics of Resistance to Immunodeficiency Virus (GRIV) cohort by excluding "elite controller" patients, who were controlling the viral load at very low levels (<100 copies/mL). The rs2234358 polymorphism in the CXCR6 gene was the strongest signal (P=2.5 x 10(-7); odds ratio, 1.85) obtained for the genomewide association study comparing the 186 GRIV LTNPs who were not elite controllers with 697 uninfected control subjects. This association was replicated in 3 additional independent European studies, reaching genomewide significance of P(combined)=9.7 x 10(-10). This association with LTNP is independent of the CCR2-CCR5 locus and the HCP5 polymorphisms. The statistical significance, the replication, and the magnitude of the association demonstrate that CXCR6 is likely involved in the molecular etiology of AIDS and, in particular, in LTNP, emphasizing the power of extreme-phenotype cohorts. CXCR6 is a chemokine receptor that is known as a minor coreceptor in human immunodeficiency virus type 1 infection but could participate in disease progression through its role as a mediator of inflammation.

Characterization In Vitro and In Vivo of a Pandemic H1N1 Influenza Virus from a Fatal Case

PubMed Central

Cuevas, Maria Teresa; Pozo, Francisco; Guerra, Susana; García-Barreno, Blanca; Martinez-Orellana, Pamela; Pérez-Breña, Pilar; Montoya, Maria; Melero, Jose Antonio; Pizarro, Manuel; Ortin, Juan; Casas, Inmaculada; Nieto, Amelia

2013-01-01

Pandemic 2009 H1N1 (pH1N1) influenza viruses caused mild symptoms in most infected patients. However, a greater rate of severe disease was observed in healthy young adults and children without co-morbid conditions. Here we tested whether influenza strains displaying differential virulence could be present among circulating pH1N1 viruses. The biological properties and the genotype of viruses isolated from a patient showing mild disease (M) or from a fatal case (F), both without known co-morbid conditions were compared in vitro and in vivo. The F virus presented faster growth kinetics and stronger induction of cytokines than M virus in human alveolar lung epithelial cells. In the murine model in vivo, the F virus showed a stronger morbidity and mortality than M virus. Remarkably, a higher proportion of mice presenting infectious virus in the hearts, was found in F virus-infected animals. Altogether, the data indicate that strains of pH1N1 virus with enhanced pathogenicity circulated during the 2009 pandemic. In addition, examination of chemokine receptor 5 (CCR5) genotype, recently reported as involved in severe influenza virus disease, revealed that the F virus-infected patient was homozygous for the deleted form of CCR5 receptor (CCR5Δ32). PMID:23326447

Low levels of SIV infection in sooty mangabey central-memory CD4+ T-cells is associated with limited CCR5 expression

PubMed Central

Paiardini, Mirko; Cervasi, Barbara; Reyes-Aviles, Elane; Micci, Luca; Ortiz, Alexandra M.; Chahroudi, Ann; Vinton, Carol; Gordon, Shari N.; Bosinger, Steven E.; Francella, Nicholas; Hallberg, Paul L.; Schlub, Timothy; Chan, Ming Liang; Riddick, Nadeene E.; Collman, Ronald G.; Apetrei, Cristian; Pandrea, Ivona; Else, James; Munch, Jan; Kirchhoff, Frank; Davenport, Miles P.; Brenchley, Jason M.; Silvestri, Guido

2011-01-01

Naturally SIV-infected sooty mangabeys (SMs) do not progress to AIDS despite high-level virus replication. We previously showed that the fraction of CD4+CCR5+ T-cells is lower in SMs compared to humans and macaques. Here we found that, after in vitro stimulation, SM CD4+ T-cells fail to up-regulate CCR5, and that this phenomenon is more pronounced in CD4+ central-memory T-cells (TCM). CD4+ T-cell activation was similarly uncoupled from CCR5 expression in SMs in vivo during (i) acute SIV infection and (ii) following antibody-mediated CD4+ T-cell depletion. Remarkably, CD4+ TCM of SMs that express low levels of CCR5 demonstrated reduced susceptibility to SIV infection both in vivo and in vitro when compared to CD4+ TCM of RMs. These data suggest that low CCR5 expression on SM CD4+ T-cells favors the preservation of CD4+ T-cell homeostasis and promotes an AIDS-free status by protecting CD4+ TCM from direct virus infection. PMID:21706028

The Central Conserved Region (CCR) of Respiratory Syncytial Virus (RSV) G Protein Modulates Host miRNA Expression and Alters the Cellular Response to Infection

PubMed Central

Haynes, Lia M.; Anderson, Larry J.

2017-01-01

Respiratory Syncytial Virus (RSV) infects respiratory epithelial cells and deregulates host gene expression by many mechanisms including expression of RSV G protein (RSV G). RSV G protein encodes a central conserved region (CCR) containing a CX3C motif that functions as a fractalkine mimic. Disruption of the CX3C motif (a.a. 182–186) located in the CCR of the G protein has been shown to affect G protein function in vitro and the severity of RSV disease pathogenesis in vivo. We show that infection of polarized Calu3 respiratory cells with recombinant RSV having point mutations in Cys173 and 176 (C173/176S) (rA2-GC12), or Cys186 (C186S) (rA2-GC4) is associated with a decline in the integrity of polarized Calu-3 cultures and decreased virus production. This is accompanied with downregulation of miRNAs let-7f and miR-24 and upregulation of interferon lambda (IFNλ), a primary antiviral cytokine for RSV in rA2-GC12/rA2-GC4 infected cells. These results suggest that residues in the cysteine noose region of RSV G protein can modulate IFN λ expression accompanied by downregulation of miRNAs, and are important for RSV G protein function and targeting. PMID:28671606

The Central Conserved Region (CCR) of Respiratory Syncytial Virus (RSV) G Protein Modulates Host miRNA Expression and Alters the Cellular Response to Infection.

PubMed

Bakre, Abhijeet A; Harcourt, Jennifer L; Haynes, Lia M; Anderson, Larry J; Tripp, Ralph A

2017-07-03

Respiratory Syncytial Virus (RSV) infects respiratory epithelial cells and deregulates host gene expression by many mechanisms including expression of RSV G protein (RSV G). RSV G protein encodes a central conserved region (CCR) containing a CX3C motif that functions as a fractalkine mimic. Disruption of the CX3C motif (a.a. 182-186) located in the CCR of the G protein has been shown to affect G protein function in vitro and the severity of RSV disease pathogenesis in vivo. We show that infection of polarized Calu3 respiratory cells with recombinant RSV having point mutations in Cys173 and 176 (C173/176S) (rA2-GC12), or Cys186 (C186S) (rA2-GC4) is associated with a decline in the integrity of polarized Calu-3 cultures and decreased virus production. This is accompanied with downregulation of miRNAs let-7f and miR-24 and upregulation of interferon lambda (IFNλ), a primary antiviral cytokine for RSV in rA2-GC12/rA2-GC4 infected cells. These results suggest that residues in the cysteine noose region of RSV G protein can modulate IFN λ expression accompanied by downregulation of miRNAs, and are important for RSV G protein function and targeting.

Flexible use of CCR5 in the absence of CXCR4 use explains the immune deficiency in HIV-1 infected children.

PubMed

Cavarelli, Mariangela; Karlsson, Ingrid; Ripamonti, Chiara; Plebani, Anna; Fenyo, Eva Maria; Scarlatti, Gabriella

2010-10-23

CCR5-using HIV-1 (R5 viruses) are usually isolated during acute infection from both adults and children. We have recently demonstrated that R5 viruses with a flexible use of CCR5 (called R5broad) can be detected in children close to birth and are predictive of a fast immunological failure. The aim of the present work was to investigate viral phenotype variation during disease progression in HIV-1 infected children, six slow and eight fast progressors. A total of 74 viral isolates obtained sequentially from 14 HIV-1 infected children were tested for their ability to infect U87.CD4 cells expressing a set of six different CCR5/CXCR4 chimeric receptors or wild-type coreceptors. The sensitivity of 35 R5 viruses to inhibition with the CC-chemokine RANTES (regulated upon activation, normal T-cell expressed and secreted) was evaluated in a peripheral blood mononuclear cells based assay. Viral evolution to R5broad or to R5X4 phenotype occurred with one exception, in all children, although at a different time point according to rate of disease progression. Immune deficiency in the children was significantly associated with the appearance of R5broad phenotype or R5X4 viruses. Analysis of the sensitivity to inhibition by RANTES revealed a significant correlation between the R5broad phenotype and an augmented resistance to this CC-chemokine. We demonstrate that the viral evolution to a more flexible CCR5-use is sufficient to explain the immunological failure in the absence of CXCR4 usage. These results warrant detailed analysis of the R5 phenotype in forthcoming clinical studies introducing CCR5 inhibitors for the treatment of pediatric HIV-1 infection.

Short communication: identification of the conformational requirement for the specificities of coreceptors for human and simian immunodeficiency viruses.

PubMed

Shimizu, Nobuaki; Tanaka, Atsushi; Jinno-Oue, Atsushi; Mori, Takahisa; Ohtsuki, Takahiro; Hoshino, Hiroo

2010-03-01

More than 10 G protein-coupled receptors (GPCRs) work as coreceptors for human and simian immunodeficiency viruses (HIVs/SIVs); however, structural features critical for coreceptor activity have not been identified. Our objective was to elucidate the structural requirement of coreceptor activities. Amino-terminal regions (NTRs), extracellular loops (ECLs), and the undecapeptidyl arch (UPA) in the second ECL have been shown to be important for coreceptor function. We made chimeric coreceptors for these regions between CCR5 and GPR1, which is genetically distant from CCR5, and analyzed their activities. The coreceptor activity and specificity of CCR5 were maintained when its NTR or UPA was replaced with GPR1. In contrast, the GPR1 chimera with CCR5 NTR was used by HIV-1 strains that can use only CCR5, but not both CCR5 and CXCR4, or GPR1. GPR1 chimera with CCR5 UPA almost lost activity. All ECL chimeras could hardly maintain activity. Thus, CCR5 is more flexibly acceptable to heterologous NTR and UPA than GPR1, suggesting the existence of conformational differences made by the integration of multiple extracellular regions. This conformation may specifically interact with HIV-1 in a strain-dependent manner. Identification of a factor that is critical to make this conformation will contribute to understanding the mechanism of coreceptor function of GPCRs. For this, the coreceptor activity of GPR1, which is genetically distant from CCR5, will be a useful tool.

The yellow fever virus vaccine induces a broad and polyfunctional human memory CD8+ T cell response.

PubMed

Akondy, Rama S; Monson, Nathan D; Miller, Joseph D; Edupuganti, Srilatha; Teuwen, Dirk; Wu, Hong; Quyyumi, Farah; Garg, Seema; Altman, John D; Del Rio, Carlos; Keyserling, Harry L; Ploss, Alexander; Rice, Charles M; Orenstein, Walter A; Mulligan, Mark J; Ahmed, Rafi

2009-12-15

The live yellow fever vaccine (YF-17D) offers a unique opportunity to study memory CD8(+) T cell differentiation in humans following an acute viral infection. We have performed a comprehensive analysis of the virus-specific CD8(+) T cell response using overlapping peptides spanning the entire viral genome. Our results showed that the YF-17D vaccine induces a broad CD8(+) T cell response targeting several epitopes within each viral protein. We identified a dominant HLA-A2-restricted epitope in the NS4B protein and used tetramers specific for this epitope to track the CD8(+) T cell response over a 2 year period. This longitudinal analysis showed the following. 1) Memory CD8(+) T cells appear to pass through an effector phase and then gradually down-regulate expression of activation markers and effector molecules. 2) This effector phase was characterized by down-regulation of CD127, Bcl-2, CCR7, and CD45RA and was followed by a substantial contraction resulting in a pool of memory T cells that re-expressed CD127, Bcl-2, and CD45RA. 3) These memory cells were polyfunctional in terms of degranulation and production of the cytokines IFN-gamma, TNF-alpha, IL-2, and MIP-1beta. 4) The YF-17D-specific memory CD8(+) T cells had a phenotype (CCR7(-)CD45RA(+)) that is typically associated with terminally differentiated cells with limited proliferative capacity (T(EMRA)). However, these cells exhibited robust proliferative potential showing that expression of CD45RA may not always associate with terminal differentiation and, in fact, may be an indicator of highly functional memory CD8(+) T cells generated after acute viral infections.

The Application of Humanized Mouse Models for the Study of Human Exclusive Viruses.

PubMed

Vahedi, Fatemeh; Giles, Elizabeth C; Ashkar, Ali A

2017-01-01

The symbiosis between humans and viruses has allowed human tropic pathogens to evolve intricate means of modulating the human immune response to ensure its survival among the human population. In doing so, these viruses have developed profound mechanisms that mesh closely with our human biology. The establishment of this intimate relationship has created a species-specific barrier to infection, restricting the virus-associated pathologies to humans. This specificity diminishes the utility of traditional animal models. Humanized mice offer a model unique to all other means of study, providing an in vivo platform for the careful examination of human tropic viruses and their interaction with human cells and tissues. These types of animal models have provided a reliable medium for the study of human-virus interactions, a relationship that could otherwise not be investigated without questionable relevance to humans.

Preferential Use of CXCR4 by R5X4 Human Immunodeficiency Virus Type 1 Isolates for Infection of Primary Lymphocytes

PubMed Central

Yi, Yanjie; Shaheen, Farida; Collman, Ronald G.

2005-01-01

Coreceptor specificity of human immunodeficiency virus type 1 (HIV-1) strains is generally defined in vitro in cell lines expressing CCR5 or CXCR4, but lymphocytes and macrophages are the principal targets in vivo. CCR5-using (R5) variants dominate early in infection, but strains that use CXCR4 emerge later in a substantial minority of subjects. Many or most CXCR4-using variants can use both CXCR4 and CCR5 (R5X4), but the pathways that are actually used to cause infection in primary cells and in vivo are unknown. We examined several R5X4 prototype and primary isolates and found that they all were largely or completely restricted to CXCR4-mediated entry in primary lymphocytes, even though lymphocytes are permissive for CCR5-mediated entry by R5 strains. In contrast, in primary macrophages R5X4 isolates used both CCR5 and CXCR4. The R5X4 strains were also more sensitive than R5 strains to CCR5 blocking, suggesting that interactions between the R5X4 strains and CCR5 are less efficient. These results indicate that coreceptor phenotyping in transformed cells does not necessarily predict utilization in primary cells, that variability exists among HIV-1 isolates in the ability to use CCR5 expressed on lymphocytes, and that many or most strains characterized as R5X4 are functionally X4 in primary lymphocytes. Less efficient interactions between R5X4 strains and CCR5 may be responsible for the inability to use CCR5 on lymphocytes, which express relatively low CCR5 levels. Since isolates that acquire CXCR4 utilization retain the capacity to use CCR5 on macrophages despite their inability to use it on lymphocytes, these results also raise the possibility that a CCR5-mediated macrophage reservoir is required for sustained infection in vivo. PMID:15650174

Induced Pluripotent Stem Cell‐Derived Endothelial Cells Overexpressing Interleukin‐8 Receptors A/B and/or C‐C Chemokine Receptors 2/5 Inhibit Vascular Injury Response

PubMed Central

Giordano, Samantha; Zhao, Xiangmin; Chen, Yiu‐Fai; Litovsky, Silvio H.; Hage, Fadi G.; Townes, Tim M.; Sun, Chiao‐Wang; Wu, Li‐Chen; Oparil, Suzanne

2017-01-01

Abstract Recruitment of neutrophils and monocytes/macrophages to the site of vascular injury is mediated by binding of chemoattractants to interleukin (IL) 8 receptors RA and RB (IL8RA/B) C‐C chemokine receptors (CCR) 2 and 5 expressed on neutrophil and monocyte/macrophage membranes. Endothelial cells (ECs) derived from rat‐induced pluripotent stem cells (RiPS) were transduced with adenovirus containing cDNA of IL8RA/B and/or CCR2/5. We hypothesized that RiPS‐ECs overexpressing IL8RA/B (RiPS‐IL8RA/B‐ECs), CCR2/5 (RiPS‐CCR2/5‐ECs), or both receptors (RiPS‐IL8RA/B+CCR2/5‐ECs) will inhibit inflammatory responses and neointima formation in balloon‐injured rat carotid artery. Twelve‐week‐old male Sprague‐Dawley rats underwent balloon injury of the right carotid artery and intravenous infusion of (a) saline vehicle, (b) control RiPS‐Null‐ECs (ECs transduced with empty virus), (c) RiPS‐IL8RA/B‐ECs, (d) RiPS‐CCR2/5‐ECs, or (e) RiPS‐IL8RA/B+CCR2/5‐ECs. Inflammatory mediator expression and leukocyte infiltration were measured in injured and uninjured arteries at 24 hours postinjury by enzyme‐linked immunosorbent assay (ELISA) and immunohistochemistry, respectively. Neointima formation was assessed at 14 days postinjury. RiPS‐ECs expressing the IL8RA/B or CCR2/5 homing device targeted the injured arteries and decreased injury‐induced inflammatory cytokine expression, neutrophil/macrophage infiltration, and neointima formation. Transfused RiPS‐ECs overexpressing IL8RA/B and/or CCR2/5 prevented inflammatory responses and neointima formation after vascular injury. Targeted delivery of iPS‐ECs with a homing device to inflammatory mediators in injured arteries provides a novel strategy for the treatment of cardiovascular diseases. Stem Cells Translational Medicine 2017;6:1168–1177 PMID:28233474

An Aptamer That Neutralizes R5 Strains of Human Immunodeficiency Virus Type 1 Blocks gp120-CCR5 Interaction

PubMed Central

Dey, Antu K.; Khati, Makobetsa; Tang, Min; Wyatt, Richard; Lea, Susan M.; James, William

2005-01-01

We recently described the isolation and structural characterization of 2′-fluoropyrimidine-substituted RNA aptamers that bind to gp120 of R5 strains of human immunodeficiency virus type 1 and thereby potently neutralize the infectivity of phylogenetically diverse R5 strains. Here we investigate the physical basis of their antiviral action. We show that both N-linked oligosaccharides and the variable loops V1/V2 and V3 are not required for binding of one aptamer, B40, to gp120. Using surface plasmon resonance binding analyses, we show that the aptamer binds to the CCR5-binding site on gp120 in a relatively CD4-independent manner, providing a mechanistic explanation for its neutralizing potency. PMID:16227301

Ultra-violet radiation is responsible for the differences in global epidemiology of chickenpox and the evolution of varicella-zoster virus as man migrated out of Africa.

PubMed

Rice, Philip S

2011-04-23

Of the eight human herpes viruses, varicella-zoster virus, which causes chickenpox and zoster, has a unique epidemiology. Primary infection is much less common in children in the tropics compared with temperate areas. This results in increased adult susceptibility causing outbreaks, for example in health-care workers migrating from tropical to temperate countries. The recent demonstration that there are different genotypes of varicella-zoster virus and their geographic segregation into tropical and temperate areas suggests a distinct, yet previously unconsidered climatic factor may be responsible for both the clinical and molecular epidemiological features of this virus infection. Unlike other human herpes viruses, varicella-zoster virus does not require intimate contact for infection to occur indicating that transmission may be interrupted by a geographically restricted climatic factor. The factor with the largest difference between tropical and temperate zones is ultra-violet radiation. This could reduce the infectiousness of chickenpox cases by inactivating virus in vesicles, before or after rupture. This would explain decreased transmissibility in the tropics and why the peak chickenpox incidence in temperate zones occurs during winter and spring, when ultra-violet radiation is at its lowest. The evolution of geographically restricted genotypes is also explained by ultra-violet radiation driving natural selection of different virus genotypes with varying degrees of resistance to inactivation, tropical genotypes being the most resistant. Consequently, temperate viruses should be more sensitive to its effects. This is supported by the observation that temperate genotypes are found in the tropics only in specific circumstances, namely where ultra-violet radiation has either been excluded or significantly reduced in intensity. The hypothesis is testable by exposing different virus genotypes to ultra-violet radiation and quantifying virus survival by plaque forming units or quantitative mRNA RT-PCR. The ancestral varicella-zoster virus, most probably a tropical genotype, co-migrated with man as he left Africa approximately 200,000 years ago. For this virus to have lost the selective advantage of resistance to ultra-violet radiation, the hypothesis would predict that the temperate, ultra-violet sensitive virus should have acquired another selective advantage as an evolutionary trade-off. One obvious advantage could be an increased reactivation rate as zoster to set up more rounds of chickenpox transmission. If this were so, the mechanism responsible for resistance to ultra-violet radiation might also be involved in reactivation and latency. This could then provide the first insight into a genetic correlate of the survival strategy of this virus.

Multiple-cohort genetic association study reveals CXCR6 as a new chemokine receptor involved in long-term nonprogression to AIDS

PubMed Central

Limou, Sophie; Coulonges, Cédric; Herbeck, Joshua T.; van Manen, Daniëlle; An, Ping; Le Clerc, Sigrid; Delaneau, Olivier; Diop, Gora; Taing, Lieng; Montes, Matthieu; van't Wout, Angélique B.; Gottlieb, Geoffrey S.; Therwath, Amu; Rouzioux, Christine; Delfraissy, Jean-François; Lelièvre, Jean-Daniel; Lévy, Yves; Hercberg, Serge; Dina, Christian; Phair, John; Donfield, Sharyne; Goedert, James J.; Buchbinder, Susan; Estaquier, Jérôme; Schächter, François; Gut, Ivo; Froguel, Philippe; Mullins, James I.; Schuitemaker, Hanneke; Winkler, Cheryl; Zagury, Jean-François

2010-01-01

Background. The compilation of previous genomewide association studies of AIDS shows a major polymorphism in the HCP5 gene associated with both control of the viral load and long-term nonprogression (LTNP) to AIDS. Methods. To look for genetic variants that affect LTNP without necessary control of the viral load, we reanalyzed the genomewide data of the unique LTNP Genomics of Resistance to Immunodeficiency Virus (GRIV) cohort by excluding “elite controller” patients, who were controlling the viral load at very low levels (<100 copies/mL). Results. The rs2234358 polymorphism in the CXCR6 gene was the strongest signal (P = 2.5 × 10−7; odds ratio, 1.85) obtained for the genomewide association study comparing the 186 GRIV LTNPs who were not elite controllers with 697 uninfected control subjects. This association was replicated in 3 additional independent European studies, reaching genomewide significance of Pcombined = 9.7 × 10−10. This association with LTNP is independent of the combined CCR2-CCR5 locus and the HCP5 polymorphisms. Conclusion. The statistical significance, the replication, and the magnitude of the association demonstrate that CXCR6 is likely involved in the molecular etiology of AIDS and, in particular, in LTNP, emphasizing the power of extreme-phenotype cohorts. CXCR6 is a chemokine receptor that is known as a minor coreceptor in human immunodeficiency virus type 1 infection but could participate in disease progression through its role as a mediator of inflammation. PMID:20704485

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kubo, Yoshinao; Yoshii, Hiroaki; Kamiyama, Haruka

Ezrin, radixin, and moesin (ERM) proteins supply functional linkage between integral membrane proteins and cytoskeleton in mammalian cells to regulate membrane protein dynamisms and cytoskeleton rearrangement. To assess potential role of the ERM proteins in HIV-1 lifecycle, we examined if suppression of ERM function in human cells expressing HIV-1 infection receptors influences HIV-1 envelope (Env)-mediated HIV-1-vector transduction and cell-cell fusion. Expression of an ezrin dominant negative mutant or knockdown of ezrin, radixin, or moesin with siRNA uniformly decreased transduction titers of HIV-1 vectors having X4-tropic Env. In contrast, transduction titers of R5-tropic Env HIV-1 vectors were decreased only by radixinmore » knockdown: ezrin knockdown had no detectable effects and moesin knockdown rather increased transduction titer. Each of the ERM suppressions had no detectable effects on cell surface expression of CD4, CCR5, and CXCR4 or VSV-Env-mediated HIV-1 vector transductions. Finally, the individual knockdown of ERM mRNAs uniformly decreased efficiency of cell-cell fusion mediated by X4- or R5-tropic Env and HIV-1 infection receptors. These results suggest that (i) the ERM proteins function as positive regulators of infection by X4-tropic HIV-1, (ii) moesin additionally functions as a negative regulator of R5-tropic HIV-1 virus infection at the early step(s) after the membrane fusion, and (iii) receptor protein dynamisms are regulated differently in R5- and X4-tropic HIV-1 infections.« less

Stromal CCR6 drives tumor growth in a murine transplantable colon cancer through recruitment of tumor-promoting macrophages

PubMed Central

Nandi, Bisweswar; Shapiro, Mia; Samur, Mehmet K.; Pai, Christine; Frank, Natasha Y.; Yoon, Charles; Prabhala, Rao H.; Munshi, Nikhil C.; Gold, Jason S.

2016-01-01

ABSTRACT Interactions between the inflammatory chemokine CCL20 and its receptor CCR6 have been implicated in promoting colon cancer; however, the mechanisms behind this effect are poorly understood. We have previously demonstrated that deficiency of CCR6 is associated with decreased tumor macrophage accumulation in a model of sporadic intestinal tumorigenesis. In this study, we aimed to determine the role of stromal CCR6 expression in a murine syngeneic transplantable colon cancer model. We show that deficiency of host CCR6 is associated with decreased growth of syngeneic CCR6-expressing colon cancers. Colon cancers adoptively transplanted into CCR6-deficient mice have decreased tumor-associated macrophages without alterations in the number of monocytes in blood or bone marrow. CCL20, the unique ligand for CCR6, promotes migration of monocytes in vitro and promotes accumulation of macrophages in vivo. Depletion of tumor-associated macrophages decreases the growth of tumors in the transplantable tumor model. Macrophages infiltrating the colon cancers in this model secrete the inflammatory mediators CCL2, IL-1α, IL-6 and TNFα. Ccl2, Il1α and Il6 are consequently downregulated in tumors from CCR6-deficient mice. CCL2, IL-1α and IL-6 also promote proliferation of colon cancer cells, linking the decreased macrophage migration into tumors mediated by CCL20–CCR6 interactions to the delay in tumor growth in CCR6-deficient hosts. The relevance of these findings in human colon cancer is demonstrated through correlation of CCR6 expression with that of the macrophage marker CD163 as well as that of CCL2, IL1α and TNFα. Our findings support the exploration of targeting the CCL20–CCR6 pathway for the treatment of colon cancer. PMID:27622061

Different Pathogenesis of CCR5-Using Primary HIV-1 Isolates from Non-Switch and Switch Virus Patients in Human Lymphoid Tissue Ex Vivo

NASA Technical Reports Server (NTRS)

Iarlsson, Ingrid; Grivel, Jean-Charles; Chen. Silvia; Karlsson, Anders; Albert, Jan; Fenyol, Eva Maria; Margolis, Leonid B.

2005-01-01

CCR5-utilizing HIV-1 variants (R5) typically transmit infection and dominate its early stages, whereas emergence of CXCR4-using (X4 or R5X4) HIV-1 is often associated with disease progression. However, such a switch in co-receptor usage can only be detected in approximately onehalf of HIV-infected patients (switch virus patients), and progression to immunodeficiency may also occur in patients without detectable switch in co-receptor usage (non-switch virus patients). Here, we used a system of ex vivo-infected tonsillar tissue to compare the pathogenesis of sequential primary R5 HIV-1 isolates from the switch and non-switch patients. Inoculation of ex vivo tissue with these R5 isolates resulted in viral replication and CCR5(+)CD4(+) T cell depletion. The levels of such depletion by HIV-1 isolated from non-switch virus patients were significantly higher than those by R5 HIV-1 isolates from switch virus patients. T cell depletion seemed to be controlled by viral factors and did not significantly vary between tissues from different donors. In contrast, viral replication did not correlate with the switch status of the patients; in tissues fiom different donors it varied 30-fold and seemed to be controlled by a combination of viral and tissue factors. Nevertheless, replication-level hierarchy among sequential isolates remained constant in tissues from various donors. Viral load in vivo was higher in switch virus patients compared to non-switch virus patients. The high cytopathogenicity of CCR5(+)CD4(+) T cells by R5 HIV-1 isolates from non-switch virus patients may explain the steady decline of CD4(+) T cells in the absence of CXCR4 using virus; elimination of target cells by these isolates may limit their own replication in vivo.

SIV Coreceptor Specificity in Natural and Non-Natural Host Infection: Implications for Cell Targeting and Differential Outcomes from Infection.

PubMed

Wetzel, Katherine S; Elliott, Sarah T C; Collman, Ronald G

2018-01-01

Pathogenic HIV-1 infection of humans and SIVmac infection of macaques are the result of zoonotic transfer of primate immunodeficiency viruses from their natural hosts into non-natural host species. Natural host infections do not result in pathogenesis despite high levels of virus replication, and evidence suggests that differences in anatomical location and specific subsets of CD4+ T cells infected may underlie distinct outcomes from infection. The coreceptor CCR5 has long been considered the sole pathway for SIV entry and the key determinant of CD4+ cell targeting, but it has also been known that natural hosts express exceedingly low levels of CCR5 despite maintaining high levels of virus replication. This review details emerging data indicating that in multiple natural host species, CCR5 is dispensable for SIV infection ex vivo and/or in vivo and, contrary to the established dogma, alternative coreceptors, particularly CXCR6, play a central role in infection and cell targeting. Infections of non-natural hosts, however, are characterized by CCR5-exclusive entry. These findings suggest that alternative coreceptor-mediated cell targeting in natural hosts, combined with low CCR5 expression, may direct the virus to distinct populations of cells that are dispensable for immune homeostasis, particularly extralymphoid and more differentiated CD4+ T cells. In contrast, CCR5-mediated entry in non-natural hosts results in targeting of CD4+ T cells that are located in lymphoid tissues, critical for immune homeostasis, or necessary for gut barrier integrity. Thus, fundamental differences in viral entry coreceptor use may be central determinants of infection outcome. These findings redefine the normal SIV/host relationship in natural host species, shed new light on key features linked to zoonotic immunodeficiency virus transfer, and highlight important questions regarding how and why this coreceptor bottleneck occurs and the coevolutionary equilibrium is lost following cross-species transfer that results in AIDS. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Analysis of Staphylococcal cassette chromosome mec in Staphylococcus haemolyticus and Staphylococcus sciuri: identification of a novel ccr gene complex with a newly identified ccrA allotype (ccrA7).

PubMed

Urushibara, Noriko; Paul, Shyamal Kumar; Hossain, Mohammad Akram; Kawaguchiya, Mitsuyo; Kobayashi, Nobumichi

2011-06-01

Methicillin resistance in staphylococci is conferred by the acquisition in its chromosome of the mecA gene, which is located on a mobile genetic element called staphylococcal cassette chromosome mec (SCCmec). Genetic type of SCCmec is defined by combination of mec gene complex class and cassette chromosome recombinase gene (ccr) allotype. In this study, we analyzed genetic diversity of the SCCmec in 11 Staphylococcus haemolyticus strains and a Staphylococcus sciuri strain, which were recently isolated from clinical specimens in Bangladesh. Among these strains, only two S. haemolyticus strains were proved to have the known types of SCCmec, that is, SCCmec V (class C2 mec-ccrC) and VII (class C1 mec-ccrC). Five S. haemolyticus strains were assigned two unique mec-ccr gene complexes combination; that is, class C1 mec-ccrA4B4 (four isolates) and class A mec-ccrC (one isolate). In the remaining four S. haemolyticus strains with class C1 mec, no known ccr allotypes could be detected. A single S. sciuri strain with class A mec complex carried a ccrA gene belonging to a novel allotype designated ccrA7, together with ccrB3. The ccrA7 gene in the S. sciuri strain showed 61.7%-82.7% sequence identity to the ccrA gene sequences published so far, and 75.3% identity to ccrA3, which is a component of the type 3 ccr complex (ccrA3-ccrB3) in methicillin-resistant Staphylococcus aureus. The results of the present study indicated that mec gene complex and ccr genes in coagulase-negative staphylococci are highly divergent, and distinct from those of common methicillin-resistant S. aureus. Identification of the novel ccrA7 allotype combined with ccrB3 suggested an occurrence of recombination between different ccr complexes in nature.

Sensitivity Changes over the Course of Infection Increases the Likelihood of Resistance Against Fusion but Not CCR5 Receptor Blockers

PubMed Central

Chatziandreou, Nikolaos; Arauz, Ana Belen; Freitas, Ines; Nyein, Phyu Hninn; Fenton, Gregory; Mehta, Shruti H.; Kirk, Gregory D.

2012-01-01

Abstract As HIV-1 evolves over the course of infection, resistance against antiretrovirals may arise in the absence of drug pressure, especially against receptor and fusion blockers because of the extensive changes observed in the envelope glycoprotein. Here we show that viruses from the chronic phase of disease are significantly less sensitive to CCR5 receptor and fusion blockers compared to early infection variants. Differences in susceptibility to CCR5 antagonists were observed in spite of no demonstrable CXCR4 receptor utilization. No significant sensitivity differences were observed to another entry blocker, soluble CD4, or to reverse transcriptase, protease, or integrase inhibitors. Chronic as compared to early phase variants demonstrated greater replication when passaged in the presence of subinhibitory concentrations of fusion but not CCR5 receptor inhibitors. Fusion antagonist resistance, however, emerged from only one chronic phase virus culture. Because sensitivity to receptor and fusion antagonists is correlated with receptor affinity and fusion capacity, respectively, changes that occur in the envelope glycoprotein over the course of infection confer greater ability to use the CCR5 receptor and increased fusion ability. Our in vitro passage studies suggest that these evolving phenotypes increase the likelihood of resistance against fusion but not CCR5 receptor blockers. PMID:22650962

Closely related, yet unique: Distinct homo- and heterodimerization patterns of G protein coupled chemokine receptors and their fine-tuning by cholesterol

PubMed Central

Gahbauer, Stefan; Pluhackova, Kristyna

2018-01-01

Chemokine receptors, a subclass of G protein coupled receptors (GPCRs), play essential roles in the human immune system, they are involved in cancer metastasis as well as in HIV-infection. A plethora of studies show that homo- and heterodimers or even higher order oligomers of the chemokine receptors CXCR4, CCR5, and CCR2 modulate receptor function. In addition, membrane cholesterol affects chemokine receptor activity. However, structural information about homo- and heterodimers formed by chemokine receptors and their interplay with cholesterol is limited. Here, we report homo- and heterodimer configurations of the chemokine receptors CXCR4, CCR5, and CCR2 at atomistic detail, as obtained from thousands of molecular dynamics simulations. The observed homodimerization patterns were similar for the closely related CC chemokine receptors, yet they differed significantly between the CC receptors and CXCR4. Despite their high sequence identity, cholesterol modulated the CC homodimer interfaces in a subtype-specific manner. Chemokine receptor heterodimers display distinct dimerization patterns for CXCR4/CCR5 and CXCR4/CCR2. Furthermore, associations between CXCR4 and CCR5 reveal an increased cholesterol-sensitivity as compared to CXCR4/CCR2 heterodimerization patterns. This work provides a first comprehensive structural overview over the complex interaction network between chemokine receptors and indicates how heterodimerization and the interaction with the membrane environment diversifies the function of closely related GPCRs. PMID:29529028

Defining Differential Genetic Signatures in CXCR4- and the CCR5-Utilizing HIV-1 Co-Linear Sequences

PubMed Central

Aiamkitsumrit, Benjamas; Dampier, Will; Martin-Garcia, Julio; Nonnemacher, Michael R.; Pirrone, Vanessa; Ivanova, Tatyana; Zhong, Wen; Kilareski, Evelyn; Aldigun, Hazeez; Frantz, Brian; Rimbey, Matthew; Wojno, Adam; Passic, Shendra; Williams, Jean W.; Shah, Sonia; Blakey, Brandon; Parikh, Nirzari; Jacobson, Jeffrey M.; Moldover, Brian; Wigdahl, Brian

2014-01-01

The adaptation of human immunodeficiency virus type-1 (HIV-1) to an array of physiologic niches is advantaged by the plasticity of the viral genome, encoded proteins, and promoter. CXCR4-utilizing (X4) viruses preferentially, but not universally, infect CD4+ T cells, generating high levels of virus within activated HIV-1-infected T cells that can be detected in regional lymph nodes and peripheral blood. By comparison, the CCR5-utilizing (R5) viruses have a greater preference for cells of the monocyte-macrophage lineage; however, while R5 viruses also display a propensity to enter and replicate in T cells, they infect a smaller percentage of CD4+ T cells in comparison to X4 viruses. Additionally, R5 viruses have been associated with viral transmission and CNS disease and are also more prevalent during HIV-1 disease. Specific adaptive changes associated with X4 and R5 viruses were identified in co-linear viral sequences beyond the Env-V3. The in silico position-specific scoring matrix (PSSM) algorithm was used to define distinct groups of X4 and R5 sequences based solely on sequences in Env-V3. Bioinformatic tools were used to identify genetic signatures involving specific protein domains or long terminal repeat (LTR) transcription factor sites within co-linear viral protein R (Vpr), trans-activator of transcription (Tat), or LTR sequences that were preferentially associated with X4 or R5 Env-V3 sequences. A number of differential amino acid and nucleotide changes were identified across the co-linear Vpr, Tat, and LTR sequences, suggesting the presence of specific genetic signatures that preferentially associate with X4 or R5 viruses. Investigation of the genetic relatedness between X4 and R5 viruses utilizing phylogenetic analyses of complete sequences could not be used to definitively and uniquely identify groups of R5 or X4 sequences; in contrast, differences in the genetic diversities between X4 and R5 were readily identified within these co-linear sequences in HIV-1-infected patients. PMID:25265194

Sequential immunization with V3 peptides from primary human immunodeficiency virus type 1 produces cross-neutralizing antibodies against primary isolates with a matching narrow-neutralization sequence motif.

PubMed

Eda, Yasuyuki; Takizawa, Mari; Murakami, Toshio; Maeda, Hiroaki; Kimachi, Kazuhiko; Yonemura, Hiroshi; Koyanagi, Satoshi; Shiosaki, Kouichi; Higuchi, Hirofumi; Makizumi, Keiichi; Nakashima, Toshihiro; Osatomi, Kiyoshi; Tokiyoshi, Sachio; Matsushita, Shuzo; Yamamoto, Naoki; Honda, Mitsuo

2006-06-01

An antibody response capable of neutralizing not only homologous but also heterologous forms of the CXCR4-tropic human immunodeficiency virus type 1 (HIV-1) MNp and CCR5-tropic primary isolate HIV-1 JR-CSF was achieved through sequential immunization with a combination of synthetic peptides representing HIV-1 Env V3 sequences from field and laboratory HIV-1 clade B isolates. In contrast, repeated immunization with a single V3 peptide generated antibodies that neutralized only type-specific laboratory-adapted homologous viruses. To determine whether the cross-neutralization response could be attributed to a cross-reactive antibody in the immunized animals, we isolated a monoclonal antibody, C25, which neutralized the heterologous primary viruses of HIV-1 clade B. Furthermore, we generated a humanized monoclonal antibody, KD-247, by transferring the genes of the complementary determining region of C25 into genes of the human V region of the antibody. KD-247 bound with high affinity to the "PGR" motif within the HIV-1 Env V3 tip region, and, among the established reference antibodies, it most effectively neutralized primary HIV-1 field isolates possessing the matching neutralization sequence motif, suggesting its promise for clinical applications involving passive immunizations. These results demonstrate that sequential immunization with B-cell epitope peptides may contribute to a humoral immune-based HIV vaccine strategy. Indeed, they help lay the groundwork for the development of HIV-1 vaccine strategies that use sequential immunization with biologically relevant peptides to overcome difficulties associated with otherwise poorly immunogenic epitopes.

Sequential Immunization with V3 Peptides from Primary Human Immunodeficiency Virus Type 1 Produces Cross-Neutralizing Antibodies against Primary Isolates with a Matching Narrow-Neutralization Sequence Motif

PubMed Central

Eda, Yasuyuki; Takizawa, Mari; Murakami, Toshio; Maeda, Hiroaki; Kimachi, Kazuhiko; Yonemura, Hiroshi; Koyanagi, Satoshi; Shiosaki, Kouichi; Higuchi, Hirofumi; Makizumi, Keiichi; Nakashima, Toshihiro; Osatomi, Kiyoshi; Tokiyoshi, Sachio; Matsushita, Shuzo; Yamamoto, Naoki; Honda, Mitsuo

2006-01-01

An antibody response capable of neutralizing not only homologous but also heterologous forms of the CXCR4-tropic human immunodeficiency virus type 1 (HIV-1) MNp and CCR5-tropic primary isolate HIV-1 JR-CSF was achieved through sequential immunization with a combination of synthetic peptides representing HIV-1 Env V3 sequences from field and laboratory HIV-1 clade B isolates. In contrast, repeated immunization with a single V3 peptide generated antibodies that neutralized only type-specific laboratory-adapted homologous viruses. To determine whether the cross-neutralization response could be attributed to a cross-reactive antibody in the immunized animals, we isolated a monoclonal antibody, C25, which neutralized the heterologous primary viruses of HIV-1 clade B. Furthermore, we generated a humanized monoclonal antibody, KD-247, by transferring the genes of the complementary determining region of C25 into genes of the human V region of the antibody. KD-247 bound with high affinity to the “PGR” motif within the HIV-1 Env V3 tip region, and, among the established reference antibodies, it most effectively neutralized primary HIV-1 field isolates possessing the matching neutralization sequence motif, suggesting its promise for clinical applications involving passive immunizations. These results demonstrate that sequential immunization with B-cell epitope peptides may contribute to a humoral immune-based HIV vaccine strategy. Indeed, they help lay the groundwork for the development of HIV-1 vaccine strategies that use sequential immunization with biologically relevant peptides to overcome difficulties associated with otherwise poorly immunogenic epitopes. PMID:16699036

Ultra-violet radiation is responsible for the differences in global epidemiology of chickenpox and the evolution of varicella-zoster virus as man migrated out of Africa

PubMed Central

2011-01-01

Background Of the eight human herpes viruses, varicella-zoster virus, which causes chickenpox and zoster, has a unique epidemiology. Primary infection is much less common in children in the tropics compared with temperate areas. This results in increased adult susceptibility causing outbreaks, for example in health-care workers migrating from tropical to temperate countries. The recent demonstration that there are different genotypes of varicella-zoster virus and their geographic segregation into tropical and temperate areas suggests a distinct, yet previously unconsidered climatic factor may be responsible for both the clinical and molecular epidemiological features of this virus infection. Presentation of the hypothesis Unlike other human herpes viruses, varicella-zoster virus does not require intimate contact for infection to occur indicating that transmission may be interrupted by a geographically restricted climatic factor. The factor with the largest difference between tropical and temperate zones is ultra-violet radiation. This could reduce the infectiousness of chickenpox cases by inactivating virus in vesicles, before or after rupture. This would explain decreased transmissibility in the tropics and why the peak chickenpox incidence in temperate zones occurs during winter and spring, when ultra-violet radiation is at its lowest. The evolution of geographically restricted genotypes is also explained by ultra-violet radiation driving natural selection of different virus genotypes with varying degrees of resistance to inactivation, tropical genotypes being the most resistant. Consequently, temperate viruses should be more sensitive to its effects. This is supported by the observation that temperate genotypes are found in the tropics only in specific circumstances, namely where ultra-violet radiation has either been excluded or significantly reduced in intensity. Testing the Hypothesis The hypothesis is testable by exposing different virus genotypes to ultra-violet radiation and quantifying virus survival by plaque forming units or quantitative mRNA RT-PCR. Implications of the hypothesis The ancestral varicella-zoster virus, most probably a tropical genotype, co-migrated with man as he left Africa approximately 200,000 years ago. For this virus to have lost the selective advantage of resistance to ultra-violet radiation, the hypothesis would predict that the temperate, ultra-violet sensitive virus should have acquired another selective advantage as an evolutionary trade-off. One obvious advantage could be an increased reactivation rate as zoster to set up more rounds of chickenpox transmission. If this were so, the mechanism responsible for resistance to ultra-violet radiation might also be involved in reactivation and latency. This could then provide the first insight into a genetic correlate of the survival strategy of this virus. PMID:21513563

A Cell Line-Based Neutralization Assay for Primary Human Immunodeficiency Virus Type 1 Isolates That Use either the CCR5 or the CXCR4 Coreceptor

PubMed Central

Trkola, Alexandra; Matthews, Jamie; Gordon, Cynthia; Ketas, Tom; Moore, John P.

1999-01-01

We describe here a cell line-based assay for the evaluation of human immunodeficiency virus type 1 (HIV-1) neutralization. The assay is based on CEM.NKR cells, transfected to express the HIV-1 coreceptor CCR5 to supplement the endogenous expression of CD4 and the CXCR4 coreceptor. The resulting CEM.NKR-CCR5 cells efficiently replicate primary HIV-1 isolates of both R5 and X4 phenotypes. A comparison of the CEM.NKR-CCR5 cells with mitogen-activated peripheral blood mononuclear cells (PBMC) in neutralization assays with sera from HIV-1-infected individuals or specific anti-HIV-1 monoclonal antibodies shows that the sensitivity of HIV-1 neutralization is similar in the two cell types. The CEM.NKR-CCR5 cell assay, however, is more convenient to perform and eliminates the donor-to-donor variation in HIV-1 replication efficiency, which is one of the principal drawbacks of the PBMC-based neutralization assay. We suggest that this new assay is suitable for the general measurement of HIV-1 neutralization by antibodies. PMID:10516002

CD4-CCR5 interaction in intracellular compartments contributes to receptor expression at the cell surface

PubMed Central

Achour, Lamia; Scott, Mark G.H.; Shirvani, Hamasseh; Thuret, Alain; Bismuth, Georges; Labbé-Jullié, Catherine; Marullo, Stefano

2009-01-01

The association of CD4, a glycoprotein involved in T cell development and antigen recognition, and CCR5, a chemotactic G protein-coupled receptor, which regulates trafficking and effector functions of immune cells, forms the main receptor for the human immunodeficiency virus HIV. We observed that the vast majority of CCR5 is maintained within the intracellular compartments of primary T lymphocytes and in a monocytic cell line, contrasting with its relative low density at the cell surface. The CCR5-CD4 association, which occurs in the endoplasmic reticulum, enhanced CCR5 export to the plasma membrane in a concentration–dependent manner, whereas inhibition of endogenous CD4 with small interfering RNAs decreased cell surface expression of endogenous CCR5. This effect was specific for CCR5, as CD4 did not affect cell distribution of CXCR4, the other HIV co-receptor. These results reveal a previously unappreciated role of CD4, which contributes to regulate CCR5 export to the plasma membrane. PMID:19064722

Distribution of simian immunodeficiency virus target cells in vaginal tissues of normal rhesus macaques: implications for virus transmission.

PubMed

Poonia, Bhawna; Wang, Xiaolei; Veazey, Ronald S

2006-12-01

Most new cases of HIV-1 infection occur as the result of vaginal transmission. Identifying the phenotype and distribution of potential viral target cells in the vagina is important for understanding events in viral transmission and for developing effective prevention strategies. For example, compounds that prevent CD4 or CCR5 binding have been demonstrated recently to prevent vaginal transmission in rhesus macaques, but the expression and distribution of CCR5 has not been examined in the macaque vagina. The objective of this study was to examine the distribution and phenotype of cells and molecules in the vagina of rhesus macaques that may be involved in HIV transmission, including CCR5, CD3, CD4, CD8, CD1a, CD28, CD95, CD123 and HLA-DR. Normal juvenile and adult female rhesus macaques were examined by multicolor immunohistochemistry and flow cytometry. Although both CD4 and CCR5 were observed in the lamina propria, essentially no CD4 or CCR5 expression was detected within the squamous or keratinized layers of the vaginal epithelium. CCR5 expression was higher in the vaginal lamina propria of mature macaques compared to 1-3-year-old juveniles. The vast majority of CD4(+)CCR5(+) lymphocytes in the vagina had a central memory (CD95(+)CD28(+)) phenotype. Numerous CCR5-expressing dendritic cells (CD123(+)) or macrophages (CD68(+)) were observed in the lamina propria, but no CCR5, CD4 or DC-SIGN expression was detectable in the epithelium. Thus, the multiple layers of squamous epithelium normally covering the vaginal mucosa may provide an effective barrier against vaginal HIV-1 transmission. Microbicides that block CD4 or CCR5 expression may act within the deeper layers of the vaginal epithelium rather than on the epithelial surface.

[Companion Diagnostics for Selecting Antiretroviral Drugs against HIV-1].

PubMed

Fukutake, Katsuyuki

2015-11-01

Currently, the treatment of human immunodeficiency virus involves combination therapy, as antiretroviral therapy(ART). The treatment has improved steadily since the advent of potent combination therapy in 1996. New drugs that offer new mechanisms of action, improvements in potency and activity even against multidrug-resistant viruses, dosing convenience, and tolerability have been approved. Among ART with useful drugs, there are two important examinations before starting the treatment using the two kinds of drug. CCR5 co-receptor antagonists, maraviroc, prevent HIV entry into target cells by binding to CCR5 receptors. Genotypic assays have been developed that can determine or predict the co-receptor tropism(i.e., CCR5, CXCR4, or both) of the patient's dominant virus population. The assay for HIV-1 co-receptor usage should be performed whenever the use of a CCR5 antagonist is being considered. One of the nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), abacavir, is an important agent to develop recommended regimens for antiretroviral therapy. Serious and sometimes fatal hypersensitivity reactions have been associated with abacavir-containing products, ZIAGEN, Epzicom, and Triumeq. Patients who carry the HLA-B*5701 allele are at high-risk of a hypersensitivity reaction to abacavir. Prior to initiating therapy with abacavir, performing a screening test for the HLA-B*5701 allele is recommended. [Review].

Dynamics of CCR5 Expression by CD4+ T Cells in Lymphoid Tissues during Simian Immunodeficiency Virus Infection

PubMed Central

Veazey, Ronald S.; Mansfield, Keith G.; Tham, Irene C.; Carville, Angela C.; Shvetz, Daniel E.; Forand, Amy E.; Lackner, Andrew A.

2000-01-01

Early viral replication and profound CD4+ T-cell depletion occur preferentially in intestinal tissues of macaques infected with simian immunodeficiency virus (SIV). Here we show that a much higher percentage of CD4+ T cells in the intestine express CCR5 compared with those found in the peripheral blood, spleen, or lymph nodes. In addition, the selectivity and extent of the CD4+ T-cell loss in SIV infection may depend upon these cells coexpressing CCR5 and having a “memory” phenotype (CD45RA−). Following intravenous infection with SIVmac251, memory CD4+ CCR5+ T cells were selectively eliminated within 14 days in all major lymphoid tissues (intestine, spleen, and lymph nodes). However, the effect on CD4+ T-cell numbers was most profound in the intestine, where cells of this phenotype predominate. The CD4+ T cells that remain after 14 days of infection lacked CCR5 and/or were naive (CD45RA+). Furthermore, when animals in the terminal stages of SIV infection (with AIDS) were examined, virtually no CCR5-expressing CD4+ T cells were found in lymphoid tissues, and all of the remaining CD4+ T cells were naive and coexpressed CXCR4. These findings suggest that chemokine receptor usage determines which cells are targeted for SIV infection and elimination in vivo. PMID:11069995

CCR5 Expression Levels in HIV-Uninfected Women Receiving Hormonal Contraception

PubMed Central

Sciaranghella, Gaia; Wang, Cuiwei; Hu, Haihong; Anastos, Kathryn; Merhi, Zaher; Nowicki, Marek; Stanczyk, Frank Z.; Greenblatt, Ruth M.; Cohen, Mardge; Golub, Elizabeth T.; Watts, D. Heather; Alter, Galit; Young, Mary A.; Tsibris, Athe M. N.

2015-01-01

Human immunodeficiency virus (HIV) infectivity increases as receptor/coreceptor expression levels increase. We determined peripheral CD4, CCR5, and CXCR4 expression levels in HIV-uninfected women who used depot medroxyprogesterone acetate (DMPA; n = 32), the levonorgestrel-releasing intrauterine device (LNG-IUD; n = 27), oral contraceptive pills (n = 32), or no hormonal contraception (n = 33). The use of LNG-IUD increased the proportion of CD4+ and CD8+ T cells that expressed CCR5; increases in the magnitude of T-cell subset CCR5 expression were observed with DMPA and LNG-IUD use (P < .01 for all comparisons). LNG-IUD and, to a lesser extent, DMPA use were associated with increased peripheral T-cell CCR5 expression. PMID:25895986

Sargassum fusiforme fraction is a potent and specific inhibitor of HIV-1 fusion and reverse transcriptase

PubMed Central

Paskaleva, Elena E; Lin, Xudong; Duus, Karen; McSharry, James J; Veille, Jean-Claude L; Thornber, Carol; Liu, Yanze; Lee, David Yu-Wei; Canki, Mario

2008-01-01

Sargassum fusiforme (Harvey) Setchell has been shown to be a highly effective inhibitor of HIV-1 infection. To identify its mechanism of action, we performed bioactivity-guided fractionation on Sargassum fusiforme mixture. Here, we report isolation of a bioactive fraction SP4-2 (S. fusiforme), which at 8 μg/ml inhibited HIV-1 infection by 86.9%, with IC50 value of 3.7 μg. That represents 230-fold enhancement of antiretroviral potency as compared to the whole extract. Inhibition was mediated against both CXCR4 (X4) and CCR5 (R5) tropic HIV-1. Specifically, 10 μg/ml SP4-2 blocked HIV-1 fusion and entry by 53%. This effect was reversed by interaction of SP4-2 with sCD4, suggesting that S. fusiforme inhibits HIV-1 infection by blocking CD4 receptor, which also explained observed inhibition of both X4 and R5-tropic HIV-1. SP4-2 also inhibited HIV-1 replication after virus entry, by directly inhibiting HIV-1 reverse transcriptase (RT) in a dose dependent manner by up to 79%. We conclude that the SP4-2 fraction contains at least two distinct and biologically active molecules, one that inhibits HIV-1 fusion by interacting with CD4 receptor, and another that directly inhibits HIV-1 RT. We propose that S. fusiforme is a lead candidate for anti-HIV-1 drug development. PMID:18197976

HIV-1 with Multiple CCR5/CXCR4 Chimeric Receptor Use Is Predictive of Immunological Failure in Infected Children

PubMed Central

Cavarelli, Mariangela; Karlsson, Ingrid; Zanchetta, Marisa; Antonsson, Liselotte; Plebani, Anna; Giaquinto, Carlo; Fenyö, Eva Maria; De Rossi, Anita; Scarlatti, Gabriella

2008-01-01

Background HIV-1 R5 viruses are characterized by a large phenotypic variation, that is reflected by the mode of coreceptor use. The ability of R5 HIV-1 to infect target cells expressing chimeric receptors between CCR5 and CXCR4 (R5broad viruses), was shown to correlate with disease stage in HIV-1 infected adults. Here, we ask the question whether phenotypic variation of R5 viruses could play a role also in mother-to-child transmission (MTCT) of HIV-1 and pediatric disease progression. Methodology/Principal Findings Viral isolates obtained from a total of 59 HIV-1 seropositive women (24 transmitting and 35 non transmitting) and 28 infected newborn children, were used to infect U87.CD4 cells expressing wild type or six different CCR5/CXCR4 chimeric receptors. HIV-1 isolates obtained from newborn infants had predominantly R5narrow phenotype (n = 20), but R5broad and R5X4 viruses were also found in seven and one case, respectively. The presence of R5broad and R5X4 phenotypes correlated significantly with a severe decline of the CD4+ T cells (CDC stage 3) or death within 2 years of age. Forty-three percent of the maternal R5 isolates displayed an R5broad phenotype, however, the presence of the R5broad virus was not predictive for MTCT of HIV-1. Of interest, while only 1 of 5 mothers with an R5X4 virus transmitted the dualtropic virus, 5 of 6 mothers carrying R5broad viruses transmitted viruses with a similar broad chimeric coreceptor usage. Thus, the maternal R5broad phenotype was largely preserved during transmission and could be predictive of the phenotype of the newborn's viral variant. Conclusions/Significance Our results show that R5broad viruses are not hampered in transmission. When transmitted, immunological failure occurs earlier than in children infected with HIV-1 of R5narrow phenotype. We believe that this finding is of utmost relevance for therapeutic interventions in pediatric HIV-1 infection. PMID:18820725

HIV-1 with multiple CCR5/CXCR4 chimeric receptor use is predictive of immunological failure in infected children.

PubMed

Cavarelli, Mariangela; Karlsson, Ingrid; Zanchetta, Marisa; Antonsson, Liselotte; Plebani, Anna; Giaquinto, Carlo; Fenyö, Eva Maria; De Rossi, Anita; Scarlatti, Gabriella

2008-09-29

HIV-1 R5 viruses are characterized by a large phenotypic variation, that is reflected by the mode of coreceptor use. The ability of R5 HIV-1 to infect target cells expressing chimeric receptors between CCR5 and CXCR4 (R5(broad) viruses), was shown to correlate with disease stage in HIV-1 infected adults. Here, we ask the question whether phenotypic variation of R5 viruses could play a role also in mother-to-child transmission (MTCT) of HIV-1 and pediatric disease progression. Viral isolates obtained from a total of 59 HIV-1 seropositive women (24 transmitting and 35 non transmitting) and 28 infected newborn children, were used to infect U87.CD4 cells expressing wild type or six different CCR5/CXCR4 chimeric receptors. HIV-1 isolates obtained from newborn infants had predominantly R5(narrow) phenotype (n = 20), but R5(broad) and R5X4 viruses were also found in seven and one case, respectively. The presence of R5(broad) and R5X4 phenotypes correlated significantly with a severe decline of the CD4+ T cells (CDC stage 3) or death within 2 years of age. Forty-three percent of the maternal R5 isolates displayed an R5(broad) phenotype, however, the presence of the R5(broad) virus was not predictive for MTCT of HIV-1. Of interest, while only 1 of 5 mothers with an R5X4 virus transmitted the dualtropic virus, 5 of 6 mothers carrying R5(broad) viruses transmitted viruses with a similar broad chimeric coreceptor usage. Thus, the maternal R5(broad) phenotype was largely preserved during transmission and could be predictive of the phenotype of the newborn's viral variant. Our results show that R5(broad) viruses are not hampered in transmission. When transmitted, immunological failure occurs earlier than in children infected with HIV-1 of R5(narrow) phenotype. We believe that this finding is of utmost relevance for therapeutic interventions in pediatric HIV-1 infection.

Targeting both viral and host determinants of human immunodeficiency virus entry, using a new lentiviral vector coexpressing the T20 fusion inhibitor and a selective CCL5 intrakine.

PubMed

Petit, Nicolas; Dorgham, Karim; Levacher, Béatrice; Burlion, Aude; Gorochov, Guy; Marodon, Gilles

2014-08-01

Numerous strategies targeting early and late steps of the HIV life cycle have been proposed for gene therapy. However, targeting viral and host determinants of HIV entry is the only strategy that would prevent viral DNA-mediated CD4(+) cell death while diminishing the possibility for the virus to escape. To this end, we devised a bicistronic lentiviral vector expressing the membrane-bound form of the T20 fusion inhibitor, referred to as the C46 peptide, and a CCR5 superagonist, modified to sequester CCR5 away from the cell surface, referred to as the P2-CCL5 intrakine. We tested the effects of the vector on HIV infection and replication, using the human CEMR5 cell line expressing CD4 and CCR5, and primary human T cells. Transduced cells expressed the C46 peptide, detected with the 2F5 monoclonal antibody by flow cytometry. Expression of the P2-CCL5 intrakine correlates with lower levels of cell surface CCR5. Complete protection against HIV infection could be observed in cells expressing the protective transgenes. Importantly, we show that the combination of the transgenes was more potent than either transgene alone, showing the interest of expressing two entry inhibitors to inhibit HIV infection. Last, genetically modified cells possessed a selective advantage over nonmodified cells on HIV challenge in vitro, showing that modified cells were protected from HIV-induced cell death. Our results demonstrate that lentiviral vectors coexpressing the T20 fusion inhibitor and the P2-CCL5 intrakine represent promising tools for HIV gene therapy.

CXCR6-Mediated Simian Immunodeficiency Virus SIVagmSab Entry into Sabaeus African Green Monkey Lymphocytes Implicates Widespread Use of Non-CCR5 Pathways in Natural Host Infections

PubMed Central

Wetzel, Katherine S.; Yi, Yanjie; Elliott, Sarah T. C.; Romero, Dino; Jacquelin, Beatrice; Hahn, Beatrice H.; Muller-Trutwin, Michaela; Apetrei, Cristian; Pandrea, Ivona

2016-01-01

ABSTRACT African green monkeys (AGM) and sooty mangabeys (SM) are well-studied natural hosts of simian immunodeficiency virus (SIV) that do not progress to AIDS when infected with their species-specific viruses. Natural hosts of SIV express very low levels of the canonical entry coreceptor CCR5, and recent studies have shown that CCR5 is dispensable for SIV infection of SM in vivo and that blocking of CCR5 does not prevent ex vivo infection of peripheral blood mononuclear cells (PBMC) from SM or vervet AGM. In both hosts, CXCR6 is an efficient entry pathway in vitro. Here we investigated the use of species-matched CXCR6 and other alternative coreceptors by SIVagmSab, which infects sabaeus AGM. We cloned sabaeus CD4 and 10 candidate coreceptors. Species-matched CXCR6, CCR5, and GPR15 mediated robust entry into transfected cells by pseudotypes carrying SIVagmSab92018ivTF Env, with lower-level entry through GPR1 and APJ. We cloned genetically divergent env genes from the plasma of two wild-infected sabaeus AGM and found similar patterns of coreceptor use. Titration experiments showed that CXCR6 and CCR5 were more efficient than other coreceptors when tested at limiting CD4/coreceptor levels. Finally, blocking of CXCR6 with its ligand CXCL16 significantly inhibited SIVagmSab replication in sabaeus PBMC and had a greater impact than did the CCR5 blocker maraviroc, confirming the use of CXCR6 in primary lymphocyte infection. These data suggest a new paradigm for SIV infection of natural host species, whereby a shared outcome of virus-host coevolution is the use of CXCR6 or other alternative coreceptors for entry, which may direct SIV toward CD4+ T cell subsets and anatomical sites that support viral replication without disrupting immune homeostasis and function. IMPORTANCE Natural hosts of SIV do not progress to AIDS, in stark contrast to pathogenic human immunodeficiency virus type 1 (HIV-1)-human and SIVmac-macaque infections. Identifying how natural hosts avoid immunodeficiency can elucidate key mechanisms of pathogenesis. It is known that despite high viral loads, natural hosts have a low frequency of CD4+ cells expressing the SIV coreceptor CCR5. In this study, we demonstrate the efficient use of the coreceptor CXCR6 by SIVagmSab to infect sabaeus African green monkey lymphocytes. In conjunction with studies of SIVsmm, which infects sooty mangabeys, and SIVagmVer, which infects vervet monkeys, our data suggest a unifying model whereby in natural hosts, in which the CCR5 expression level is low, the use of CXCR6 or other coreceptors to mediate infection may target SIV toward distinct cell populations that are able to support high-level viral replication without causing a loss of CD4+ T cell homeostasis and lymphoid tissue damage that lead to AIDS in HIV-1 and SIVmac infections. PMID:27903799

CXCR6-Mediated Simian Immunodeficiency Virus SIVagmSab Entry into Sabaeus African Green Monkey Lymphocytes Implicates Widespread Use of Non-CCR5 Pathways in Natural Host Infections.

PubMed

Wetzel, Katherine S; Yi, Yanjie; Elliott, Sarah T C; Romero, Dino; Jacquelin, Beatrice; Hahn, Beatrice H; Muller-Trutwin, Michaela; Apetrei, Cristian; Pandrea, Ivona; Collman, Ronald G

2017-02-15

African green monkeys (AGM) and sooty mangabeys (SM) are well-studied natural hosts of simian immunodeficiency virus (SIV) that do not progress to AIDS when infected with their species-specific viruses. Natural hosts of SIV express very low levels of the canonical entry coreceptor CCR5, and recent studies have shown that CCR5 is dispensable for SIV infection of SM in vivo and that blocking of CCR5 does not prevent ex vivo infection of peripheral blood mononuclear cells (PBMC) from SM or vervet AGM. In both hosts, CXCR6 is an efficient entry pathway in vitro Here we investigated the use of species-matched CXCR6 and other alternative coreceptors by SIVagmSab, which infects sabaeus AGM. We cloned sabaeus CD4 and 10 candidate coreceptors. Species-matched CXCR6, CCR5, and GPR15 mediated robust entry into transfected cells by pseudotypes carrying SIVagmSab92018ivTF Env, with lower-level entry through GPR1 and APJ. We cloned genetically divergent env genes from the plasma of two wild-infected sabaeus AGM and found similar patterns of coreceptor use. Titration experiments showed that CXCR6 and CCR5 were more efficient than other coreceptors when tested at limiting CD4/coreceptor levels. Finally, blocking of CXCR6 with its ligand CXCL16 significantly inhibited SIVagmSab replication in sabaeus PBMC and had a greater impact than did the CCR5 blocker maraviroc, confirming the use of CXCR6 in primary lymphocyte infection. These data suggest a new paradigm for SIV infection of natural host species, whereby a shared outcome of virus-host coevolution is the use of CXCR6 or other alternative coreceptors for entry, which may direct SIV toward CD4 + T cell subsets and anatomical sites that support viral replication without disrupting immune homeostasis and function. Natural hosts of SIV do not progress to AIDS, in stark contrast to pathogenic human immunodeficiency virus type 1 (HIV-1)-human and SIVmac-macaque infections. Identifying how natural hosts avoid immunodeficiency can elucidate key mechanisms of pathogenesis. It is known that despite high viral loads, natural hosts have a low frequency of CD4 + cells expressing the SIV coreceptor CCR5. In this study, we demonstrate the efficient use of the coreceptor CXCR6 by SIVagmSab to infect sabaeus African green monkey lymphocytes. In conjunction with studies of SIVsmm, which infects sooty mangabeys, and SIVagmVer, which infects vervet monkeys, our data suggest a unifying model whereby in natural hosts, in which the CCR5 expression level is low, the use of CXCR6 or other coreceptors to mediate infection may target SIV toward distinct cell populations that are able to support high-level viral replication without causing a loss of CD4 + T cell homeostasis and lymphoid tissue damage that lead to AIDS in HIV-1 and SIVmac infections. Copyright © 2017 American Society for Microbiology.

Protection of rhesus macaques from vaginal infection by vaginally delivered maraviroc, an inhibitor of HIV-1 entry via the CCR5 co-receptor.

PubMed

Veazey, Ronald S; Ketas, Thomas J; Dufour, Jason; Moroney-Rasmussen, Terri; Green, Linda C; Klasse, P J; Moore, John P

2010-09-01

An effective vaginal microbicide could reduce human immunodeficiency virus type 1 (HIV-1) transmission to women. Among microbicide candidates in clinical development is Maraviroc (MVC), a small-molecule drug that binds the CCR5 co-receptor and impedes HIV-1 entry into cells. Delivered systemically, MVC reduces viral load in HIV-1-infected individuals, but its ability to prevent transmission is untested. We have now evaluated MVC as a vaginal microbicide with use of a stringent model that involves challenge of rhesus macaques with a high-dose of a CCR5-using virus, SHIV-162P3. Gel-formulated, prescription-grade MVC provided dose-dependent protection, half-maximally at 0.5 mM (0.25 mg/mL). The duration of protection was transient; the longer the delay between MVC application and virus challenge, the less protection (half life of approximately 4 h). As expected, MVC neither protected against challenge with a CXCR4-using virus, SHIV-KU1, nor exacerbated postinfection viremia. These findings validate MVC development as a vaginal microbicide for women and should guide clinical programs.

A Novel Role for the Receptor of the Complement Cleavage Fragment C5a, C5aR1, in CCR5-Mediated Entry of HIV into Macrophages.

PubMed

Moreno-Fernandez, Maria E; Aliberti, Julio; Groeneweg, Sander; Köhl, Jörg; Chougnet, Claire A

2016-04-01

The complement system is an ancient pattern recognition system that becomes activated during all stages of HIV infection. Previous studies have shown that C5a can enhance the infection of monocyte-derived macrophages and T cells indirectly through the production of interleukin (IL)-6 and tumor necrosis factor (TNF)-α and the attraction of dendritic cells. C5a exerts its multiple biologic functions mainly through activation of C5a receptor 1 (C5aR1). Here, we assessed the role of C5aR1 as an enhancer of CCR5-mediated HIV infection. We determined CCR5 and C5aR1 heterodimer formation in myeloid cells and the impact of C5aR1 blockade on HIV entry and genomic integration. C5aR1/CCR5 heterodimer formation was identified by immunoprecipitation and western blotting. THP-1 cells and monocyte-derived macrophages (MDM) were infected by R5 laboratory strains or HIV pseudotyped for the vesicular stomatitis virus (VSV) envelope. Levels of integrated HIV were measured by quantitative PCR after targeting of C5aR1 by a C5aR antagonist, neutralizing C5aR1 monoclonal antibody (mAb) or hC5a. C5aR1 was also silenced by specific siRNA prior to viral entry. We found that C5aR1 forms heterodimers with the HIV coreceptor CCR5 in myeloid cells. Targeting C5aR1 significantly decreased integration by R5 viruses but not by VSV-pseudotyped viruses, suggesting that C5aR1 is critical for viral entry. The level of inhibition achieved with C5aR1-blocking reagents was comparable to that of CCR5 antagonists. Mechanistically, C5aR1 targeting decreased CCR5 expression. MDM from CCR5Δ32 homozygous subjects expressed levels of C5aR1 similar to CCR5 WT individuals, suggesting that mere C5aR1 expression is not sufficient for HIV infection. HIV appeared to preferentially enter THP-1 cells expressing high levels of both C5aR1 and CCR5. Targeted reduction of C5aR1 expression in such cells reduced HIV infection by ~50%. Our data thus suggest that C5aR1 acts as an enhancer of CCR5-mediated HIV entry into macrophages, the targeting of which may prove useful to reduce HIV infection by R5 strains.

The puzzling role of CXCR4 in human immunodeficiency virus infection.

PubMed

Vicenzi, Elisa; Liò, Pietro; Poli, Guido

2013-01-01

The human immunodeficiency virus type-1 (HIV-1) is the etiological agent of the acquired immunodeficiency syndrome (AIDS), a disease highly lethal in the absence of combination antiretroviral therapy. HIV infects CD4(+) cells of the immune system (T cells, monocyte-macrophages and dendritic cells) via interaction with a universal primary receptor, the CD4 molecule, followed by a mandatory interaction with a second receptor (co-receptor) belonging to the chemokine receptor family. Apart from some rare cases, two chemokine receptors have been evolutionarily selected to accomplish this need for HIV-1: CCR5 and CXCR4. Yet, usage of these two receptors appears to be neither casual nor simply explained by their levels of cell surface expression. While CCR5 use is the universal rule at the start of every infection regardless of the transmission route (blood-related, sexual or mother to child), CXCR4 utilization emerges later in disease coinciding with the immunological deficient phase of infection. Moreover, in most instances CXCR4 use as viral entry co-receptor is associated with maintenance of CCR5 use. Since antiviral agents preventing CCR5 utilization by the virus are already in use, while others targeting either CCR5 or CXCR4 (or both) are under investigation, understanding the biological correlates of this "asymmetrical" utilization of HIV entry co-receptors bears relevance for the clinical choice of which therapeutics should be administered to infected individuals. We will here summarize the basic knowledge and the hypotheses underlying the puzzling and yet unequivocal role of CXCR4 in HIV-1 infection.

CCR4 frameshift mutation identifies a distinct group of adult T cell leukaemia/lymphoma with poor prognosis.

PubMed

Yoshida, Noriaki; Miyoshi, Hiroaki; Kato, Takeharu; Sakata-Yanagimoto, Mamiko; Niino, Daisuke; Taniguchi, Hiroaki; Moriuchi, Yukiyoshi; Miyahara, Masaharu; Kurita, Daisuke; Sasaki, Yuya; Shimono, Joji; Kawamoto, Keisuke; Utsunomiya, Atae; Imaizumi, Yoshitaka; Seto, Masao; Ohshima, Koichi

2016-04-01

Adult T cell leukaemia/lymphoma (ATLL) is an intractable T cell neoplasm caused by human T cell leukaemia virus type 1. Next-generation sequencing-based comprehensive mutation studies have revealed recurrent somatic CCR4 mutations in ATLL, although clinicopathological findings associated with CCR4 mutations remain to be delineated. In the current study, 184 cases of peripheral T cell lymphoma, including 113 cases of ATLL, were subjected to CCR4 mutation analysis. This sequence analysis identified mutations in 27% (30/113) of cases of ATLL and 9% (4/44) of cases of peripheral T cell lymphoma not otherwise specified. Identified mutations included nonsense (NS) and frameshift (FS) mutations. No significant differences in clinicopathological findings were observed between ATLL cases stratified by presence of CCR4 mutation. All ATLL cases with CCR4 mutations exhibited cell-surface CCR4 positivity. Semi-quantitative CCR4 protein analysis of immunohistochemical sections revealed higher CCR4 expression in cases with NS mutations of CCR4 than in cases with wild-type (WT) CCR4. Furthermore, among ATLL cases, FS mutation was significantly associated with a poor prognosis, compared with NS mutation and WT CCR4. These results suggest that CCR4 mutation is an important determinant of the clinical course in ATLL cases, and that NS and FS mutations of CCR4 behave differently with respect to ATLL pathophysiology. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Type III interferon is a critical regulator of innate antifungal immunity.

PubMed

Espinosa, Vanessa; Dutta, Orchi; McElrath, Constance; Du, Peicheng; Chang, Yun-Juan; Cicciarelli, Bryan; Pitler, Amy; Whitehead, Ian; Obar, Joshua J; Durbin, Joan E; Kotenko, Sergei V; Rivera, Amariliz

2017-10-06

Type III interferons (IFN-λs) are the most recently found members of the IFN cytokine family and engage IFNLR1 and IL10R2 receptor subunits to activate innate responses against viruses. We have identified IFN-λs as critical instructors of antifungal neutrophil responses. Using Aspergillus fumigatus ( Af ) as a model to study antifungal immune responses, we found that depletion of CCR2 + monocytes compromised the ability of neutrophils to control invasive fungal growth. Using an unbiased approach, we identified type I and III IFNs as critical regulators of the interplay between monocytes and neutrophils responding to Af We found that CCR2 + monocytes are an important early source of type I IFNs that prime optimal expression of IFN-λ. Type III IFNs act directly on neutrophils to activate their antifungal response, and mice with neutrophil-specific deletion of IFNLR1 succumb to invasive aspergillosis. Dysfunctional neutrophil responses in CCR2-depleted mice were rescued by adoptive transfer of pulmonary CCR2 + monocytes or by exogenous administration of IFN-α and IFN-λ. Thus, CCR2 + monocytes promote optimal activation of antifungal neutrophils by initiating a coordinated IFN response. We have identified type III IFNs as critical regulators of neutrophil activation and type I IFNs as early stimulators of IFN-λ expression. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Infection of rhesus macaques with a pool of simian immunodeficiency virus with the envelope genes from acute HIV-1 infections.

PubMed

Krebs, Kendall C; Tian, Meijuan; Asmal, Mohammed; Ling, Binhua; Nelson, Kenneth; Henry, Kenneth; Gibson, Richard; Li, Yuejin; Han, Weining; Shattock, Robin J; Veazey, Ronald S; Letvin, Norman; Arts, Eric J; Gao, Yong

2016-11-25

New simian-human immunodeficiency chimeric viruses with an HIV-1 env (SHIVenv) are critical for studies on HIV pathogenesis, vaccine development, and microbicide testing. Macaques are typically exposed to single CCR5-using SHIVenv which in most instances does not reflect the conditions during acute/early HIV infection (AHI) in humans. Instead of individual and serial testing new SHIV constructs, a pool of SHIVenv_B derived from 16 acute HIV-1 infections were constructed using a novel yeast-based SHIV cloning approach and then used to infect macaques. Even though none of the 16 SHIVenvs contained the recently reported mutations in env genes that could significantly enhance their binding affinity to RhCD4, one SHIVenv (i.e. SHIVenv_B3-PRB926) established infection in macaques exposed to this pool. AHI SHIVenv_B viruses as well as their HIVenv_B counterparts were analyzed for viral protein content, function, and fitness to identify possible difference between SHIVenv_B3-PRB926 and the other 15 SHIVenvs in the pool. All of the constructs produced SHIV or HIV chimeric with wild type levels of capsid (p27 and p24) content, reverse transcriptase (RT) activity, and expressed envelope glycoproteins that could bind to cell receptors CD4/CCR5 and mediate virus entry. HIV-1env_B chimeric viruses were propagated in susceptible cell lines but the 16 SHIVenv_B variants showed only limited replication in macaque peripheral blood mononuclear cells (PBMCs) and 174×CEM.CCR5 cell line. AHI chimeric viruses including HIVenv_B3 showed only minor variations in cell entry efficiency and kinetics as well as replicative fitness in human PBMCs. Reduced number of N-link glycosylation sites and slightly greater CCR5 affinity/avidity was the only distinguishing feature of env_B3 versus other AHI env's in the pool, a feature also observed in the HIV establishing new infections in humans. Despite the inability to propagate in primary cells and cell lines, a pool of 16 SHIVenv viruses could establish infection but only one virus, SHIVenv_B3 was isolated in the macaque and then shown to repeatedly infected macaques. This SHIVenv_B3 virus did not show any distinct phenotypic property from the other 15 SHIVenv viruses but did have the fewest N-linked glycosylation sites.

Elucidating a Key Anti-HIV-1 and Cancer-Associated Axis: The Structure of CCL5 (Rantes) in Complex with CCR5

NASA Astrophysics Data System (ADS)

Tamamis, Phanourios; Floudas, Christodoulos A.

2014-06-01

CCL5 (RANTES) is an inflammatory chemokine which binds to chemokine receptor CCR5 and induces signaling. The CCL5:CCR5 associated chemotactic signaling is of critical biological importance and is a potential HIV-1 therapeutic axis. Several studies provided growing evidence for the expression of CCL5 and CCR5 in non-hematological malignancies. Therefore, the delineation of the CCL5:CCR5 complex structure can pave the way for novel CCR5-targeted drugs. We employed a computational protocol which is primarily based on free energy calculations and molecular dynamics simulations, and report, what is to our knowledge, the first computationally derived CCL5:CCR5 complex structure which is in excellent agreement with experimental findings and clarifies the functional role of CCL5 and CCR5 residues which are associated with binding and signaling. A wealth of polar and non-polar interactions contributes to the tight CCL5:CCR5 binding. The structure of an HIV-1 gp120 V3 loop in complex with CCR5 has recently been derived through a similar computational protocol. A comparison between the CCL5 : CCR5 and the HIV-1 gp120 V3 loop : CCR5 complex structures depicts that both the chemokine and the virus primarily interact with the same CCR5 residues. The present work provides insights into the blocking mechanism of HIV-1 by CCL5.

Sexually-Transmitted/Founder HIV-1 Cannot Be Directly Predicted from Plasma or PBMC-Derived Viral Quasispecies in the Transmitting Partner

PubMed Central

Frange, Pierre; Meyer, Laurence; Jung, Matthieu; Goujard, Cecile; Zucman, David; Abel, Sylvie; Hochedez, Patrick; Gousset, Marine; Gascuel, Olivier; Rouzioux, Christine; Chaix, Marie-Laure

2013-01-01

Objective Characterization of HIV-1 sequences in newly infected individuals is important for elucidating the mechanisms of viral sexual transmission. We report the identification of transmitted/founder viruses in eight pairs of HIV-1 sexually-infected patients enrolled at the time of primary infection (“recipients”) and their transmitting partners (“donors”). Methods Using a single genome-amplification approach, we compared quasispecies in donors and recipients on the basis of 316 and 376 C2V5 env sequences amplified from plasma viral RNA and PBMC-associated DNA, respectively. Results Both DNA and RNA sequences indicated very homogeneous viral populations in all recipients, suggesting transmission of a single variant, even in cases of recent sexually transmitted infections (STIs) in donors (n = 2) or recipients (n = 3). In all pairs, the transmitted/founder virus was derived from an infrequent variant population within the blood of the donor. The donor variant sequences most closely related to the recipient sequences were found in plasma samples in 3/8 cases and/or in PBMC samples in 6/8 cases. Although donors were exclusively (n = 4) or predominantly (n = 4) infected by CCR5-tropic (R5) strains, two recipients were infected with highly homogeneous CXCR4/dual-mixed-tropic (X4/DM) viral populations, identified in both DNA and RNA. The proportion of X4/DM quasispecies in donors was higher in cases of X4/DM than R5 HIV transmission (16.7–22.0% versus 0–2.6%), suggesting that X4/DM transmission may be associated with a threshold population of X4/DM circulating quasispecies in donors. Conclusions These suggest that a severe genetic bottleneck occurs during subtype B HIV-1 heterosexual and homosexual transmission. Sexually-transmitted/founder virus cannot be directly predicted by analysis of the donor’s quasispecies in plasma and/or PBMC. Additional studies are required to fully understand the traits that confer the capacity to transmit and establish infection, and determine the role of concomitant STIs in mitigating the genetic bottleneck in mucosal HIV transmission. PMID:23874894

Comparison of Human Immunodeficiency Virus Type 1 Tropism Profiles in Clinical Samples by the Trofile and MT-2 Assays▿

PubMed Central

Coakley, Eoin; Reeves, Jacqueline D.; Huang, Wei; Mangas-Ruiz, Marga; Maurer, Irma; Harskamp, Agnes M.; Gupta, Soumi; Lie, Yolanda; Petropoulos, Christos J.; Schuitemaker, Hanneke; van 't Wout, Angélique B.

2009-01-01

The recent availability of CCR5 antagonists as anti-human immunodeficiency virus (anti-HIV) therapeutics has highlighted the need to accurately identify CXCR4-using variants in patient samples when use of this new drug class is considered. The Trofile assay (Monogram Biosciences) has become the method that is the most widely used to define tropism in the clinic prior to the use of a CCR5 antagonist. By comparison, the MT-2 assay has been used since early in the HIV epidemic to define tropism in clinical specimens. Given that there are few data from direct comparisons of these two assays, we evaluated the performance of the plasma-based Trofile assay and the peripheral blood mononuclear cell (PBMC)-based MT-2 assay for the detection of CXCR4 use in defining the tropism of HIV isolates derived from clinical samples. The various samples used for this comparison were derived from participants of the Amsterdam Cohort Studies on HIV infection and AIDS who underwent consecutive MT-2 assay testing of their PBMCs at approximately 3-month intervals. This unique sample set was specifically selected because consecutive MT-2 assays had demonstrated a shift from negative to positive in PBMCs, reflecting the first emergence of CXCR4-using virus in PBMCs above the level of detection of the assay in these individuals. Trofile testing was performed with clonal HIV type 1 (HIV-1) variants (n = 21), MT-2 cell culture-derived cells (n = 20) and supernatants (n = 42), and plasma samples (n = 76). Among the clonal HIV-1 variants and MT-2 cell culture-derived samples, the results of the Trofile and MT-2 assays demonstrated a high degree of concordance (95% to 98%). Among consecutive plasma samples, detection of CXCR4-using virus was at or before the time of first detection by the MT-2 assay in 5/10 patients by the original Trofile assay and in 9/10 patients by the enhanced-sensitivity Trofile assay. Differences in the time to the first detection of CXCR4 use between the MT-2 assay (PBMCs) and the original Trofile assay (plasma) were greatly reduced by the enhanced-sensitivity Trofile assay, suggesting that sensitivity for the detection of minor CXCR4-using variants may be a more important determinant of discordant findings than compartmentalization. The similarities in performance of the enhanced-sensitivity Trofile and MT-2 assays suggest that either may be an appropriate methodology to define tropism in patient specimens. PMID:19687240

Comparison of human immunodeficiency virus type 1 tropism profiles in clinical samples by the Trofile and MT-2 assays.

PubMed

Coakley, Eoin; Reeves, Jacqueline D; Huang, Wei; Mangas-Ruiz, Marga; Maurer, Irma; Harskamp, Agnes M; Gupta, Soumi; Lie, Yolanda; Petropoulos, Christos J; Schuitemaker, Hanneke; van 't Wout, Angélique B

2009-11-01

The recent availability of CCR5 antagonists as anti-human immunodeficiency virus (anti-HIV) therapeutics has highlighted the need to accurately identify CXCR4-using variants in patient samples when use of this new drug class is considered. The Trofile assay (Monogram Biosciences) has become the method that is the most widely used to define tropism in the clinic prior to the use of a CCR5 antagonist. By comparison, the MT-2 assay has been used since early in the HIV epidemic to define tropism in clinical specimens. Given that there are few data from direct comparisons of these two assays, we evaluated the performance of the plasma-based Trofile assay and the peripheral blood mononuclear cell (PBMC)-based MT-2 assay for the detection of CXCR4 use in defining the tropism of HIV isolates derived from clinical samples. The various samples used for this comparison were derived from participants of the Amsterdam Cohort Studies on HIV infection and AIDS who underwent consecutive MT-2 assay testing of their PBMCs at approximately 3-month intervals. This unique sample set was specifically selected because consecutive MT-2 assays had demonstrated a shift from negative to positive in PBMCs, reflecting the first emergence of CXCR4-using virus in PBMCs above the level of detection of the assay in these individuals. Trofile testing was performed with clonal HIV type 1 (HIV-1) variants (n = 21), MT-2 cell culture-derived cells (n = 20) and supernatants (n = 42), and plasma samples (n = 76). Among the clonal HIV-1 variants and MT-2 cell culture-derived samples, the results of the Trofile and MT-2 assays demonstrated a high degree of concordance (95% to 98%). Among consecutive plasma samples, detection of CXCR4-using virus was at or before the time of first detection by the MT-2 assay in 5/10 patients by the original Trofile assay and in 9/10 patients by the enhanced-sensitivity Trofile assay. Differences in the time to the first detection of CXCR4 use between the MT-2 assay (PBMCs) and the original Trofile assay (plasma) were greatly reduced by the enhanced-sensitivity Trofile assay, suggesting that sensitivity for the detection of minor CXCR4-using variants may be a more important determinant of discordant findings than compartmentalization. The similarities in performance of the enhanced-sensitivity Trofile and MT-2 assays suggest that either may be an appropriate methodology to define tropism in patient specimens.

Long-term effect of depot medroxyprogesterone acetate on vaginal microbiota, epithelial thickness and HIV target cells.

PubMed

Mitchell, Caroline M; McLemore, Leslie; Westerberg, Katharine; Astronomo, Rena; Smythe, Kimberly; Gardella, Carolyn; Mack, Matthias; Magaret, Amalia; Patton, Dorothy; Agnew, Kathy; McElrath, M Juliana; Hladik, Florian; Eschenbach, David

2014-08-15

Depot medroxyprogesterone acetate (DMPA) has been linked to human immunodeficiency virus type 1 (HIV-1) acquisition. Vaginal microbiota of women using DMPA for up to 2 years were cultured. Mucosal immune cell populations were measured by immunohistological staining. Over 12 months, the proportion with H2O2-positive lactobacilli decreased (n = 32; 53% vs 27%; P = .03). Median vaginal CD3(+) cells also decreased (n = 15; 355 vs 237 cells/mm(2); P = .03), as did CD3(+)CCR5(+) cells (195 vs 128 cells/mm(2); P = .04), HLA-DR(+) cells (130 vs 96 cells/mm(2); P = .27), and HLA-DR(+)CCR5(+) cells (18 vs 10 cells/mm(2); P = .33). DMPA contraception does not increase vaginal mucosal CCR5(+) HIV target cells but does decrease CD3(+) T lymphocytes and vaginal H2O2-producing lactobacilli. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

PSC-RANTES Blocks R5 Human Immunodeficiency Virus Infection of Langerhans Cells Isolated from Individuals with a Variety of CCR5 Diplotypes

PubMed Central

Kawamura, Tatsuyoshi; Bruce, Shannon E.; Abraha, Awet; Sugaya, Makoto; Hartley, Oliver; Offord, Robin E.; Arts, Eric J.; Zimmerman, Peter A.; Blauvelt, Andrew

2004-01-01

Topical microbicides that effectively block interactions between CCR5+ immature Langerhans cells (LC) residing within genital epithelia and R5 human immunodeficiency virus (HIV) may decrease sexual transmission of HIV. Here, we investigated the ability of synthetic RANTES analogues (AOP-, NNY-, and PSC-RANTES) to block R5 HIV infection of human immature LC by using a skin explant model. In initial experiments using activated peripheral blood mononuclear cells, each analogue compound demonstrated marked antiviral activity against two R5 HIV isolates. Next, we found that 20-min preincubation of skin explants with each RANTES analogue blocked R5 HIV infection of LC in a dose-dependent manner (1 to 100 nM) and that PSC-RANTES was the most potent of these compounds. Similarly, preincubation of LC with each analogue was able to block LC-mediated infection of cocultured CD4+ T cells. Competition experiments between primary R5 and X4 HIV isolates showed blocking of R5 HIV by PSC-RANTES and no evidence of increased propagation of X4 HIV, data that are consistent with the specificity of PSC-RANTES for CCR5 and the CCR5+ CXCR4− phenotype of immature LC. Finally, when CCR5 genetic polymorphism data were integrated with results from the in vitro LC infection studies, PSC-RANTES was found to be equally effective in inhibiting R5 HIV in LC isolated from individuals with CCR5 diplotypes known to be associated with low, intermediate, and high cell surface levels of CCR5. In summary, PSC-RANTES is a potent inhibitor of R5 HIV infection in immature LC, suggesting that it may be useful as a topical microbicide to block sexual transmission of HIV. PMID:15220435

HbAHP-25, an In-Silico Designed Peptide, Inhibits HIV-1 Entry by Blocking gp120 Binding to CD4 Receptor.

PubMed

Bashir, Tahir; Patgaonkar, Mandar; Kumar, Selvaa C; Pasi, Achhelal; Reddy, Kudumula Venkata Rami

2015-01-01

Human Immunodeficiency Virus (HIV-1) poses a serious threat to the developing world and sexual transmission continues to be the major source of new infections. Therefore, the development of molecules, which prevent new HIV-1 infections, is highly warranted. In the present study, a panel of human hemoglobin (Hb)-α subunit derived peptides and their analogues, with an ability to bind gp120, were designed in-silico and their anti-HIV-1 activity was evaluated. Of these peptides, HbAHP-25, an analogue of Hb-α derived peptide, demonstrated significant anti-HIV-1 activity. HbAHP-25 was found to be active against CCR5-tropic HIV-1 strains (ADA5 and BaL) and CXCR4-tropic HIV-1 strains (IIIB and NL4-3). Surface plasmon resonance (SPR) and ELISA revealed direct interaction between HbAHP-25 and HIV-1 envelope protein, gp120. The peptide prevented binding of CD4 to gp120 and blocked subsequent steps leading to entry and/or fusion or both. Anti-HIV activity of HbAHP-25 appeared to be specific as it failed to inhibit the entry of HIV-1 pseudotyped virus (HIV-1 VSV). Further, HbAHP-25 was found to be non-cytotoxic to TZM-bl cells, VK2/E6E7 cells, CEM-GFP cells and PBMCs, even at higher concentrations. Moreover, HbAHP-25 retained its anti-HIV activity in presence of seminal plasma and vaginal fluid. In brief, the study identified HbAHP-25, a novel anti-HIV peptide, which directly interacts with gp120 and thus has a potential to inhibit early stages of HIV-1 infection.

HbAHP-25, an In-Silico Designed Peptide, Inhibits HIV-1 Entry by Blocking gp120 Binding to CD4 Receptor

PubMed Central

Bashir, Tahir; Patgaonkar, Mandar; Kumar C, Selvaa; Pasi, Achhelal; Reddy, Kudumula Venkata Rami

2015-01-01

Human Immunodeficiency Virus (HIV-1) poses a serious threat to the developing world and sexual transmission continues to be the major source of new infections. Therefore, the development of molecules, which prevent new HIV-1 infections, is highly warranted. In the present study, a panel of human hemoglobin (Hb)-α subunit derived peptides and their analogues, with an ability to bind gp120, were designed in-silico and their anti-HIV-1 activity was evaluated. Of these peptides, HbAHP-25, an analogue of Hb-α derived peptide, demonstrated significant anti-HIV-1 activity. HbAHP-25 was found to be active against CCR5-tropic HIV-1 strains (ADA5 and BaL) and CXCR4-tropic HIV-1 strains (IIIB and NL4-3). Surface plasmon resonance (SPR) and ELISA revealed direct interaction between HbAHP-25 and HIV-1 envelope protein, gp120. The peptide prevented binding of CD4 to gp120 and blocked subsequent steps leading to entry and/or fusion or both. Anti-HIV activity of HbAHP-25 appeared to be specific as it failed to inhibit the entry of HIV-1 pseudotyped virus (HIV-1 VSV). Further, HbAHP-25 was found to be non-cytotoxic to TZM-bl cells, VK2/E6E7 cells, CEM-GFP cells and PBMCs, even at higher concentrations. Moreover, HbAHP-25 retained its anti-HIV activity in presence of seminal plasma and vaginal fluid. In brief, the study identified HbAHP-25, a novel anti-HIV peptide, which directly interacts with gp120 and thus has a potential to inhibit early stages of HIV-1 infection. PMID:25915507

P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions

PubMed Central

Giroud, Charline; Marin, Mariana; Hammonds, Jason; Spearman, Paul

2015-01-01

ABSTRACT HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env. IMPORTANCE Inhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF279, is due to the blocking of virus interactions with both the CXCR4 and CCR5 coreceptors. The ability of NF279 to abrogate cellular calcium signaling induced by the respective chemokines showed that this compound acts as a dual-coreceptor antagonist. P2X1 receptor antagonists could thus represent a new class of dual-coreceptor inhibitors with a structure and a mechanism of action that are distinct from those of known HIV-1 coreceptor antagonists. PMID:26136569

P2X1 Receptor Antagonists Inhibit HIV-1 Fusion by Blocking Virus-Coreceptor Interactions.

PubMed

Giroud, Charline; Marin, Mariana; Hammonds, Jason; Spearman, Paul; Melikyan, Gregory B

2015-09-01

HIV-1 Env glycoprotein-mediated fusion is initiated upon sequential binding of Env to CD4 and the coreceptor CXCR4 or CCR5. Whereas these interactions are thought to be necessary and sufficient to promote HIV-1 fusion, other host factors can modulate this process. Previous studies reported potent inhibition of HIV-1 fusion by selective P2X1 receptor antagonists, including NF279, and suggested that these receptors play a role in HIV-1 entry. Here we investigated the mechanism of antiviral activity of NF279 and found that this compound does not inhibit HIV-1 fusion by preventing the activation of P2X1 channels but effectively blocks the binding of the virus to CXCR4 or CCR5. The notion of an off-target effect of NF279 on HIV-1 fusion is supported by the lack of detectable expression of P2X1 receptors in cells used in fusion experiments and by the fact that the addition of ATP or the enzymatic depletion of ATP in culture medium does not modulate viral fusion. Importantly, NF279 fails to inhibit HIV-1 fusion with cell lines and primary macrophages when added at an intermediate stage downstream of Env-CD4-coreceptor engagement. Conversely, in the presence of NF279, HIV-1 fusion is arrested downstream of CD4 binding but prior to coreceptor engagement. NF279 also antagonizes the signaling function of CCR5, CXCR4, and another chemokine receptor, as evidenced by the suppression of calcium responses elicited by specific ligands and by recombinant gp120. Collectively, our results demonstrate that NF279 is a dual HIV-1 coreceptor inhibitor that interferes with the functional engagement of CCR5 and CXCR4 by Env. Inhibition of P2X receptor activity suppresses HIV-1 fusion and replication, suggesting that P2X signaling is involved in HIV-1 entry. However, mechanistic experiments conducted in this study imply that P2X1 receptor is not expressed in target cells or involved in viral fusion. Instead, we found that inhibition of HIV-1 fusion by a specific P2X1 receptor antagonist, NF279, is due to the blocking of virus interactions with both the CXCR4 and CCR5 coreceptors. The ability of NF279 to abrogate cellular calcium signaling induced by the respective chemokines showed that this compound acts as a dual-coreceptor antagonist. P2X1 receptor antagonists could thus represent a new class of dual-coreceptor inhibitors with a structure and a mechanism of action that are distinct from those of known HIV-1 coreceptor antagonists. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Plasmacytoid Dendritic Cell Infection and Sensing Capacity during Pathogenic and Nonpathogenic Simian Immunodeficiency Virus Infection

PubMed Central

Jochems, Simon P.; Jacquelin, Beatrice; Chauveau, Lise; Huot, Nicolas; Petitjean, Gaël; Lepelley, Alice; Liovat, Anne-Sophie; Ploquin, Mickaël J.; Cartwright, Emily K.; Bosinger, Steven E.; Silvestri, Guido; Barré-Sinoussi, Françoise; Lebon, Pierre; Schwartz, Olivier

2015-01-01

ABSTRACT Human immunodeficiency virus (HIV) in humans and simian immunodeficiency virus (SIV) in macaques (MAC) lead to chronic inflammation and AIDS. Natural hosts, such as African green monkeys (AGM) and sooty mangabeys (SM), are protected against SIV-induced chronic inflammation and AIDS. Here, we report that AGM plasmacytoid dendritic cells (pDC) express extremely low levels of CD4, unlike MAC and human pDC. Despite this, AGM pDC efficiently sensed SIVagm, but not heterologous HIV/SIV isolates, indicating a virus-host adaptation. Moreover, both AGM and SM pDC were found to be, in contrast to MAC pDC, predominantly negative for CCR5. Despite such limited CD4 and CCR5 expression, lymphoid tissue pDC were infected to a degree similar to that seen with CD4+ T cells in both MAC and AGM. Altogether, our finding of efficient pDC infection by SIV in vivo identifies pDC as a potential viral reservoir in lymphoid tissues. We discovered low expression of CD4 on AGM pDC, which did not preclude efficient sensing of host-adapted viruses. Therefore, pDC infection and efficient sensing are not prerequisites for chronic inflammation. The high level of pDC infection by SIVagm suggests that if CCR5 paucity on immune cells is important for nonpathogenesis of natural hosts, it is possibly not due to its role as a coreceptor. IMPORTANCE The ability of certain key immune cell subsets to resist infection might contribute to the asymptomatic nature of simian immunodeficiency virus (SIV) infection in its natural hosts, such as African green monkeys (AGM) and sooty mangabeys (SM). This relative resistance to infection has been correlated with reduced expression of CD4 and/or CCR5. We show that plasmacytoid dendritic cells (pDC) of natural hosts display reduced CD4 and/or CCR5 expression, unlike macaque pDC. Surprisingly, this did not protect AGM pDC, as infection levels were similar to those found in MAC pDC. Furthermore, we show that AGM pDC did not consistently produce type I interferon (IFN-I) upon heterologous SIVmac/HIV type 1 (HIV-1) encounter, while they sensed autologous SIVagm isolates. Pseudotyping SIVmac/HIV-1 overcame this deficiency, suggesting that reduced uptake of heterologous viral strains underlays this lack of sensing. The distinct IFN-I responses depending on host species and HIV/SIV isolates reveal the host/virus species specificity of pDC sensing. PMID:25903334

77 FR 69715 - Federal Acquisition Regulation; Updates to Contract Reporting and Central Contractor Registration

Federal Register 2010, 2011, 2012, 2013, 2014

2012-11-20

... Contractor Registration AGENCY: Department of Defense (DoD), General Services Administration (GSA), and... changes the clauses requiring contractor registration in the Central Contractor Registration (CCR... unique identifier for contractors. It is used (1) to uniquely identify a contractor entity, and (2) to...

Protection of macaques from vaginal SHIV challenge by vaginally delivered inhibitors of virus-cell fusion.

PubMed

Veazey, Ronald S; Klasse, Per Johan; Schader, Susan M; Hu, Qinxue; Ketas, Thomas J; Lu, Min; Marx, Preston A; Dufour, Jason; Colonno, Richard J; Shattock, Robin J; Springer, Martin S; Moore, John P

2005-11-03

Human immunodeficiency virus type 1 (HIV-1) continues to spread, principally by heterosexual sex, but no vaccine is available. Hence, alternative prevention methods are needed to supplement educational and behavioural-modification programmes. One such approach is a vaginal microbicide: the application of inhibitory compounds before intercourse. Here, we have evaluated the microbicide concept using the rhesus macaque 'high dose' vaginal transmission model with a CCR5-receptor-using simian-human immunodeficiency virus (SHIV-162P3) and three compounds that inhibit different stages of the virus-cell attachment and entry process. These compounds are BMS-378806, a small molecule that binds the viral gp120 glycoprotein and prevents its attachment to the CD4 and CCR5 receptors, CMPD167, a small molecule that binds to CCR5 to inhibit gp120 association, and C52L, a bacterially expressed peptide inhibitor of gp41-mediated fusion. In vitro, all three compounds inhibit infection of T cells and cervical tissue explants, and C52L acts synergistically with CMPD167 or BMS-378806 to inhibit infection of cell lines. In vivo, significant protection was achieved using each compound alone and in combinations. CMPD167 and BMS-378806 were protective even when applied 6 h before challenge.

CD4+ Memory Stem Cells Are Infected by HIV-1 in a Manner Regulated in Part by SAMHD1 Expression

PubMed Central

Tabler, Caroline O.; Lucera, Mark B.; Haqqani, Aiman A.; McDonald, David J.; Migueles, Stephen A.; Connors, Mark

2014-01-01

ABSTRACT CD4+ and CD8+ memory T cells with stem cell-like properties (TSCM cells) have been identified in mice, humans, and nonhuman primates and are being investigated for antitumor and antiviral vaccines and immunotherapies. Whether CD4+ TSCM cells are infected by human immunodeficiency virus (HIV) was investigated by using a combination HIV reporter virus system in vitro and by direct staining for HIV p24 antigen ex vivo. A proportion of TSCM cells were found to express the HIV coreceptors CCR5 and CXCR4 and were infected by HIV both in vitro and in vivo. Analysis of viral outcome following fusion using the combination reporter virus system revealed that TSCM cells can become productively or latently infected, although the vast majority of TSCM cells are abortively infected. Knockdown of the HIV restriction factor SAMHD1 using Vpx-containing simian immunodeficiency virus (SIV) virion-like particles enhanced the productive infection of TSCM cells, indicating that SAMHD1 contributes to abortive infection in these cells. These results demonstrate that CD4+ TSCM cells are targets for HIV infection, that they become productively or latently infected at low levels, and that SAMHD1 expression promotes abortive infection of this important memory cell subset. IMPORTANCE Here we demonstrate the susceptibility of CD4+ memory stem cells (TSCM cells) to infection by HIV in vitro and in vivo, provide an in-depth analysis of coreceptor expression, demonstrate the infection of naïve and memory CD4+ T cell subsets with both CCR5- and CXCR4-tropic HIV, and also perform outcome analysis to calculate the percentage of cells that are productively, latently, or abortively infected. Through these outcome studies, we determined that the vast majority of TSCM cells are abortively infected by HIV, and we demonstrate that knockdown of SAMHD1 significantly increases the frequency of infection of this CD4+ T cell subset, indicating that SAMHD1 is an active restriction factor in TSCM cells. PMID:24554663

Decreased HIV diversity after allogeneic stem cell transplantation of an HIV-1 infected patient: a case report

PubMed Central

2010-01-01

The human immunodeficiency virus type 1 (HIV-1) coreceptor use and viral evolution were analyzed in blood samples from an HIV-1 infected patient undergoing allogeneic stem cell transplantation (SCT). Coreceptor use was predicted in silico from sequence data obtained from the third variable loop region of the viral envelope gene with two software tools. Viral diversity and evolution was evaluated on the same samples by Bayesian inference and maximum likelihood methods. In addition, phenotypic analysis was done by comparison of viral growth in peripheral blood mononuclear cells and in a CCR5 (R5)-deficient T-cell line which was controlled by a reporter assay confirming viral tropism. In silico coreceptor predictions did not match experimental determinations that showed a consistent R5 tropism. Anti-HIV directed antibodies could be detected before and after the SCT. These preexisting antibodies did not prevent viral rebound after the interruption of antiretroviral therapy during the SCT. Eventually, transplantation and readministration of anti-retroviral drugs lead to sustained increase in CD4 counts and decreased viral load to undetectable levels. Unexpectedly, viral diversity decreased after successful SCT. Our data evidence that only R5-tropic virus was found in the patient before and after transplantation. Therefore, blocking CCR5 receptor during stem cell transplantation might have had beneficial effects and this might apply to more patients undergoing allogeneic stem cell transplantation. Furthermore, we revealed a scenario of HIV-1 dynamic different from the commonly described ones. Analysis of viral evolution shows the decrease of viral diversity even during episodes with bursts in viral load. PMID:20210988

Structural Studies of Cinnamoyl-CoA Reductase and Cinnamyl-Alcohol Dehydrogenase, Key Enzymes of Monolignol Biosynthesis[C][W

PubMed Central

Pan, Haiyun; Zhou, Rui; Louie, Gordon V.; Mühlemann, Joëlle K.; Bomati, Erin K.; Bowman, Marianne E.; Dudareva, Natalia; Dixon, Richard A.; Noel, Joseph P.; Wang, Xiaoqiang

2014-01-01

The enzymes cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) catalyze the two key reduction reactions in the conversion of cinnamic acid derivatives into monolignol building blocks for lignin polymers in plant cell walls. Here, we describe detailed functional and structural analyses of CCRs from Medicago truncatula and Petunia hybrida and of an atypical CAD (CAD2) from M. truncatula. These enzymes are closely related members of the short-chain dehydrogenase/reductase (SDR) superfamily. Our structural studies support a reaction mechanism involving a canonical SDR catalytic triad in both CCR and CAD2 and an important role for an auxiliary cysteine unique to CCR. Site-directed mutants of CAD2 (Phe226Ala and Tyr136Phe) that enlarge the phenolic binding site result in a 4- to 10-fold increase in activity with sinapaldehyde, which in comparison to the smaller coumaraldehyde and coniferaldehyde substrates is disfavored by wild-type CAD2. This finding demonstrates the potential exploitation of rationally engineered forms of CCR and CAD2 for the targeted modification of monolignol composition in transgenic plants. Thermal denaturation measurements and structural comparisons of various liganded and unliganded forms of CCR and CAD2 highlight substantial conformational flexibility of these SDR enzymes, which plays an important role in the establishment of catalytically productive complexes of the enzymes with their NADPH and phenolic substrates. PMID:25217505

CCR5-edited gene therapies for HIV cure: Closing the door to viral entry.

PubMed

Haworth, Kevin G; Peterson, Christopher W; Kiem, Hans-Peter

2017-11-01

Human immunodeficiency virus (HIV) was first reported and characterized more than three decades ago. Once thought of as a death sentence, HIV infection has become a chronically manageable disease. However, it is estimated that a staggering 0.8% of the world's population is infected with HIV, with more than 1 million deaths reported in 2015 alone. Despite the development of effective anti-retroviral drugs, a permanent cure has only been documented in one patient to date. In 2007, an HIV-positive patient received a bone marrow transplant to treat his leukemia from an individual who was homozygous for a mutation in the CCR5 gene. This mutation, known as CCR5Δ32, prevents HIV replication by inhibiting the early stage of viral entry into cells, resulting in resistance to infection from the majority of HIV isolates. More than 10 years after his last dose of anti-retroviral therapy, the transplant recipient remains free of replication-competent virus. Multiple groups are now attempting to replicate this success through the use of other CCR5-negative donor cell sources. Additionally, developments in the use of lentiviral vectors and targeted nucleases have opened the doors of precision medicine and enabled new treatment methodologies to combat HIV infection through targeted ablation or down-regulation of CCR5 expression. Here, we review historical cases of CCR5-edited cell-based therapies, current clinical trials and future benefits and challenges associated with this technology. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Virucidal activity of the dendrimer microbicide SPL7013 against HIV-1.

PubMed

Telwatte, Sushama; Moore, Katie; Johnson, Adam; Tyssen, David; Sterjovski, Jasminka; Aldunate, Muriel; Gorry, Paul R; Ramsland, Paul A; Lewis, Gareth R; Paull, Jeremy R A; Sonza, Secondo; Tachedjian, Gilda

2011-06-01

Topical microbicides for use by women to prevent the transmission of human immunodeficiency virus (HIV) and other sexually transmitted infections are urgently required. Dendrimers are highly branched nanoparticles being developed as microbicides. SPL7013 is a dendrimer with broad-spectrum activity against HIV type I (HIV-1) and -2 (HIV-2), herpes simplex viruses type-1 (HSV-1) and -2 (HSV-2) and human papillomavirus. SPL7013 [3% (w/w)] has been formulated in a mucoadhesive carbopol gel (VivaGel®) for use as a topical microbicide. Previous studies showed that SPL7013 has similar potency against CXCR4-(X4) and CCR5-using (R5) strains of HIV-1 and that it blocks viral entry. However, the ability of SPL7013 to directly inactivate HIV-1 is unknown. We examined whether SPL7013 demonstrates virucidal activity against X4 (NL4.3, MBC200, CMU02 clade EA and 92UG046 clade D), R5 (Ba-L, NB25 and 92RW016 clade A) and dual-tropic (R5X4; MACS1-spln) HIV-1 using a modified HLA-DR viral capture method and by polyethylene glycol precipitation. Evaluation of virion integrity was determined by ultracentrifugation through a sucrose cushion and detection of viral proteins by Western blot analysis. SPL7013 demonstrated potent virucidal activity against X4 and R5X4 strains, although virucidal activity was less potent for the 92UG046 X4 clade D isolate. Where potent virucidal activity was observed, the 50% virucidal concentrations were similar to the 50% effective concentrations previously reported in drug susceptibility assays, indicating that the main mode of action of SPL7013 is by direct viral inactivation for these strains. In contrast, SPL7013 lacked potent virucidal activity against R5 HIV-1 strains. Evaluation of the virucidal mechanism showed that SPL7013-treated NL4.3, 92UG046 and MACS1-spln virions were intact with no significant decrease in gp120 surface protein with respect to p24 capsid content compared to the corresponding untreated virus. These studies demonstrate that SPL7013 is virucidal against HIV-1 strains that utilize the CXCR4 coreceptor but not viruses tested in this study that solely use CCR5 by a mechanism that is distinct from virion disruption or loss of gp120. In addition, the mode of action by which SPL7013 prevents infection of cells with X4 and R5X4 strains is likely to differ from R5 strains of HIV-1. Copyright © 2011 Elsevier B.V. All rights reserved.

75 FR 51434 - Hazardous and Solid Waste Management System; Identification and Listing of Special Wastes...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-20

... characters, any form of encryption, and be free of any defects or viruses. For additional information about... hearings can be found on EPA's Web site at: http://www.epa.gov/epawaste/nonhaz/industrial/special/fossil... information on the Web site: http://www.epa.gov/osw/nonhaz/industrial/special/fossil/ccr-rule/ccr-webinar.htm...

Moderate restriction of macrophage-tropic human immunodeficiency virus type 1 by SAMHD1 in monocyte-derived macrophages.

PubMed

Taya, Kahoru; Nakayama, Emi E; Shioda, Tatsuo

2014-01-01

Macrophage-tropic human immunodeficiency virus type 1 (HIV-1) strains are able to grow to high titers in human monocyte-derived macrophages. However, it was recently reported that cellular protein SAMHD1 restricts HIV-1 replication in human cells of the myeloid lineage, including monocyte-derived macrophages. Here we show that degradation of SAMHD1 in monocyte-derived macrophages was associated with moderately enhanced growth of the macrophage-tropic HIV-1 strain. SAMHD1 degradation was induced by treating target macrophages with vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 2 (HIV-2) particles containing viral protein X. For undifferentiated monocytes, HIV-2 particle treatment allowed undifferentiated monocytes to be fully permissive for productive infection by the macrophage-tropic HIV-1 strain. In contrast, untreated monocytes were totally resistant to HIV-1 replication. These results indicated that SAMHD1 moderately restricts even a macrophage-tropic HIV-1 strain in monocyte-derived macrophages, whereas the protein potently restricts HIV-1 replication in undifferentiated monocytes.

Nonproductive human immunodeficiency virus type 1 infection of human fetal astrocytes: independence from CD4 and major chemokine receptors.

PubMed

Sabri, F; Tresoldi, E; Di Stefano, M; Polo, S; Monaco, M C; Verani, A; Fiore, J R; Lusso, P; Major, E; Chiodi, F; Scarlatti, G

1999-11-25

Human immunodeficiency virus type 1 (HIV-1) infection of the brain is associated with neurological manifestations both in adults and in children. The primary target for HIV-1 infection in the brain is the microglia, but astrocytes can also be infected. We tested 26 primary HIV-1 isolates for their capacity to infect human fetal astrocytes in culture. Eight of these isolates, independent of their biological phenotype and chemokine receptor usage, were able to infect astrocytes. Although no sustained viral replication could be demonstrated, the virus was recovered by coculture with receptive cells such as macrophages or on stimulation with interleukin-1beta. To gain knowledge into the molecular events that regulate attachment and penetration of HIV-1 in astrocytes, we investigated the expression of several chemokine receptors. Fluorocytometry and calcium-mobilization assay did not provide evidence of expression of any of the major HIV-1 coreceptors, including CXCR4, CCR5, CCR3, and CCR2b, as well as the CD4 molecule on the cell surface of human fetal astrocytes. However, mRNA transcripts for CXCR4, CCR5, Bonzo/STRL33/TYMSTR, and APJ were detected by RT-PCR. Furthermore, infection of astrocytes by HIV-1 isolates with different chemokine receptor usage was not inhibited by the chemokines SDF-1beta, RANTES, MIP-1beta, or MCP-1 or by antibodies directed against the third variable region or the CD4 binding site of gp120. These data show that astrocytes can be infected by primary HIV-1 isolates via a mechanism independent of CD4 or major chemokine receptors. Furthermore, astrocytes are potential carriers of latent HIV-1 and on activation may be implicated in spreading the infection to other neighbouring cells, such as microglia or macrophages. Copyright 1999 Academic Press.

Thymic pathogenicity of an HIV-1 envelope is associated with increased CXCR4 binding efficiency and V5-gp41-dependent activity, but not V1/V2-associated CD4 binding efficiency and viral entry

DOE Office of Scientific and Technical Information (OSTI.GOV)

Meissner, Eric G.; Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599; Coffield, Vernon M.

2005-06-05

We previously described a thymus-tropic HIV-1 envelope (R3A Env) from a rapid progressor obtained at the time of transmission. An HIV-1 molecular recombinant with the R3A Env supported extensive replication and pathogenesis in the thymus and did not require Nef. Another Env from the same patient did not display the same thymus-tropic pathogenesis (R3B Env). Here, we show that relative to R3B Env, R3A Env enhances viral entry of T cells, increases fusion-induced cytopathicity, and shows elevated binding efficiency for both CD4 and CXCR4, but not CCR5, in vitro. We created chimeric envelopes to determine the region(s) responsible for eachmore » in vitro phenotype and for thymic pathogenesis. Surprisingly, while V1/V2 contributed to enhanced viral entry, CD4 binding efficiency, and cytopathicity in vitro, it made no contribution to thymic pathogenesis. Rather, CXCR4 binding efficiency and V5-gp41-associated activity appear to independently contribute to thymic pathogenesis of the R3A Env. These data highlight the contribution of unique HIV pathogenic factors in the thymic microenvironment and suggest that novel mechanisms may be involved in Env pathogenic activity in vivo.« less

Lower prevalence of human immunodeficiency virus type 1 Brazilian subtype B found in northeastern Brazil with slower progression to AIDS.

PubMed

Araujo, Adriano Fernando; Brites, Carlos; Monteiro-Cunha, Joana; Santos, Luciane Amorim; Galvao-Castro, Bernardo; Alcantara, Luiz Carlos Junior

2010-11-01

Besides being extremely useful in measuring the level of HIV-1 diversity and prevalence in populations, the molecular analysis of genomic sequences provides crucial surveillance support and aids in the development of new therapies and effective vaccines. The present study focused on gag and env DNA and amino acid sequences that were generated from samples taken from 61 infected patients in the City of Salvador, Bahia, located in northeastern Brazil. In order to determine selective pressure and predict coreceptor usage, Bioinformatics tools were employed in phylogeny reconstruction. Fifty-six (91.8%) viruses were classified as belonging to subtype B, three (4.9%) from F1, and two (3.3%) from BF1 recombinants. Based on the characterization of the V3 region, the subtype B strains were represented by eight (18.2%) Brazilian variants (B'-GWGR), 20 (46.5%) European/EUA B variants (GPGR), and 15 (34.9%) GXGX variants. The mean time elapsed since diagnosis was 13 years among subtype B' and 9 years in subtype B. The mean dN/dS ratios from the GWGR, GPGR, and GXGX groups, when compared to an HXB2 reference, were 0.72, 0.77, and 0.67, respectively. Seventy-six percent of the viruses studied were predicted to use the CCR5 coreceptor for cell entry (R5 viruses), while 24% were predicted to use the CXCR4 or were classified as dual tropic viruses. The prevalence of subtypes B' and recombinant B/F1 was shown to be lower than findings from previous studies performed both in Brazil (B') and in Bahia (B/F1). The association between subtype B' and a lengthy period of time since diagnosis can be correlated with a slower disease progression in infected patients, when compared with those infected with subtype B.

Neutralization Properties of Simian Immunodeficiency Viruses Infecting Chimpanzees and Gorillas

PubMed Central

Barbian, Hannah J.; Decker, Julie M.; Bibollet-Ruche, Frederic; Galimidi, Rachel P.; West, Anthony P.; Learn, Gerald H.; Parrish, Nicholas F.; Iyer, Shilpa S.; Li, Yingying; Pace, Craig S.; Song, Ruijiang; Huang, Yaoxing; Denny, Thomas N.; Mouquet, Hugo; Martin, Loic; Acharya, Priyamvada; Zhang, Baoshan; Kwong, Peter D.; Mascola, John R.; Verrips, C. Theo; Strokappe, Nika M.; Rutten, Lucy; McCoy, Laura E.; Weiss, Robin A.; Brown, Corrine S.; Jackson, Raven; Silvestri, Guido; Connors, Mark; Burton, Dennis R.; Shaw, George M.; Nussenzweig, Michel C.; Bjorkman, Pamela J.; Ho, David D.; Farzan, Michael

2015-01-01

ABSTRACT Broadly cross-reactive neutralizing antibodies (bNabs) represent powerful tools to combat human immunodeficiency virus type 1 (HIV-1) infection. Here, we examined whether HIV-1-specific bNabs are capable of cross-neutralizing distantly related simian immunodeficiency viruses (SIVs) infecting central (Pan troglodytes troglodytes) (SIVcpzPtt) and eastern (Pan troglodytes schweinfurthii) (SIVcpzPts) chimpanzees (n = 11) as well as western gorillas (Gorilla gorilla gorilla) (SIVgor) (n = 1). We found that bNabs directed against the CD4 binding site (n = 10), peptidoglycans at the base of variable loop 3 (V3) (n = 5), and epitopes at the interface of surface (gp120) and membrane-bound (gp41) envelope glycoproteins (n = 5) failed to neutralize SIVcpz and SIVgor strains. In addition, apex V2-directed bNabs (n = 3) as well as llama-derived (heavy chain only) antibodies (n = 6) recognizing both the CD4 binding site and gp41 epitopes were either completely inactive or neutralized only a fraction of SIVcpzPtt strains. In contrast, one antibody targeting the membrane-proximal external region (MPER) of gp41 (10E8), functional CD4 and CCR5 receptor mimetics (eCD4-Ig, eCD4-Igmim2, CD4-218.3-E51, and CD4-218.3-E51-mim2), as well as mono- and bispecific anti-human CD4 (iMab and LM52) and CCR5 (PRO140, PRO140-10E8) receptor antibodies neutralized >90% of SIVcpz and SIVgor strains with low-nanomolar (0.13 to 8.4 nM) potency. Importantly, the latter antibodies blocked virus entry not only in TZM-bl cells but also in Cf2Th cells expressing chimpanzee CD4 and CCR5 and neutralized SIVcpz in chimpanzee CD4+ T cells, with 50% inhibitory concentrations (IC50s) ranging from 3.6 to 40.5 nM. These findings provide new insight into the protective capacity of anti-HIV-1 bNabs and identify candidates for further development to combat SIVcpz infection. PMID:25900654

Correlations between polymorphisms in the uridine diphosphate-glucuronosyltransferase 1A and C-C motif chemokine receptor 5 genes and infection with the hepatitis B virus in three ethnic groups in China.

PubMed

Zhang, Chan; He, Yan; Shan, Ke-Ren; Tan, Kui; Zhang, Ting; Wang, Chan-Juan; Guan, Zhi-Zhong

2018-02-01

Objective To determine whether genetic polymorphisms in the uridine diphosphate-glucuronosyltransferase 1A ( UGT1A) and the C-C motif chemokine receptor 5 ( CCR5) genes are associated with hepatitis B virus (HBV) infection in Yi, Yao and Han ethnic groups in the Guizhou Province of China. Methods The study enrolled subjects with and without HBV infection. Whole blood was used for DNA genotyping using standard techniques. The study determined the frequencies of several polymorphic alleles ( UGT1A6 [rs2070959], UGT1A1 [rs8175347], CCR5-59029 [rs1799987] and CCR5Δ32 [rs333]) and then characterized their relationship with HBV infection. Results A total of 404 subjects were enrolled in the study: 138 from the Yao group, 101 from the Yi group and 165 from the Han group. There was a significant difference in the frequency of UGT1A1 rs8175347 polymorphisms among the three groups. The rates of 7TA carriers of UGT1A1 rs8175347 in all three groups were significantly higher than the other genotypes. Individuals with genotype AA of UGT1A6 rs2070959 in the Yi group had a higher risk for HBV infection than in the Yao and Han groups. The frequency of genotype GG in CCR5-59029 in the Yao group was significantly higher than in the Yi group. The genotypes of CCR5Δ32 were not associated with HBV infection. Conclusion These findings provide genetic and epidemiological evidence for an association of UGT1A and CCR5-59029 polymorphisms with HBV infection in Chinese Yi and Yao populations.

Structural studies of cinnamoyl-CoA reductase and cinnamyl-alcohol dehydrogenase, key enzymes of monolignol biosynthesis.

PubMed

Pan, Haiyun; Zhou, Rui; Louie, Gordon V; Mühlemann, Joëlle K; Bomati, Erin K; Bowman, Marianne E; Dudareva, Natalia; Dixon, Richard A; Noel, Joseph P; Wang, Xiaoqiang

2014-09-01

The enzymes cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) catalyze the two key reduction reactions in the conversion of cinnamic acid derivatives into monolignol building blocks for lignin polymers in plant cell walls. Here, we describe detailed functional and structural analyses of CCRs from Medicago truncatula and Petunia hybrida and of an atypical CAD (CAD2) from M. truncatula. These enzymes are closely related members of the short-chain dehydrogenase/reductase (SDR) superfamily. Our structural studies support a reaction mechanism involving a canonical SDR catalytic triad in both CCR and CAD2 and an important role for an auxiliary cysteine unique to CCR. Site-directed mutants of CAD2 (Phe226Ala and Tyr136Phe) that enlarge the phenolic binding site result in a 4- to 10-fold increase in activity with sinapaldehyde, which in comparison to the smaller coumaraldehyde and coniferaldehyde substrates is disfavored by wild-type CAD2. This finding demonstrates the potential exploitation of rationally engineered forms of CCR and CAD2 for the targeted modification of monolignol composition in transgenic plants. Thermal denaturation measurements and structural comparisons of various liganded and unliganded forms of CCR and CAD2 highlight substantial conformational flexibility of these SDR enzymes, which plays an important role in the establishment of catalytically productive complexes of the enzymes with their NADPH and phenolic substrates. © 2014 American Society of Plant Biologists. All rights reserved.

Paucity of CD4+CCR5+ T cells is a typical feature of natural SIV hosts

PubMed Central

Pandrea, Ivona; Apetrei, Cristian; Gordon, Shari; Barbercheck, Joseph; Dufour, Jason; Bohm, Rudolf; Sumpter, Beth; Roques, Pierre; Marx, Preston A.; Hirsch, Vanessa M.; Kaur, Amitinder; Lackner, Andrew A.; Veazey, Ronald S.; Silvestri, Guido

2007-01-01

In contrast to lentiviral infections of humans and macaques, simian immunodeficiency virus (SIV) infection of natural hosts is nonpathogenic despite high levels of viral replication. However, the mechanisms underlying this absence of disease are unknown. Here we report that natural hosts for SIV infection express remarkably low levels of CCR5 on CD4+ T cells isolated from blood, lymph nodes, and mucosal tissues. Given that this immunologic feature is found in 5 different species of natural SIV hosts (sooty mangabeys, African green monkeys, mandrills, sun-tailed monkeys, and chimpanzees) but is absent in 5 nonnatural/recent hosts (humans, rhesus, pigtail, cynomolgus macaques, and baboons), it may represent a key feature of the coevolution between the virus and its natural hosts that led to a nonpathogenic infection. Beneficial effects of low CCR5 expression on CD4+ T cells may include the reduction of target cells for viral replication and a decreased homing of activated CD4+ T cells to inflamed tissue. PMID:17003371

Vpx complementation of 'non-macrophage tropic' R5 viruses reveals robust entry of infectious HIV-1 cores into macrophages.

PubMed

Mlcochova, Petra; Watters, Sarah A; Towers, Greg J; Noursadeghi, Mahdad; Gupta, Ravindra K

2014-03-21

It is now known that clinically derived viruses are most commonly R5 tropic with very low infectivity in macrophages. As these viruses utilize CD4 inefficiently, defective entry has been assumed to be the dominant restriction. The implication is that macrophages are not an important reservoir for the majority of circulating viruses. Macrophage infection by clinical transmitted/founder isolates was 10-100 and 30-450 fold less efficient as compared to YU-2 and BaL respectively. Vpx complementation augmented macrophage infection by non-macrophage tropic viruses to the level of infectivity observed for YU-2 in the absence of Vpx. Augmentation was evident even when Vpx was provided 24 hours post-infection. The entry defect was measured as 2.5-5 fold, with a further 3.5-10 fold block at strong stop and subsequent stages of reverse transcription as compared to YU-2. The overall block to infection was critically dependent on the mechanism of entry as demonstrated by rescue of infection after pseudotyping with VSV-G envelope. Reverse transcription in macrophages could not be enhanced using a panel of cytokines or lipopolysaccharide (LPS). Although the predominant block to clinical transmitted/founder viruses is post-entry, infectivity is determined by Env-CD4 interactions and can be rescued with VSV-G pseudotyping. This suggests a functional link between the optimal entry pathway taken by macrophage tropic viruses and downstream events required for reverse transcription. Consistent with a predominantly post-entry block, replication of R5 using viruses can be greatly enhanced by Vpx. We conclude therefore that entry is not the limiting step and that macrophages represent clinically relevant reservoirs for 'non-macrophage tropic' viruses.

Determinants for Sensitivity of Human Immunodeficiency Virus Coreceptor CXCR4 to the Bicyclam AMD3100

PubMed Central

Labrosse, Béatrice; Brelot, Anne; Heveker, Nikolaus; Sol, Nathalie; Schols, Dominique; De Clercq, Erik; Alizon, Marc

1998-01-01

The bicyclam AMD3100 is a potent and selective inhibitor of the replication of human immunodeficiency virus type 1 and type 2 (HIV-1 and HIV-2). It was recently demonstrated that the compound inhibited HIV entry through CXCR4 but not through CCR5. Selectivity of AMD3100 for CXCR4 was further indicated by its lack of effect on HIV-1 and HIV-2 infection mediated by the CCR5, CCR3, Bonzo, BOB, and US28, coreceptors. AMD3100 completely blocked HIV-1 infection mediated by a mutant CXCR4 bearing a deletion of most of the amino-terminal extracellular domain. In contrast, relative resistance to AMD3100 was conferred by different single amino acid substitutions in the second extracellular loop (ECL2) or in the adjacent membrane-spanning domain, TM4. Only substitutions of a neutral residue for aspartic acid and of a nonaromatic residue for phenylalanine (Phe) were associated with drug resistance. This suggests a direct interaction of AMD3100 with these amino acids rather than indirect effects of their mutation on the CXCR4 structure. The interaction of aspartic acids of ECL2 and TM4 with AMD3100 is consistent with the positive charge of bicyclams, which might block HIV-1 entry by preventing electrostatic interactions between CXCR4 and the HIV-1 envelope protein gp120. Other features of AMD3100 must account for its high antiviral activity, in particular the presence of an aromatic linker between the cyclam units. This aromatic group might engage in hydrophobic interactions with the Phe-X-Phe motifs of ECL2 or TM4. These results confirm the importance of ECL2 for the HIV coreceptor activity of CXCR4. PMID:9658078

CCL26/eotaxin-3 is more effective to induce the migration of eosinophils of asthmatics than CCL11/eotaxin-1 and CCL24/eotaxin-2.

PubMed

Provost, Véronique; Larose, Marie-Chantal; Langlois, Anick; Rola-Pleszczynski, Marek; Flamand, Nicolas; Laviolette, Michel

2013-08-01

CCL11, CCL24, and CCL26 are chemokines involved in the recruitment of eosinophils into tissues and mainly activate CCR3. Whereas the genomic or pharmacological inhibition of CCR3 prevents the development of experimental asthma in rodents, it only impairs the recruitment of eosinophils by ∼40% in humans. As humans, but not rodents, express CCL26, we investigated the impact of CCL11, CCL24, and CCL26 on human eosinophils recruitment and evaluated the involvement of CCR3. The migration of eosinophils of healthy volunteers was similar for the three eotaxins. Eosinophils of mild asthmatics had a greater response to CCL11 and a much greater response to CCL26. Whereas all eotaxins induced the migration of eosinophil of asthmatics from 0 to 6 h, CCL26 triggered a second phase of migration between 12 and 18 h. Given that the CCR3 antagonists SB 328437 and SB 297006 inhibited the 5-oxo-eicosatetraenoate-induced migration of eosinophils and that the CCR3 antagonist UCB 35625 was not specific for CCR3, CCR3 blockade was performed with the CCR3 mAb. This antibody completely blocked the effect of all eotaxins on eosinophils of healthy subjects and the effect of CCL24 on the eosinophils of asthmatics. Interestingly, CCR3 blockade did not affect the second migration phase induced by CCL26 on eosinophils of asthmatics. In conclusion, CCL26 is a more effective chemoattractant than CCL11 and CCL24 for eosinophils of asthmatics. The mechanism of this greater efficiency is not yet defined. However, these results suggest that CCL26 may play a unique and important role in the recruitment of eosinophils in persistent asthma.

Almendravirus: A Proposed New Genus of Rhabdoviruses Isolated from Mosquitoes in Tropical Regions of the Americas.

PubMed

Contreras, Maria Angelica; Eastwood, Gillian; Guzman, Hilda; Popov, Vsevolod; Savit, Chelsea; Uribe, Sandra; Kramer, Laura D; Wood, Thomas G; Widen, Steven G; Fish, Durland; Tesh, Robert B; Vasilakis, Nikos; Walker, Peter J

2017-01-11

The Rhabdoviridae is a diverse family of negative-sense single-stranded RNA viruses, many of which infect vertebrate hosts and are transmitted by hematophagous arthropods. Others appear to be arthropod specific, circulating only within arthropod populations. Herein, we report the isolation and characterization of three novel viruses from mosquitoes collected from the Americas. Coot Bay virus was isolated from Anopheles quadrimaculatus mosquitoes collected in the Everglades National Park, Florida; Rio Chico virus was isolated from Anopheles triannulatus mosquitoes collected in Panama; and Balsa virus was isolated from two pools of Culex erraticus mosquitoes collected in Colombia. Sequence analysis indicated that the viruses share a similar genome organization to Arboretum virus and Puerto Almendras virus that had previously been isolated from mosquitoes collected in Peru. Each genome features the five canonical rhabdovirus structural protein genes as well as a gene encoding a class 1A viroporin-like protein (U1) located between the G and L genes (3'-N-P-M-G-U1-L-5'). Phylogenetic analysis of complete L protein sequences indicated that all five viruses cluster in a unique clade that is relatively deeply rooted in the ancestry of animal rhabdoviruses. The failure of all viruses in this clade to grow in newborn mice or vertebrate cells in culture suggests that they may be poorly adapted to replication in vertebrates. © The American Society of Tropical Medicine and Hygiene.

Unique Phenotypic Characteristics of Recently Transmitted HIV-1 Subtype C Envelope Glycoprotein gp120: Use of CXCR6 Coreceptor by Transmitted Founder Viruses.

PubMed

Ashokkumar, Manickam; Aralaguppe, Shambhu G; Tripathy, Srikanth P; Hanna, Luke Elizabeth; Neogi, Ujjwal

2018-05-01

Adequate information on the precise molecular and biological composition of the viral strains that establish HIV infection in the human host will provide effective means of immunization against HIV infection. In an attempt to identify the transmitted founder (TF) virus and differentiate the biological properties and infectious potential of the TF virus from those of the population of the early transmitted viruses, 250 patient-derived gp120 envelope glycoproteins were cloned in pMN-K7-Luc-IRESs-NefΔgp120 to obtain chimeric viruses. Samples were obtained from eight infants who had recently become infected with HIV through mother-to-child transmission (MTCT) and two adults who acquired infection through the heterosexual route and were in the chronic stage of infection. Among the 250 clones tested, 65 chimeric viruses were infectious, and all belonged to HIV-1 subtype C. The 65 clones were analyzed for molecular features of the envelope, per-infectious-particle infectivity, coreceptor tropism, drug sensitivity, and sensitivity to broadly neutralizing antibodies. Based on genotypic and phenotypic analysis of the viral clones, we identified 10 TF viruses from the eight infants. The TF viruses were characterized by shorter V1V2 regions, a reduced number of potential N-linked glycosylation sites, and a higher infectivity titer compared to the virus variants from the adults in the chronic stage of infection. CXCR6 coreceptor usage, in addition to that of the CCR5 coreceptor, which was used by all 65 chimeric viruses, was identified in 13 viruses. The sensitivity of the TF variants to maraviroc and a standard panel of neutralizing monoclonal antibodies (VRC01, PG09, PG16, and PGT121) was found to be much lower than that of the virus variants from the adults in the chronic stage of infection. IMPORTANCE Tremendous progress has been made during the last three and half decades of HIV research, but some significant gaps continue to exist. One of the frontier areas of HIV research which has not seen a breakthrough yet is vaccine research, which is because of the enormous genetic diversity of HIV-1 and the unique infectious fitness of the virus. Among the repertoire of viral variants, the virus that establishes successful infection (transmitted founder [TF] virus) has not been well characterized yet. An insight into the salient features of the TF virus would go a long way toward helping with the design of an effective vaccine against HIV. Here we studied the biological properties of recently transmitted viruses isolated from infants who acquired infection from the mother and have come up with unique characterizations for the TF virus that establishes infection in the human host. Copyright © 2018 American Society for Microbiology.

Oral keratinocytes support non-replicative infection and transfer of harbored HIV-1 to permissive cells.

PubMed

Vacharaksa, Anjalee; Asrani, Anil C; Gebhard, Kristin H; Fasching, Claudine E; Giacaman, Rodrigo A; Janoff, Edward N; Ross, Karen F; Herzberg, Mark C

2008-07-17

Oral keratinocytes on the mucosal surface are frequently exposed to HIV-1 through contact with infected sexual partners or nursing mothers. To determine the plausibility that oral keratinocytes are primary targets of HIV-1, we tested the hypothesis that HIV-1 infects oral keratinocytes in a restricted manner. To study the fate of HIV-1, immortalized oral keratinocytes (OKF6/TERT-2; TERT-2 cells) were characterized for the fate of HIV-specific RNA and DNA. At 6 h post inoculation with X4 or R5-tropic HIV-1, HIV-1gag RNA was detected maximally within TERT-2 cells. Reverse transcriptase activity in TERT-2 cells was confirmed by VSV-G-mediated infection with HIV-NL4-3Deltaenv-EGFP. AZT inhibited EGFP expression in a dose-dependent manner, suggesting that viral replication can be supported if receptors are bypassed. Within 3 h post inoculation, integrated HIV-1 DNA was detected in TERT-2 cell nuclei and persisted after subculture. Multiply spliced and unspliced HIV-1 mRNAs were not detectable up to 72 h post inoculation, suggesting that HIV replication may abort and that infection is non-productive. Within 48 h post inoculation, however, virus harbored by CD4 negative TERT-2 cells trans infected co-cultured peripheral blood mononuclear cells (PBMCs) or MOLT4 cells (CD4+ CCR5+) by direct cell-to-cell transfer or by releasing low levels of infectious virions. Primary tonsil epithelial cells also trans infected HIV-1 to permissive cells in a donor-specific manner. Oral keratinocytes appear, therefore, to support stable non-replicative integration, while harboring and transmitting infectious X4- or R5-tropic HIV-1 to permissive cells for up to 48 h.

Binding site exploration of CCR5 using in silico methodologies: a 3D-QSAR approach.

PubMed

Gadhe, Changdev G; Kothandan, Gugan; Cho, Seung Joo

2013-01-01

Chemokine receptor 5 (CCR5) is an important receptor used by human immunodeficiency virus type 1 (HIV-1) to gain viral entry into host cell. In this study, we used a combined approach of comparative modeling, molecular docking, and three dimensional quantitative structure activity relationship (3D-QSAR) analyses to elucidate detailed interaction of CCR5 with their inhibitors. Docking study of the most potent inhibitor from a series of compounds was done to derive the bioactive conformation. Parameters such as random selection, rational selection, different charges and grid spacing were utilized in the model development to check their performance on the model predictivity. Final comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) models were chosen based on the rational selection method, Gasteiger-Hückel charges and a grid spacing of 0.5 Å. Rational model for CoMFA (q(2) = 0.722, r(2) = 0.884, Q(2) = 0.669) and CoMSIA (q(2) = 0.712, r(2) = 0.825, Q(2) = 0.522) was obtained with good statistics. Mapping of contour maps onto CCR5 interface led us to better understand of the ligand-protein interaction. Docking analysis revealed that the Glu283 is crucial for interaction. Two new amino acid residues, Tyr89 and Thr167 were identified as important in ligand-protein interaction. No site directed mutagenesis studies on these residues have been reported.

Pathogenic infection of Rhesus macaques by an evolving SIV-HIV derived from CCR5-using envelope genes of acute HIV-1 infections

PubMed Central

Asmal, Mohammed; Lane, Sophie; Tian, Meijuan; Nickle, Gabrielle; Venner, Colin; Dirk, Brennan; Dikeakos, Jimmy; Luedemann, Corinne; Mach, Linh; Balachandran, Harikrishnan; Buzby, Adam; Rao, Srinivas; Letvin, Norman; Gao, Yong; Arts, Eric J.

2016-01-01

For studies on vaccines and therapies for HIV disease, SIV-HIV chimeric viruses harboring the HIV-1 env gene (SHIVenv) remain the best virus in non-human primate models. However, there are still very few SHIVenv viruses that can cause AIDS in non-CD8-depleted animals. In the present study, a recently created CCR5-using SHIVenv_B3 virus with env gene derived from acute/early HIV-1 infections (AHI) successfully established pathogenic infection in macaques. Through a series of investigations on the evolution, mutational profile, and phenotype of the virus and the resultant humoral immune response in infected rhesus macaques, we found that the E32K mutation in the Env C1 domain was associated with macaque pathogenesis, and that the electrostatic interactions in Env may favor E32K at the gp120 N terminus and “lock” the binding to heptad repeat 1 of gp41 in the trimer and produce a SHIVenv with increased fitness and pathogenesis during macaque infections. PMID:27723488

Increase in chemokines CXCL10 and CCL2 in blood from pigs infected with high compared to low virulence African swine fever virus isolates.

PubMed

Fishbourne, Emma; Hutet, Evelyne; Abrams, Charles; Cariolet, Roland; Le Potier, Marie-Frédérique; Takamatsu, Haru-H; Dixon, Linda K

2013-10-01

Modulation of the expression of chemokines and chemokine receptors in whole blood was compared following infection of pigs with high and low virulence isolates of African swine fever virus. Levels of mRNAs for CCL2, CCL3L1, CCL4, CXCL10, CCR1 and CCR5 were significantly increased in at least one time point following infection in two experiments and CCL5, CCR9 and CXCR4 mRNA were significantly increased in one of the experiments. The results showed that greatest fold increases in mRNAs for CXCL10 and CCL2 were observed following infection of pigs. CXCL10 mRNA was increased by up to 15 fold in infected compared to uninfected pigs. CXCL10 protein was also detected in serum from pigs infected with the high virulence Benin 97/1 isolate. Levels of CCL2 mRNA were increased in pigs infected with high virulence Benin 97/1 isolate compared to low virulence OURT88/3 isolate and this correlated with an increase of greater than 30 fold in levels of CCL2 protein detected in serum from pigs infected with this isolate. An increase in overall chemotaxis active compounds in defibrinated plasma samples from Benin 97/1 infected pigs was observed at 3 days post-infection (dpi) and a decrease by 7 dpi as measured by chemotaxis assay using normal pig leucocytes in vitro. Increased levels of CXCL10 may either contribute to the activation of lymphocyte priming toward the Th1 phenotype or induction of T lymphocyte apoptosis. Increased levels of CCL2, a chemoattractant for macrophages, may result in increased recruitment of monocytes from bone marrow thus increasing the pool of cells susceptible to infection.

CC chemokine receptor 4 is required for experimental autoimmune encephalomyelitis by regulating GM-CSF and IL-23 production in dendritic cells

PubMed Central

Poppensieker, Karola; Otte, David-Marian; Schürmann, Britta; Limmer, Andreas; Dresing, Philipp; Drews, Eva; Schumak, Beatrix; Klotz, Luisa; Raasch, Jennifer; Mildner, Alexander; Waisman, Ari; Scheu, Stefanie; Knolle, Percy; Förster, Irmgard; Prinz, Marco; Maier, Wolfgang; Zimmer, Andreas; Alferink, Judith

2012-01-01

Dendritic cells (DCs) are pivotal for the development of experimental autoimmune encephalomyelitis (EAE). However, the mechanisms by which they control disease remain to be determined. This study demonstrates that expression of CC chemokine receptor 4 (CCR4) by DCs is required for EAE induction. CCR4−/− mice presented enhanced resistance to EAE associated with a reduction in IL-23 and GM-CSF expression in the CNS. Restoring CCR4 on myeloid cells in bone marrow chimeras or intracerebral microinjection of CCR4-competent DCs, but not macrophages, restored EAE in CCR4−/− mice, indicating that CCR4+ DCs are cellular mediators of EAE development. Mechanistically, CCR4−/− DCs were less efficient in GM-CSF and IL-23 production and also TH-17 maintenance. Intraspinal IL-23 reconstitution restored EAE in CCR4−/− mice, whereas intracerebral inoculation using IL-23−/− DCs or GM-CSF−/− DCs failed to induce disease. Thus, CCR4-dependent GM-CSF production in DCs required for IL-23 release in these cells is a major component in the development of EAE. Our study identified a unique role for CCR4 in regulating DC function in EAE, harboring therapeutic potential for the treatment of CNS autoimmunity by targeting CCR4 on this specific cell type. PMID:22355103

Productive Entry of HIV-1 during Cell-to-Cell Transmission via Dynamin-Dependent Endocytosis

PubMed Central

Sloan, Richard D.; Kuhl, Björn D.; Mesplède, Thibault; Münch, Jan; Donahue, Daniel A.

2013-01-01

HIV-1 can be transmitted as cell-free virus or via cell-to-cell contacts. Cell-to-cell transmission between CD4+ T cells is the more efficient mode of transmission and is predominant in lymphoid tissue, where the majority of virus resides. Yet the cellular mechanisms underlying productive cell-to-cell transmission in uninfected target cells are unclear. Although it has been demonstrated that target cells can take up virus via endocytosis, definitive links between this process and productive infection remain undefined, and this route of transmission has been proposed to be nonproductive. Here, we report that productive cell-to-cell transmission can occur via endocytosis in a dynamin-dependent manner and is sensitive to clathrin-associated antagonists. These data were obtained in a number of CD4+ T-cell lines and in primary CD4+ T cells, using both CXCR4- and CCR5-tropic virus. However, we also found that HIV-1 demonstrated flexibility in its use of such endocytic pathways as certain allogeneic transmissions were seen to occur in a dynamin-dependent manner but were insensitive to clathrin-associated antagonists. Also, depleting cells of the clathrin accessory protein AP180 led to a viral uptake defect associated with enhanced infection. Collectively, these data demonstrate that endosomal uptake of HIV-1 during cell-to-cell transmission leads to productive infection, but they are also indicative of a flexible model of viral entry during cell-to-cell transmission, in which the virus can alter its entry route according to the pressures that it encounters. PMID:23678185

Plasmacytoid Dendritic Cells Suppress HIV-1 Replication but Contribute to HIV-1 Induced Immunopathogenesis in Humanized Mice

PubMed Central

Li, Guangming; Cheng, Menglan; Nunoya, Jun-ichi; Cheng, Liang; Guo, Haitao; Yu, Haisheng; Liu, Yong-jun; Su, Lishan; Zhang, Liguo

2014-01-01

The role of plasmacytoid dendritic cells (pDC) in human immunodeficiency virus type 1 (HIV-1) infection and pathogenesis remains unclear. HIV-1 infection in the humanized mouse model leads to persistent HIV-1 infection and immunopathogenesis, including type I interferons (IFN-I) induction, immune-activation and depletion of human leukocytes, including CD4 T cells. We developed a monoclonal antibody that specifically depletes human pDC in all lymphoid organs in humanized mice. When pDC were depleted prior to HIV-1 infection, the induction of IFN-I and interferon-stimulated genes (ISGs) were abolished during acute HIV-1 infection with either a highly pathogenic CCR5/CXCR4-dual tropic HIV-1 or a standard CCR5-tropic HIV-1 isolate. Consistent with the anti-viral role of IFN-I, HIV-1 replication was significantly up-regulated in pDC-depleted mice. Interestingly, the cell death induced by the highly pathogenic HIV-1 isolate was severely reduced in pDC-depleted mice. During chronic HIV-1 infection, depletion of pDC also severely reduced the induction of IFN-I and ISGs, associated with elevated HIV-1 replication. Surprisingly, HIV-1 induced depletion of human immune cells including T cells in lymphoid organs, but not the blood, was reduced in spite of the increased viral replication. The increased cell number in lymphoid organs was associated with a reduced level of HIV-induced cell death in human leukocytes including CD4 T cells. We conclude that pDC play opposing roles in suppressing HIV-1 replication and in promoting HIV-1 induced immunopathogenesis. These findings suggest that pDC-depletion and IFN-I blockade will provide novel strategies for treating those HIV-1 immune non-responsive patients with persistent immune activation despite effective anti-retrovirus treatment. PMID:25077616

Precursor Forms of Vitamin D Reduce HIV-1 Infection In Vitro.

PubMed

Aguilar-Jimenez, Wbeimar; Villegas-Ospina, Simon; Gonzalez, Sandra; Zapata, Wildeman; Saulle, Irma; Garziano, Micaela; Biasin, Mara; Clerici, Mario; Rugeles, Maria T

2016-12-15

Although the anti-HIV-1 effects of vitamin D (VitD) have been reported, mechanisms behind such protection remain largely unexplored. The effects of two precursor forms (cholecalciferol/calciol at 0.01, 1 and 100 nM and calcidiol at 100 and 250 nM) on HIV-1 infection, immune activation, and gene expression were analyzed in vitro in cells of Colombian and Italian healthy donors. We quantified levels of released p24 by enzyme-linked immunosorbent assay, of intracellular p24 and cell-surface expression of CD38 and HLA-DR by flow cytometry, and mRNA expression of antiviral and immunoregulatory genes by real-time reverse transcription-polymerase chain reaction. Cholecalciferol decreased the frequency of HIV-1-infected p24CD4 T cells and levels of p24 in supernatants in a dose-dependent manner. Moreover, the CD4CD38HLA-DR and CD4CD38HLA-DR subpopulations were more susceptible to infection but displayed the greatest cholecalciferol-induced decreases in infection rate by an X4-tropic strain. Likewise, cholecalciferol at its highest concentration decreased the frequency of CD38HLA-DR but not of CD38HLA-DR T-cell subsets. Analyzing the effects of calcidiol, the main VitD source for immune cells and an R5-tropic strain as the most frequently transmitted virus, a reduction in HIV-1 productive infection was also observed. In addition, an increase in mRNA expression of APOBEC3G and PI3 and a reduction of TRIM22 and CCR5 expression, this latter positively correlated with p24 levels, was noted. VitD reduces HIV-1 infection in T cells possibly by inducing antiviral gene expression, reducing the viral co-receptor CCR5 and, at least at the highest cholecalciferol concentration, by promoting an HIV-1-restrictive CD38HLA-DR immunophenotype.

Complexity and dynamics of HIV-1 chemokine receptor usage in a multidrug-resistant adolescent.

PubMed

Cavarelli, Mariangela; Mainetti, Lara; Pignataro, Angela Rosa; Bigoloni, Alba; Tolazzi, Monica; Galli, Andrea; Nozza, Silvia; Castagna, Antonella; Sampaolo, Michela; Boeri, Enzo; Scarlatti, Gabriella

2014-12-01

Maraviroc (MVC) is licensed in clinical practice for patients with R5 virus and virological failure; however, in anecdotal reports, dual/mixed viruses were also inhibited. We retrospectively evaluated the evolution of HIV-1 coreceptor tropism in plasma and peripheral blood mononuclear cells (PBMCs) of an infected adolescent with a CCR5/CXCR4 Trofile profile who experienced an important but temporary immunological and virological response during a 16-month period of MVC-based therapy. Coreceptor usage of biological viral clones isolated from PBMCs was investigated in U87.CD4 cells expressing wild-type or chimeric CCR5 and CXCR4. Plasma and PBMC-derived viral clones were sequenced to predict coreceptor tropism using the geno2pheno algorithm from the V3 envelope sequence and pol gene-resistant mutations. From start to 8.5 months of MVC treatment only R5X4 viral clones were observed, whereas at 16 months the phenotype enlarged to also include R5 and X4 clones. Chimeric receptor usage suggested the preferential usage of the CXCR4 coreceptor by the R5X4 biological clones. According to phenotypic data, R5 viruses were susceptible, whereas R5X4 and X4 viruses were resistant to RANTES and MVC in vitro. Clones at 16 months, but not at baseline, showed an amino acidic resistance pattern in protease and reverse transcription genes, which, however, did not drive their tropisms. The geno2pheno algorithm predicted at baseline R5 viruses in plasma, and from 5.5 months throughout follow-up only CXCR4-using viruses. An extended methodological approach is needed to unravel the complexity of the phenotype and variation of viruses resident in the different compartments of an infected individual. The accurate evaluation of the proportion of residual R5 viruses may guide therapeutic intervention in highly experienced patients with limited therapeutic options.

Complexity and Dynamics of HIV-1 Chemokine Receptor Usage in a Multidrug-Resistant Adolescent

PubMed Central

Mainetti, Lara; Pignataro, Angela Rosa; Bigoloni, Alba; Tolazzi, Monica; Galli, Andrea; Nozza, Silvia; Castagna, Antonella; Sampaolo, Michela; Boeri, Enzo; Scarlatti, Gabriella

2014-01-01

Abstract Maraviroc (MVC) is licensed in clinical practice for patients with R5 virus and virological failure; however, in anecdotal reports, dual/mixed viruses were also inhibited. We retrospectively evaluated the evolution of HIV-1 coreceptor tropism in plasma and peripheral blood mononuclear cells (PBMCs) of an infected adolescent with a CCR5/CXCR4 Trofile profile who experienced an important but temporary immunological and virological response during a 16-month period of MVC-based therapy. Coreceptor usage of biological viral clones isolated from PBMCs was investigated in U87.CD4 cells expressing wild-type or chimeric CCR5 and CXCR4. Plasma and PBMC-derived viral clones were sequenced to predict coreceptor tropism using the geno2pheno algorithm from the V3 envelope sequence and pol gene-resistant mutations. From start to 8.5 months of MVC treatment only R5X4 viral clones were observed, whereas at 16 months the phenotype enlarged to also include R5 and X4 clones. Chimeric receptor usage suggested the preferential usage of the CXCR4 coreceptor by the R5X4 biological clones. According to phenotypic data, R5 viruses were susceptible, whereas R5X4 and X4 viruses were resistant to RANTES and MVC in vitro. Clones at 16 months, but not at baseline, showed an amino acidic resistance pattern in protease and reverse transcription genes, which, however, did not drive their tropisms. The geno2pheno algorithm predicted at baseline R5 viruses in plasma, and from 5.5 months throughout follow-up only CXCR4-using viruses. An extended methodological approach is needed to unravel the complexity of the phenotype and variation of viruses resident in the different compartments of an infected individual. The accurate evaluation of the proportion of residual R5 viruses may guide therapeutic intervention in highly experienced patients with limited therapeutic options. PMID:25275490

Postdoctoral Fellow | Center for Cancer Research

Cancer.gov

A postdoctoral position is available in the Viral Recombination Section (VRS), HIV Dynamics and Replication Program, CCR.  The VRS studies retroviral replication using human immunodeficiency viruses and other retroviruses, with a particular emphasis on the mechanisms of viral RNA biology, specific RNA packaging, virus assembly, and HIV replication.  Molecular tools and

Punica granatum (Pomegranate) juice provides an HIV-1 entry inhibitor and candidate topical microbicide

PubMed Central

Neurath, A Robert; Strick, Nathan; Li, Yun-Yao; Debnath, Asim K

2004-01-01

Background For ≈ 24 years the AIDS pandemic has claimed ≈ 30 million lives, causing ≈ 14,000 new HIV-1 infections daily worldwide in 2003. About 80% of infections occur by heterosexual transmission. In the absence of vaccines, topical microbicides, expected to block virus transmission, offer hope for controlling the pandemic. Antiretroviral chemotherapeutics have decreased AIDS mortality in industrialized countries, but only minimally in developing countries. To prevent an analogous dichotomy, microbicides should be: acceptable; accessible; affordable; and accelerative in transition from development to marketing. Already marketed pharmaceutical excipients or foods, with established safety records and adequate anti-HIV-1 activity, may provide this option. Methods Fruit juices were screened for inhibitory activity against HIV-1 IIIB using CD4 and CXCR4 as cell receptors. The best juice was tested for inhibition of: (1) infection by HIV-1 BaL, utilizing CCR5 as the cellular coreceptor; and (2) binding of gp120 IIIB and gp120 BaL, respectively, to CXCR4 and CCR5. To remove most colored juice components, the adsorption of the effective ingredient(s) to dispersible excipients and other foods was investigated. A selected complex was assayed for inhibition of infection by primary HIV-1 isolates. Results HIV-1 entry inhibitors from pomegranate juice adsorb onto corn starch. The resulting complex blocks virus binding to CD4 and CXCR4/CCR5 and inhibits infection by primary virus clades A to G and group O. Conclusion These results suggest the possibility of producing an anti-HIV-1 microbicide from inexpensive, widely available sources, whose safety has been established throughout centuries, provided that its quality is adequately standardized and monitored. PMID:15485580

Role of Abl kinase and the Wave2 signaling complex in HIV-1 entry at a post-hemifusion step.

PubMed

Harmon, Brooke; Campbell, Nancy; Ratner, Lee

2010-06-17

Entry of human immunodeficiency virus type 1 (HIV-1) commences with binding of the envelope glycoprotein (Env) to the receptor CD4, and one of two coreceptors, CXCR4 or CCR5. Env-mediated signaling through coreceptor results in Galphaq-mediated Rac activation and actin cytoskeleton rearrangements necessary for fusion. Guanine nucleotide exchange factors (GEFs) activate Rac and regulate its downstream protein effectors. In this study we show that Env-induced Rac activation is mediated by the Rac GEF Tiam-1, which associates with the adaptor protein IRSp53 to link Rac to the Wave2 complex. Rac and the tyrosine kinase Abl then activate the Wave2 complex and promote Arp2/3-dependent actin polymerization. Env-mediated cell-cell fusion, virus-cell fusion and HIV-1 infection are dependent on Tiam-1, Abl, IRSp53, Wave2, and Arp3 as shown by attenuation of fusion and infection in cells expressing siRNA targeted to these signaling components. HIV-1 Env-dependent cell-cell fusion, virus-cell fusion and infection were also inhibited by Abl kinase inhibitors, imatinib, nilotinib, and dasatinib. Treatment of cells with Abl kinase inhibitors did not affect cell viability or surface expression of CD4 and CCR5. Similar results with inhibitors and siRNAs were obtained when Env-dependent cell-cell fusion, virus-cell fusion or infection was measured, and when cell lines or primary cells were the target. Using membrane curving agents and fluorescence microscopy, we showed that inhibition of Abl kinase activity arrests fusion at the hemifusion (lipid mixing) step, suggesting a role for Abl-mediated actin remodeling in pore formation and expansion. These results suggest a potential utility of Abl kinase inhibitors to treat HIV-1 infected patients.

HIV envelope-mediated, CCR5/α4β7-dependent killing of CD4-negative γδ T cells which are lost during progression to AIDS.

PubMed

Li, Haishan; Pauza, C David

2011-11-24

HIV infects and replicates in CD4+ T cells but effects on host immunity and disease also involve depletion, hyper-activation, and modification of CD4-negative cell populations. In particular, the depletion of CD4-negative γδ T cells is common to all HIV+ individuals. We found that soluble or cell-associated envelope glycoproteins from CCR5-tropic strains of HIV could bind, activates the p38-caspase pathway, and induce the death of γδ cells. Envelope binding requires integrin α4β7 and chemokine receptor CCR5 which are at high levels and form a complex on the γδ T cell membrane. This receptor complex facilitated V3 loop binding to CCR5 in the absence of CD4-induced conformational changes. Cell death was increased by antigen stimulation after exposure to envelope glycoprotein. Direct signaling by envelope glycoprotein killed CD4-negative γδ T cells and reproduced a defect observed in all patients with HIV disease.

Development of a rapid cell-fusion-based phenotypic HIV-1 tropism assay

PubMed Central

Teeranaipong, Phairote; Hosoya, Noriaki; Kawana-Tachikawa, Ai; Fujii, Takeshi; Koibuchi, Tomohiko; Nakamura, Hitomi; Koga, Michiko; Kondo, Naoyuki; Gao, George F; Hoshino, Hiroo; Matsuda, Zene; Iwamoto, Aikichi

2013-01-01

Introduction A dual split reporter protein system (DSP), recombining Renilla luciferase (RL) and green fluorescent protein (GFP) split into two different constructs (DSP1–7 and DSP8–11), was adapted to create a novel rapid phenotypic tropism assay (PTA) for HIV-1 infection (DSP-Pheno). Methods DSP1–7 was stably expressed in the glioma-derived NP-2 cell lines, which expressed CD4/CXCR4 (N4X4) or CD4/CCR5 (N4R5), respectively. An expression vector with DSP8–11 (pRE11) was constructed. The HIV-1 envelope genes were subcloned in pRE11 (pRE11-env) and transfected into 293FT cells. Transfected 293FT cells were incubated with the indicator cell lines independently. In developing the assay, we selected the DSP1–7-positive clones that showed the highest GFP activity after complementation with DSP8–11. These cell lines, designated N4R5-DSP1–7, N4X4-DSP1–7 were used for subsequent assays. Results The env gene from the reference strains (BaL for R5 virus, NL4-3 for X4 virus, SF2 for dual tropic virus) subcloned in pRE11 and tested, was concordant with the expected co-receptor usage. Assay results were available in two ways (RL or GFP). The assay sensitivity by RL activity was comparable with those of the published phenotypic assays using pseudovirus. The shortest turnaround time was 5 days after obtaining the patient's plasma. All clinical samples gave positive RL signals on R5 indicator cells in the fusion assay. Median RLU value of the low CD4 group was significantly higher on X4 indicator cells and suggested the presence of more dual or X4 tropic viruses in this group of patients. Comparison of representative samples with Geno2Pheno [co-receptor] assay was concordant. Conclusions A new cell-fusion-based, high-throughput PTA for HIV-1, which would be suitable for in-house studies, was developed. Equipped with two-way reporter system, RL and GFP, DSP-Pheno is a sensitive test with short turnaround time. Although maintenance of cell lines and laboratory equipment is necessary, it provides a safe assay system without infectious viruses. With further validation against other conventional analyses, DSP-Pheno may prove to be a useful laboratory tool. The assay may be useful especially for the research on non-B subtype HIV-1 whose co-receptor usage has not been studied much. PMID:24050252

Blood CXCR3+ CD4 T Cells Are Enriched in Inducible Replication Competent HIV in Aviremic Antiretroviral Therapy-Treated Individuals

PubMed Central

Banga, Riddhima; Procopio, Francesco A.; Ruggiero, Alessandra; Noto, Alessandra; Ohmiti, Khalid; Cavassini, Matthias; Corpataux, Jean-Marc; Paxton, William A.; Pollakis, Georgios; Perreau, Matthieu

2018-01-01

We recently demonstrated that lymph nodes (LNs) PD-1+/T follicular helper (Tfh) cells from antiretroviral therapy (ART)-treated HIV-infected individuals were enriched in cells containing replication competent virus. However, the distribution of cells containing inducible replication competent virus has been only partially elucidated in blood memory CD4 T-cell populations including the Tfh cell counterpart circulating in blood (cTfh). In this context, we have investigated the distribution of (1) total HIV-infected cells and (2) cells containing replication competent and infectious virus within various blood and LN memory CD4 T-cell populations of conventional antiretroviral therapy (cART)-treated HIV-infected individuals. In the present study, we show that blood CXCR3-expressing memory CD4 T cells are enriched in cells containing inducible replication competent virus and contributed the most to the total pool of cells containing replication competent and infectious virus in blood. Interestingly, subsequent proviral sequence analysis did not indicate virus compartmentalization between blood and LN CD4 T-cell populations, suggesting dynamic interchanges between the two compartments. We then investigated whether the composition of blood HIV reservoir may reflect the polarization of LN CD4 T cells at the time of reservoir seeding and showed that LN PD-1+ CD4 T cells of viremic untreated HIV-infected individuals expressed significantly higher levels of CXCR3 as compared to CCR4 and/or CCR6, suggesting that blood CXCR3-expressing CD4 T cells may originate from LN PD-1+ CD4 T cells. Taken together, these results indicate that blood CXCR3-expressing CD4 T cells represent the major blood compartment containing inducible replication competent virus in treated aviremic HIV-infected individuals. PMID:29459864

Blood CXCR3+ CD4 T Cells Are Enriched in Inducible Replication Competent HIV in Aviremic Antiretroviral Therapy-Treated Individuals.

PubMed

Banga, Riddhima; Procopio, Francesco A; Ruggiero, Alessandra; Noto, Alessandra; Ohmiti, Khalid; Cavassini, Matthias; Corpataux, Jean-Marc; Paxton, William A; Pollakis, Georgios; Perreau, Matthieu

2018-01-01

We recently demonstrated that lymph nodes (LNs) PD-1 + /T follicular helper (Tfh) cells from antiretroviral therapy (ART)-treated HIV-infected individuals were enriched in cells containing replication competent virus. However, the distribution of cells containing inducible replication competent virus has been only partially elucidated in blood memory CD4 T-cell populations including the Tfh cell counterpart circulating in blood (cTfh). In this context, we have investigated the distribution of (1) total HIV-infected cells and (2) cells containing replication competent and infectious virus within various blood and LN memory CD4 T-cell populations of conventional antiretroviral therapy (cART)-treated HIV-infected individuals. In the present study, we show that blood CXCR3-expressing memory CD4 T cells are enriched in cells containing inducible replication competent virus and contributed the most to the total pool of cells containing replication competent and infectious virus in blood. Interestingly, subsequent proviral sequence analysis did not indicate virus compartmentalization between blood and LN CD4 T-cell populations, suggesting dynamic interchanges between the two compartments. We then investigated whether the composition of blood HIV reservoir may reflect the polarization of LN CD4 T cells at the time of reservoir seeding and showed that LN PD-1 + CD4 T cells of viremic untreated HIV-infected individuals expressed significantly higher levels of CXCR3 as compared to CCR4 and/or CCR6, suggesting that blood CXCR3-expressing CD4 T cells may originate from LN PD-1 + CD4 T cells. Taken together, these results indicate that blood CXCR3-expressing CD4 T cells represent the major blood compartment containing inducible replication competent virus in treated aviremic HIV-infected individuals.

Synthesis and circularization of N- and B-tropic retroviral DNA in Fv-1 permissive and restrictive mouse cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, W.K.; Kiggans, J.O.; Yang, D.M.

1980-05-01

Production of various forms of nonintegrated viral DNA was measured in cultured mouse cells carrying different Fv-1 alleles early after infection with N-tropic or B-tropic retroviruses. Quantitative analyses were performed by agarose gel electrophoresis, transfer to diazobenzyloxymethyl-paper, and molecular hybridization. In permissive infection of Fv-1/sup n/ cells (NIH Swiss and DBA mouse strains) with N-tropic virus and of Fv-1/sup b/ cells (BALB/c and C57BL/6 strains) with B-tropic virus, form III (double-stranded linear) DNA first appeared at 3 to 4 hr and reached a maximum at 8 to 10 hr; two form I (closed circle) DNAs appeared at 7 to 8more » hr and reached a maximum at or beyond 12 hr. In the two Fv-1/sup b/ cells infected with N-tropic virus and in DBA (Fv-1/sup n/) cells infected with B-tropic virus, formation of the two form I DNAs was quantitatively restricted but formation of form III DNA was unaltered. In Fv-1/sup n/ NIH Swiss mouse embryo cells infected with B-tropic virus, the level of form III DNA was markedly depressed and hence the two form I DNAs were not detectable. In C57BL/6 cells as well as in DBA/2 cells 12 hr after infection, the quantity of form III DNA varied directly with the amount of restricted virus, whereas the quantity of form I DNA varied according to the square of the amount of restricted virus. The significance of these results for understanding the molecular basis of retrovirus replication and its restriction by the Fv-1 gene is discussed.« less

Relationship between white spot syndrome virus (WSSV) loads and characterizations of water quality in Litopenaeus vannamei culture ponds during the tropical storm.

PubMed

Zhang, J S; Li, Z J; Wen, G L; Wang, Y L; Luo, L; Zhang, H J; Dong, H B

2016-01-01

An in-situ experiment was conducted to investigate the effect of tropical storm on the white spot syndrome virus (WSSV) loads in Litopenaeus vannamei rearing ponds. White spot syndrome virus loads, heterotrophic bacteria, Vibrio and water quality (including temperature, dissolved oxygen (DO), salinity, pH, NH 4 -N, and NO 2 -N) were continually monitored through one tropical storm. The WSSV loads decreased when tropical storm made landfall, and substantially increased when typhoon passed. The variation of WSSV loads was correlated with DO, temperature, heterotrophic bacteria count, and ammonia-N concentrations. These results suggested that maintaining high level DO and promoting heterotrophic bacteria growth in the shrimp ponds might prevent the diseases' outbreak after the landfall of tropical storm.

Relationship between white spot syndrome virus (WSSV) loads and characterizations of water quality in Litopenaeus vannamei culture ponds during the tropical storm

PubMed Central

Zhang, J. S.; Li, Z. J.; Wen, G. L.; Wang, Y. L.; Luo, L.; Zhang, H. J.; Dong, H. B.

2016-01-01

An in-situ experiment was conducted to investigate the effect of tropical storm on the white spot syndrome virus (WSSV) loads in Litopenaeus vannamei rearing ponds. White spot syndrome virus loads, heterotrophic bacteria, Vibrio and water quality (including temperature, dissolved oxygen (DO), salinity, pH, NH4-N, and NO2-N) were continually monitored through one tropical storm. The WSSV loads decreased when tropical storm made landfall, and substantially increased when typhoon passed. The variation of WSSV loads was correlated with DO, temperature, heterotrophic bacteria count, and ammonia-N concentrations. These results suggested that maintaining high level DO and promoting heterotrophic bacteria growth in the shrimp ponds might prevent the diseases’ outbreak after the landfall of tropical storm. PMID:27822254

Recombinant Human T-Cell Leukemia Virus Types 1 and 2 Tax Proteins Induce High Levels of CC-Chemokines and Downregulate CCR5 in Human Peripheral Blood Mononuclear Cells

PubMed Central

Barrios, Christy S.; Abuerreish, Muna; Lairmore, Michael D.; Castillo, Laura; Giam, Chou-Zen

2011-01-01

Abstract Human T-cell leukemia viruses types 1 (HTLV-1) and 2 (HTLV-2) produce key transcriptional regulatory gene products, known as Tax1 and Tax2, respectively. Tax1 and Tax2 transactivate multiple host genes involved in cellular immune responses within the cellular microenvironment, including induction of genes encoding expression of CC-chemokines. It is speculated that HTLV Tax proteins may act as immune modulators. In this study, recombinant Tax1 and Tax2 proteins were tested for their effects on the viability of cultured peripheral blood mononuclear cells (PBMCs), and their ability to induce expression of CC-chemokines and to downregulate the level of CCR5 expression in PBMCs. PBMCs obtained from uninfected donors were cultured in a range of Tax1 and Tax2 concentrations (10–100 pM), and supernatant fluids were harvested at multiple time points for quantitative determinations of MIP-1α/CCL3, MIP-1β/CCL4, and RANTES/CCL5. Treatment of PBMCs with Tax1 and Tax2 proteins (100 pM) resulted in a significant increase in viability over a 7-d period compared to controls (p<0.01). Both Tax1 and Tax2 induced high levels of all three CC-chemokines over the dosing range compared to mock-treated controls (p<0.05). The gated population of lymphocytes treated with Tax2, as well as lymphocytes from HTLV-2-infected donors, showed a significantly lower percentage of CCR5-positive cells compared to those of uninfected donors and from mock-treated lymphocytes, respectively (p<0.05). These results suggest that Tax1 and Tax2 could promote innate immunity in the extracellular environment during HTLV-1 and HTLV-2 infections via CC-chemokine ligands and receptors. PMID:22111594

Role for Macrophage Metalloelastase in Glomerular Basement Membrane Damage Associated with Alport Syndrome

PubMed Central

Rao, Velidi H.; Meehan, Daniel T.; Delimont, Duane; Nakajima, Motowo; Wada, Takashi; Ann Gratton, Michael; Cosgrove, Dominic

2006-01-01

Alport syndrome is a glomerular basement membrane (GBM) disease caused by mutations in type IV collagen genes. A unique irregular thickening and thinning of the GBM characterizes the progressive glomerular pathology. The metabolic imbalances responsible for these GBM irregularities are not known. Here we show that macrophage metalloelastase (MMP-12) expression is >40-fold induced in glomeruli from Alport mice and is markedly induced in glomeruli of both humans and dogs with Alport syndrome. Treatment of Alport mice with MMI270 (CGS27023A), a broad spectrum MMP inhibitor that blocks MMP-12 activity, results in largely restored GBM ultrastructure and function. Treatment with BAY-129566, a broad spectrum MMP inhibitor that does not inhibit MMP-12, had no effect. We show that inhibition of CC chemokine receptor 2 (CCR2) receptor signaling with propagermanium blocks induction of MMP-12 mRNA and prevents GBM damage. CCR2 receptor is expressed in glomerular podocytes of Alport mice, suggesting MCP-1 activation of CCR2 on podocytes may underlie induction of MMP-12. These data indicate that the irregular GBM that characterizes Alport syndrome may be mediated, in part, by focal degradation of the GBM due to MMP dysregulation, in particular, MMP-12. Thus, MMP-12/CCR2 inhibitors may provide a novel and effective therapeutic strategy for Alport glomerular disease. PMID:16816359

Distinct T and NK cell populations may serve as immune correlates of protection against symptomatic pandemic influenza A(H1N1) virus infection during pregnancy

PubMed Central

Dembinski, Jennifer L.; Laake, Ida; Hungnes, Olav; Cox, Rebecca; Oftung, Fredrik; Trogstad, Lill; Mjaaland, Siri

2017-01-01

Maternal influenza infection during pregnancy is associated with increased risk of morbidity and mortality. However, the link between the anti-influenza immune responses and health-related risks during infection is not well understood. We have analyzed memory T and NK cell mediated immunity (CMI) responses in pandemic influenza A(H1N1)pdm09 (pdm09) virus infected non-vaccinated pregnant women participating in the Norwegian Influenza Pregnancy Cohort (NorFlu). The cohort includes information on immunization, self-reported health and disease status, and biological samples (plasma and PBMC). Infected cases (N = 75) were defined by having a serum hemagglutination inhibition (HI) titer > = 20 to influenza pdm09 virus at the time of delivery, while controls (N = 75) were randomly selected among non-infected pregnant women (HI titer <10). In ELISpot assays cases had higher frequencies of IFNγ+ CD8+ T cells responding to pdm09 virus or conserved CD8 T cell-restricted influenza A virus epitopes, compared to controls. Within this T cell population, frequencies of CD95+ late effector (CD45RA+CCR7-) and naive (CD45RA+CCR7+) CD8+ memory T cells correlated inversely with self-reported influenza illness (ILI) symptoms. ILI symptoms in infected women were also associated with lower numbers of poly-functional (IFNγ+TNFα+, IL2+IFNγ+, IL2+IFNγ+TNFα+) CD4+ T cells and increased frequencies of IFNγ+CD3-CD7+ NK cells compared to asymptomatic cases, or controls, after stimulation with the pdm09 virus. Taken together, virus specific and functionally distinct T and NK cell populations may serve as cellular immune correlates of clinical outcomes of pandemic influenza disease in pregnant women. Our results may provide information important for future universal influenza vaccine design. PMID:29145441

Distinct T and NK cell populations may serve as immune correlates of protection against symptomatic pandemic influenza A(H1N1) virus infection during pregnancy.

PubMed

Savic, Miloje; Dembinski, Jennifer L; Laake, Ida; Hungnes, Olav; Cox, Rebecca; Oftung, Fredrik; Trogstad, Lill; Mjaaland, Siri

2017-01-01

Maternal influenza infection during pregnancy is associated with increased risk of morbidity and mortality. However, the link between the anti-influenza immune responses and health-related risks during infection is not well understood. We have analyzed memory T and NK cell mediated immunity (CMI) responses in pandemic influenza A(H1N1)pdm09 (pdm09) virus infected non-vaccinated pregnant women participating in the Norwegian Influenza Pregnancy Cohort (NorFlu). The cohort includes information on immunization, self-reported health and disease status, and biological samples (plasma and PBMC). Infected cases (N = 75) were defined by having a serum hemagglutination inhibition (HI) titer > = 20 to influenza pdm09 virus at the time of delivery, while controls (N = 75) were randomly selected among non-infected pregnant women (HI titer <10). In ELISpot assays cases had higher frequencies of IFNγ+ CD8+ T cells responding to pdm09 virus or conserved CD8 T cell-restricted influenza A virus epitopes, compared to controls. Within this T cell population, frequencies of CD95+ late effector (CD45RA+CCR7-) and naive (CD45RA+CCR7+) CD8+ memory T cells correlated inversely with self-reported influenza illness (ILI) symptoms. ILI symptoms in infected women were also associated with lower numbers of poly-functional (IFNγ+TNFα+, IL2+IFNγ+, IL2+IFNγ+TNFα+) CD4+ T cells and increased frequencies of IFNγ+CD3-CD7+ NK cells compared to asymptomatic cases, or controls, after stimulation with the pdm09 virus. Taken together, virus specific and functionally distinct T and NK cell populations may serve as cellular immune correlates of clinical outcomes of pandemic influenza disease in pregnant women. Our results may provide information important for future universal influenza vaccine design.

A database of human genes and a gene network involved in response to tick-borne encephalitis virus infection.

PubMed

Ignatieva, Elena V; Igoshin, Alexander V; Yudin, Nikolay S

2017-12-28

Tick-borne encephalitis is caused by the neurotropic, positive-sense RNA virus, tick-borne encephalitis virus (TBEV). TBEV infection can lead to a variety of clinical manifestations ranging from slight fever to severe neurological illness. Very little is known about genetic factors predisposing to severe forms of disease caused by TBEV. The aims of the study were to compile a catalog of human genes involved in response to TBEV infection and to rank genes from the catalog based on the number of neighbors in the network of pairwise interactions involving these genes and TBEV RNA or proteins. Based on manual review and curation of scientific publications a catalog comprising 140 human genes involved in response to TBEV infection was developed. To provide access to data on all genes, the TBEVhostDB web resource ( http://icg.nsc.ru/TBEVHostDB/ ) was created. We reconstructed a network formed by pairwise interactions between TBEV virion itself, viral RNA and viral proteins and 140 genes/proteins from TBEVHostDB. Genes were ranked according to the number of interactions in the network. Two genes/proteins (CCR5 and IFNAR1) that had maximal number of interactions were revealed. It was found that the subnetworks formed by CCR5 and IFNAR1 and their neighbors were a fragments of two key pathways functioning during the course of tick-borne encephalitis: (1) the attenuation of interferon-I signaling pathway by the TBEV NS5 protein that targeted peptidase D; (2) proinflammation and tissue damage pathway triggered by chemokine receptor CCR5 interacting with CD4, CCL3, CCL4, CCL2. Among nine genes associated with severe forms of TBEV infection, three genes/proteins (CCR5, IL10, ARID1B) were found to have protein-protein interactions within the network, and two genes/proteins (IFNL3 and the IL10, that was just mentioned) were up- or down-regulated in response to TBEV infection. Based on this finding, potential mechanisms for participation of CCR5, IL10, ARID1B, and IFNL3 in the host response to TBEV infection were suggested. A database comprising 140 human genes involved in response to TBEV infection was compiled and the TBEVHostDB web resource, providing access to all genes was created. This is the first effort of integrating and unifying data on genetic factors that may predispose to severe forms of diseases caused by TBEV. The TBEVHostDB could potentially be used for assessment of risk factors for severe forms of tick-borne encephalitis and for the design of personalized pharmacological strategies for the treatment of TBEV infection.

Diet-induced obesity prolongs neuroinflammation and recruits CCR2+ monocytes to the brain following herpes simplex virus (HSV)-1 latency in mice

PubMed Central

White, Katherine A.; Hutton, Scott R.; Weimer, Jill M.

2016-01-01

Herpes simplex virus (HSV)-1 is a ubiquitous human infection, with increased prevalence in obese populations. Obesity has been linked to increased inflammation, susceptibility to infection, and higher rates of anxiety disorder and cognitive impairment. To determine how obesity alters neuroinflammation and behavior following infection, we infected weanling C57BL/6 or CCR2RFP/+/CX3CR1GFP/+ mice with a very low dose of HSV-1. Following viral latency (14 days post infection (d p.i.)), mice were randomly assigned to remain on the low fat (LF) diet or switched to a 45% high fat (HF) diet. Eight weeks post diet shift, latently infected mice on the HF diet (HSV-HF) had greater microglial activation and infiltration of inflammatory CCR2+ monocytes in the hypothalamus and dentate gyrus, in comparison to both HSV-LF mice and uninfected mice on LF and HF diets. VCAM staining was present in hypothalamus and hippocampus of the HSV-HF mice in the areas of monocyte infiltration. Infiltrating monocytes also produced proinflammatory cytokines demonstrating that, along with activated microglia, monocytes contribute to sustained neuroinflammation in latently infected obese mice. Utilizing a light-dark preference test, we found that HSV-HF mice had increased anxiety-like behavior. In the marble-burying test, HF diet and HSV infection resulted in increased numbers of buried marbles. Together, these mice provide a useful, testable model to study the biobehavioral effects of obesity and latent HSV-1 infection in regards to anxiety and may provide a tool for studying diet intervention programs in the future. PMID:27311830

Genetic diversity and linkage disequilibrium in the chemokine receptor CCR2-CCR5 region among individuals and populations

PubMed Central

Lawhorn, Collene; Yuferov, Vadim; Randesi, Matthew; Ho, Ann; Morgello, Susan; Kreek, Mary Jeanne; Levran, Orna

2013-01-01

Background Chemokine receptors CCR2 and CCR5 play a key role in immune and inflammatory responses and have been associated with several diseases, including AIDS. In order to comprehend health disparities it is important to understand the nature of genetic variation in specific genes of interest in different populations. Current studies of the CCR2 and CCR5 receptor genes are primarily focused on the CCR5-Δ32, and CCR2-V64I SNPs. Methods Sanger sequencing was used to sequence the regions containing 16 SNPs in the adjacent CCR2 and CCR5 genes (including CCR5-Δ32, and CCR2-V64I) in 249 subjects of African, European and Hispanic ancestry. Linkage disequilibrium (LD) and haplotypes were determined using Haploview. Results The data revealed large differences in allele frequencies of several SNPs and LD patterns among the ethnic groups, including SNPs that were restricted to Africans or Europeans. Seven known CCR5 haplotypes and six novel CCR2 haplotypes were identified. A rare case of an HIV+ subject with the CCR5-Δ32/Δ32 was identified. Conclusions These data demonstrate a LD between CCR2 and CCR5 at several loci and provide new information about CCR2 that contributes to our understanding of its population-specific genetic variability. The data indicate that in addition to CCR5-Δ32 and CCR2-V64I, other SNPs and haplotypes may be important genetic determinants of disease and should be investigated. PMID:24011637

Defining the chemokine basis for leukocyte recruitment during viral encephalitis.

PubMed

Michlmayr, Daniela; McKimmie, Clive S; Pingen, Marieke; Haxton, Ben; Mansfield, Karen; Johnson, Nicholas; Fooks, Anthony R; Graham, Gerard J

2014-09-01

The encephalitic response to viral infection requires local chemokine production and the ensuing recruitment of immune and inflammatory leukocytes. Accordingly, chemokine receptors present themselves as plausible therapeutic targets for drugs aimed at limiting encephalitic responses. However, it remains unclear which chemokines are central to this process and whether leukocyte recruitment is important for limiting viral proliferation and survival in the brain or whether it is predominantly a driver of coincident inflammatory pathogenesis. Here we examine chemokine expression and leukocyte recruitment in the context of avirulent and virulent Semliki Forest virus (SFV) as well as West Nile virus infection and demonstrate rapid and robust expression of a variety of inflammatory CC and CXC chemokines in all models. On this basis, we define a chemokine axis involved in leukocyte recruitment to the encephalitic brain during SFV infection. CXCR3 is the most active; CCR2 is also active but less so, and CCR5 plays only a modest role in leukocyte recruitment. Importantly, inhibition of each of these receptors individually and the resulting suppression of leukocyte recruitment to the infected brain have no effect on viral titer or survival following infection with a virulent SFV strain. In contrast, simultaneous blockade of CXCR3 and CCR2 results in significantly reduced mortality in response to virulent SFV infection. In summary, therefore, our data provide an unprecedented level of insight into chemokine orchestration of leukocyte recruitment in viral encephalitis. Our data also highlight CXCR3 and CCR2 as possible therapeutic targets for limiting inflammatory damage in response to viral infection of the brain. Brain inflammation (encephalitis) in response to viral infection can lead to severe illness and even death. This therefore represents an important clinical problem and one that requires the development of new therapeutic approaches. Central to the pathogenesis of encephalitis is the recruitment of inflammatory leukocytes to the infected brain, a process driven by members of the chemokine family. Here we provide an in-depth analysis of the chemokines involved in leukocyte recruitment to the virally infected brain and demonstrate that simultaneous blockade of two of these receptors, namely, CXCR3 and CCR2, does not alter viral titers within the brain but markedly reduces inflammatory leukocyte recruitment and enhances survival in a murine model of lethal viral encephalitis. Our results therefore highlight chemokine receptors as plausible therapeutic targets in treating viral encephalitis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Rhesus macaque model of chronic opiate dependence and neuro-AIDS: longitudinal assessment of auditory brainstem responses and visual evoked potentials

PubMed Central

Riazi, Mariam; Marcario, Joanne K; Samson, Frank K.; Kenjale, Himanshu; Adany, Istvan; Staggs, Vincent; Ledford, Emily; Marquis, Janet; Narayan, Opendra; Cheney, Paul D.

2013-01-01

Our work characterizes the effects of opiate (morphine) dependence on auditory brainstem and visual evoked responses in a rhesus macaque model of neuro-AIDS utilizing a chronic continuous drug delivery paradigm. The goal of this study was to clarify whether morphine is protective, or if it exacerbates simian immunodeficiency virus (SIV) related systemic and neurological disease. Our model employs a macrophage tropic CD4/CCR5 co-receptor virus, SIVmac239 (R71/E17), which crosses the blood brain barrier shortly after inoculation and closely mimics the natural disease course of human immunodeficiency virus (HIV) infection. The cohort was divided into 3 groups: morphine only, SIV only, and SIV + morphine. Evoked potential (EP) abnormalities in sub-clinically infected macaques were evident as early as eight weeks post-inoculation. Prolongations in EP latencies were observed in SIV-infected macaques across all modalities. Animals with the highest CSF viral loads and clinical disease showed more abnormalities than those with sub-clinical disease, confirming our previous work (Raymond et al, 1998, 1999, 2000). Although some differences were observed in auditory and visual evoked potentials in morphine treated compared to untreated SIV-infected animals, the effects were relatively small and not consistent across evoked potential type. However, morphine treated animals with subclinical disease had a clear tendency toward higher virus loads in peripheral and CNS tissues (Marcario et al., 2008) suggesting that if had been possible to follow all animals to end-stage disease, a clearer pattern of evoked potential abnormality might have emerged. PMID:19283490

Macrophage-tropic variants initiate human immunodeficiency virus type 1 infection after sexual, parenteral, and vertical transmission.

PubMed Central

van't Wout, A B; Kootstra, N A; Mulder-Kampinga, G A; Albrecht-van Lent, N; Scherpbier, H J; Veenstra, J; Boer, K; Coutinho, R A; Miedema, F; Schuitemaker, H

1994-01-01

Macrophage-tropic, non-syncytium-inducing, HIV-1 variants predominate in the asymptomatic phase of infection and may be responsible for establishing infection in an individual exposed to the mixture of HIV-1 variants. Here, genotypical and phenotypical characteristics of virus populations, present in sexual, parenteral, or vertical donor-recipient pairs, were studied. Sequence analysis of the V3 domain confirmed the presence of a homogeneous virus population in recently infected individuals. Biological HIV-1 clones were further characterized for syncytium inducing capacity on the MT2 cell line and for macrophage tropism as defined by the appearance of proviral DNA upon inoculation of monocyte-derived macrophages. Both sexual and parenteral transmission cases revealed a selective outgrowth in the recipient of the most macrophage-tropic variant(s) present in the donor. In three out of five vertical transmission cases, more than one highly macrophage-tropic virus variant was present in the child shortly after birth, suggestive of transmission of multiple variants. In three primary infection cases, homogeneous virus populations of macrophage-tropic, non-syncytium-inducing variants were present prior to seroconversion, thus excluding humoral immunity as the selective pressure in favour of macrophage-tropic variants. These observations may have important implications for vaccine development. PMID:7962552

Efficient modification of CCR5 in primary human hematopoietic cells using a megaTAL nuclease and AAV donor template.

PubMed

Sather, Blythe D; Romano Ibarra, Guillermo S; Sommer, Karen; Curinga, Gabrielle; Hale, Malika; Khan, Iram F; Singh, Swati; Song, Yumei; Gwiazda, Kamila; Sahni, Jaya; Jarjour, Jordan; Astrakhan, Alexander; Wagner, Thor A; Scharenberg, Andrew M; Rawlings, David J

2015-09-30

Genetic mutations or engineered nucleases that disrupt the HIV co-receptor CCR5 block HIV infection of CD4(+) T cells. These findings have motivated the engineering of CCR5-specific nucleases for application as HIV therapies. The efficacy of this approach relies on efficient biallelic disruption of CCR5, and the ability to efficiently target sequences that confer HIV resistance to the CCR5 locus has the potential to further improve clinical outcomes. We used RNA-based nuclease expression paired with adeno-associated virus (AAV)-mediated delivery of a CCR5-targeting donor template to achieve highly efficient targeted recombination in primary human T cells. This method consistently achieved 8 to 60% rates of homology-directed recombination into the CCR5 locus in T cells, with over 80% of cells modified with an MND-GFP expression cassette exhibiting biallelic modification. MND-GFP-modified T cells maintained a diverse repertoire and engrafted in immune-deficient mice as efficiently as unmodified cells. Using this method, we integrated sequences coding chimeric antigen receptors (CARs) into the CCR5 locus, and the resulting targeted CAR T cells exhibited antitumor or anti-HIV activity. Alternatively, we introduced the C46 HIV fusion inhibitor, generating T cell populations with high rates of biallelic CCR5 disruption paired with potential protection from HIV with CXCR4 co-receptor tropism. Finally, this protocol was applied to adult human mobilized CD34(+) cells, resulting in 15 to 20% homologous gene targeting. Our results demonstrate that high-efficiency targeted integration is feasible in primary human hematopoietic cells and highlight the potential of gene editing to engineer T cell products with myriad functional properties. Copyright © 2015, American Association for the Advancement of Science.

Incomplete APOBEC3G/F Neutralization by HIV-1 Vif Mutants Facilitates the Genetic Evolution from CCR5 to CXCR4 Usage

PubMed Central

Alteri, Claudia; Surdo, Matteo; Bellocchi, Maria Concetta; Saccomandi, Patrizia; Continenza, Fabio; Armenia, Daniele; Parrotta, Lucia; Carioti, Luca; Costa, Giosuè; Fourati, Slim; Di Santo, Fabiola; Scutari, Rossana; Barbaliscia, Silvia; Fedele, Valentina; Carta, Stefania; Balestra, Emanuela; Alcaro, Stefano; Marcelin, Anne Genevieve; Calvez, Vincent; Ceccherini-Silberstein, Francesca; Artese, Anna

2015-01-01

Incomplete APOBEC3G/F neutralization by a defective HIV-1Vif protein can promote genetic diversification by inducing G-to-A mutations in the HIV-1 genome. The HIV-1 Env V3 loop, critical for coreceptor usage, contains several putative APOBEC3G/F target sites. Here, we determined if APOBEC3G/F, in the presence of Vif-defective HIV-1 virus, can induce G-to-A mutations at V3 positions critical to modulation of CXCR4 usage. Peripheral blood mononuclear cells (PBMC) and monocyte-derived macrophages (MDM) from 2 HIV-1-negative donors were infected with CCR5-using 81.A-VifWT virus (i.e., with wild-type [WT] Vif protein), 81.A-VifE45G, or 81.A-VifK22E (known to incompletely/partially neutralize APOBEC3G/F). The rate of G-toA mutations was zero or extremely low in 81.A-VifWT- and 81.A-VifE45G-infected PBMC from both donors. Conversely, G-to-A enrichment was detected in 81.A-VifK22E-infected PBMC (prevalence ranging from 2.18% at 7 days postinfection [dpi] to 3.07% at 21 dpi in donor 1 and from 10.49% at 7 dpi to 8.69% at 21 dpi in donor 2). A similar scenario was found in MDM. G-to-A mutations occurred at 8 V3 positions, resulting in nonsynonymous amino acid substitutions. Of them, G24E and E25K strongly correlated with phenotypically/genotypically defined CXCR4-using viruses (P = 0.04 and 5.5e−7, respectively) and increased the CXCR4 N-terminal binding affinity for V3 (WT, −40.1 kcal/mol; G24E, −510 kcal/mol; E25K, −522 kcal/mol). The analysis of paired V3 and Vif DNA sequences from 84 HIV-1-infected patients showed that the presence of a Vif-defective virus correlated with CXCR4 usage in proviral DNA (P = 0.04). In conclusion, incomplete APOBEC3G/F neutralization by a single Vif amino acid substitution seeds a CXCR4-using proviral reservoir. This can have implications for the success of CCR5 antagonist-based therapy, as well as for the risk of disease progression. PMID:26055363

CCR5 polymorphism and plague resistance in natural populations of the black rat in Madagascar.

PubMed

Tollenaere, C; Rahalison, L; Ranjalahy, M; Rahelinirina, S; Duplantier, J-M; Brouat, C

2008-12-01

Madagascar remains one of the world's largest plague foci. The black rat, Rattus rattus, is the main reservoir of plague in rural areas. This species is highly susceptible to plague in plague-free areas (low-altitude regions), whereas rats from the plague focus areas (central highlands) have evolved a disease-resistance polymorphism. We used the candidate gene CCR5 to investigate the genetic basis of plague resistance in R. rattus. We found a unique non-synonymous substitution (H184R) in a functionally important region of the gene. We then compared (i) CCR5 genotypes of dying and surviving plague-challenged rats and (ii) CCR5 allelic frequencies in plague focus and plague-free populations. Our results suggested a higher prevalence of the substitution in resistant animals compared to susceptible individuals, and a tendency for higher frequencies in plague focus areas compared to plague-free areas. Therefore, the CCR5 polymorphism may be involved in Malagasy black rat plague resistance. CCR5 and other undetermined plague resistance markers may provide useful biological information about host evolution and disease dynamics.

48 CFR 4.605 - Procedures.

Code of Federal Regulations, 2010 CFR

2010-10-01

... unique identifiers on the FPDS website, and must validate their use in all transactions. The PIID shall... Registration (CCR) database in accordance with the clause at 52.204-7, Central Contractor Registration. The...)(2) of this section, the contracting officer shall use one of the generic DUNS numbers identified in...

The CCL3L1-CCR5 genotype influences the development of AIDS, but not HIV susceptibility or the response to HAART

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhattacharya, Tanmoy; Stanton, Jennifer; Kim, Eun - Young

2008-01-01

A selective advantage against infectious diseases such as HIV/AIDS is associated with differences in the genes relevant to immunity and virus replication. The CC chemokine receptor 5 (CCR5), the principal coreceptor for HIV, and its chemokine ligands, including CCL3L1, influences the CD4+ target cells susceptibility to infection. The CCL3L1 gene is in a region of segmental duplication on the q-arm of human chromosome 17. Increased numbers of CCL3L1 gene copies that affect the gene expression phenotype might have substantial protective effects. Here we show that the population-specific CCL3L1 gene copy number and the CCR5 {Delta}32 protein-inactivating deletion that categorizes themore » CCL3L1-CCR5 genotype do not influence HIV/AIDS susceptibility or the robustness of immune recovery after the initiation of highly active antiretroviral therapy (HAART).« less

Week 96 results of the randomized, multicentre Maraviroc Switch (MARCH) study.

PubMed

Pett, S L; Amin, J; Horban, A; Andrade-Villanueva, J; Losso, M; Porteiro, N; Madero, J S; Belloso, W; Tu, E; Silk, D; Kelleher, A; Harrigan, R; Clark, A; Sugiura, W; Wolff, M; Gill, J; Gatell, J; Clarke, A; Ruxrungtham, K; Prazuck, T; Kaiser, R; Woolley, I; Alberto Arnaiz, J; Cooper, D; Rockstroh, J K; Mallon, P; Emery, S

2018-01-01

The Maraviroc Switch (MARCH) study week 48 data demonstrated that maraviroc, a chemokine receptor-5 (CCR5) inhibitor, was a safe and effective switch for the ritonavir-boosted protease inhibitor (PI/r) component of a two nucleos(t)ide reverse transcriptase inhibitor [N(t)RTI] plus PI/r-based antiretroviral regimen in patients with R5-tropic virus. Here we report the durability of this finding. MARCH, an international, multicentre, randomized, 96-week open-label switch study, enrolled HIV-1-infected adults with R5-tropic virus who were stable (> 24 weeks) and virologically suppressed [plasma viral load (pVL) < 50 HIV-1 RNA copies/mL]. Participants were randomized to continue their current PI/r-based regimen (PI/r) or to switch to MVC plus two N(t)RTIs (MVC) (1:2 randomization). The primary endpoint was the difference in the proportion with pVL < 200 copies/mL at 96 weeks. The switch arm was defined as noninferior if the lower limit of the 95% confidence interval (CI) for the difference was < -12% in the intention-to-treat (ITT) population. Safety endpoints (the difference in the mean change from baseline or a comparison of proportions) were analysed as key secondary endpoints. Eighty-two (PI/r) and 156 (MVC) participants were randomized and included in the ITT analysis; 71 (87%) and 130 (83%) were in follow-up and on therapy at week 96. At week 96, 89.0% and 90.4% in the PI/r and MVC arms, respectively, had pVL < 50 copies/mL (95% CI -6.6, 10.2). Moreover, in those switching away from PI/r, there were significant reductions in mean total cholesterol (differences 0.31 mmol/L; P = 0.02) and triglycerides (difference 0.44 mmol/L; P < 0.001). Changes in CD4 T-cell count, renal function, and serious and nonserious adverse events were similar in the two arms. MVC as a switch for a PI/r is safe and effective at maintaining virological suppression while having significant lipid benefits over 96 weeks. © 2017 British HIV Association.

Investigation of Inhibition Mechanism of Chemokine Receptor CCR5 by Micro-second Molecular Dynamics Simulations.

PubMed

Salmas, Ramin Ekhteiari; Yurtsever, Mine; Durdagi, Serdar

2015-08-24

Chemokine receptor 5 (CCR5) belongs to G protein coupled receptors (GPCRs) and plays an important role in treatment of human immunodeficiency virus (HIV) infection since HIV uses CCR5 protein as a co-receptor. Recently, the crystal structure of CCR5-bound complex with an approved anti-retroviral drug (maroviroc) was resolved. During the crystallization procedure, amino acid residues (i.e., Cys224, Arg225, Asn226 and Glu227) at the third intra-cellular loop were replaced by the rubredoxin for stability reasons. In the current study, we aimed to understand the impact of the incorporated rubredoxin on the conformations of TM domains of the target protein. For this reason, rubredoxin was deleted from the crystal structure and the missing amino acids were engineered. The resultant structure was subjected to long (μs) molecular dynamics (MD) simulations to shed light into the inhibitory mechanism. The derived model structure displayed a significant deviation in the cytoplasmic domain of TM5 and IC3 in the absence of rubredoxin. The principal component analyses (PCA) and MD trajectory analyses revealed important structural and dynamical differences at apo and holo forms of the CCR5.

Silencing CCR2 in Macrophages Alleviates Adipose Tissue Inflammation and the Associated Metabolic Syndrome in Dietary Obese Mice.

PubMed

Kim, Jongkil; Chung, Kunho; Choi, Changseon; Beloor, Jagadish; Ullah, Irfan; Kim, Nahyeon; Lee, Kuen Yong; Lee, Sang-Kyung; Kumar, Priti

2016-01-26

Adipose tissue macrophage (ATM)-mediated inflammation is a key feature contributing to the adverse metabolic outcomes of dietary obesity. Recruitment of macrophages to obese adipose tissues (AT) can occur through the engagement of CCR2, the receptor for MCP-1 (monocyte chemoattractant protein-1), which is expressed on peripheral monocytes/macrophages. Here, we show that i.p. administration of a rabies virus glycoprotein-derived acetylcholine receptor-binding peptide effectively delivers complexed siRNA into peritoneal macrophages and ATMs in a mouse model of high-fat diet-induced obesity. Treatment with siRNA against CCR2 inhibited macrophage infiltration and accumulation in AT and, therefore, proinflammatory cytokines produced by macrophages. Consequently, the treatment significantly improved glucose tolerance and insulin sensitivity profiles, and also alleviated the associated symptoms of hepatic steatosis and reduced hepatic triglyceride production. These results demonstrate that disruption of macrophage chemotaxis to the AT through cell-targeted gene knockdown strategies can provide a therapeutic intervention for obesity-related metabolic diseases. The study also highlights a siRNA delivery approach for targeting specific monocyte subsets that contribute to obesity-associated inflammation without affecting the function of other tissue-resident macrophages that are essential for host homeostasis and survival.

Structural map of Kaposi sarcoma-associated herpesvirus RNA provides clues to molecular interactions | Center for Cancer Research

Cancer.gov

Scientists from CCR have generated a comprehensive structural map of Kaposi sarcoma-associated herpesvirus polyadenylated nuclear (PAN) RNA, a long non-coding RNA that helps the virus evade detection by its host’s immune system. The findings open new oppportunites to study the life cycle of this cancer-causing virus.  Learn more...

A rare subset of skin-tropic regulatory T cells expressing Il10/Gzmb inhibits the cutaneous immune response.

PubMed

Ikebuchi, Ryoyo; Teraguchi, Shunsuke; Vandenbon, Alexis; Honda, Tetsuya; Shand, Francis H W; Nakanishi, Yasutaka; Watanabe, Takeshi; Tomura, Michio

2016-10-19

Foxp3 + regulatory T cells (Tregs) migrating from the skin to the draining lymph node (dLN) have a strong immunosuppressive effect on the cutaneous immune response. However, the subpopulations responsible for their inhibitory function remain unclear. We investigated single-cell gene expression heterogeneity in Tregs from the dLN of inflamed skin in a contact hypersensitivity model. The immunosuppressive genes Ctla4 and Tgfb1 were expressed in the majority of Tregs. Although Il10-expressing Tregs were rare, unexpectedly, the majority of Il10-expressing Tregs co-expressed Gzmb and displayed Th1-skewing. Single-cell profiling revealed that CD43 + CCR5 + Tregs represented the main subset within the Il10/Gzmb-expressing cell population in the dLN. Moreover, CD43 + CCR5 + CXCR3 - Tregs expressed skin-tropic chemokine receptors, were preferentially retained in inflamed skin and downregulated the cutaneous immune response. The identification of a rare Treg subset co-expressing multiple immunosuppressive molecules and having tissue-remaining capacity offers a novel strategy for the control of skin inflammatory responses.

Mapping global environmental suitability for Zika virus

PubMed Central

Messina, Jane P; Kraemer, Moritz UG; Brady, Oliver J; Pigott, David M; Shearer, Freya M; Weiss, Daniel J; Golding, Nick; Ruktanonchai, Corrine W; Gething, Peter W; Cohn, Emily; Brownstein, John S; Khan, Kamran; Tatem, Andrew J; Jaenisch, Thomas; Murray, Christopher JL; Marinho, Fatima; Scott, Thomas W; Hay, Simon I

2016-01-01

Zika virus was discovered in Uganda in 1947 and is transmitted by Aedes mosquitoes, which also act as vectors for dengue and chikungunya viruses throughout much of the tropical world. In 2007, an outbreak in the Federated States of Micronesia sparked public health concern. In 2013, the virus began to spread across other parts of Oceania and in 2015, a large outbreak in Latin America began in Brazil. Possible associations with microcephaly and Guillain-Barré syndrome observed in this outbreak have raised concerns about continued global spread of Zika virus, prompting its declaration as a Public Health Emergency of International Concern by the World Health Organization. We conducted species distribution modelling to map environmental suitability for Zika. We show a large portion of tropical and sub-tropical regions globally have suitable environmental conditions with over 2.17 billion people inhabiting these areas. DOI: http://dx.doi.org/10.7554/eLife.15272.001 PMID:27090089

Influence of CCR5 and CCR2 Genetic Variants in the Resistance/Susceptibility to HIV in Serodiscordant Couples from Colombia

PubMed Central

Zapata, Wildeman; Aguilar-Jiménez, Wbeimar; Pineda-Trujillo, Nicolás; Rojas, Winston; Estrada, Hernando

2013-01-01

Abstract The main genetic factor related to HIV-1 resistance is the CCR5-Δ32 mutation; however, the homozygous genotype is uncommon. The CCR5-Δ32 mutation along with single nucleotide polymorphisms (SNPs) in the CCR5 promoter and the CCR2-V64I mutation have been included in seven human haplogroups (HH) previously associated with resistance/susceptibility to HIV-1 infection and different rates of AIDS progression. Here, we determined the association of the CCR5 promoter SNPs, the CCR5-Δ32 mutation, CCR2-V64I SNP, and HH frequencies with resistance/susceptibility to HIV-1 infection in a cohort of HIV-1-serodiscordant couples from Colombia. Seventy HIV-1-exposed, but seronegative (HESN) individuals, 57 seropositives (SP), and 112 healthy controls (HC) were included. The CCR5-Δ32 mutation and CCR2-V64I SNP were identified by PCR, and the CCR5 promoter SNPs were evaluated by sequencing. None of the individuals exhibited a homozygous Δ32 genotype; the CCR2-I allele was more frequent in HESN (34%) than HC (23%) (p=0.039, OR=1.672). The frequency of the 29G allele was higher in SP than HC (p=0.003, OR=3). HHF2 showed a higher frequency in HC (19%) than SP (9%) (p=0.027), while HHG1 was more frequent in SP (11.1%) than in HC (4.2%) (p=0.019). The AGACCAC-CCR2-I-CCR5 wild-type haplotype showed a higher frequency in SP (14.2%) than in HC (3.7%) (p=0.001). In conclusion, the CCR5-Δ32 allele is not responsible for HIV-1 resistance in this HESN group; however, the CCR2-I allele could be protective, while the 29G allele might increase the likelihood of acquiring HIV-1 infection. HHG1 and the AGACCAC-CCR2-I-CCR5 wild-type haplotype might promote HIV-1 infection while HHF2 might be related to resistance. However, additional studies are required to evaluate the implications of these findings. PMID:24098976

Adult T-cell leukemia: molecular basis for clonal expansion and transformation of HTLV-1-infected T cells.

PubMed

Watanabe, Toshiki

2017-03-02

Adult T-cell leukemia (ATL) is an aggressive T-cell malignancy caused by human T-cell leukemia virus type 1 (HTLV-1) that develops through a multistep carcinogenesis process involving 5 or more genetic events. We provide a comprehensive overview of recently uncovered information on the molecular basis of leukemogenesis in ATL. Broadly, the landscape of genetic abnormalities in ATL that include alterations highly enriched in genes for T-cell receptor-NF-κB signaling such as PLCG1 , PRKCB , and CARD11 and gain-of function mutations in CCR4 and CCR7 Conversely, the epigenetic landscape of ATL can be summarized as polycomb repressive complex 2 hyperactivation with genome-wide H3K27 me3 accumulation as the basis of the unique transcriptome of ATL cells. Expression of H3K27 methyltransferase enhancer of zeste 2 was shown to be induced by HTLV-1 Tax and NF-κB. Furthermore, provirus integration site analysis with high-throughput sequencing enabled the analysis of clonal composition and cell number of each clone in vivo, whereas multicolor flow cytometric analysis with CD7 and cell adhesion molecule 1 enabled the identification of HTLV-1-infected CD4 + T cells in vivo. Sorted immortalized but untransformed cells displayed epigenetic changes closely overlapping those observed in terminally transformed ATL cells, suggesting that epigenetic abnormalities are likely earlier events in leukemogenesis. These new findings broaden the scope of conceptualization of the molecular mechanisms of leukemogenesis, dissecting them into immortalization and clonal progression. These recent findings also open a new direction of drug development for ATL prevention and treatment because epigenetic marks can be reprogrammed. Mechanisms underlying initial immortalization and progressive accumulation of these abnormalities remain to be elucidated. © 2017 by The American Society of Hematology.

Type 1 Responses of Human Vγ9Vδ2 T Cells to Influenza A Viruses▿

PubMed Central

Qin, Gang; Liu, Yinping; Zheng, Jian; Ng, Iris H. Y.; Xiang, Zheng; Lam, Kwok-Tai; Mao, Huawei; Li, Hong; Peiris, J. S. Malik; Lau, Yu-Lung; Tu, Wenwei

2011-01-01

γδ T cells are essential constituents of antimicrobial and antitumor defenses. We have recently reported that phosphoantigen isopentenyl pyrophosphate (IPP)-expanded human Vγ9Vδ2 T cells participated in anti-influenza virus immunity by efficiently killing both human and avian influenza virus-infected monocyte-derived macrophages (MDMs) in vitro. However, little is known about the noncytolytic responses and trafficking program of γδ T cells to influenza virus. In this study, we found that Vγ9Vδ2 T cells expressed both type 1 cytokines and chemokine receptors during influenza virus infection, and IPP-expanded cells had a higher capacity to produce gamma interferon (IFN-γ). Besides their potent cytolytic activity against pandemic H1N1 virus-infected cells, IPP-activated γδ T cells also had noncytolytic inhibitory effects on seasonal and pandemic H1N1 viruses via IFN-γ but had no such effects on avian H5N1 or H9N2 virus. Avian H5N1 and H9N2 viruses induced significantly higher CCL3, CCL4, and CCL5 production in Vγ9Vδ2 T cells than human seasonal H1N1 virus. CCR5 mediated the migration of Vγ9Vδ2 T cells toward influenza virus-infected cells. Our findings suggest a novel therapeutic strategy of using phosphoantigens to boost the antiviral activities of human Vγ9Vδ2 T cells against influenza virus infection. PMID:21752902

Cross-Desensitization of CCR1, But Not CCR2, Following Activation of the Formyl Peptide Receptor (FPR1)1

PubMed Central

Bednar, Filip; Song, Changcheng; Bardi, Giuseppe; Cornwell, William; Rogers, Thomas J.

2014-01-01

The cross-regulation of G protein-coupled receptors (GPCRs) plays an important role in the immune response. Studies from several laboratories have suggested that a hierarchy of sensitivities to cross-desensitization exists for the chemoattractant GPCRs. We carried out experiments to study the capacity of the formyl peptide receptor-1 (FPR1) to desensitize chemokine receptors CCR1 and CCR2. Our results show that activation of FPR1 resulted in the desensitization and partial internalization of CCR1, but not CCR2, in both primary human monocytes and HEK293 cells co-expressing CCR1, CCR2, and FPR1 (HR1R2F cells). The desensitization of CCR1 by FPR1 stimulation was not due to the simple depletion of the Ca2+ stores, but was dependent on activation of PKC. Furthermore, we found that the cross-desensitization of CCR1 by FPR1 was associated with CCR1 phosphorylation, and moderate reduction of CCR1 cell surface expression. In contrast, CCR2 was not phosphorylated or internalized following FPR1 activation. Additional studies showed that optimal cross-talk between FPR1 and CCR1 were dependent on the functional activity of PKCβ. These results provide a mechanistic basis for the capacity of certain GPCR ligands to exert rapid and selective cross-inactivation of other chemoattractant receptors, and suggest that FPR1 is able to exert “traffic control” in the migration of inflammatory cells by rapidly inhibiting the cell responses to potentially “low priority” chemoattractants such as CCR1 agonists without inhibiting the response to “higher priority” CCR2 chemoattractants. PMID:24778447

Natural Killer Cell Recruitment to the Lung During Influenza A Virus Infection Is Dependent on CXCR3, CCR5, and Virus Exposure Dose

PubMed Central

Carlin, Lindsey E.; Hemann, Emily A.; Zacharias, Zeb R.; Heusel, Jonathan W.; Legge, Kevin L.

2018-01-01

Natural killer (NK) cells are vital components of the antiviral immune response, but their contributions in defense against influenza A virus (IAV) are not well understood. To better understand NK cell responses during IAV infections, we examined the magnitude, kinetics, and contribution of NK cells to immunity and protection during high- and low-dose IAV infections. Herein, we demonstrate an increased accumulation of NK cells in the lung in high-dose vs. low-dose infections. In part, this increase is due to the local proliferation of pulmonary NK cells. However, the majority of NK cell accumulation within the lungs and airways during an IAV infection is due to recruitment that is partially dependent upon CXCR3 and CCR5, respectively. Therefore, altogether, our results demonstrate that NK cells are actively recruited to the lungs and airways during IAV infection and that the magnitude of the recruitment may relate to the inflammatory environment found within the tissues during high- and low-dose IAV infections. PMID:29719539

Visualizing High-Efficiency HIV Transfer | Center for Cancer Research

Cancer.gov

The Human Immunodeficiency Virus (HIV), the causative agent of Acquired Immunodeficiency Syndrome (AIDS), infects and eventually kills CD4 receptor-expressing T cells, which are critical for proper immune system function. The gp120 protein on the surface of HIV particles is known to bind CD4 and a co-receptor, either CCR5 or CXCR4, leading to fusion of the virus and T cell

Potential cancer immunotherapy drug shows promise against HIV | Center for Cancer Research

Cancer.gov

An immunotherapy currently being tested in a clinical trial as a treatment for metastatic cancer has now shown potential in an animal study to reduce recalcitrant pools of SHIV, a laboratory-designed virus used to study human immunodeficiency virus (HIV) infection. The study, published online February 23 in PLoS Pathogens, is the result of a collaborative effort by CCR, the

CCR-2 neutralization augments murine fresh BMC activation by Staphylococcus aureus via two distinct mechanisms: at the level of ROS production and cytokine response.

PubMed

Nandi, Ajeya; Bishayi, Biswadev

2017-05-01

CCR-2 signaling regulates recruitment of monocytes from the bone marrow into the bloodstream and then to sites of infection. We sought to determine whether CCL-2/CCR-2 signaling is involved in the killing of Staphylococcus aureus by murine bone marrow cells (BMCs). The intermittent link of reactive oxygen species (ROS)-NF-κB/p38-MAPK-mediated CCL-2 production in CCR-2 signaling prompted us to determine whether neutralization of CCR-2 augments the response of murine fresh BMCs (FBMCs) after S. aureus infection. It was observed that anti-CCR-2 Ab-treated FBMCs released fewer ROS on encountering S. aureus infection than CCR-2 non-neutralized FBMCs, also correlating with reduced killing of S. aureus in CCR-2 neutralized FBMCs. Staphylococcal catalase and SOD were also found to play a role in protecting S. aureus from the ROS-mediated killing of FBMC. S. aureus infection of CCR-2 intact FBMCs pre-treated with either NF-κB or p-38-MAPK blocker induced less CCL-2, suggesting that NF-κB or p-38-MAPK is required for CCL-2 production by FBMCs. Moreover, blocking of CCR-2 along with NF-κB or p-38-MAPK resulted in elevated CCL-2 production and reduced CCR-2 expression. Inhibition of CCR-2 impairs the response of murine BMCs to S. aureus infection by attenuation ROS production and modulating the cytokine response.

Podocyte-specific chemokine (C-C motif) receptor 2 overexpression mediates diabetic renal injury in mice

PubMed Central

You, Hanning; Gao, Ting; Raup-Konsavage, Wesley M.; Cooper, Timothy K.; Bronson, Sarah K.; Reeves, W. Brian; Awad, Alaa S.

2016-01-01

Inflammation is a central pathophysiologic mechanism that contributes to diabetes mellitus and diabetic nephropathy. Recently, we showed that macrophages directly contribute to diabetic renal injury, and that pharmacological blockade or genetic deficiency of chemokine (C-C motif) receptor 2 (CCR2) confers kidney protection in diabetic nephropathy. However, the direct role of CCR2 in kidney-derived cells such as podocytes in diabetic nephropathy remains unclear. To study this, we developed a transgenic mouse model expressing CCR2 specifically in podocytes (Tg(NPHS2-Ccr2)) on a nephropathy prone (DBA/2J) and CCR2 deficient (Ccr2−/−) background with heterozygous Ccr2+/− littermate controls. Diabetes was induced by streptozotocin. As expected, absence of CCR2 conferred kidney protection after nine weeks of diabetes. In contrast, transgenic CCR2 over expression in the podocytes of Ccr2−/− mice resulted in significantly increased albuminuria, blood urea nitrogen, histopathologic changes, kidney fibronectin and type-1 collagen expression, podocyte loss, and glomerular apoptosis after nine weeks of streptozotocin-induced diabetes. Interestingly, there was no concurrent increase in kidney macrophage recruitment or inflammatory cytokine levels in the mice. These findings support a direct role for CCR2 expression in podocytes to mediate diabetic renal injury, independent of monocyte/macrophage recruitment. Thus, targeting the CCR2 signaling cascade in podocytes could be a novel therapeutic approach for treatment of diabetic nephropathy. PMID:27914709

Analysis of Social and Genetic Factors Influencing Heterosexual Transmission of HIV within Serodiscordant Couples in the Henan Cohort

PubMed Central

Zhang, Yilei; Li, Jie; Ma, Xuejun; Li, Ning; Wang, Qi; Xue, Xiujuan; Luo, Le; Li, Zizhao; Ring, Huijun Z.; Ring, Brian Z.; Su, Li

2015-01-01

There is considerable variability between individuals in susceptibility to infection by human immunodeficiency virus (HIV). Many social, clinical and genetic factors are known to contribute to the likelihood of HIV transmission, but there is little consensus on the relative importance and potential interaction of these factors. Additionally, recent studies of several variants in chemokine receptors have identified alleles that may be predictive of HIV transmission and disease progression; however the strengths and directions of the associations of these genetic markers with HIV transmission have markedly varied between studies. To better identify factors that predict HIV transmission in a Chinese population, 180 cohabiting serodiscordant couples were enrolled for study by the Henan Center for Disease Prevention and Control, and transmission and progression of HIV infection were regularly measured. We found that anti-retroviral therapy, education level, and condom use were the most significant factors in determining likelihood of HIV transmission in this study. We also assessed ten variants in three genes (CXCL12, CCR2, and CCR5) that have been shown to influence HIV transmission. We found two tightly linked variants in CCR2 and CCR5, rs1799864 and rs1800024, have a significant positive association with transmission as recessive models (OR>10, P value=0.011). Mixed effects models showed that these genetic variants both retained significance when assessed with either treatment or condom use. These markers of transmission susceptibility may therefore serve to help stratify individuals by risk for HIV transmission. PMID:26068906

New respiratory virus (chicken pox, influenza and respiratory syncytial virus) vaccines: efficacy, necessity and policy for the tropical world at present.

PubMed

Wiwanitkit, Viroj

2009-09-01

Several respiratory viruses are documented in medicine. Several infectious diseases due to these viruses are current global public health problems. Prevention of respiratory viral infections becomes the focus of the public health ministries of many tropical countries. Presently, there are many new vaccines for respiratory viruses. These vaccines include chicken pox vaccine, influenza vaccine and respiratory syncytial virus vaccine. In this article, the author will briefly discuss on these quoted vaccines focusing on efficacy, necessity and policy for tropical world at present.

Genetic diversity and prevalence of CCR2-CCR5 gene polymorphisms in the Omani population

PubMed Central

Al-Mahruqi, Samira H.; Zadjali, Fahad; Beja-Pereira, Albano; Koh, Crystal Y.; Balkhair, Abdullah; Al-Jabri, Ali A.

2014-01-01

Polymorphisms in the regulatory region of the CCR5 gene affect protein expression and modulate the progress of HIV-1 disease. Because of this prominent role, variations in this gene have been under differential pressure and their frequencies vary among human populations. The CCR2V64I mutation is tightly linked to certain polymorphisms in the CCR5 gene. The current Omani population is genetically diverse, a reflection of their history as traders who ruled extensive regions around the Indian Ocean. In this study, we examined the CCR2-CCR5 haplotypes in Omanis and compared the patterns of genetic diversity with those of other populations. Blood samples were collected from 115 Omani adults and genomic DNA was screened to identify the polymorphic sites in the CCR5 gene and the CCR2V64I mutation. Four minor alleles were common: CCR5-2554T and CCR5-2086G showed frequencies of 49% and 46%, respectively, whereas CCR5-2459A and CCR5-2135C both had a frequency of 36%. These alleles showed moderate levels of heterozygosity, indicating that they were under balancing selection. However, the well-known allele CCR5Δ32 was relatively rare. Eleven haplotypes were identified, four of which were common: HHC (46%), HHE (20%), HHA (14%) and HHF*2 (12%). PMID:24688285

Incomplete APOBEC3G/F Neutralization by HIV-1 Vif Mutants Facilitates the Genetic Evolution from CCR5 to CXCR4 Usage.

PubMed

Alteri, Claudia; Surdo, Matteo; Bellocchi, Maria Concetta; Saccomandi, Patrizia; Continenza, Fabio; Armenia, Daniele; Parrotta, Lucia; Carioti, Luca; Costa, Giosuè; Fourati, Slim; Di Santo, Fabiola; Scutari, Rossana; Barbaliscia, Silvia; Fedele, Valentina; Carta, Stefania; Balestra, Emanuela; Alcaro, Stefano; Marcelin, Anne Genevieve; Calvez, Vincent; Ceccherini-Silberstein, Francesca; Artese, Anna; Perno, Carlo Federico; Svicher, Valentina

2015-08-01

Incomplete APOBEC3G/F neutralization by a defective HIV-1Vif protein can promote genetic diversification by inducing G-to-A mutations in the HIV-1 genome. The HIV-1 Env V3 loop, critical for coreceptor usage, contains several putative APOBEC3G/F target sites. Here, we determined if APOBEC3G/F, in the presence of Vif-defective HIV-1 virus, can induce G-to-A mutations at V3 positions critical to modulation of CXCR4 usage. Peripheral blood mononuclear cells (PBMC) and monocyte-derived macrophages (MDM) from 2 HIV-1-negative donors were infected with CCR5-using 81.A-VifWT virus (i.e., with wild-type [WT] Vif protein), 81.A-VifE45G, or 81.A-VifK22E (known to incompletely/partially neutralize APOBEC3G/F). The rate of G-toA mutations was zero or extremely low in 81.A-VifWT- and 81.A-VifE45G-infected PBMC from both donors. Conversely, G-to-A enrichment was detected in 81.A-VifK22E-infected PBMC (prevalence ranging from 2.18% at 7 days postinfection [dpi] to 3.07% at 21 dpi in donor 1 and from 10.49% at 7 dpi to 8.69% at 21 dpi in donor 2). A similar scenario was found in MDM. G-to-A mutations occurred at 8 V3 positions, resulting in nonsynonymous amino acid substitutions. Of them, G24E and E25K strongly correlated with phenotypically/genotypically defined CXCR4-using viruses (P = 0.04 and 5.5e-7, respectively) and increased the CXCR4 N-terminal binding affinity for V3 (WT, -40.1 kcal/mol; G24E, -510 kcal/mol; E25K, -522 kcal/mol). The analysis of paired V3 and Vif DNA sequences from 84 HIV-1-infected patients showed that the presence of a Vif-defective virus correlated with CXCR4 usage in proviral DNA (P = 0.04). In conclusion, incomplete APOBEC3G/F neutralization by a single Vif amino acid substitution seeds a CXCR4-using proviral reservoir. This can have implications for the success of CCR5 antagonist-based therapy, as well as for the risk of disease progression. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The complete nucleotide sequence and genomic characterization of tropical soda apple mosaic virus

USDA-ARS?s Scientific Manuscript database

Tropical soda apple mosaic virus (TSAMV) was first identified in tropical soda apple (Solanum viarum), a noxious weed, in Florida in 2002. This report provides the first full genome sequence of TSAMV. The full genome sequence of this virus will enable research scientists to develop additional spec...

Role of novel type I interferon epsilon in viral infection and mucosal immunity

PubMed Central

Xi, Yang; Day, Stephanie L; Jackson, Ronald J; Ranasinghe, Charani

2012-01-01

Intranasal infection with vaccinia virus co-expressing interferon epsilon (VV-HIV-IFN-ɛ) was used to evaluate the role of IFN-ɛ in mucosal immunity. VV-HIV- IFN-ɛ infection induced a rapid VV clearance in lung that correlated with (i) an elevated lung VV-specific CD8+CD107a+IFN-γ+ population expressing activation markers CD69/CD103, (ii) enhanced lymphocyte recruitment to lung alveoli with reduced inflammation, and (iii) an heightened functional/cytotoxic CD8+CD4+ T-cell subset (CD3hiCCR7hiCD62Llo) in lung lymph nodes. These responses were different to that observed with intranasal VV-HA-IFN-α4 or VV-HA-IFN-β infections. When IFN-ɛ was used in an intranasal/intramuscular heterologous HIV prime-boost immunization, elevated HIV-specific effector, but not memory CD8+T cells responses, were observed in spleen, genito-rectal nodes, and Peyer's patch. Homing marker α4β7 and CCR9 analysis indicated that unlike other type I IFNs, IFN-ɛ could promote migration of antigen-specific CD8+T cells to the gut. Our results indicate that IFN-ɛ has a unique role in the mucosae and most likely can be used to control local lung and/or gut infections (i.e., microbicide) such as tuberculosis, HIV-1, or sexually transmitted diseases. PMID:22617838

Proof of activity with AMD11070, an orally bioavailable inhibitor of CXCR4-tropic HIV type 1.

PubMed

Moyle, Graeme; DeJesus, Edwin; Boffito, Marta; Wong, Rebecca S; Gibney, Colleen; Badel, Karin; MacFarland, Ron; Calandra, Gary; Bridger, Gary; Becker, Stephen

2009-03-15

The X4 Antagonist Concept Trial investigates the safety and antiviral activity of AMD11070, a potent inhibitor of X4-tropic human immunodeficiency virus (HIV) in vitro in HIV-infected patients harboring X4-tropic virus. Patients enrolled in the study had an X4 virus population 2000 relative luminescence units (rlu; by the Monogram Trofile Assay) and an HIV-1 RNA level 5000 copies/mL. Patients received AMD11070 monotherapy for 10 days. Coreceptor tropism, plasma HIV-1 RNA level, and CD4 cell count were measured at study entry, on day 5, and on day 10. Daily predose and serial samples on the last day of treatment were obtained for determination of plasma AMD11070 concentration. Ten patients were given AMD11070 monotherapy (200 mg to 8 patients and 100 mg to 2 patients) twice daily for 10 days. The median baseline CD4 cell count was 160 cells/mm(3), and the median HIV-1 RNA level was 91,447 copies/mL. Four of 9 evaluable patients achieved a reduction in X4 virus population of >or= rlu. The median change in X4 virus population at the end of treatment was -0.22 log(10) rlu (range, -1.90 to 0.23 log(10) rlu). Three of 4 patients who responded to therapy showed a tropism shift from dual- or mixed-tropic viruses to exclusively R5 virus by day 10. There were no drug-related serious adverse events, adverse events of greater than grade 2, or laboratory abnormalities. These results demonstrate the activity of AMD11070, the first oral CXCR4 antagonist, against X4-tropic HIV-1. The drug was well tolerated, with no serious safety concerns. AMD11070 is on clinical hold because of histologic changes to the liver observed in long-term animal studies; additional preclinical safety assessments are pending.

HIV-1 Pathogenesis: The Virus

PubMed Central

Swanstrom, Ronald; Coffin, John

2012-01-01

Transmission of HIV-1 results in the establishment of a new infection, typically starting from a single virus particle. That virion replicates to generate viremia and persistent infection in all of the lymphoid tissue in the body. HIV-1 preferentially infects T cells with high levels of CD4 and those subsets of T cells that express CCR5, particularly memory T cells. Most of the replicating virus is in the lymphoid tissue, yet most of samples studied are from blood. For the most part the tissue and blood viruses represent a well-mixed population. With the onset of immunodeficiency, the virus evolves to infect new cell types. The tropism switch involves switching from using CCR5 to CXCR4 and corresponds to an expansion of infected cells to include naïve CD4+ T cells. Similarly, the virus evolves the ability to enter cells with low levels of CD4 on the surface and this potentiates the ability to infect macrophages, although the scope of sites where infection of macrophages occurs and the link to pathogenesis is only partly known and is clear only for infection of the central nervous system. A model linking viral evolution to these two pathways has been proposed. Finally, other disease states related to immunodeficiency may be the result of viral infection of additional tissues, although the evidence for a direct role for the virus is less strong. Advancing immunodeficiency creates an environment in which viral evolution results in viral variants that can target new cell types to generate yet another class of opportunistic infections (i.e., HIV-1 with altered tropism). PMID:23143844

Analysis of Proteins of Mouse Sarcoma Pseudotype Viruses: Type-Specific Radioimmunoassays for Ecotropic Virus p30's

PubMed Central

Kennel, Stephen J.; Tennant, Raymond W.

1979-01-01

Murine sarcoma virus pseudotypes were prepared by infection of nonproducer cells (A1-2), which were transformed by the Gazdar strain of mouse sarcoma virus, with Gross (N-tropic), WN1802B (B-tropic), or Moloney (NB-tropic) viruses. The respective host range pseudotype sarcoma viruses were defined by the titration characteristics on cells with the appropriate Fv-1 genotype. Proteins from virus progeny were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Bands present in both the 65,000- and the 10,000- to 20,000- molecular-weight regions of the gel distinguished the pseudotype viruses from their respective helpers. Furthermore, two protein bands were noted in the p30 region of murine sarcoma virus (Gross), one corresponding to Gross virus p30, and another of slightly slower mobility. However, since the mobility of the putative sarcoma p30 is nearly indentical to that of WN1802B, its presence could not be established by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Type-specific radioimmunoassays for Gross virus p30 and for WN1802B p30 were applied for analysis of pseudotype preparations, and among several ecotropic viruses tested, only the homologous virus scored in the respective assay. By use of these assays, pseudotype viruses were found to contain only 8 to 48% helper-specific p30's; the remainder is presumably derived from the sarcoma virus. Images PMID:90164

A role for MCP-1/CCR2 in interstitial lung disease in children

PubMed Central

Hartl, Dominik; Griese, Matthias; Nicolai, Thomas; Zissel, Gernot; Prell, Christine; Reinhardt, Dietrich; Schendel, Dolores J; Krauss-Etschmann, Susanne

2005-01-01

Background Interstitial lung diseases (ILD) are chronic inflammatory disorders leading to pulmonary fibrosis. Monocyte chemotactic protein 1 (MCP-1) promotes collagen synthesis and deletion of the MCP-1 receptor CCR2 protects from pulmonary fibrosis in ILD mouse models. We hypothesized that pulmonary MCP-1 and CCR2+ T cells accumulate in pediatric ILD and are related to disease severity. Methods Bronchoalveolar lavage fluid was obtained from 25 children with ILD and 10 healthy children. Levels of pulmonary MCP-1 and Th1/Th2-associated cytokines were quantified at the protein and the mRNA levels. Pulmonary CCR2+, CCR4+, CCR3+, CCR5+ and CXCR3+ T cells were quantified by flow-cytometry. Results CCR2+ T cells and MCP-1 levels were significantly elevated in children with ILD and correlated with forced vital capacity, total lung capacity and ILD disease severity scores. Children with lung fibrosis had significantly higher MCP-1 levels and CCR2+ T cells in bronchoalveolar lavage fluid compared to non-fibrotic children. Conclusion The results indicate that pulmonary CCR2+ T cells and MCP-1 contribute to the pathogenesis of pediatric ILD and might provide a novel target for therapeutic strategies. PMID:16095529

Fv1-like restriction of N-tropic replication-competent murine leukaemia viruses in mCAT-1-expressing human cells.

PubMed

Aagaard, Lars; Mikkelsen, Jacob Giehm; Warming, Søren; Duch, Mogens; Pedersen, Finn Skou

2002-02-01

To study the replication of murine leukaemia viruses in human cells we have used full-length as well as EGFP-tagged ecotropic viruses in combination with mCAT-1-expressing human cells. We present results showing that N-tropic murine leukaemia viruses are restricted in both infection and replication in such cells while B-tropic viruses, modified at capsid position 110, escape restriction. These results support a recently reported Fv1-like restriction in mammalian cells. We extend the analysis of Fv1-like restriction by demonstrating that NB-tropic viruses also escape restriction and human mCAT-1-expressing cells are thus similar to murine Fv1(b) cells with respect to infection though the ecotropic receptor pathway.

Three-Year Safety and Efficacy of Vicriviroc, a CCR5 Antagonist, in HIV-1-Infected, Treatment-Experienced Patients

PubMed Central

Wilkin, Timothy J.; Su, Zhaohui; Krambrink, Amy; Long, Jianmin; Greaves, Wayne; Gross, Robert; Hughes, Michael D.; Flexner, Charles; Skolnik, Paul R.; Coakley, Eoin; Godfrey, Catherine; Hirsch, Martin; Kuritzkes, Daniel R.; Gulick, Roy M.

2010-01-01

Background Vicriviroc, an investigational CCR5 antagonist, demonstrated short-term safety and antiretroviral activity. Methods Phase 2, double-blind, randomized study of vicriviroc in treatment-experienced subjects with CCR5-using HIV-1. Vicriviroc (5, 10 or 15 mg) or placebo was added to a failing regimen with optimization of background antiretroviral medications at day 14. Subjects experiencing virologic failure and subjects completing 48 weeks were offered open-label vicriviroc. Results 118 subjects were randomized. Virologic failure (<1 log10 decline in HIV-1 RNA ≥16 weeks post-randomization) occurred by week 48 in 24/28 (86%), 12/30 (40%), 8/30 (27%), 10/30 (33%) of subjects randomized to placebo, 5, 10 and 15 mg respectively. Overall, 113 subjects received vicriviroc at randomization or after virologic failure, and 52 (46%) achieved HIV-1 RNA <50 copies/mL within 24 weeks. Through 3 years, 49% of those achieving suppression did not experience confirmed viral rebound. Dual or mixed-tropic HIV-1 was detected in 33 (29%). Vicriviroc resistance (progressive decrease in maximal percentage inhibition on phenotypic testing) was detected in 6 subjects. Nine subjects discontinued vicriviroc due to adverse events. Conclusions Vicriviroc appears safe and demonstrates sustained virologic suppression through 3 years of follow-up. Further trials of vicriviroc will establish its clinical utility for the treatment of HIV-1 infection. PMID:20672447

Infection with dengue-2 virus alters proteins in naturally expectorated saliva of Aedes aegypti mosquitoes

PubMed Central

2014-01-01

Background Dengue virus (DENV) is responsible for up to approximately 300 million infections and an increasing number of deaths related to severe manifestations each year in affected countries throughout the tropics. It is critical to understand the drivers of this emergence, including the role of vector-virus interactions. When a DENV-infected Aedes aegypti mosquito bites a vertebrate, the virus is deposited along with a complex mixture of salivary proteins. However, the influence of a DENV infection upon the expectorated salivary proteome of its vector has yet to be determined. Methods Therefore, we conducted a proteomic analysis using 2-D gel electrophoresis coupled with mass spectrometry based protein identification comparing the naturally expectorated saliva of Aedes aegypti infected with DENV-2 relative to that of uninfected Aedes aegypti. Results Several proteins were found to be differentially expressed in the saliva of DENV-2 infected mosquitoes, in particular proteins with anti-hemostatic and pain inhibitory functions were significantly reduced. Hypothetical consequences of these particular protein reductions include increased biting rates and transmission success, and lead to alteration of transmission potential as calculated in our vectorial capacity model. Conclusions We present our characterizations of these changes with regards to viral transmission and mosquito blood-feeding success. Further, we conclude that our proteomic analysis of Aedes aegypti saliva altered by DENV infection provides a unique opportunity to identify pro-viral impacts key to virus transmission. PMID:24886023

CCR2, CCR5, and CXCL12 variation and HIV/AIDS in Papua New Guinea

PubMed Central

Mehlotra, Rajeev K.; Zikursh, Melinda J. Blood; Stein, Catherine M.; Siba, Peter M.; Zimmerman, Peter A.

2015-01-01

Polymorphisms in chemokine receptors, serving as HIV co-receptors, and their ligands are among the well-known host genetic factors associated with susceptibility to HIV infection and/or disease progression. Papua New Guinea (PNG) has one of the highest adult HIV prevalences in the Asia-Pacific region. However, information regarding the distribution of polymorphisms in chemokine receptor (CCR5, CCR2) and chemokine (CXCL12) genes in PNG is very limited. In this study, we genotyped a total of nine CCR2-CCR5 polymorphisms, including CCR2 190G>A, CCR5 −2459G>A and Δ32, and CXCL12 801G>A in PNG (n = 258), North America (n = 184), and five countries in West Africa (n = 178). Using this data, we determined previously characterized CCR5 haplotypes. In addition, based on the previously reported associations of CCR2 190, CCR5 −2459, CCR5 open reading frame, and CXCL12 801 genotypes with HIV acquisition and/or disease progression, we calculated composite full risk scores, considering both protective as well as susceptibility effects of the CXCL12 801 AA genotype. We observed a very high frequency of the CCR5 −2459A allele (0.98) in the PNG population, which together with the absence of Δ32 resulted in a very high frequency of the HHE haplotype (0.92). These frequencies were significantly higher than in any other population (all P-values < 0.001). Regardless of whether we considered the CXCL12 801 AA genotype protective or susceptible, the risk scores were significantly higher in the PNG population compared with any other population (all P-values < 0.001). The results of this study provide new insights regarding CCR5 variation in the PNG population, and suggest that the collective variation in CCR2, CCR5, and CXCL12 may increase the risk of HIV/AIDS in a large majority of Papua New Guineans. PMID:26397046

Exaggerated Hepatic Injury Due to Acetaminophen Challenge in Mice Lacking C-C Chemokine Receptor 2

PubMed Central

Hogaboam, Cory M.; Bone-Larson, Cynthia L.; Steinhauser, Matthew L.; Matsukawa, Akihiro; Gosling, Jennifa; Boring, Landin; Charo, Israel F.; Simpson, Kenneth J.; Lukacs, Nicholas W.; Kunkel, Steven L.

2000-01-01

Monocyte chemoattractant protein-1 is one of the major C-C chemokines that has been implicated in liver injury. The C-C chemokine receptor, CCR2, has been identified as the primary receptor that mediates monocyte chemoattractant protein-1 (MCP-1) responses in the mouse. Accordingly, the present study addressed the role of CCR2 in mice acutely challenged with acetaminophen (APAP). Mice genetically deficient in CCR2 (CCR2−/−) and their wild-type counterparts (CCR2+/+) were fasted for 10 hours before receiving an intraperitoneal injection of APAP (300 mg/kg). Liver and serum samples were removed from both groups of mice before and at 24 and 48 hours post APAP. Significantly elevated levels of MCP-1 were detected in liver samples from CCR2+/+ and CCR2−/− mice at 24 hours post-APAP. Although CCR2+/+ mice exhibited no liver injury at any time after receiving APAP, CCR2−/− mice exhibited marked evidence of necrotic and TUNEL-positive cells in the liver, particularly at 24 hours post-APAP. Enzyme-linked immunosorbent assay analysis of liver homogenates from both groups of mice at the 24 hours time point revealed that liver tissue from CCR2−/− mice contained significantly greater amounts of immunoreactive IFN-γ and TNF-α. The in vivo immunoneutralization of IFN-γ or TNF-α significantly attenuated APAP-induced liver injury in CCR2−/− mice and increased hepatic IL-13 levels. Taken together, these findings demonstrate that CCR2 expression in the liver provides a hepatoprotective effect through its regulation of cytokine generation during APAP challenge. PMID:10751350

Factors secreted by human T lymphotropic virus type I (HTLV-I)-infected cells can enhance or inhibit replication of HIV-1 in HTLV-I-uninfected cells: implications for in vivo coinfection with HTLV-I and HIV-1.

PubMed

Moriuchi, H; Moriuchi, M; Fauci, A S

1998-05-18

It remains controversial whether human T lymphotropic virus type I (HTLV-I) coinfection leads to more rapid progression of human immunodeficiency virus (HIV) disease in dually infected individuals. To investigate whether HTLV-I infection of certain cells can modulate HIV-1 infection of surrounding cells, primary CD4(+) T cells were treated with cell-free supernatants from HTLV-I-infected MT-2 cell cultures. The primary CD4+ T cells became resistant to macrophage (M)-tropic HIV-1 but highly susceptible to T cell (T)-tropic HIV-1. The CC chemokines RANTES (regulated on activation, normal T cell expressed and secreted), macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta in the MT-2 cell supernatants were identified as the major suppressive factors for M-tropic HIV-1 as well as the enhancers of T-tropic HIV-1 infection, whereas soluble Tax protein increased susceptibility to both M- and T-tropic HIV-1. The effect of Tax or CC chemokines on T-tropic HIV-1 was mediated, at least in part, by increasing HIV Env-mediated fusogenicity. Our data suggest that the net effect of HTLV-I coinfection in HIV-infected individuals favors the transition from M- to T-tropic HIV phenotype, which is generally indicative of progressive HIV disease.

Pulmonary CCR2+CD4+ T cells are immune regulatory and attenuate lung fibrosis development.

PubMed

Milger, Katrin; Yu, Yingyan; Brudy, Eva; Irmler, Martin; Skapenko, Alla; Mayinger, Michael; Lehmann, Mareike; Beckers, Johannes; Reichenberger, Frank; Behr, Jürgen; Eickelberg, Oliver; Königshoff, Melanie; Krauss-Etschmann, Susanne

2017-11-01

Animal models have suggested that CCR2-dependent signalling contributes to the pathogenesis of pulmonary fibrosis, but global blockade of CCL2 failed to improve the clinical course of patients with lung fibrosis. However, as levels of CCR2 + CD4 + T cells in paediatric lung fibrosis had previously been found to be increased, correlating with clinical symptoms, we hypothesised that distinct CCR2 + cell populations might either increase or decrease disease pathogenesis depending on their subtype. To investigate the role of CCR2 + CD4 + T cells in experimental lung fibrosis and in patients with idiopathic pulmonary fibrosis and other fibrosis. Pulmonary CCR2 + CD4 + T cells were analysed using flow cytometry and mRNA profiling, followed by in silico pathway analysis, in vitro assays and adoptive transfer experiments. Frequencies of CCR2 + CD4 + T cells were increased in experimental fibrosis-specifically the CD62L - CD44 + effector memory T cell phenotype, displaying a distinct chemokine receptor profile. mRNA profiling of isolated CCR2 + CD4 + T cells from fibrotic lungs suggested immune regulatory functions, a finding that was confirmed in vitro using suppressor assays. Importantly, adoptive transfer of CCR2 + CD4 + T cells attenuated fibrosis development. The results were partly corroborated in patients with lung fibrosis, by showing higher percentages of Foxp3 + CD25 + cells within bronchoalveolar lavage fluid CCR2 + CD4 + T cells as compared with CCR2 - CD4 + T cells. Pulmonary CCR2 + CD4 + T cells are immunosuppressive, and could attenuate lung inflammation and fibrosis. Therapeutic strategies completely abrogating CCR2-dependent signalling will therefore also eliminate cell populations with protective roles in fibrotic lung disease. This emphasises the need for a detailed understanding of the functions of immune cell subsets in fibrotic lung disease. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Could FIV zoonosis responsible of the breakdown of the pathocenosis which has reduced the European CCR5-Delta32 allele frequencies?

PubMed Central

Faure, Eric

2008-01-01

Background In Europe, the north-south downhill cline frequency of the chemokine receptor CCR5 allele with a 32-bp deletion (CCR5-Δ32) raises interesting questions for evolutionary biologists. We had suggested first that, in the past, the European colonizers, principally Romans, might have been instrumental of a progressively decrease of the frequencies southwards. Indeed, statistical analyses suggested strong negative correlations between the allele frequency and historical parameters including the colonization dates by Mediterranean civilisations. The gene flows from colonizers to native populations were extremely low but colonizers are responsible of the spread of several diseases suggesting that the dissemination of parasites in naive populations could have induced a breakdown rupture of the fragile pathocenosis changing the balance among diseases. The new equilibrium state has been reached through a negative selection of the null allele. Results Most of the human diseases are zoonoses and cat might have been instrumental in the decrease of the allele frequency, because its diffusion through Europe was a gradual process, due principally to Romans; and that several cat zoonoses could be transmitted to man. The possible implication of a feline lentivirus (FIV) which does not use CCR5 as co-receptor is discussed. This virus can infect primate cells in vitro and induces clinical signs in macaque. Moreover, most of the historical regions with null or low frequency of CCR5-Δ32 allele coincide with historical range of the wild felid species which harbor species-specific FIVs. Conclusion We proposed the hypothesis that the actual European CCR5 allelic frequencies are the result of a negative selection due to a disease spreading. A cat zoonosis, could be the most plausible hypothesis. Future studies could provide if CCR5 can play an antimicrobial role in FIV pathogenesis. Moreover, studies of ancient DNA could provide more evidences regarding the implications of zoonoses in the actual CCR5-Δ32 distribution. PMID:18925940

Effects of CCR5-delta32 and CCR2-64I alleles on disease progression of perinatally HIV-1-infected children: an international meta-analysis.

PubMed

Ioannidis, John P A; Contopoulos-Ioannidis, Despina G; Rosenberg, Philip S; Goedert, James J; De Rossi, Anita; Espanol, Teresa; Frenkel, Lisa; Mayaux, Marie-Jeanne; Newell, Marie-Louise; Pahwa, Savita G; Rousseau, Christine; Scarlatti, Gabriella; Sei, Shizuko; Sen, Luisa; O'Brien, Thomas R

2003-07-25

Among perinatally infected children, the effects of certain alleles of the CCR5 and CCR2 genes on the rate of disease progression remain unclear. We addressed the effects of CCR5-delta32 and CCR2-64I in an international meta-analysis. Genotype data were contributed from 10 studies with 1317 HIV-1-infected children (7263 person-years of follow-up). Time-to-event analyses were performed stratified by study and racial group. Endpoints included progression to clinical AIDS, death, and death after the diagnosis of clinical AIDS. The time-dependence of the genetic effects was specifically investigated. There was large heterogeneity in the observed rates of disease progression between different cohorts. For progression to clinical AIDS, both CCR5-delta32 and CCR2-64I showed overall non-significant trends for protection [hazard ratios 0.84, 95% confidence interval (CI) 0.58-1.23; and 0.87, 95% CI 0.67-1.14, respectively]. However, analyses of survival showed statistically significant time-dependence. No deaths occurred among CCR5-delta32 carriers in the first 3 years of life, whereas there was no protective effect (hazard ratio 0.95; 95% CI 0.43-2.10) in later years (P=0.01 for the time-dependent model). For CCR2-64I, the hazard ratio for death was 0.69 (95% CI 0.39-1.21) in the first 6 years of life and 2.56 (95% CI 1.26-5.20) in subsequent years (P<0.01 for the time-dependent model). CCR5-delta32 and CCR2-64I offered no clear protection after clinical AIDS had developed. The CCR5-delta32 and CCR2-64I alleles are associated with a decreased risk of death among perinatally infected children, but only for the first years of life.

Donor single nucleotide polymorphism in the CCR9 gene affects the incidence of skin GVHD.

PubMed

Inamoto, Y; Murata, M; Katsumi, A; Kuwatsuka, Y; Tsujimura, A; Ishikawa, Y; Sugimoto, K; Onizuka, M; Terakura, S; Nishida, T; Kanie, T; Taji, H; Iida, H; Suzuki, R; Abe, A; Kiyoi, H; Matsushita, T; Miyamura, K; Kodera, Y; Naoe, T

2010-02-01

The interactions between chemokines and their receptors may have an important role in initiating GVHD after allogeneic hematopoietic SCT (allo-HSCT). CCL25 and CCR9 are unique because they are exclusively expressed in epithelial cells and in Peyer's patches of the small intestine. We focused on rs12721497 (G926A), one of the non-synonymous single nucleotide polymorphisms (SNPs) in the CCR9 gene, and analyzed the SNP of donors in 167 consecutive patients who received allo-HSCT from an HLA-identical sibling donor. Genotypes were tested for associations with acute and chronic GVHD in each organ and transplant outcome. Multivariate analyses showed that the genotype 926AG was significantly associated with the incidence of acute stage > or =2 skin GVHD (hazard ratio: 3.2; 95% confidence interval (95% CI): 1.1-9.1; P=0.032) and chronic skin GVHD (hazard ratio: 4.1; 95% CI: 1.1-15; P=0.036), but not with GVHD in other organs or with relapse, non-relapse mortality or OS. To clarify the functional differences between genotypes, each SNP in retroviral vectors was transfected into Jurkat cells. In chemotaxis assays, the 926G transfectant showed greater response to CCL25 than the 926A transfectant. In conclusion, more active homing of CCR9-926AG T cells to Peyer's patches may produce changes in Ag presentation and result in increased incidence of skin GVHD.

CCR2 V64I polymorphism in rifampicin resistant tuberculosis patients in Moewardi General Hospital Surakarta, Indonesia

NASA Astrophysics Data System (ADS)

Marwoto; Agung Prasetyo, Afiono; Reviono; Suradi

2018-05-01

CC chemokine receptor-2 (CCR2) play important roles in inflammation. The CCR2 V64I polymorphism already reported associated with many diseases; however, the association of CCR2 V64I polymorphism with tuberculosis is still unknown. Also, there is no report about the presentation of CCR2 V64I polymorphisms in Indonesian tuberculosis patients with rifampicin-mono resistant status has ever been published, to the best of our knowledge. This study evaluated the presence of CCR2 V64I polymorphisms in Javanese rifampicin-mono resistant tuberculosis patients. In an ongoing molecular epidemiology study of human genomic polymorphisms and infection, 51 Javanese rifampicin-mono resistant tuberculosis patients in Dr. Moewardi General Hospital in Surakarta were enrolled in the study. The blood samples were aliquoted and fractionated. The nucleic acids were extracted from all blood samples and subjected to the CCR2 V64I polymorphisms detection by a polymerase chain reaction-sequence-specific primer (PCR-SSP) technique. PCR products were analyzed in 3% agarose. CCR2 64V and 64I homozygote were found in 23.5% (12/51) and 23.5% (12/51) blood samples, respectively. The CCR2 VI genotype was found in 52.9% (27/51) blood samples. The CCR2 VI genotype was found predominant in Javanese rifampicin-mono resistant tuberculosis patients and may have anassociation with the clinical progression.

Chemokine receptor antagonists: part 2.

PubMed

Pease, James E; Horuk, Richard

2009-02-01

The first part of this two-part review discussed approaches to generating antagonists for some of the CC chemokine receptors, including CCR1, CCR2, CCR3, and CCR4. This second part of the series concludes the review by describing antagonists for CCR5, CCR8, CCR9, CXCR3, CXCR4, and promiscuous antagonists. Chemokine receptor antagonists have found mixed success as therapeutics. Although one antagonist--maraviroc, a CCR5 inhibitor to treat AIDS--has been registered as an approved drug, this is the only success so far. There have been many failures in the clinic and we discuss the idea of promiscuous receptor antagonists as an alternative approach.

Characterization of Novel Simian Immunodeficiency Viruses from Red-Capped Mangabeys from Nigeria (SIVrcmNG409 and -NG411)

PubMed Central

Beer, Brigitte E.; Foley, Brian T.; Kuiken, Carla L.; Tooze, Zena; Goeken, Robert M.; Brown, Charles R.; Hu, Jinjie; Claire, Marisa St.; Korber, Bette T.; Hirsch, Vanessa M.

2001-01-01

Two novel simian immunodeficiency virus (SIV) strains from wild-caught red-capped mangabeys (Cercocebus torquatus torquatus) from Nigeria were characterized. Sequence analysis of the fully sequenced SIV strain rcmNG411 (SIVrcmNG411) and gag and pol sequence of SIVrcmNG409 revealed that they were genetically most closely related to the recently characterized SIVrcm from Gabon (SIVrcmGB1). Thus, red-capped mangabeys from distant geographic locations harbor a common lineage of SIV. SIVrcmNG411 carried a vpx gene in addition to vpr, suggesting a common evolutionary ancestor with SIVsm (from sooty mangabeys). However, SIVrcm was only marginally closer to SIVsm in that region than to any of the other lentiviruses. SIVrcm showed the highest similarity in pol with SIVdrl, isolated from a drill, a primate that is phylogenetically distinct from mangabey monkeys, and clustered with other primate lentiviruses (primarily SIVcpz [from chimpanzees] and SIVagmSab [from African green monkeys]) discordantly in different regions of the genome, suggesting a history of recombination. Despite the genetic relationship to SIVcpz in the pol gene, SIVrcmNG411 did not replicate in chimpanzee peripheral blood mononuclear cells (PBMC), although two other viruses unrelated to SIVcpz, SIVmndGB1 (from mandrills) and SIVlhoest (from L'Hoest monkeys), were able to grow in chimpanzee PBMC. The CCR5 24-bp deletion previously described in red-capped mangabeys from Gabon was also observed in Nigerian red-capped mangabeys, and SIVrcmNG411, like SIVrcmGB1, used CCR2B and STRL33 as coreceptors for virus entry. SIVrcm, SIVsm, SIVmndGB1, and all four SIVlhoest isolates but not SIVsun (from sun-tailed monkeys) replicated efficiently in human PBMC, suggesting that the ability to infect the human host can vary within one lineage. PMID:11711592

Cyclooxygenase-2 up-regulates CCR7 expression via AKT-mediated phosphorylation and activation of Sp1 in breast cancer cells.

PubMed

Chuang, Chun-Wei; Pan, Mei-Ren; Hou, Ming-Feng; Hung, Wen-Chun

2013-02-01

Up-regulation of cyclooxygenase-2 (COX-2) is frequently found in human cancers and is significantly associated with tumor metastasis. Our previous results demonstrate that COX-2 and its metabolite prostaglandin E2 (PGE2) stimulate the expression of CCR7 chemokine receptor via EP2/EP4 receptors to promote lymphatic invasion in breast cancer cells. In this study, we address the underlying mechanism of COX-2/PGE2-induced CCR7 expression. We find that COX-2/PGE2 increase CCR7 expression via the AKT signaling pathway in breast cancer cells. Promoter deletion and mutation assays identify the Sp1 site located at the -60/-57 region of CCR7 gene promoter is critical for stimulation. Chromatin immunoprecipitation (ChIP) assay confirms that in vivo binding of Sp1 to human CCR7 promoter is increased by COX-2 and PGE2. Knockdown of Sp1 by shRNA reduces the induction of CCR7 by PGE2. We demonstrate for the first time that AKT may directly phosphorylate Sp1 at S42, T679, and S698. Phosphorylation-mimic Sp1 protein harboring S42D, T679D, and S698D mutation strongly activates CCR7 expression. In contrast, change of these three residues to alanine completely blocks the induction of CCR7 by PGE2. Pathological investigation demonstrates that CCR7 expression is strongly associated with phospho-AKT and Sp1 in 120 breast cancer tissues. Collectively, our results demonstrate that COX-2 up-regulates CCR7 expression via AKT-mediated phosphorylation and activation of Sp1 and this pathway is highly activated in metastatic breast cancer. Copyright © 2012 Wiley Periodicals, Inc.

Use of four next-generation sequencing platforms to determine HIV-1 coreceptor tropism.

PubMed

Archer, John; Weber, Jan; Henry, Kenneth; Winner, Dane; Gibson, Richard; Lee, Lawrence; Paxinos, Ellen; Arts, Eric J; Robertson, David L; Mimms, Larry; Quiñones-Mateu, Miguel E

2012-01-01

HIV-1 coreceptor tropism assays are required to rule out the presence of CXCR4-tropic (non-R5) viruses prior treatment with CCR5 antagonists. Phenotypic (e.g., Trofile™, Monogram Biosciences) and genotypic (e.g., population sequencing linked to bioinformatic algorithms) assays are the most widely used. Although several next-generation sequencing (NGS) platforms are available, to date all published deep sequencing HIV-1 tropism studies have used the 454™ Life Sciences/Roche platform. In this study, HIV-1 co-receptor usage was predicted for twelve patients scheduled to start a maraviroc-based antiretroviral regimen. The V3 region of the HIV-1 env gene was sequenced using four NGS platforms: 454™, PacBio® RS (Pacific Biosciences), Illumina®, and Ion Torrent™ (Life Technologies). Cross-platform variation was evaluated, including number of reads, read length and error rates. HIV-1 tropism was inferred using Geno2Pheno, Web PSSM, and the 11/24/25 rule and compared with Trofile™ and virologic response to antiretroviral therapy. Error rates related to insertions/deletions (indels) and nucleotide substitutions introduced by the four NGS platforms were low compared to the actual HIV-1 sequence variation. Each platform detected all major virus variants within the HIV-1 population with similar frequencies. Identification of non-R5 viruses was comparable among the four platforms, with minor differences attributable to the algorithms used to infer HIV-1 tropism. All NGS platforms showed similar concordance with virologic response to the maraviroc-based regimen (75% to 80% range depending on the algorithm used), compared to Trofile (80%) and population sequencing (70%). In conclusion, all four NGS platforms were able to detect minority non-R5 variants at comparable levels suggesting that any NGS-based method can be used to predict HIV-1 coreceptor usage.

Identification of the Single Immunodominant Region of the Native Human CC Chemokine Receptor 6 Recognized by Mouse Monoclonal Antibodies.

PubMed

Dorgham, Karim; Dejou, Cécile; Piesse, Christophe; Gorochov, Guy; Pène, Jérôme; Yssel, Hans

2016-01-01

Chemokines and their receptors play an important role in cell trafficking and recruitment. The CCR6 chemokine receptor, selectively expressed on leukocyte populations, has been shown to play a deleterious role in the pathogenesis of various chronic inflammatory diseases and, as such, may constitute a prime target in the development of immunotherapeutic treatment. However, to date no neutralizing mouse monoclonal antibodies (mAbs) specific for this chemokine receptor have been reported, whereas information on small molecules capable of interfering with the interaction of CCR6 and its ligands is scant. Here, we report the failure to generate neutralizing mouse mAbs specific for human (hu)CCR6. Immunization of mice with peptides mimicking extracellular domains, potentially involved in CCR6 function, failed to induce Abs reactive with the native receptor. Although the use of NIH-3T3 cells expressing huCCR6 resulted in the isolation of mAbs specific for this receptor, they were not able to block the interaction between huCCR6 and huCCL20. Investigation of the anti-huCCR6 mAbs generated in the present study, as well as those commercially available, show that all mAbs invariably recognize a unique, non-neutralizing, immunodominant region in the first part of its N-terminal domain. Together, these results indicate that the generation of potential neutralizing anti-huCCR6 mAbs in the mouse is unlikely to succeed and that alternative techniques, such as the use of other animal species for immunization, might constitute a better approach to generate such a potentially therapeutic tool for the treatment of inflammatory disease.

Biological and genetic evolution of HIV type 1 in two siblings with different patterns of disease progression.

PubMed

Ripamonti, Chiara; Leitner, Thomas; Laurén, Anna; Karlsson, Ingrid; Pastore, Angela; Cavarelli, Mariangela; Antonsson, Liselotte; Plebani, Anna; Fenyö, Eva Maria; Scarlatti, Gabriella

2007-12-01

To investigate the immunological and virological factors that may lead to different patterns of disease progression characteristic of HIV-1-infected children, two HIV-1-infected siblings, a slow and a fast progressor, were followed prospectively before the onset of highly active antiretroviral therapy. Viral coreceptor usage, including the use of CCR5/CXCR4 chimeric receptors, macrophage tropism, and sensitivity to the CC-chemokine RANTES, has been studied. An autologous and heterologous neutralizing antibody response has been documented using peripheral blood mononuclear cells- and GHOST(3) cell line-based assays. Viral evolution was investigated by env C2-V3 region sequence analysis. Although both siblings were infected with HIV-1 of the R5 phenotype, their viruses showed important biological differences. In the fast progressor there was a higher RANTES sensitivity of the early virus, an increased trend to change the mode of CCR5 receptor use, and a larger genetic evolution. Both children developed an autologous neutralizing antibody response starting from the second year with evidence of the continuous emergence of resistant variants. A marked viral genetic and phenotypic evolution was documented in the fast progressor sibling, which is accompanied by a high viral RANTES sensitivity and persistent neutralizing antibodies.

Increased expression of RANTES, CCR3 and CCR5 in the lesional skin of patients with atopic eczema.

PubMed

Kato, Yoko; Pawankar, Ruby; Kimura, Yoichi; Kawana, Seiji

2006-01-01

Atopic eczema (AE) is a relapsing inflammatory disease based on IgE sensitization and characterized by peripheral blood eosinophilia and eosinophil infiltration into the lesional skin. In the patch test reaction of AE by allergens, an increased infiltration of activated eosinophils has been demonstrated peaking at 24-48 h. Regulated on activation normal T cell expressed and secreted (RANTES/CCL5) is a chemokine that induces eosinophil migration, and CCR3 and CCR5 are the receptors of RANTES. In order to further clarify the pathomechanisms of eosinophil infiltration in ongoing chronic inflammation in the skin of patients with AE and its relation to disease severity, we examined the expression of RANTES and its receptors CCR3 and CCR5 in challenged and unchallenged lesional skin of AE. We examined the number of RANTES+ cells, CCR3+ cells, CCR5+cells, activated (EG2+) eosinophils and CD3+ T cells in normal skin of healthy volunteers, and in challenged lesional skin (24 h after mite patch test) as well as unchallenged lesional skin of AE patients by immunohistochemistry. The cellular source of RANTES, CCR3 and CCR5 was analyzed by double immunohistochemistry using specific antibodies to RANTES, CCR3 or CCR5, and antibodies to ECP (EG2) or CD3. The numbers of RANTES+ cells, CCR3+ cells, CCR5+ cells, EG2+ cells and CD3+ cells were all significantly increased in challenged (mite patch-tested) lesional skin of AE patients as compared to those in unchallenged lesional skin and normal skin. The numbers of these cells in unchallenged lesional skin were greater than those in normal skin. The number of EG2+ cells in the unchallenged lesional skin correlated with both the peripheral blood eosinophil count and the SCORAD index. The number of EG2+ cells in challenged lesional skin correlated with the number of CCR5+ cells. Activated eosinophils and T cells expressed RANTES and various proportions of these cells were CCR3+ and CCR5+ in both challenged and unchallenged lesional skin. Taken together, these results suggest that RANTES as well as its receptors CCR3 and CCR5 may play important roles in the orchestration of eosinophil infiltration in ongoing chronic inflammation in AE, and also reflect the severity of disease. Copyright (c) 2006 S. Karger AG, Basel.

Clinical Case Registries: Simultaneous Local and National Disease Registries for Population Quality Management

PubMed Central

Backus, Lisa I.; Gavrilov, Sergey; Loomis, Timothy P.; Halloran, James P.; Phillips, Barbara R.; Belperio, Pamela S.; Mole, Larry A.

2009-01-01

The Department of Veterans Affairs (VA) has a system-wide, patient-centric electronic medical record system (EMR) within which the authors developed the Clinical Case Registries (CCR) to support population-centric delivery and evaluation of VA medical care. To date, the authors have applied the CCR to populations with human immunodeficiency virus (HIV) and hepatitis C virus (HCV). Local components use diagnosis codes and laboratory test results to identify patients who may have HIV or HCV and support queries on local care delivery with customizable reports. For each patient in a local registry, key EMR data are transferred via HL7 messaging to a single national registry. From 128 local registry systems, over 60,000 and 320,000 veterans in VA care have been identified as having HIV and HCV, respectively, and entered in the national database. Local and national reports covering demographics, resource usage, quality of care metrics and medication safety issues have been generated. PMID:19717794

A highly reproducible quantitative viral outgrowth assay for the measurement of the replication-competent latent HIV-1 reservoir.

PubMed

Fun, Axel; Mok, Hoi Ping; Wills, Mark R; Lever, Andrew M

2017-02-24

Cure of Human Immunodeficiency Virus (HIV) infection remains elusive due to the persistence of HIV in a latent reservoir. Strategies to eradicate latent infection can only be evaluated with robust, sensitive and specific assays to quantitate reactivatable latent virus. We have taken the standard peripheral blood mononuclear cell (PBMC) based viral outgrowth methodology and from it created a logistically simpler and more highly reproducible assay to quantify replication-competent latent HIV in resting CD4 + T cells, both increasing accuracy and decreasing cost and labour. Purification of resting CD4 + T cells from whole PBMC is expedited and achieved in 3 hours, less than half the time of conventional protocols. Our indicator cell line, SupT1-CCR5 cells (a clonal cell line expressing CD4, CXCR4 and CCR5) provides a readily available standardised readout. Reproducibility compares favourably to other published assays but with reduced cost, labour and assay heterogeneity without compromising sensitivity.

Th17 Cells Are Preferentially Infected Very Early after Vaginal Transmission of SIV in Macaques.

PubMed

Stieh, Daniel J; Matias, Edgar; Xu, Huanbin; Fought, Angela J; Blanchard, James L; Marx, Preston A; Veazey, Ronald S; Hope, Thomas J

2016-04-13

The difficulty in detecting rare infected cells immediately after mucosal HIV transmission has hindered our understanding of the initial cells targeted by the virus. Working with the macaque simian immunodeficiency virus (SIV) vaginal challenge model, we developed methodology to identify discrete foci of SIV (mac239) infection 48 hr after vaginal inoculation. We find infectious foci throughout the reproductive tract, from labia to ovary. Phenotyping infected cells reveals that SIV has a significant bias for infection of CCR6+ CD4+ T cells. SIV-infected cells expressed the transcriptional regulator RORγt, confirming that the initial target cells are specifically of the Th17 lineage. Furthermore, we detect host responses to infection, as evidenced by apoptosis, cell lysis, and phagocytosis of infected cells. Thus, our analysis identifies Th17-lineage CCR6+ CD4+ T cells as primary targets of SIV during vaginal transmission. This opens new opportunities for interventions to protect these cells and prevent HIV transmission. Copyright © 2016 Elsevier Inc. All rights reserved.

Pseudogenization of the MCP-2/CCL8 chemokine gene in European rabbit (genus Oryctolagus), but not in species of Cottontail rabbit (Sylvilagus) and Hare (Lepus)

PubMed Central

2012-01-01

Background Recent studies in human have highlighted the importance of the monocyte chemotactic proteins (MCP) in leukocyte trafficking and their effects in inflammatory processes, tumor progression, and HIV-1 infection. In European rabbit (Oryctolagus cuniculus) one of the prime MCP targets, the chemokine receptor CCR5 underwent a unique structural alteration. Until now, no homologue of MCP-2/CCL8a, MCP-3/CCL7 or MCP-4/CCL13 genes have been reported for this species. This is interesting, because at least the first two genes are expressed in most, if not all, mammals studied, and appear to be implicated in a variety of important chemokine ligand-receptor interactions. By assessing the Rabbit Whole Genome Sequence (WGS) data we have searched for orthologs of the mammalian genes of the MCP-Eotaxin cluster. Results We have localized the orthologs of these chemokine genes in the genome of European rabbit and compared them to those of leporid genera which do (i.e. Oryctolagus and Bunolagus) or do not share the CCR5 alteration with European rabbit (i.e. Lepus and Sylvilagus). Of the Rabbit orthologs of the CCL8, CCL7, and CCL13 genes only the last two were potentially functional, although showing some structural anomalies at the protein level. The ortholog of MCP-2/CCL8 appeared to be pseudogenized by deleterious nucleotide substitutions affecting exon1 and exon2. By analyzing both genomic and cDNA products, these studies were extended to wild specimens of four genera of the Leporidae family: Oryctolagus, Bunolagus, Lepus, and Sylvilagus. It appeared that the anomalies of the MCP-3/CCL7 and MCP-4/CCL13 proteins are shared among the different species of leporids. In contrast, whereas MCP-2/CCL8 was pseudogenized in every studied specimen of the Oryctolagus - Bunolagus lineage, this gene was intact in species of the Lepus - Sylvilagus lineage, and was, at least in Lepus, correctly transcribed. Conclusion The biological function of a gene was often revealed in situations of dysfunction or gene loss. Infections with Myxoma virus (MYXV) tend to be fatal in European rabbit (genus Oryctolagus), while being harmless in Hares (genus Lepus) and benign in Cottontail rabbit (genus Sylvilagus), the natural hosts of the virus. This communication should stimulate research on a possible role of MCP-2/CCL8 in poxvirus related pathogenicity. PMID:22894773

Antiviral evaluation of plants from Brazilian Atlantic Tropical Forest.

PubMed

Andrighetti-Fröhner, C R; Sincero, T C M; da Silva, A C; Savi, L A; Gaido, C M; Bettega, J M R; Mancini, M; de Almeida, M T R; Barbosa, R A; Farias, M R; Barardi, C R M; Simões, C M O

2005-06-01

The antiviral activity of six medicinal plants from Brazilian Atlantic Tropical Forest was investigated against two viruses: herpes simplex virus type 1 (HSV-1) and poliovirus type 2 (PV-2). Cuphea carthagenensis and Tillandsia usneoides extracts showed the best antiherpes activity. T. usneoides dichloromethane, ethyl acetate and n-butanol extracts, and Lippia alba n-butanol extract showed inhibition of HSV-1, strain 29R/acyclovir resistant. In addition, only L. alba ethyl acetate extract showed antipoliovirus activity. These results corroborate that medicinal plants can be a rich source of potential antiviral compounds.

Role of inflammation in the development of renal damage and dysfunction in Angiotensin II-induced hypertension

PubMed Central

Liao, Tang-Dong; Yang, Xiao-Ping; Liu, Yun-He; Shesely, Edward G.; Cavasin, Maria A.; Kuziel, William A.; Pagano, Patrick J.; Carretero, Oscar A.

2008-01-01

Angiotensin II (Ang II)-induced hypertension is associated with an inflammatory response that may contribute to development of target organ damage. We tested the hypothesis that in Angiotensin II-induced hypertension, CC chemokine receptor 2 (CCR2) activation plays an important role in development of renal fibrosis, damage and dysfunction by causing: a) oxidative stress, b) macrophage infiltration, and c) cell proliferation. To test this hypothesis we used CCR2 knockout mice (CCR2−/−). The natural ligand of CCR2 is monocyte chemoattractant protein-1 (MCP-1), a chemokine important for macrophage recruitment and activation. CCR2−/− and age-matched wild-type (CCR2+/+) C57BL/6J mice were infused continuously with either Ang II (5.2 ng/10 g/min) or vehicle via osmotic mini-pumps for 2 or 4 weeks. Ang II infusion caused similar increases in systolic blood pressure and left ventricular hypertrophy in both strains of mice. However, in CCR2−/− mice with Ang II-induced hypertension, oxidative stress, macrophage infiltration, albuminuria and renal damage were significantly decreased and glomerular filtration rate was significantly higher than in CCR2+/+ mice. We concluded that in Ang II-induced hypertension, CCR2 activation plays an important role in development of hypertensive nephropathy via increased oxidative stress and inflammation. PMID:18541733

Molecular Characterization of a Novel Temperate Sinorhizobium Bacteriophage, ФLM21, Encoding DNA Methyltransferase with CcrM-Like Specificity

PubMed Central

Dziewit, Lukasz; Oscik, Karolina; Bartosik, Dariusz

2014-01-01

ABSTRACT ΦLM21 is a temperate phage isolated from Sinorhizobium sp. strain LM21 (Alphaproteobacteria). Genomic analysis and electron microscopy suggested that ΦLM21 is a member of the family Siphoviridae. The phage has an isometric head and a long noncontractile tail. The genome of ΦLM21 has 50,827 bp of linear double-stranded DNA encoding 72 putative proteins, including proteins responsible for the assembly of the phage particles, DNA packaging, transcription, replication, and lysis. Virion proteins were characterized using mass spectrometry, leading to the identification of the major capsid and tail components, tape measure, and a putative portal protein. We have confirmed the activity of two gene products, a lytic enzyme (a putative chitinase) and a DNA methyltransferase, sharing sequence specificity with the cell cycle-regulating methyltransferase (CcrM) of the bacterial host. Interestingly, the genome of Sinorhizobium phage ΦLM21 shows very limited similarity to other known phage genome sequences and is thus considered unique. IMPORTANCE Prophages are known to play an important role in the genomic diversification of bacteria via horizontal gene transfer. The influence of prophages on pathogenic bacteria is very well documented. However, our knowledge of the overall impact of prophages on the survival of their lysogenic, nonpathogenic bacterial hosts is still limited. In particular, information on prophages of the agronomically important Sinorhizobium species is scarce. In this study, we describe the isolation and molecular characterization of a novel temperate bacteriophage, ΦLM21, of Sinorhizobium sp. LM21. Since we have not found any similar sequences, we propose that this bacteriophage is a novel species. We conducted a functional analysis of selected proteins. We have demonstrated that the phage DNA methyltransferase has the same sequence specificity as the cell cycle-regulating methyltransferase CcrM of its host. We point out that this phenomenon of mimicking the host regulatory mechanisms by viruses is quite common in bacteriophages. PMID:25187538

Initiation of Antiretroviral Therapy Restores CD4+ T Memory Stem Cell Homeostasis in Simian Immunodeficiency Virus-Infected Macaques.

PubMed

Cartwright, Emily K; Palesch, David; Mavigner, Maud; Paiardini, Mirko; Chahroudi, Ann; Silvestri, Guido

2016-08-01

Treatment of human immunodeficiency virus (HIV) infection with antiretroviral therapy (ART) has significantly improved prognosis. Unfortunately, interruption of ART almost invariably results in viral rebound, attributed to a pool of long-lived, latently infected cells. Based on their longevity and proliferative potential, CD4(+) T memory stem cells (TSCM) have been proposed as an important site of HIV persistence. In a previous study, we found that in simian immunodeficiency virus (SIV)-infected rhesus macaques (RM), CD4(+) TSCM are preserved in number but show (i) a decrease in the frequency of CCR5(+) cells, (ii) an expansion of the fraction of proliferating Ki-67(+) cells, and (iii) high levels of SIV DNA. To understand the impact of ART on both CD4(+) TSCM homeostasis and virus persistence, we conducted a longitudinal analysis of these cells in the blood and lymph nodes of 25 SIV-infected RM. We found that ART induced a significant restoration of CD4(+) CCR5(+) TSCM both in blood and in lymph nodes and a reduction in the fraction of proliferating CD4(+) Ki-67(+) TSCM in blood (but not lymph nodes). Importantly, we found that the level of SIV DNA in CD4(+) transitional memory (TTM) and effector memory (TEM) T cells declined ∼100-fold after ART in both blood and lymph nodes, while the level of SIV DNA in CD4(+) TSCM and central memory T cells (TCM-) did not significantly change. These data suggest that ART is effective at partially restoring CD4(+) TSCM homeostasis, and the observed stable level of virus in TSCM supports the hypothesis that these cells are a critical contributor to SIV persistence. Understanding the roles of various CD4(+) T cell memory subsets in immune homeostasis and HIV/SIV persistence during antiretroviral therapy (ART) is critical to effectively treat and cure HIV infection. T memory stem cells (TSCM) are a unique memory T cell subset with enhanced self-renewal capacity and the ability to differentiate into other memory T cell subsets, such as central and transitional memory T cells (TCM and TTM, respectively). CD4(+) TSCM are disrupted but not depleted during pathogenic SIV infection. We find that ART is partially effective at restoring CD4(+) TSCM homeostasis and that SIV DNA harbored within this subset contracts more slowly than virus harbored in shorter-lived subsets, such as TTM and effector memory (TEM). Because of their ability to persist long-term in an individual, understanding the dynamics of virally infected CD4(+) TSCM during suppressive ART is important for future therapeutic interventions aimed at modulating immune activation and purging the HIV reservoir. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Initiation of Antiretroviral Therapy Restores CD4+ T Memory Stem Cell Homeostasis in Simian Immunodeficiency Virus-Infected Macaques

PubMed Central

Cartwright, Emily K.; Palesch, David; Mavigner, Maud; Paiardini, Mirko; Chahroudi, Ann

2016-01-01

ABSTRACT Treatment of human immunodeficiency virus (HIV) infection with antiretroviral therapy (ART) has significantly improved prognosis. Unfortunately, interruption of ART almost invariably results in viral rebound, attributed to a pool of long-lived, latently infected cells. Based on their longevity and proliferative potential, CD4+ T memory stem cells (TSCM) have been proposed as an important site of HIV persistence. In a previous study, we found that in simian immunodeficiency virus (SIV)-infected rhesus macaques (RM), CD4+ TSCM are preserved in number but show (i) a decrease in the frequency of CCR5+ cells, (ii) an expansion of the fraction of proliferating Ki-67+ cells, and (iii) high levels of SIV DNA. To understand the impact of ART on both CD4+ TSCM homeostasis and virus persistence, we conducted a longitudinal analysis of these cells in the blood and lymph nodes of 25 SIV-infected RM. We found that ART induced a significant restoration of CD4+ CCR5+ TSCM both in blood and in lymph nodes and a reduction in the fraction of proliferating CD4+ Ki-67+ TSCM in blood (but not lymph nodes). Importantly, we found that the level of SIV DNA in CD4+ transitional memory (TTM) and effector memory (TEM) T cells declined ∼100-fold after ART in both blood and lymph nodes, while the level of SIV DNA in CD4+ TSCM and central memory T cells (TCM-) did not significantly change. These data suggest that ART is effective at partially restoring CD4+ TSCM homeostasis, and the observed stable level of virus in TSCM supports the hypothesis that these cells are a critical contributor to SIV persistence. IMPORTANCE Understanding the roles of various CD4+ T cell memory subsets in immune homeostasis and HIV/SIV persistence during antiretroviral therapy (ART) is critical to effectively treat and cure HIV infection. T memory stem cells (TSCM) are a unique memory T cell subset with enhanced self-renewal capacity and the ability to differentiate into other memory T cell subsets, such as central and transitional memory T cells (TCM and TTM, respectively). CD4+ TSCM are disrupted but not depleted during pathogenic SIV infection. We find that ART is partially effective at restoring CD4+ TSCM homeostasis and that SIV DNA harbored within this subset contracts more slowly than virus harbored in shorter-lived subsets, such as TTM and effector memory (TEM). Because of their ability to persist long-term in an individual, understanding the dynamics of virally infected CD4+ TSCM during suppressive ART is important for future therapeutic interventions aimed at modulating immune activation and purging the HIV reservoir. PMID:27170752

Assessment of kidney function in children by enzymatic determination of 2- or 24-h creatinine clearance: comparison with inulin clearance.

PubMed

Uemura, Osamu; Nagai, Takuhito; Yamakawa, Satoshi; Kaneko, Tetsuji; Hibi, Yoshiko; Yamasaki, Yasuhito; Yamamoto, Masaki; Nakano, Masaru; Iwata, Naoyuki; Hibino, Satoshi

2016-06-01

Although renal inulin clearance (Cin) is the gold standard for evaluation of kidney function, it cannot be measured easily. Therefore, creatinine clearance (Ccr) is often used clinically to evaluate kidney function. Enzymatically measured Ccr was recently found to be much higher than Cin because of the tubular secretion of creatinine (Cr). This study compared three measures of renal clearance, inulin, 2-h Ccr, and 24-h Ccr, in children. Kidney function was evaluated in 76 children (51 males and 25 females) aged 1 month to 18 years with chronic kidney disease (CKD) by three renal clearance methods at almost the same time. Correlations between each pair of three renal clearance measurements were determined. Approximate glomerular filtration rate (GFR) was equal to 62 % of 2-h Ccr or 76 % of 24-h Ccr. Cr secretion by renal tubules was approximately 50 % of the GFR. In this study, we indicate that the measurements of 2-h Ccr or 24-h Ccr do not show true GFR but we could infer approximate GFR from the values. The use of 2- or 24-h Ccr might contribute to the treatment of pediatric CKD patients.

Quantifying design trade-offs of beryllium targets on NIF

NASA Astrophysics Data System (ADS)

Yi, S. A.; Zylstra, A. B.; Kline, J. L.; Loomis, E. N.; Kyrala, G. A.; Shah, R. C.; Perry, T. S.; Kanzleiter, R. J.; Batha, S. H.; MacLaren, S. A.; Ralph, J. E.; Masse, L. P.; Salmonson, J. D.; Tipton, R. E.; Callahan, D. A.; Hurricane, O. A.

2017-10-01

An important determinant of target performance is implosion kinetic energy, which scales with the capsule size. The maximum achievable performance for a given laser is thus related to the largest capsule that can be imploded symmetrically, constrained by drive uniformity. A limiting factor for symmetric radiation drive is the ratio of hohlraum to capsule radii, or case-to-capsule ratio (CCR). For a fixed laser energy, a larger hohlraum allows for driving bigger capsules symmetrically at the cost of reduced peak radiation temperature (Tr). Beryllium ablators may thus allow for unique target design trade-offs due to their higher ablation efficiency at lower Tr. By utilizing larger hohlraum sizes than most modern NIF designs, beryllium capsules thus have the potential to operate in unique regions of the target design parameter space. We present design simulations of beryllium targets with a large CCR = 4.3 3.7 . These are scaled surrogates of large hohlraum low Tr beryllium targets, with the goal of quantifying symmetry tunability as a function of CCR. This work performed under the auspices of the U.S. DOE by LANL under contract DE-AC52- 06NA25396, and by LLNL under Contract DE-AC52-07NA27344.

Structure of CC chemokine receptor 2 with orthosteric and allosteric antagonists

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zheng, Yi; Qin, Ling; Zacarías, Natalia V. Ortiz

CC chemokine receptor 2 (CCR2) is one of 19 members of the chemokine receptor subfamily of human class A G-protein-coupled receptors. CCR2 is expressed on monocytes, immature dendritic cells, and T-cell subpopulations, and mediates their migration towards endogenous CC chemokine ligands such as CCL2 (ref. 1). CCR2 and its ligands are implicated in numerous inflammatory and neurodegenerative diseases2 including atherosclerosis, multiple sclerosis, asthma, neuropathic pain, and diabetic nephropathy, as well as cancer3. These disease associations have motivated numerous preclinical studies and clinical trials4 (see http://www.clinicaltrials.gov) in search of therapies that target the CCR2–chemokine axis. To aid drug discovery efforts5, heremore » we solve a structure of CCR2 in a ternary complex with an orthosteric (BMS-681 (ref. 6)) and allosteric (CCR2-RA-[R]7) antagonist. BMS-681 inhibits chemokine binding by occupying the orthosteric pocket of the receptor in a previously unseen binding mode. CCR2-RA-[R] binds in a novel, highly druggable pocket that is the most intracellular allosteric site observed in class A G-protein-coupled receptors so far; this site spatially overlaps the G-protein-binding site in homologous receptors. CCR2-RA-[R] inhibits CCR2 non-competitively by blocking activation-associated conformational changes and formation of the G-protein-binding interface. The conformational signature of the conserved microswitch residues observed in double-antagonist-bound CCR2 resembles the most inactive G-protein-coupled receptor structures solved so far. Like other protein–protein interactions, receptor–chemokine complexes are considered challenging therapeutic targets for small molecules, and the present structure suggests diverse pocket epitopes that can be exploited to overcome obstacles in drug design.« less

Structure of CC Chemokine Receptor 2 with Orthosteric and Allosteric Antagonists

PubMed Central

Zheng, Yi; Qin, Ling; Ortiz Zacarías, Natalia V.; de Vries, Henk; Han, Gye Won; Gustavsson, Martin; Dabros, Marta; Zhao, Chunxia; Cherney, Robert J.; Carter, Percy; Stamos, Dean; Abagyan, Ruben; Cherezov, Vadim; Stevens, Raymond C.; IJzerman, Adriaan P.; Heitman, Laura H.; Tebben, Andrew; Kufareva, Irina; Handel, Tracy M.

2016-01-01

Summary CC chemokine receptor 2 (CCR2) is one of 19 members of the chemokine receptor subfamily of human Class A G protein-coupled receptors (GPCRs). CCR2 is expressed on monocytes, immature dendritic cells and T cell subpopulations, and mediates their migration towards endogenous CC chemokine ligands such as CCL21. CCR2 and its ligands are implicated in numerous inflammatory and neurodegenerative diseases2 including atherosclerosis, multiple sclerosis, asthma, neuropathic pain, and diabetic nephropathy, as well as cancer3. These disease associations have motivated numerous preclinical studies and clinical trials4 (see ClinicalTrials.gov) in search of therapies that target the CCR2:chemokine axis. To aid drug discovery efforts5, we solved a structure of CCR2 in a ternary complex with an orthosteric (BMS-6816) and allosteric (CCR2-RA-[R]7) antagonist. BMS-681 inhibits chemokine binding by occupying the orthosteric pocket of the receptor in a previously unseen binding mode. CCR2-RA-[R] binds in a novel, highly druggable pocket that is the most intracellular allosteric site observed in Class A GPCRs to date; this site spatially overlaps the G protein-binding site in homologous receptors. CCR2-RA-[R] inhibits CCR2 non-competitively by blocking activation-associated conformational changes and formation of the G protein-binding interface. The conformational signature of the conserved microswitch residues observed in double-antagonist-bound CCR2 resembles the most inactive GPCR structures solved to date. Like other protein:protein interactions, receptor:chemokine complexes are considered challenging therapeutic targets for small molecules, and the present structure suggests diverse pocket epitopes that can be exploited to overcome drug design obstacles. PMID:27926736

1,25-dihydroxyvitamin D3 induces CCR10 expression in terminally differentiating human B cells.

PubMed

Shirakawa, Aiko-Konno; Nagakubo, Daisuke; Hieshima, Kunio; Nakayama, Takashi; Jin, Zhe; Yoshie, Osamu

2008-03-01

In the B cell lineage, CCR10 is known to be selectively expressed by plasma cells, especially those secreting IgA. In this study, we examined the regulation of CCR10 expression in terminally differentiating human B cells. As reported previously, IL-21 efficiently induced the differentiation of activated human CD19+ B cells into IgD-CD38+ plasma cells in vitro. A minor proportion of the resulting CD19+IgD-CD38+ cells expressed CCR10 at low levels. 1,25-Dihydroxyvitamin D3 (1,25-(OH)2D3), the active metabolite of vitamine D3, dramatically increased the proportion of CD19+IgD-CD38+ cells expressing high levels of CCR10. The 1,25-(OH)2D3 also increased the number of CCR10+ cells expressing surface IgA, although the majority of CCR10+ cells remained negative for surface IgA. Thus, 1,25-(OH)2D3 alone may not be sufficient for the induction of IgA expression in terminally differentiating human B cells. To further determine whether 1,25-(OH)2D3 directly induces CCR10 expression in terminally differentiating B cells, we next performed the analysis on the human CCR10 promoter. We identified a proximal Ets-1 site and an upstream potential vitamin D response element to be critical for the inducible expression of CCR10 by 1,25-(OH)2D3. We confirmed the specific binding of Ets-1 and 1,25-(OH)2D3-activated vitamin D receptor to the respective sites. In conclusion, 1,25-(OH)2D3 efficiently induces CCR10 expression in terminally differentiating human B cells in vitro. Furthermore, the human CCR10 promoter is cooperatively activated by Ets-1 and vitamin D receptor in the presence of 1,25-(OH)2D3.

Purification and crystallization of dengue and West Nile virus NS2B-NS3 complexes.

PubMed

D'Arcy, Allan; Chaillet, Maxime; Schiering, Nikolaus; Villard, Frederic; Lim, Siew Pheng; Lefeuvre, Peggy; Erbel, Paul

2006-02-01

Both dengue and West Nile virus infections are an increasing risk to humans, not only in tropical and subtropical areas, but also in North America and parts of Europe. These viral infections are generally transmitted by mosquitoes, but may also be tick-borne. Infection usually results in mild flu-like symptoms, but can also cause encephalitis and fatalities. Approximately 2799 severe West Nile virus cases were reported this year in the United States, resulting in 102 fatalities. With this alarming increase in the number of West Nile virus infections in western countries and the fact that dengue virus already affects millions of people per year in tropical and subtropical climates, there is a real need for effective medicines. A possible therapeutic target to combat these viruses is the protease, which is essential for virus replication. In order to provide structural information to help to guide a lead identification and optimization program, crystallizations of the NS2B-NS3 protease complexes from both dengue and West Nile viruses have been initiated. Crystals that diffract to high resolution, suitable for three-dimensional structure determinations, have been obtained.

Potential cancer immunotherapy drug shows promise against HIV | Center for Cancer Research

Cancer.gov

An immunotherapy currently being tested in a clinical trial as a treatment for metastatic cancer has now shown potential in an animal study to reduce recalcitrant pools of SHIV, a laboratory-designed virus used to study human immunodeficiency virus (HIV) infection. The study, published online February 23 in PLoS Pathogens, is the result of a collaborative effort by CCR, the National Institute of Allergy and Infectious Diseases and the Frederick National Laboratory for Cancer Research. Read more…

Postdoctoral Fellow | Center for Cancer Research

Cancer.gov

A postdoctoral position is available in the Viral Recombination Section (VRS), HIV Dynamics and Replication Program, CCR.  The VRS studies retroviral replication using human immunodeficiency viruses and other retroviruses, with a particular emphasis on the mechanisms of viral RNA biology, specific RNA packaging, virus assembly, and HIV replication.  Molecular tools and advanced imaging approaches are used to dissect various aspects of viral replication mechanisms.  A more complete description of the projects can be found at http://home.ncifcrf.gov/hivdrp/Hu_res.html.

Dengue Infection Increases the Locomotor Activity of Aedes aegypti Females

PubMed Central

Luz, Paula M.; Castro, Márcia G.; Lourenço-de-Oliveira, Ricardo; Sorgine, Marcos H. F.; Peixoto, Alexandre A.

2011-01-01

Background Aedes aegypti is the main vector of the virus causing Dengue fever, a disease that has increased dramatically in importance in recent decades, affecting many tropical and sub-tropical areas of the globe. It is known that viruses and other parasites can potentially alter vector behavior. We investigated whether infection with Dengue virus modifies the behavior of Aedes aegypti females with respect to their activity level. Methods/Principal Findings We carried out intrathoracic Dengue 2 virus (DENV-2) infections in Aedes aegypti females and recorded their locomotor activity behavior. We observed an increase of up to ∼50% in the activity of infected mosquitoes compared to the uninfected controls. Conclusions Dengue infection alters mosquito locomotor activity behavior. We speculate that the higher levels of activity observed in infected Aedes aegypti females might involve the circadian clock. Further studies are needed to assess whether this behavioral change could have implications for the dynamics of Dengue virus transmission. PMID:21408119

Chemokine CCL2–CCR2 Signaling Induces Neuronal Cell Death via STAT3 Activation and IL-1β Production after Status Epilepticus

PubMed Central

Tian, Dai-Shi; Feng, Li-Jie; Liu, Jun-Li

2017-01-01

Elevated levels of chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 have been reported in patients with temporal lobe epilepsy and in experimental seizures. However, the functional significance and molecular mechanism underlying CCL2–CCR2 signaling in epileptic brain remains largely unknown. In this study, we found that the upregulated CCL2 was mainly expressed in hippocampal neurons and activated microglia from mice 1 d after kainic acid (KA)-induced seizures. Taking advantage of CX3CR1GFP/+:CCR2RFP/+ double-transgenic mice, we demonstrated that CCL2–CCR2 signaling has a role in resident microglial activation and blood-derived monocyte infiltration. Moreover, seizure-induced degeneration of neurons in the hippocampal CA3 region was attenuated in mice lacking CCL2 or CCR2. We further showed that CCR2 activation induced STAT3 (signal transducer and activator of transcription 3) phosphorylation and IL-1β production, which are critical for promoting neuronal cell death after status epilepticus. Consistently, pharmacological inhibition of STAT3 by WP1066 reduced seizure-induced IL-1β production and subsequent neuronal death. Two weeks after KA-induced seizures, CCR2 deficiency not only reduced neuronal loss, but also attenuated seizure-induced behavioral impairments, including anxiety, memory decline, and recurrent seizure severity. Together, we demonstrated that CCL2–CCR2 signaling contributes to neurodegeneration via STAT3 activation and IL-1β production after status epilepticus, providing potential therapeutic targets for the treatment of epilepsy. SIGNIFICANCE STATEMENT Epilepsy is a global concern and epileptic seizures occur in many neurological conditions. Neuroinflammation associated with microglial activation and monocyte infiltration are characteristic of epileptic brains. However, molecular mechanisms underlying neuroinflammation in neuronal death following epilepsy remain to be elucidated. Here we demonstrate that CCL2–CCR2 signaling is required for monocyte infiltration, which in turn contributes to kainic acid (KA)-induced neuronal cell death. The downstream of CCR2 activation involves STAT3 (signal transducer and activator of transcription 3) phosphorylation and IL-1β production. Two weeks after KA-induced seizures, CCR2 deficiency not only reduced neuronal loss, but also attenuated seizure-induced behavioral impairments, including anxiety, memory decline, and recurrent seizure severity. The current study provides a novel insight on the function and mechanisms of CCL2–CCR2 signaling in KA-induced neurodegeneration and behavioral deficits. PMID:28716963

The Monocyte Chemoattractant Protein-1/Cognate CC Chemokine Receptor 2 System Affects Cell Motility in Cultured Human Podocytes

PubMed Central

Burt, Davina; Salvidio, Gennaro; Tarabra, Elena; Barutta, Federica; Pinach, Silvia; Dentelli, Patrizia; Camussi, Giovanni; Perin, Paolo Cavallo; Gruden, Gabriella

2007-01-01

In crescentic glomerulonephritis (GN), monocyte chemoattractant protein-1 (MCP-1) is overexpressed within the glomeruli, and MCP-1 blockade has renoprotective effects. Adult podocytes are in a quiescent state, but acquisition of a migratory/proliferative phenotype has been described in crescentic GN and implicated in crescent formation. The cognate CC chemokine receptor 2 (CCR2), the MCP-1 receptor, is expressed by other cell types besides monocytes and has been implicated in both cell proliferation and migration. We investigated whether MCP-1 binding to CCR2 can induce a migratory/proliferative response in cultured podocytes. MCP-1 binding to CCR2 enhanced podocyte chemotaxis/haptotaxis in a concentration-dependent manner and had a modest effect on cell proliferation. Closure of a wounded podocyte monolayer was delayed by CCR2 blockade, and CCR2 was overexpressed at the wound edge, suggesting a role for CCR2 in driving podocyte migration. Immunohistochemical analysis of kidney biopsies from patients with crescentic GN demonstrated CCR2 expression in both podocytes and cellular crescents, confirming the clinical relevance of our in vitro findings. In conclusion, the MCP-1/CCR2 system is functionally active in podocytes and may be implicated in the migratory events triggered by podocyte injury in crescentic GN and other glomerular diseases. PMID:18055544

Switch to maraviroc with darunavir/r, both QD, in patients with suppressed HIV-1 was well tolerated but virologically inferior to standard antiretroviral therapy: 48-week results of a randomized trial

PubMed Central

Gagliardini, Roberta; Meini, Genny; Sterrantino, Gaetana; Colangeli, Vincenzo; Re, Maria Carla; Latini, Alessandra; Colafigli, Manuela; Vignale, Francesca; Rusconi, Stefano; Micheli, Valeria; Di Biagio, Antonio; Orofino, Giancarlo; Ghisetti, Valeria; Fantauzzi, Alessandra; Vullo, Vincenzo; Grima, Pierfrancesco; Francisci, Daniela; Mastroianni, Claudio; Antinori, Andrea; Trezzi, Michele; Lisi, Lucia; Navarra, Pierluigi; Canovari, Benedetta; D’Arminio Monforte, Antonella; Lamonica, Silvia; D’Avino, Alessandro; Zazzi, Maurizio; Di Giambenedetto, Simona; De Luca, Andrea

2017-01-01

Objectives Primary study outcome was absence of treatment failure (virological failure, VF, or treatment interruption) per protocol at week 48. Methods Patients on 3-drug ART with stable HIV-1 RNA <50 copies/mL and CCR5-tropic virus were randomized 1:1 to maraviroc with darunavir/ritonavir qd (study arm) or continue current ART (continuation arm). Results In June 2015, 115 patients were evaluable for the primary outcome (56 study, 59 continuation arm). The study was discontinued due to excess of VF in the study arm (7 cases, 12.5%, vs 0 in the continuation arm, p = 0.005). The proportion free of treatment failure was 73.2% in the study and 59.3% in the continuation arm. Two participants in the study and 10 in the continuation arm discontinued therapy due to adverse events (p = 0.030). At VF, no emergent drug resistance was detected. Co-receptor tropism switched to non-R5 in one patient. Patients with VF reported lower adherence and had lower plasma drug levels. Femoral bone mineral density was significantly improved in the study arm. Conclusion Switching to maraviroc with darunavir/ritonavir qd in virologically suppressed patients was associated with improved tolerability but was virologically inferior to 3-drug therapy. PMID:29161288

Pace of Coreceptor Tropism Switch in HIV-1-Infected Individuals after Recent Infection.

PubMed

Arif, Muhammad Shoaib; Hunter, James; Léda, Ana Rachel; Zukurov, Jean Paulo Lopes; Samer, Sadia; Camargo, Michelle; Galinskas, Juliana; Kallás, Esper Georges; Komninakis, Shirley Vasconcelos; Janini, Luiz Mario; Sucupira, Maria Cecilia; Diaz, Ricardo Sobhie

2017-10-01

HIV-1 entry into target cells influences several aspects of HIV-1 pathogenesis, including viral tropism, HIV-1 transmission and disease progression, and response to entry inhibitors. The evolution from CCR5- to CXCR4-using strains in a given human host is still unpredictable. Here we analyzed timing and predictors for coreceptor evolution among recently HIV-1-infected individuals. Proviral DNA was longitudinally evaluated in 66 individuals using Geno2pheno [coreceptor] Demographics, viral load, CD4 + and CD8 + T cell counts, CCR5Δ32 polymorphisms, GB virus C (GBV-C) coinfection, and HLA profiles were also evaluated. Ultradeep sequencing was performed on initial samples from 11 selected individuals. A tropism switch from CCR5- to CXCR4-using strains was identified in 9/49 (18.4%) individuals. Only a low baseline false-positive rate (FPR) was found to be a significant tropism switch predictor. No minor CXCR4-using variants were identified in initial samples of 4 of 5 R5/non-R5 switchers. Logistic regression analysis showed that patients with an FPR of >40.6% at baseline presented a stable FPR over time whereas lower FPRs tend to progressively decay, leading to emergence of CXCR4-using strains, with a mean evolution time of 27.29 months (range, 8.90 to 64.62). An FPR threshold above 40.6% determined by logistic regression analysis may make it unnecessary to further determine tropism for prediction of disease progression related to emergence of X4 strains or use of CCR5 antagonists. The detection of variants with intermediate FPRs and progressive FPR decay over time not only strengthens the power of Geno2pheno in predicting HIV tropism but also indirectly confirms a continuous evolution from earlier R5 variants toward CXCR4-using strains. IMPORTANCE The introduction of CCR5 antagonists in the antiretroviral arsenal has sparked interest in coreceptors utilized by HIV-1. Despite concentrated efforts, viral and human host features predicting tropism switch are still poorly understood. Limited longitudinal data are available to assess the influence that these factors have on predicting tropism switch and disease progression. The present study describes longitudinal tropism evolution in a group of recently HIV-infected individuals to determine the prevalence and potential correlates of tropism switch. We demonstrated here that a low baseline FPR determined by the Geno2pheno [coreceptor] algorithm can predict tropism evolution from CCR5 to CXCR4 coreceptor use. Copyright © 2017 American Society for Microbiology.

A CCR2+ myeloid cell niche required for pancreatic β cell growth

PubMed Central

Mussar, Kristin; Pardike, Stephanie; Hohl, Tobias M.; Hardiman, Gary; Cirulli, Vincenzo

2017-01-01

Organ-specific patterns of myeloid cells may contribute tissue-specific growth and/or regenerative potentials. The perinatal stage of pancreas development marks a time characterized by maximal proliferation of pancreatic islets, ensuring the maintenance of glucose homeostasis throughout life. Ontogenically distinct CX3CR1+ and CCR2+ macrophage populations have been reported in the adult pancreas, but their functional contribution to islet cell growth at birth remains unknown. Here, we uncovered a temporally restricted requirement for CCR2+ myeloid cells in the perinatal proliferation of the endocrine pancreatic epithelium. CCR2+ macrophages are transiently enriched over CX3CR1+ subsets in the neonatal pancreas through both local expansion and recruitment of immature precursors. Using CCR2-specific depletion models, we show that loss of this myeloid population leads to a striking reduction in β cell proliferation, dysfunctional islet phenotypes, and glucose intolerance in newborns. Replenishment of pancreatic CCR2+ myeloid compartments by adoptive transfer rescues these defects. Gene profiling identifies pancreatic CCR2+ myeloid cells as a prominent source of IGF2, which contributes to IGF1R-mediated islet proliferation. These findings uncover proproliferative functions of CCR2+ myeloid subsets and identify myeloid-dependent regulation of IGF signaling as a local cue supporting pancreatic proliferation. PMID:28768911

Methamphetamine abuse affects gene expression in brain-derived microglia of SIV-infected macaques to enhance inflammation and promote virus targets.

PubMed

Najera, Julia A; Bustamante, Eduardo A; Bortell, Nikki; Morsey, Brenda; Fox, Howard S; Ravasi, Timothy; Marcondes, Maria Cecilia Garibaldi

2016-04-23

Methamphetamine (Meth) abuse is a major health problem linked to the aggravation of HIV- associated complications, especially within the Central Nervous System (CNS). Within the CNS, Meth has the ability to modify the activity/function of innate immune cells and increase brain viral loads. Here, we examined changes in the gene expression profile of neuron-free microglial cell preparations isolated from the brain of macaques infected with the Simian Immunodeficiency Virus (SIV), a model of neuroAIDS, and exposed to Meth. We aimed to identify molecular patterns triggered by Meth that could explain the detection of higher brain viral loads and the development of a pro-inflammatory CNS environment in the brain of infected drug abusers. We found that Meth alone has a strong effect on the transcription of genes associated with immune pathways, particularly inflammation and chemotaxis. Systems analysis led to a strong correlation between Meth exposure and enhancement of molecules associated with chemokines and chemokine receptors, especially CXCR4 and CCR5, which function as co-receptors for viral entry. The increase in CCR5 expression was confirmed in the brain in correlation with increased brain viral load. Meth enhances the availability of CCR5-expressing cells for SIV in the brain, in correlation with increased viral load. This suggests that Meth is an important factor in the susceptibility to the infection and to the aggravated CNS inflammatory pathology associated with SIV in macaques and HIV in humans.

A CCL chemokine-derived peptide (CDIP-2) exerts anti-inflammatory activity via CCR1, CCR2 and CCR3 chemokine receptors: Implications as a potential therapeutic treatment of asthma.

PubMed

Méndez-Enríquez, E; Medina-Tamayo, J; Soldevila, G; Fortoul, T I; Anton, B; Flores-Romo, L; García-Zepeda, E A

2014-05-01

Allergic asthma is a chronic inflammatory disease characterized by the accumulation of eosinophils, Th2 cells and mononuclear cells in the airways, leading to changes in lung architecture and subsequently reduced respiratory function. We have previously demonstrated that CDIP-2, a chemokine derived peptide, reduced in vitro chemotaxis and decreased cellular infiltration in a murine model of allergic airway inflammation. However, the mechanisms involved in this process have not been identified yet. Now, we found that CDIP-2 reduces chemokine-mediated functions via interactions with CCR1, CCR2 and CCR3. Moreover, using bone marrow-derived eosinophils, we demonstrated that CDIP-2 modifies the calcium fluxes induced by CCL11 and down-modulated CCR3 expression. Finally, CDIP-2 treatment in a murine model of OVA-induced allergic airway inflammation reduced leukocyte recruitment and decreases production of cytokines. These data suggest that chemokine-derived peptides represent new therapeutic tools to generate more effective antiinflammatory drugs. Copyright © 2014 Elsevier B.V. All rights reserved.

Inflammatory Monocytes Orchestrate Innate Antifungal Immunity in the Lung

PubMed Central

Dutta, Orchi; Kasahara, Shinji; Donnelly, Robert; Du, Peicheng; Rosenfeld, Jeffrey; Leiner, Ingrid; Chen, Chiann-Chyi; Ron, Yacov; Hohl, Tobias M.; Rivera, Amariliz

2014-01-01

Aspergillus fumigatus is an environmental fungus that causes invasive aspergillosis (IA) in immunocompromised patients. Although -CC-chemokine receptor-2 (CCR2) and Ly6C-expressing inflammatory monocytes (CCR2+Mo) and their derivatives initiate adaptive pulmonary immune responses, their role in coordinating innate immune responses in the lung remain poorly defined. Using conditional and antibody-mediated cell ablation strategies, we found that CCR2+Mo and monocyte-derived dendritic cells (Mo-DCs) are essential for innate defense against inhaled conidia. By harnessing fluorescent Aspergillus reporter (FLARE) conidia that report fungal cell association and viability in vivo, we identify two mechanisms by which CCR2+Mo and Mo-DCs exert innate antifungal activity. First, CCR2+Mo and Mo-DCs condition the lung inflammatory milieu to augment neutrophil conidiacidal activity. Second, conidial uptake by CCR2+Mo temporally coincided with their differentiation into Mo-DCs, a process that resulted in direct conidial killing. Our findings illustrate both indirect and direct functions for CCR2+Mo and their derivatives in innate antifungal immunity in the lung. PMID:24586155

Inhibition of highly productive HIV-1 infection in T cells, primary human macrophages, microglia, and astrocytes by Sargassum fusiforme

PubMed Central

Paskaleva, Elena E; Lin, Xudong; Li, Wen; Cotter, Robin; Klein, Michael T; Roberge, Emily; Yu, Er K; Clark, Bruce; Veille, Jean-Claude; Liu, Yanze; Lee, David Y-W; Canki, Mario

2006-01-01

Background The high rate of HIV-1 mutation and increasing resistance to currently available antiretroviral (ART) therapies highlight the need for new antiviral agents. Products derived from natural sources have been shown to inhibit HIV-1 replication during various stages of the virus life cycle, and therefore represent a potential source of novel therapeutic agents. To expand our arsenal of therapeutics against HIV-1 infection, we investigated aqueous extract from Sargassum fusiforme (S. fusiforme) for ability to inhibit HIV-1 infection in the periphery, in T cells and human macrophages, and for ability to inhibit in the central nervous system (CNS), in microglia and astrocytes. Results S. fusiforme extract blocked HIV-1 infection and replication by over 90% in T cells, human macrophages and microglia, and it also inhibited pseudotyped HIV-1 (VSV/NL4-3) infection in human astrocytes by over 70%. Inhibition was mediated against both CXCR4 (X4) and CCR5 (R5)-tropic HIV-1, was dose dependant and long lasting, did not inhibit cell growth or viability, was not toxic to cells, and was comparable to inhibition by the nucleoside analogue 2', 3'-didoxycytidine (ddC). S. fusiforme treatment blocked direct cell-to-cell infection spread. To investigate at which point of the virus life cycle this inhibition occurs, we infected T cells and CD4-negative primary human astrocytes with HIV-1 pseudotyped with envelope glycoprotein of vesicular stomatitis virus (VSV), which bypasses the HIV receptor requirements. Infection by pseudotyped HIV-1 (VSV/NL4-3) was also inhibited in a dose dependant manner, although up to 57% less, as compared to inhibition of native NL4-3, indicating post-entry interferences. Conclusion This is the first report demonstrating S. fusiforme to be a potent inhibitor of highly productive HIV-1 infection and replication in T cells, in primary human macrophages, microglia, and astrocytes. Results with VSV/NL4-3 infection, suggest inhibition of both entry and post-entry events of the virus life cycle. Absence of cytotoxicity and high viability of treated cells also suggest that S. fusiforme is a potential source of novel naturally occurring antiretroviral compounds that inhibit HIV-1 infection and replication at more than one site of the virus life cycle. PMID:16725040

Benzimidazoles as benzamide replacements within cyclohexane-based CC chemokine receptor 2 (CCR2) antagonists.

PubMed

Cherney, Robert J; Mo, Ruowei; Meyer, Dayton T; Pechulis, Anthony D; Guaciaro, Michael A; Lo, Yvonne C; Yang, Gengjie; Miller, Persymphonie B; Scherle, Peggy A; Zhao, Qihong; Cvijic, Mary Ellen; Barrish, Joel C; Decicco, Carl P; Carter, Percy H

2012-10-01

We describe the design, synthesis, and evaluation of benzimidazoles as benzamide replacements within a series of trisubstituted cyclohexane CCR2 antagonists. 7-Trifluoromethylbenzimidazoles displayed potent binding and functional antagonism of CCR2 while being selective over CCR3. These benzimidazoles were also incorporated into lactam-containing antagonists, thus completely eliminating the customary bis-amide. Copyright © 2012 Elsevier Ltd. All rights reserved.

Repression of Ccr9 transcription in mouse T lymphocyte progenitors by the Notch signaling pathway.

PubMed

Krishnamoorthy, Veena; Carr, Tiffany; de Pooter, Renee F; Emanuelle, Akinola Olumide; Akinola, Emanuelle Olumide; Gounari, Fotini; Kee, Barbara L

2015-04-01

The chemokine receptor CCR9 controls the immigration of multipotent hematopoietic progenitor cells into the thymus to sustain T cell development. Postimmigration, thymocytes downregulate CCR9 and migrate toward the subcapsular zone where they recombine their TCR β-chain and γ-chain gene loci. CCR9 is subsequently upregulated and participates in the localization of thymocytes during their selection for self-tolerant receptor specificities. Although the dynamic regulation of CCR9 is essential for early T cell development, the mechanisms controlling CCR9 expression have not been determined. In this article, we show that key regulators of T cell development, Notch1 and the E protein transcription factors E2A and HEB, coordinately control the expression of Ccr9. E2A and HEB bind at two putative enhancers upstream of Ccr9 and positively regulate CCR9 expression at multiple stages of T cell development. In contrast, the canonical Notch signaling pathway prevents the recruitment of p300 to the putative Ccr9 enhancers, resulting in decreased acetylation of histone H3 and a failure to recruit RNA polymerase II to the Ccr9 promoter. Although Notch signaling modestly modulates the binding of E proteins to one of the two Ccr9 enhancers, we found that Notch signaling represses Ccr9 in T cell lymphoma lines in which Ccr9 transcription is independent of E protein function. Our data support the hypothesis that activation of Notch1 has a dominant-negative effect on Ccr9 transcription and that Notch1 and E proteins control the dynamic expression of Ccr9 during T cell development. Copyright © 2015 by The American Association of Immunologists, Inc.

Human liver-resident CD56(bright)/CD16(neg) NK cells are retained within hepatic sinusoids via the engagement of CCR5 and CXCR6 pathways.

PubMed

Hudspeth, Kelly; Donadon, Matteo; Cimino, Matteo; Pontarini, Elena; Tentorio, Paolo; Preti, Max; Hong, Michelle; Bertoletti, Antonio; Bicciato, Silvio; Invernizzi, Pietro; Lugli, Enrico; Torzilli, Guido; Gershwin, M Eric; Mavilio, Domenico

2016-01-01

The liver-specific natural killer (NK) cell population is critical for local innate immune responses, but the mechanisms that lead to their selective homing and the definition of their functionally relevance remain enigmatic. We took advantage of the availability of healthy human liver to rigorously define the mechanisms regulating the homing of NK cells to liver and the repertoire of receptors that distinguish liver-resident NK (lr-NK) cells from circulating counterparts. Nearly 50% of the entire liver NK cell population is composed of functionally relevant CD56(bright) lr-NK cells that localize within hepatic sinusoids. CD56(bright) lr-NK cells express CD69, CCR5 and CXCR6 and this unique repertoire of chemokine receptors is functionally critical as it determines selective migration in response to the chemotactic stimuli exerted by CCL3, CCL5 and CXCL16. Here, we also show that hepatic sinusoids express CCL3(pos) Kupffer cells, CXCL16(pos) endothelial cells and CCL5(pos) T and NK lymphocytes. The selective presence of these chemokines in sinusoidal spaces creates a unique tissue niche for lr-CD56(bright) NK cells that constitutively express CCR5 and CXCR6. CD56(bright) lr-NK cells co-exist with CD56(dim) conventional NK (c-NK) cells that are, interestingly, transcriptionally and phenotypically similar to their peripheral circulating counterparts. Indeed, CD56(dim) c-NK cells lack expression of CD69, CCR5, and CXCR6 but express selectins, integrins and CX3CR1. Our findings disclosing the phenotypic and functional differences between lr-Nk cells and c-NK cells are critical to distinguish liver-specific innate immune responses. Hence, any therapeutic attempts at modifying the large population of CD56(bright) lr-NK cells will require modification of hepatic CCR5 and CXCR6. Copyright © 2015 Elsevier Ltd. All rights reserved.

The positive correlation of the CCL2-CCR2 axis with the disease activity may indicate the fundamental role in the pathogenesis of oral lichen planus.

PubMed

Yin, Jingfang; Yang, Xi; Zeng, Qi; Yang, Linglan; Cheng, Bin; Tao, Xiaoan

2016-01-01

The important roles of CCL2 and its receptor CCR2 had been reported in a series of inflammatory disorders. However, few studies investigated the potential role of CCL2/CCR2 axis in oral lichen planus (OLP). Therefore, this study aimed to detect the expression of CCL2 and CCR2 in OLP lesions and compare their changes before and after treatment. CCL2 and CCR2 expression was investigated using immunohistochemical staining and real-time RT-PCR in 32 patients with OLP and eight controls. Moreover, changes in their expression after treatment with triamcinolone acetonide were assessed in lesions from three patients. CCL2+ and CCR2+ cells were few in the controls and remarkably increased in the epithelial and subepithelial layers of lesions (n = 32, all P < 0.001). However, the densities of CCL2+ and CCR2+ cells were not significantly different between reticular (n = 12) and erythematous/erosive lesions (n = 20), although they significantly decreased after treatment (627.7 ± 108.2 vs. 258.3 ± 148.3, P = 0.017; 1034.7 ± 74.6 vs. 648 ± 77.6, P = 0.003, respectively). CCL2+/CCR2+ cell numbers were positively correlated with disease activity (correlation coefficient, 0.588; P < 0.001; correlation coefficient, 0.409; P = 0.02, respectively). The results of this study indicated that the CCL2-CCR2 axis was involved in the pathogenesis of OLP and was positively correlated with disease activity. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

CCR2 antagonist CCX140-B provides renal and glycemic benefits in diabetic transgenic human CCR2 knockin mice

PubMed Central

Sullivan, Timothy; Miao, Zhenhua; Dairaghi, Daniel J.; Krasinski, Antoni; Wang, Yu; Zhao, Bin N.; Baumgart, Trageen; Ertl, Linda S.; Pennell, Andrew; Seitz, Lisa; Powers, Jay; Zhao, Ruiping; Ungashe, Solomon; Wei, Zheng; Boring, Landin; Tsou, Chia-Lin; Charo, Israel; Schall, Thomas J.; Jaen, Juan C.

2013-01-01

Chemokine (C-C motif) receptor 2 (CCR2) is central for the migration of monocytes into inflamed tissues. The novel CCR2 antagonist CCX140-B, which is currently in two separate phase 2 clinical trials in diabetic nephropathy, has recently been shown to reduce hemoglobin A1c and fasting blood glucose levels in type 2 diabetics. In this report, we describe the effects of this compound on glycemic and renal function parameters in diabetic mice. Since CCX140-B has a low affinity for mouse CCR2, transgenic human CCR2 knockin mice were generated and rendered diabetic with either a high-fat diet (diet-induced obesity) or by deletion of the leptin receptor gene (db/db). CCX140-B treatment in both models resulted in decreased albuminuria, which was associated with decreased glomerular hypertrophy and increased podocyte density. Moreover, treatment of diet-induced obese mice with CCX140-B resulted in decreased levels of fasting blood glucose and insulin, normalization of homeostatic model assessment of insulin resistance values, and decreased numbers of adipose tissue inflammatory macrophages. Unlike other CCR2 antagonists, CCX140-B had no effect on plasma levels of the CCR2 ligand CCL2 or on the numbers of blood monocytes. These results support the ongoing evaluation of this molecule in diabetic subjects with impaired renal function. PMID:23986513

Significance of bacteria and viruses in the carbon flow of tropical freshwater impoundments

NASA Astrophysics Data System (ADS)

Peduzzi, P.; Schiemer, F.

2003-04-01

In two types of tropical freshwater impoundments, free and particle-attached bacterial abundance and production as well as virus abundance, frequency of viral infection and virus production were investigated together with a set of environmental factors during two characteristic seasons. Organic nitrogen, phosphorus species, dissolved organic carbon and suspended solids were elevated in the wind-mixed water body of a shallow reservoir during the dry season, whereas a deeper reservoir type exhibited no obvious seasonality in these parameters. In SYBR GREEN-stained samples, bacterial abundance showed no seasonal pattern in either reservoir type. A large proportion of the overall bacterial production was associated with particulate material. Highest densities of virus particles and elevated frequency of bacteria containing mature phages were observed in the shallow reservoir during the dry season. The specific bacterial production was related to the abundance of particulate organic matter, phosphorus species and organic nitrogen. Most virus parameters were positively linked to bacterial density, production and to organic nitrogen. We calculated that between 13.2 and 46.1% of the bacterial standing stocks would be subjected to virus-mediated mortality. Carbon budgets for the microbial and organic matter compartments of these tropical freshwater reservoirs indicate prevailing autotrophy and a substantial pathway through the viral shunt. During the dry season the shallow, wind-mixed reservoir provided favorable conditions for bacterial growth and virus propagation.

Factors involved in the generation of memory CD8+ T cells in patients with X-linked lymphoproliferative disease (XLP)

PubMed Central

Belmonte, L; Parodi, C; Baré, P; Malbrán, A; Ruibal-Ares, B; de E de Bracco, María M

2007-01-01

We have analysed the phenotype of T lymphocytes in two X-linked lymphoproliferative disease (XLP) patients with the same SH2D1A mutation differing in initial exposure to Epstein–Barr virus (EBV) and treatment. While memory T lymphocytes (with low CCR7 and CD62L expression) prevailed in both XLP patients, in patient 9, who developed acute infectious mononucleosis (AIM) and received B cell ablative treatment, the predominant phenotype was that of late effector CD8 T cells (CD27–, CD28–, CCR7–, CD62L–, CD45 RA+, perforin+), while in patient 4 (who did not suffer AIM) the prevalent phenotype of CD8 T lymphocytes was similar to that of normal controls (N) or to that of adult individuals who recovered from AIM: CD27+, CD28+, CCR7–, CD62L–, CD45 RO+ and perforin–. CD57 expression (related to senescence) was also higher in CD8 T cells from patient 9 than in patient 4, AIM or N. Persistently high EBV viral load was observed in patient 9. The results obtained from this limited number of XLP patients suggest that events related to the initial EBV encounter (antigen load, treatment, cytokine environment) may have more weight than lack of SH2D1A in determining the long-term differentiation pattern of CD8 memory T cells. PMID:17302894

Differential Roles of Chemokines CCL2 and CCL7 in Monocytosis and Leukocyte Migration during West Nile Virus Infection.

PubMed

Bardina, Susana V; Michlmayr, Daniela; Hoffman, Kevin W; Obara, Christopher J; Sum, Janet; Charo, Israel F; Lu, Wuyuan; Pletnev, Alexander G; Lim, Jean K

2015-11-01

West Nile virus (WNV) is a re-emerging pathogen and the leading cause of epidemic encephalitis in the United States. Inflammatory monocytes are a critical component of the cellular infiltrate found in the CNS during WNV encephalitis, although the molecular cues involved in their migration are not fully understood. In mice, we previously showed that WNV infection induces a CCR2-dependent monocytosis that precedes monocyte migration into the CNS. Currently, the relative contribution of the CCR2 ligands, chemokines CCL2 and CCL7, in directing monocyte mobilization and leukocyte migration into the CNS is unclear. In this study, we demonstrate that, although both CCL2 and CCL7 are required for efficient monocytosis and monocyte accumulation in the CNS, only CCL7 deficiency resulted in increased viral burden in the brain and enhanced mortality. The enhanced susceptibility in the absence of CCL7 was associated with the delayed migration of neutrophils and CD8(+) T cells into the CNS compared with WT or Ccl2(-/-) mice. To determine whether CCL7 reconstitution could therapeutically alter the survival outcome of WNV infection, we administered exogenous CCL7 i.v. to WNV-infected Ccl7(-/-) mice and observed a significant increase in monocytes and neutrophils, but not CD8(+) T cells, within the CNS, as well as an enhancement in survival compared with Ccl7(-/-) mice treated with a linear CCL7 control peptide. Our experiments suggest that CCL7 is an important protective signal involved in leukocyte trafficking during WNV infection, and it may have therapeutic potential for the treatment of acute viral infections of the CNS. Copyright © 2015 by The American Association of Immunologists, Inc.

Human T-cell leukemia virus type 1 infects multiple lineage hematopoietic cells in vivo

PubMed Central

Sugata, Kenji; Ueno, Takaharu; Koh, Ki-Ryang; Higuchi, Yusuke; Matsuda, Fumihiko; Melamed, Anat; Bangham, Charles R.

2017-01-01

Human T-cell leukemia virus type 1 (HTLV-1) infects mainly CD4+CCR4+ effector/memory T cells in vivo. However, it remains unknown whether HTLV-1 preferentially infects these T cells or this virus converts infected precursor cells to specialized T cells. Expression of viral genes in vivo is critical to study viral replication and proliferation of infected cells. Therefore, we first analyzed viral gene expression in non-human primates naturally infected with simian T-cell leukemia virus type 1 (STLV-1), whose virological attributes closely resemble those of HTLV-1. Although the tax transcript was detected only in certain tissues, Tax expression was much higher in the bone marrow, indicating the possibility of de novo infection. Furthermore, Tax expression of non-T cells was suspected in bone marrow. These data suggest that HTLV-1 infects hematopoietic cells in the bone marrow. To explore the possibility that HTLV-1 infects hematopoietic stem cells (HSCs), we analyzed integration sites of HTLV-1 provirus in various lineages of hematopoietic cells in patients with HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) and a HTLV-1 carrier using the high-throughput sequencing method. Identical integration sites were detected in neutrophils, monocytes, B cells, CD8+ T cells and CD4+ T cells, indicating that HTLV-1 infects HSCs in vivo. We also detected Tax protein in myeloperoxidase positive neutrophils. Furthermore, dendritic cells differentiated from HTLV-1 infected monocytes caused de novo infection to T cells, indicating that infected monocytes are implicated in viral spreading in vivo. Certain integration sites were re-detected in neutrophils from HAM/TSP patients at different time points, indicating that infected HSCs persist and differentiate in vivo. This study demonstrates that HTLV-1 infects HSCs, and infected stem cells differentiate into diverse cell lineages. These data indicate that infection of HSCs can contribute to the persistence and spread of HTLV-1 in vivo. PMID:29186194

Aspergillus antigen induces robust Th2 cytokine production, inflammation, airway hyperreactivity and fibrosis in the absence of MCP-1 or CCR2.

PubMed

Koth, Laura L; Rodriguez, Madeleine W; Bernstein, Xin Liu; Chan, Salina; Huang, Xiaozhu; Charo, Israel F; Rollins, Barrett J; Erle, David J

2004-09-15

Asthma is characterized by type 2 T-helper cell (Th2) inflammation, goblet cell hyperplasia, airway hyperreactivity, and airway fibrosis. Monocyte chemoattractant protein-1 (MCP-1 or CCL2) and its receptor, CCR2, have been shown to play important roles in the development of Th2 inflammation. CCR2-deficient mice have been found to have altered inflammatory and physiologic responses in some models of experimental allergic asthma, but the role of CCR2 in contributing to inflammation and airway hyperreactivity appears to vary considerably between models. Furthermore, MCP-1-deficient mice have not previously been studied in models of experimental allergic asthma. To test whether MCP-1 and CCR2 are each required for the development of experimental allergic asthma, we applied an Aspergillus antigen-induced model of Th2 cytokine-driven allergic asthma associated with airway fibrosis to mice deficient in either MCP-1 or CCR2. Previous studies with live Aspergillus conidia instilled into the lung revealed that MCP-1 and CCR2 play a role in anti-fungal responses; in contrast, we used a non-viable Aspergillus antigen preparation known to induce a robust eosinophilic inflammatory response. We found that wild-type C57BL/6 mice developed eosinophilic airway inflammation, goblet cell hyperplasia, airway hyperreactivity, elevations in serum IgE, and airway fibrosis in response to airway challenge with Aspergillus antigen. Surprisingly, mice deficient in either MCP-1 or CCR2 had responses to Aspergillus antigen similar to those seen in wild-type mice, including production of Th2 cytokines. We conclude that robust Th2-mediated lung pathology can occur even in the complete absence of MCP-1 or CCR2.

Aspergillus antigen induces robust Th2 cytokine production, inflammation, airway hyperreactivity and fibrosis in the absence of MCP-1 or CCR2

PubMed Central

Koth, Laura L; Rodriguez, Madeleine W; Bernstein, Xin Liu; Chan, Salina; Huang, Xiaozhu; Charo, Israel F; Rollins, Barrett J; Erle, David J

2004-01-01

Background Asthma is characterized by type 2 T-helper cell (Th2) inflammation, goblet cell hyperplasia, airway hyperreactivity, and airway fibrosis. Monocyte chemoattractant protein-1 (MCP-1 or CCL2) and its receptor, CCR2, have been shown to play important roles in the development of Th2 inflammation. CCR2-deficient mice have been found to have altered inflammatory and physiologic responses in some models of experimental allergic asthma, but the role of CCR2 in contributing to inflammation and airway hyperreactivity appears to vary considerably between models. Furthermore, MCP-1-deficient mice have not previously been studied in models of experimental allergic asthma. Methods To test whether MCP-1 and CCR2 are each required for the development of experimental allergic asthma, we applied an Aspergillus antigen-induced model of Th2 cytokine-driven allergic asthma associated with airway fibrosis to mice deficient in either MCP-1 or CCR2. Previous studies with live Aspergillus conidia instilled into the lung revealed that MCP-1 and CCR2 play a role in anti-fungal responses; in contrast, we used a non-viable Aspergillus antigen preparation known to induce a robust eosinophilic inflammatory response. Results We found that wild-type C57BL/6 mice developed eosinophilic airway inflammation, goblet cell hyperplasia, airway hyperreactivity, elevations in serum IgE, and airway fibrosis in response to airway challenge with Aspergillus antigen. Surprisingly, mice deficient in either MCP-1 or CCR2 had responses to Aspergillus antigen similar to those seen in wild-type mice, including production of Th2 cytokines. Conclusion We conclude that robust Th2-mediated lung pathology can occur even in the complete absence of MCP-1 or CCR2. PMID:15377395

Surveillance of enteric viruses and coliphages in a tropical urban catchment.

PubMed

Rezaeinejad, S; Vergara, G G R V; Woo, C H; Lim, T T; Sobsey, M D; Gin, K Y H

2014-07-01

An assessment of the occurrence and concentration of enteric viruses and coliphages was carried out in highly urbanized catchment waters in the tropical city-state of Singapore. Target enteric viruses in this study were noroviruses, adenoviruses, astroviruses and rotaviruses. In total, 65 water samples were collected from canals and the reservoir of the Marina catchment on a monthly basis over a period of a year. Quantitative PCR (qPCR) and single agar layer plaque assay (SAL) were used to enumerate target enteric viruses and coliphages in water samples, respectively. The most prevalent pathogen were noroviruses, detected in 37 samples (57%), particularly norovirus genogroup II (48%), with a mean concentration of 3.7 × 10(2) gene copies per liter. Rotavirus was the second most prevalent virus (40%) with a mean concentration of 2.5 × 10(2) GC/L. The mean concentrations of somatic and male-specific coliphages were 2.2 × 10(2) and 1.1 × 10(2) PFU/100 ml, respectively. The occurrence and concentration of each target virus and the ratio of somatic to male-specific coliphages varied at different sampling sites in the catchment. For sampling sites with higher frequency of occurrence and concentration of viruses, the ratio of somatic to male-specific coliphages was generally much lower than other sampling sites with lower incidences of enteric viruses. Overall, higher statistical correlation was observed between target enteric viruses than between enteric viruses and coliphages. However, male-specific coliphages were positively correlated with norovirus concentrations. A multi-level integrated surveillance system, which comprises the monitoring of bacterial indicators, coliphages and selected enteric viruses, could help to meet recreational and surface water quality criteria in a complex urbanized catchment. Copyright © 2014 Elsevier Ltd. All rights reserved.

Sexually transmitted diseases diagnosed among travelers returning from the tropics.

PubMed

Ansart, Séverine; Hochedez, Patrick; Perez, Lucia; Bricaire, François; Caumes, Eric

2009-01-01

Data are lacking on the spectrum of sexually transmitted diseases (STDs) diagnosed in returning travelers. All consecutive travelers consulting our tropical unit between November 1, 2002 and October 31, 2003 were included if they presented within 1 month after their return from the tropics, with mucocutaneous signs suggesting STDs. Forty-nine patients (12 women and 37 men; median age 36.4 y, 35 heterosexuals) were included. Four patients had traveled with their usual sexual partner and 45 patients had casual sex while abroad (31 with locals and 14 with other tourists). The main diagnoses were gonococcal urethritis (n = 18), herpes simplex virus 2 infection (n = 12), urethritis of undetermined origin (n = 9), Chlamydia trachomatis infection (n = 4), primary syphilis (n = 4), and primary human immunodeficiency virus infection (n = 2). These results illustrate the broad spectrum of STDs contracted by travelers to the tropics. They suggest the need to also inform travelers of the risks of STD and to promote the use of condoms in case of casual sex while abroad.

CCR5 chemokine receptor gene polymorphisms in ocular toxoplasmosis.

PubMed

de Faria Junior, Geraldo M; Ayo, Christiane M; de Oliveira, Amanda P; Lopes, Alessandro G; Frederico, Fábio B; Silveira-Carvalho, Aparecida P; Previato, Mariana; Barbosa, Amanda P; Murata, Fernando H A; de Almeida Junior, Gildásio Castello; Siqueira, Rubens Camargo; de Mattos, Luiz C; Brandão de Mattos, Cinara C

2018-02-01

CC chemokine receptor type 5 (CCR5) is a chemokine receptor that influences the immune response to infectious and parasitic diseases. This study aimed to determine whether the CCR5Δ32 and CCR5 59029 A/G polymorphisms are associated with the development of ocular toxoplasmosis in humans. Patients with positive serology for Toxoplasma gondii were analyzed and grouped as 'with ocular toxoplasmosis' (G1: n=160) or 'without ocular toxoplasmosis' (G2: n=160). A control group (G3) consisted of 160 individuals with negative serology. The characterization of the CCR5Δ32 and CCR5 59029 A/G polymorphisms was by PCR and by PCR-RFLP, respectively. The difference between the groups with respect to the mean age (G1: mean age: 47.3, SD±19.3, median: 46 [range: 18-95]; G2: mean age: 61.3, SD±13.7, median: 61 [range: 21-87]; G3: mean age: 38.8, SD±17.9, median: 34 [range: 18-80]) was statistically significant (G1 vs.G2: p-value <0.0001; t=7.21; DF=318; G1 vs.G3: p-value <0.0001; t=4.32; DF=318; G2 vs. G3: p-value <0.0001; t=9.62; DF=318). The Nagelkerke r 2 value was 0.040. There were statistically significant differences for the CCR5/CCR5 (p-value=0.008; OR=0.261), AA (p-value=0.007; OR=2.974) and AG genotypes (p-value=0.018; OR=2.447) between G1 and G2. Individuals with the CCR5/CCR5 genotype and simultaneously the CCR5-59029 AA or AG genotypes have a greater risk of developing ocular toxoplasmosis (4% greater), which may be associated with a strong and persistent inflammatory response in ocular tissue. Copyright © 2017 Elsevier B.V. All rights reserved.

Human Beta Defensin 2 Selectively Inhibits HIV-1 in Highly Permissive CCR6⁺CD4⁺ T Cells.

PubMed

Lafferty, Mark K; Sun, Lingling; Christensen-Quick, Aaron; Lu, Wuyuan; Garzino-Demo, Alfredo

2017-05-16

Chemokine receptor type 6 (CCR6)⁺CD4⁺ T cells are preferentially infected and depleted during HIV disease progression, but are preserved in non-progressors. CCR6 is expressed on a heterogeneous population of memory CD4⁺ T cells that are critical to mucosal immunity. Preferential infection of these cells is associated, in part, with high surface expression of CCR5, CXCR4, and α4β7. In addition, CCR6⁺CD4⁺ T cells harbor elevated levels of integrated viral DNA and high levels of proliferation markers. We have previously shown that the CCR6 ligands MIP-3α and human beta defensins inhibit HIV replication. The inhibition required CCR6 and the induction of APOBEC3G. Here, we further characterize the induction of apolipoprotein B mRNA editing enzyme (APOBEC3G) by human beta defensin 2. Human beta defensin 2 rapidly induces transcriptional induction of APOBEC3G that involves extracellular signal-regulated kinases 1/2 (ERK1/2) activation and the transcription factors NFATc2, NFATc1, and IRF4. We demonstrate that human beta defensin 2 selectively protects primary CCR6⁺CD4⁺ T cells infected with HIV-1. The selective protection of CCR6⁺CD4⁺ T cell subsets may be critical in maintaining mucosal immune function and preventing disease progression.

On the etiology of tropical spastic paraparesis and human T-cell lymphotropic virus-I-associated myelopathy.

PubMed

Zaninovic, V

1999-01-01

The purpose of this review is to present some concepts on the etiology of tropical spastic paraparesis or human T-cell lymphotropic virus-I (HTLV-I)-associated myelopathy (TSP/HAM). The large number of syndromes that have been associated with HTLV-I (60 to date), the existence of TSP/HAM cases associated with other retroviruses (human immunodeficiency virus-2 [HIV-2], HTLV-II), the existence of many TSPs without HTLV-I, and the evidence of clear epidemiologic contradictions in TSP/HAM indicate that the etiopathogenesis of TSP/HAM is not yet clear. Tropical spastic paraparesis/HAM affects patients of all human ethnic groups, but usually in well localized and relatively isolated geographic regions where HTLV-I has been endemic for a long time. Environmental factors and geographic locations appear to be critical factors. Because the neuropathology of TSP/HAM suggests a toxometabolic, rather than a viral cause, it is proposed that an intoxication similar to neurolathyrism could account for some of TSP/HAM cases, mainly in tropical and subtropical countries. If this were the case, HTLV-I could be a cofactor or act as a bystander. it is possible that co-infection with another agent is necessary to produce TSP/HAM and most of the syndromes associated with HTLV-I.

Cytokines Elevated in HIV Elite Controllers Reduce HIV Replication In Vitro and Modulate HIV Restriction Factor Expression

PubMed Central

Jacobs, Evan S.; Abdel-Mohsen, Mohamed; Gibb, Stuart L.; Heitman, John W.; Inglis, Heather C.; Martin, Jeffrey N.; Zhang, Jinbing; Kaidarova, Zhanna; Deng, Xutao; Wu, Shiquan; Anastos, Kathryn; Crystal, Howard; Villacres, Maria C.; Young, Mary; Greenblatt, Ruth M.; Landay, Alan L.; Gange, Stephen J.; Deeks, Steven G.; Golub, Elizabeth T.; Pillai, Satish K.

2017-01-01

ABSTRACT A subset of HIV-infected individuals termed elite controllers (ECs) maintain CD4+ T cell counts and control viral replication in the absence of antiretroviral therapy (ART). Systemic cytokine responses may differentiate ECs from subjects with uncontrolled viral replication or from those who require ART to suppress viral replication. We measured 87 cytokines in four groups of women: 73 ECs, 42 with pharmacologically suppressed viremia (ART), 42 with uncontrolled viral replication (noncontrollers [NCs]), and 48 HIV-uninfected (NEG) subjects. Four cytokines were elevated in ECs but not NCs or ART subjects: CCL14, CCL21, CCL27, and XCL1. In addition, median stromal cell-derived factor-1 (SDF-1) levels were 43% higher in ECs than in NCs. The combination of the five cytokines suppressed R5 and X4 virus replication in resting CD4+ T cells, and individually SDF-1β, CCL14, and CCL27 suppressed R5 virus replication, while SDF-1β, CCL21, and CCL14 suppressed X4 virus replication. Functional studies revealed that the combination of the five cytokines upregulated CD69 and CCR5 and downregulated CXCR4 and CCR7 on CD4+ T cells. The CD69 and CXCR4 effects were driven by SDF-1, while CCL21 downregulated CCR7. The combination of the EC-associated cytokines induced expression of the anti-HIV host restriction factors IFITM1 and IFITM2 and suppressed expression of RNase L and SAMHD1. These results identify a set of cytokines that are elevated in ECs and define their effects on cellular activation, HIV coreceptor expression, and innate restriction factor expression. This cytokine pattern may be a signature characteristic of HIV-1 elite control, potentially important for HIV therapeutic and curative strategies. IMPORTANCE Approximately 1% of people infected with HIV control virus replication without taking antiviral medications. These subjects, termed elite controllers (ECs), are known to have stronger immune responses targeting HIV than the typical HIV-infected subject, but the exact mechanisms of how their immune responses control infection are not known. In this study, we identified five soluble immune signaling molecules (cytokines) in the blood that were higher in ECs than in subjects with typical chronic HIV infection. We demonstrated that these cytokines can activate CD4+ T cells, the target cells for HIV infection. Furthermore, these five EC-associated cytokines could change expression levels of intrinsic resistance factors, or molecules inside the target cell that fight HIV infection. This study is significant in that it identified cytokines elevated in subjects with a good immune response against HIV and defined potential mechanisms as to how these cytokines could induce resistance to the virus in target cells. PMID:28053103

Cytokines Elevated in HIV Elite Controllers Reduce HIV Replication In Vitro and Modulate HIV Restriction Factor Expression.

PubMed

Jacobs, Evan S; Keating, Sheila M; Abdel-Mohsen, Mohamed; Gibb, Stuart L; Heitman, John W; Inglis, Heather C; Martin, Jeffrey N; Zhang, Jinbing; Kaidarova, Zhanna; Deng, Xutao; Wu, Shiquan; Anastos, Kathryn; Crystal, Howard; Villacres, Maria C; Young, Mary; Greenblatt, Ruth M; Landay, Alan L; Gange, Stephen J; Deeks, Steven G; Golub, Elizabeth T; Pillai, Satish K; Norris, Philip J

2017-03-15

A subset of HIV-infected individuals termed elite controllers (ECs) maintain CD4 + T cell counts and control viral replication in the absence of antiretroviral therapy (ART). Systemic cytokine responses may differentiate ECs from subjects with uncontrolled viral replication or from those who require ART to suppress viral replication. We measured 87 cytokines in four groups of women: 73 ECs, 42 with pharmacologically suppressed viremia (ART), 42 with uncontrolled viral replication (noncontrollers [NCs]), and 48 HIV-uninfected (NEG) subjects. Four cytokines were elevated in ECs but not NCs or ART subjects: CCL14, CCL21, CCL27, and XCL1. In addition, median stromal cell-derived factor-1 (SDF-1) levels were 43% higher in ECs than in NCs. The combination of the five cytokines suppressed R5 and X4 virus replication in resting CD4 + T cells, and individually SDF-1β, CCL14, and CCL27 suppressed R5 virus replication, while SDF-1β, CCL21, and CCL14 suppressed X4 virus replication. Functional studies revealed that the combination of the five cytokines upregulated CD69 and CCR5 and downregulated CXCR4 and CCR7 on CD4 + T cells. The CD69 and CXCR4 effects were driven by SDF-1, while CCL21 downregulated CCR7. The combination of the EC-associated cytokines induced expression of the anti-HIV host restriction factors IFITM1 and IFITM2 and suppressed expression of RNase L and SAMHD1. These results identify a set of cytokines that are elevated in ECs and define their effects on cellular activation, HIV coreceptor expression, and innate restriction factor expression. This cytokine pattern may be a signature characteristic of HIV-1 elite control, potentially important for HIV therapeutic and curative strategies. IMPORTANCE Approximately 1% of people infected with HIV control virus replication without taking antiviral medications. These subjects, termed elite controllers (ECs), are known to have stronger immune responses targeting HIV than the typical HIV-infected subject, but the exact mechanisms of how their immune responses control infection are not known. In this study, we identified five soluble immune signaling molecules (cytokines) in the blood that were higher in ECs than in subjects with typical chronic HIV infection. We demonstrated that these cytokines can activate CD4 + T cells, the target cells for HIV infection. Furthermore, these five EC-associated cytokines could change expression levels of intrinsic resistance factors, or molecules inside the target cell that fight HIV infection. This study is significant in that it identified cytokines elevated in subjects with a good immune response against HIV and defined potential mechanisms as to how these cytokines could induce resistance to the virus in target cells. Copyright © 2017 American Society for Microbiology.

Purification and crystallization of dengue and West Nile virus NS2B–NS3 complexes

DOE Office of Scientific and Technical Information (OSTI.GOV)

D’Arcy, Allan, E-mail: allan.darcy@novartis.com; Chaillet, Maxime; Schiering, Nikolaus

Crystals of dengue serotype 2 and West Nile virus NS2B–NS3 protease complexes have been obtained and the crystals of both diffract to useful resolution. Sample homogeneity was essential for obtaining X-ray-quality crystals of the dengue protease. Controlled proteolysis produced a crystallizable fragment of the apo West Nile virus NS2B–NS3 and crystals were also obtained in the presence of a peptidic inhibitor. Both dengue and West Nile virus infections are an increasing risk to humans, not only in tropical and subtropical areas, but also in North America and parts of Europe. These viral infections are generally transmitted by mosquitoes, but maymore » also be tick-borne. Infection usually results in mild flu-like symptoms, but can also cause encephalitis and fatalities. Approximately 2799 severe West Nile virus cases were reported this year in the United States, resulting in 102 fatalities. With this alarming increase in the number of West Nile virus infections in western countries and the fact that dengue virus already affects millions of people per year in tropical and subtropical climates, there is a real need for effective medicines. A possible therapeutic target to combat these viruses is the protease, which is essential for virus replication. In order to provide structural information to help to guide a lead identification and optimization program, crystallizations of the NS2B–NS3 protease complexes from both dengue and West Nile viruses have been initiated. Crystals that diffract to high resolution, suitable for three-dimensional structure determinations, have been obtained.« less

A novel CCR2 antagonist inhibits atherogenesis in apoE deficient mice by achieving high receptor occupancy.

PubMed

Bot, Ilze; Ortiz Zacarías, Natalia V; de Witte, Wilhelmus E A; de Vries, Henk; van Santbrink, Peter J; van der Velden, Daniël; Kröner, Mara J; van der Berg, Dirk-Jan; Stamos, Dean; de Lange, Elizabeth C M; Kuiper, Johan; IJzerman, Adriaan P; Heitman, Laura H

2017-03-03

CC Chemokine Receptor 2 (CCR2) and its endogenous ligand CCL2 are involved in a number of diseases, including atherosclerosis. Several CCR2 antagonists have been developed as potential therapeutic agents, however their in vivo clinical efficacy was limited. In this report, we aimed to determine whether 15a, an antagonist with a long residence time on the human CCR2, is effective in inhibiting the development of atherosclerosis in a mouse disease model. First, radioligand binding assays were performed to determine affinity and binding kinetics of 15a on murine CCR2. To assess the in vivo efficacy, western-type diet fed apoE -/- mice were treated daily with 15a or vehicle as control. Treatment with 15a reduced the amount of circulating CCR2 + monocytes and the size of the atherosclerotic plaques in both the carotid artery and the aortic root. We then showed that the long pharmacokinetic half-life of 15a combined with the high drug concentrations ensured prolonged CCR2 occupancy. These data render 15a a promising compound for drug development and confirms high receptor occupancy as a key parameter when targeting chemokine receptors.

Effects of Therapy with Maraviroc on the Carotid Intima Media Thickness in HIV-1/HCV Co-infected Patients

PubMed Central

MAGGI, PAOLO; BRUNO, GIUSEPPE; PERILLI, FRANCESCO; SARACINO, ANNALISA; VOLPE, ANNA; SANTORO, CARMEN; LADISA, NICOLETTA; ANGARANO, GIOACCHINO

2017-01-01

Aim: To evaluate, in human immunodeficiency virus-hepatitis C virus co-infected patients, the impact of C-C chemokine receptor type 5 (CCR5) antagonist maraviroc-based antiretroviral therapy on the carotid intima media thickness and on atheromasic plaques. Patients and Methods: In this pilot prospective study, 12 HIV-HCV co-infected patients underwent color-Doppler ultrasonography before and 48 weeks after switching to a dual therapy based on maraviroc plus protease inhibitors boosted with ritonavir. Changes of intima media thickness, inflammatory and endothelial adhesion biomarkers levels, Veterans Aging Cohort Study index and Framingham risk score were evaluated. Results: At baseline 11 (91.6%) patients showed pathological ultrasonographic findings. After 48 weeks, two patients showed an amelioration of intima media thickness. Of the remaining patients with plaques, four showed a reduction of the previously diagnosed plaque; no patients worsened. Conclusion: Our data suggest that CCR5 inhibition could reduce the development of atherosclerosis especially in the non-calcific stage and could play an important role in the blockade of atheromasic plaque progression. PMID:28064231

Effects of Therapy with Maraviroc on the Carotid Intima Media Thickness in HIV-1/HCV Co-infected Patients.

PubMed

Maggi, Paolo; Bruno, Giuseppe; Perilli, Francesco; Saracino, Annalisa; Volpe, Anna; Santoro, Carmen; Ladisa, Nicoletta; Angarano, Gioacchino

2017-01-02

To evaluate, in human immunodeficiency virus-hepatitis C virus co-infected patients, the impact of C-C chemokine receptor type 5 (CCR5) antagonist maraviroc-based antiretroviral therapy on the carotid intima media thickness and on atheromasic plaques. In this pilot prospective study, 12 HIV-HCV co-infected patients underwent color-Doppler ultrasonography before and 48 weeks after switching to a dual therapy based on maraviroc plus protease inhibitors boosted with ritonavir. Changes of intima media thickness, inflammatory and endothelial adhesion biomarkers levels, Veterans Aging Cohort Study index and Framingham risk score were evaluated. At baseline 11 (91.6%) patients showed pathological ultrasonographic findings. After 48 weeks, two patients showed an amelioration of intima media thickness. Of the remaining patients with plaques, four showed a reduction of the previously diagnosed plaque; no patients worsened. Our data suggest that CCR5 inhibition could reduce the development of atherosclerosis especially in the non-calcific stage and could play an important role in the blockade of atheromasic plaque progression. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

CCR2 deficiency leads to increased eosinophils, alternative macrophage activation, and type 2 cytokine expression in adipose tissue.

PubMed

Bolus, W Reid; Gutierrez, Dario A; Kennedy, Arion J; Anderson-Baucum, Emily K; Hasty, Alyssa H

2015-10-01

Adipose tissue (AT) inflammation during obesity is mediated by immune cells and closely correlates with systemic insulin resistance. In lean AT, eosinophils are present in low but significant numbers and capable of promoting alternative macrophage activation in an IL-4/IL-13-dependent manner. In WT mice, obesity causes the proportion of AT eosinophils to decline, concomitant with inflammation and classical activation of AT macrophages. In this study, we show that CCR2 deficiency leads to increased eosinophil accumulation in AT. Furthermore, in contrast to WT mice, the increase in eosinophils in CCR2(-/-) AT is sustained and even amplified during obesity. Interestingly, a significant portion of eosinophils is found in CLSs in AT of obese CCR2(-/-) mice, which is the first time eosinophils have been shown to localize to these inflammatory hot spots. CCR2(-/-) bone marrow precursors displayed increased expression of various key eosinophil genes during in vitro differentiation to eosinophils, suggesting a potentially altered eosinophil phenotype in the absence of CCR2. In addition, the proportion of eosinophils in AT positively correlated with local expression of Il5, a potent eosinophil stimulator. The increase in eosinophils in CCR2(-/-) mice was detected in all white fat pads analyzed and in the peritoneal cavity but not in bone marrow, blood, spleen, or liver. In AT of CCR2(-/-) mice, an increased eosinophil number positively correlated with M2-like macrophages, expression of the Treg marker Foxp3, and type 2 cytokines, Il4, Il5, and Il13. This is the first study to link CCR2 function with regulation of AT eosinophil accumulation. © Society for Leukocyte Biology.

CCR2 deficiency leads to increased eosinophils, alternative macrophage activation, and type 2 cytokine expression in adipose tissue

PubMed Central

Bolus, W. Reid; Gutierrez, Dario A.; Kennedy, Arion J.; Anderson-Baucum, Emily K.; Hasty, Alyssa H.

2015-01-01

Adipose tissue (AT) inflammation during obesity is mediated by immune cells and closely correlates with systemic insulin resistance. In lean AT, eosinophils are present in low but significant numbers and capable of promoting alternative macrophage activation in an IL-4/IL-13-dependent manner. In WT mice, obesity causes the proportion of AT eosinophils to decline, concomitant with inflammation and classical activation of AT macrophages. In this study, we show that CCR2 deficiency leads to increased eosinophil accumulation in AT. Furthermore, in contrast to WT mice, the increase in eosinophils in CCR2−/− AT is sustained and even amplified during obesity. Interestingly, a significant portion of eosinophils is found in CLSs in AT of obese CCR2−/− mice, which is the first time eosinophils have been shown to localize to these inflammatory hot spots. CCR2−/− bone marrow precursors displayed increased expression of various key eosinophil genes during in vitro differentiation to eosinophils, suggesting a potentially altered eosinophil phenotype in the absence of CCR2. In addition, the proportion of eosinophils in AT positively correlated with local expression of Il5, a potent eosinophil stimulator. The increase in eosinophils in CCR2−/− mice was detected in all white fat pads analyzed and in the peritoneal cavity but not in bone marrow, blood, spleen, or liver. In AT of CCR2−/− mice, an increased eosinophil number positively correlated with M2-like macrophages, expression of the Treg marker Foxp3, and type 2 cytokines, Il4, Il5, and Il13. This is the first study to link CCR2 function with regulation of AT eosinophil accumulation. PMID:25934927

Immune response CC chemokines CCL2 and CCL5 are associated with pulmonary sarcoidosis

PubMed Central

2011-01-01

Background Pulmonary sarcoidosis involves an intense leukocyte infiltration of the lung with the formation of non-necrotizing granulomas. CC chemokines (chemokine (C-C motif) ligand 2 (CCL2)-CCL5) are chemoattractants of mononuclear cells and act through seven transmembrane G-coupled receptors. Previous studies have demonstrated conflicting results with regard to the associations of these chemokines with sarcoidosis. In an effort to clarify previous discrepancies, we performed the largest observational study to date of CC chemokines in bronchoalveolar lavage fluid (BALF) from patients with pulmonary sarcoidosis. Results BALF chemokine levels from 72 patients affected by pulmonary sarcoidosis were analyzed by enzyme-linked immunosorbent assay (ELISA) and compared to 8 healthy volunteers. BALF CCL3 and CCL4 levels from pulmonary sarcoidosis patients were not increased compared to controls. However, CCL2 and CCL5 levels were elevated, and subgroup analysis showed higher levels of both chemokines in all stages of pulmonary sarcoidosis. CCL2, CCL5, CC chemokine receptor type 1 (CCR1), CCR2 and CCR3 were expressed from mononuclear cells forming the lung granulomas, while CCR5 was only found on mast cells. Conclusions These data suggest that CCL2 and CCL5 are important mediators in recruiting CCR1, CCR2, and CCR3 expressing mononuclear cells as well as CCR5-expressing mast cells during all stages of pulmonary sarcoidosis. PMID:21463523

Immune response CC chemokines CCL2 and CCL5 are associated with pulmonary sarcoidosis.

PubMed

Palchevskiy, Vyacheslav; Hashemi, Nastran; Weigt, Stephen S; Xue, Ying Ying; Derhovanessian, Ariss; Keane, Michael P; Strieter, Robert M; Fishbein, Michael C; Deng, Jane C; Lynch, Joseph P; Elashoff, Robert; Belperio, John A

2011-04-04

Pulmonary sarcoidosis involves an intense leukocyte infiltration of the lung with the formation of non-necrotizing granulomas. CC chemokines (chemokine (C-C motif) ligand 2 (CCL2)-CCL5) are chemoattractants of mononuclear cells and act through seven transmembrane G-coupled receptors. Previous studies have demonstrated conflicting results with regard to the associations of these chemokines with sarcoidosis. In an effort to clarify previous discrepancies, we performed the largest observational study to date of CC chemokines in bronchoalveolar lavage fluid (BALF) from patients with pulmonary sarcoidosis. BALF chemokine levels from 72 patients affected by pulmonary sarcoidosis were analyzed by enzyme-linked immunosorbent assay (ELISA) and compared to 8 healthy volunteers. BALF CCL3 and CCL4 levels from pulmonary sarcoidosis patients were not increased compared to controls. However, CCL2 and CCL5 levels were elevated, and subgroup analysis showed higher levels of both chemokines in all stages of pulmonary sarcoidosis. CCL2, CCL5, CC chemokine receptor type 1 (CCR1), CCR2 and CCR3 were expressed from mononuclear cells forming the lung granulomas, while CCR5 was only found on mast cells. These data suggest that CCL2 and CCL5 are important mediators in recruiting CCR1, CCR2, and CCR3 expressing mononuclear cells as well as CCR5-expressing mast cells during all stages of pulmonary sarcoidosis.

Characterization of Developmental- and Stress-Mediated Expression of Cinnamoyl-CoA Reductase in Kenaf (Hibiscus cannabinus L.)

PubMed Central

Lim, Hyoun-Sub; Park, Sang-Un; Bae, Hyeun-Jong; Natarajan, Savithiry

2014-01-01

Cinnamoyl-CoA reductase (CCR) is an important enzyme for lignin biosynthesis as it catalyzes the first specific committed step in monolignol biosynthesis. We have cloned a full length coding sequence of CCR from kenaf (Hibiscus cannabinus L.), which contains a 1,020-bp open reading frame (ORF), encoding 339 amino acids of 37.37 kDa, with an isoelectric point (pI) of 6.27 (JX524276, HcCCR2). BLAST result found that it has high homology with other plant CCR orthologs. Multiple alignment with other plant CCR sequences showed that it contains two highly conserved motifs: NAD(P) binding domain (VTGAGGFIASWMVKLLLEKGY) at N-terminal and probable catalytic domain (NWYCYGK). According to phylogenetic analysis, it was closely related to CCR sequences of Gossypium hirsutum (ACQ59094) and Populus trichocarpa (CAC07424). HcCCR2 showed ubiquitous expression in various kenaf tissues and the highest expression was detected in mature flower. HcCCR2 was expressed differentially in response to various stresses, and the highest expression was observed by drought and NaCl treatments. PMID:24723816

A Quantitative Comparison of Anti-HIV Gene Therapy Delivered to Hematopoietic Stem Cells versus CD4+ T Cells

PubMed Central

Savkovic, Borislav; Nichols, James; Birkett, Donald; Applegate, Tanya; Ledger, Scott; Symonds, Geoff; Murray, John M.

2014-01-01

Gene therapy represents an alternative and promising anti-HIV modality to highly active antiretroviral therapy. It involves the introduction of a protective gene into a cell, thereby conferring protection against HIV. While clinical trials to date have delivered gene therapy to CD4+T cells or to CD34+ hematopoietic stem cells (HSC), the relative benefits of each of these two cellular targets have not been conclusively determined. In the present analysis, we investigated the relative merits of delivering a dual construct (CCR5 entry inhibitor + C46 fusion inhibitor) to either CD4+T cells or to CD34+ HSC. Using mathematical modelling, we determined the impact of each scenario in terms of total CD4+T cell counts over a 10 year period, and also in terms of inhibition of CCR5 and CXCR4 tropic virus. Our modelling determined that therapy delivery to CD34+ HSC generally resulted in better outcomes than delivery to CD4+T cells. An early one-off therapy delivery to CD34+ HSC, assuming that 20% of CD34+ HSC in the bone marrow were gene-modified (G+), resulted in total CD4+T cell counts ≥180 cells/ µL in peripheral blood after 10 years. If the uninfected G+ CD4+T cells (in addition to exhibiting lower likelihood of becoming productively infected) also exhibited reduced levels of bystander apoptosis (92.5% reduction) over non gene-modified (G-) CD4+T cells, then total CD4+T cell counts of ≥350 cells/ µL were observed after 10 years, even if initially only 10% of CD34+ HSC in the bone marrow received the protective gene. Taken together our results indicate that: 1.) therapy delivery to CD34+ HSC will result in better outcomes than delivery to CD4+T cells, and 2.) a greater impact of gene therapy will be observed if G+ CD4+T cells exhibit reduced levels of bystander apoptosis over G- CD4+T cells. PMID:24945407

CCL17 and CCL22/CCR4 signaling is a strong candidate for novel targeted therapy against nasal natural killer/T-cell lymphoma

PubMed Central

Kumai, Takumi; Kobayashi, Hiroya; Komabayashi, Yuki; Ueda, Seigo; Kishibe, Kan; Ohkuri, Takayuki; Takahara, Miki; Celis, Esteban; Harabuchi, Yasuaki

2015-01-01

Nasal natural killer/T-cell lymphoma (NNKTL) is associated with Epstein–Barr virus and has a poor prognosis because of local invasion and/or multiple dissemination. Various chemokines play a role in tumor proliferation and invasion, and chemokine receptors including the C-C chemokine receptor 4 (CCR4) are recognized as potential targets for treating hematologic malignancies. The aim of the present study was to determine whether specific chemokines are produced by NNKTL. We compared chemokine expression patterns in culture supernatants of NNKTL cell lines with those of other lymphoma or leukemia cell lines using chemokine protein array and ELISA. Chemokine (C-C motif) ligand (CCL) 17 and CCL22 were highly produced by NNKTL cell lines as compared to the other cell lines. In addition, CCL17 and CCL22 were readily observed in the sera of NNKTL patients. The levels of these chemokines were significantly higher in patients than in healthy controls. Furthermore, we detected the expression of CCR4 (the receptor for CCL17 and CCL22) on the surface of NNKTL cell lines and in tissues of NNKTL patients. Anti-CCR4 monoclonal antibody (mAb) efficiently induced antibody-dependent cellular cytotoxicity mediated by natural killer cells against NNKTL cell lines. Our results suggest that CCL17 and CCL22 may be important factors in the development of NNKTL and open up the possibility of immunotherapy of this lymphoma using anti-CCR4 mAb. PMID:25754123

Polymorphisms in chemokine and receptor genes and gastric cancer risk and survival in a high risk Polish population.

PubMed

Gawron, Andrew J; Fought, Angela J; Lissowska, Jolanta; Ye, Weimin; Zhang, Xiao; Chow, Wong-Ho; Beane Freeman, Laura E; Hou, Lifang

2011-03-01

To examine if genetic variations in chemokine receptor and ligand genes are associated with gastric cancer risk and survival. The study included 298 cases and 417 controls from a population-based study of gastric cancer conducted in Warsaw, Poland in 1994-1996. We investigated seven single nucleotide polymorphisms in a chemokine ligand (CXCL12) and chemokine receptor (CCR2, CCR5, CX3CR1) genes and one frameshift deletion (CCR5) in blood leukocyte DNA in relation to gastric cancer risk and survival. Genotyping was conducted at the NCI Core Genotyping Facility. Odds ratios and 95% confidence intervals were computed using univariate and multivariate logistic regression models. Survival analysis was performed using Cox proportional hazards models. Gastric cancer risk was not associated with single chemokine polymorphisms. A CCR5 haplotype that contained the common alleles of IVS1+151 G>T (rs2734648), IVS2+80 C>T (rs1800024) and minor allele of IVS1+246 A>G (rs1799987) was associated with a borderline significantly increased risk (OR = 1.5, 95% CI: 1.0?2.2). For gastric cancer cases, there was a greater risk of death for carriers of the minor alleles of CCR2 Ex2+241 G>A (rs1799864) (HR = 1.5, 95% CI: 1.1-2.1) and CCR5 IVS2+80 C>T (rs1800024) (HR = 1.5, 95% CI: 1.1-2.1). Carriers of the CCR5 minor allele of IVS1+151 G>T (rs2734648) had a decreased risk of death compared to homozygote carriers of the common allele (HR = 0.8, 95% CI: 0.6-1.0). Our findings do not support an association between gastric cancer risk and single chemokine genetic variation. The observed associations between cancer risk and a CCR5 haplotype and between survival and polymorphisms in CCR2 and CCR5 need replication in future studies.

Sensitive cell-based assay for determination of human immunodeficiency virus type 1 coreceptor tropism.

PubMed

Weber, Jan; Vazquez, Ana C; Winner, Dane; Gibson, Richard M; Rhea, Ariel M; Rose, Justine D; Wylie, Doug; Henry, Kenneth; Wright, Alison; King, Kevin; Archer, John; Poveda, Eva; Soriano, Vicente; Robertson, David L; Olivo, Paul D; Arts, Eric J; Quiñones-Mateu, Miguel E

2013-05-01

CCR5 antagonists are a powerful new class of antiretroviral drugs that require a companion assay to evaluate the presence of CXCR4-tropic (non-R5) viruses prior to use in human immunodeficiency virus (HIV)-infected individuals. In this study, we have developed, characterized, verified, and prevalidated a novel phenotypic test to determine HIV-1 coreceptor tropism (VERITROP) based on a sensitive cell-to-cell fusion assay. A proprietary vector was constructed containing a near-full-length HIV-1 genome with the yeast uracil biosynthesis (URA3) gene replacing the HIV-1 env coding sequence. Patient-derived HIV-1 PCR products were introduced by homologous recombination using an innovative yeast-based cloning strategy. The env-expressing vectors were then used in a cell-to-cell fusion assay to determine the presence of R5 and/or non-R5 HIV-1 variants within the viral population. Results were compared with (i) the original version of Trofile (Monogram Biosciences, San Francisco, CA), (ii) population sequencing, and (iii) 454 pyrosequencing, with the genotypic data analyzed using several bioinformatics tools, i.e., the 11/24/25 rule, Geno2Pheno (2% to 5.75%, 3.5%, or 10% false-positive rate [FPR]), and webPSSM. VERITROP consistently detected minority non-R5 variants from clinical specimens, with an analytical sensitivity of 0.3%, with viral loads of ≥1,000 copies/ml, and from B and non-B subtypes. In a pilot study, a 73.7% (56/76) concordance was observed with the original Trofile assay, with 19 of the 20 discordant results corresponding to non-R5 variants detected using VERITROP and not by the original Trofile assay. The degree of concordance of VERITROP and Trofile with population and deep sequencing results depended on the algorithm used to determine HIV-1 coreceptor tropism. Overall, VERITROP showed better concordance with deep sequencing/Geno2Pheno at a 0.3% detection threshold (67%), whereas Trofile matched better with population sequencing (79%). However, 454 sequencing using Geno2Pheno at a 10% FPR and 0.3% threshold and VERITROP more accurately predicted the success of a maraviroc-based regimen. In conclusion, VERITROP may promote the development of new HIV coreceptor antagonists and aid in the treatment and management of HIV-infected individuals prior to and/or during treatment with this class of drugs.

Reduced toll-like receptor 4 and substance P gene expression is associated with airway bacterial colonization in children.

PubMed

Grissell, Terry V; Chang, Anne B; Gibson, Peter G

2007-04-01

Neuro-immune interactions are increasingly relevant to human health and disease. The neuropeptide Substance P also has antibacterial activity and bears similarities to the innate immune antibacterial defensins. This suggests possible co-regulation of neuropeptide and innate immune mediators. In this study, non-bronchoscopic bronchoalveolar lavage (BAL) was performed on 69 children. BAL was examined for cellular profile, microbiology (bacteria, virus) and gene expression for TLRs 2, 3, 4; chemokine receptors (CCR3, CCR5, CXCR1); neurotrophins and neurokinin genes (TAC1, TAC3, CGRP, NGF). In children with bacterial colonization (n=10) there was an airway inflammatory response with increased BAL neutrophils, IL-8 protein, and CXCR1 expression. Substance P (TAC1) and TLR4 RNA expression were reduced in children with bacterial colonization. TLR3 mRNA was increased in 7.2% (n=5) children with rhinovirus, and there was a non-significant trend to increased TLR2. There is evidence for co-regulation of neurokinin (TAC1) and TLR4 gene expression in airway cells from children with airway bacterial colonization and their reduced expression may be associated with an impaired bacterial clearance. (c) 2007 Wiley-Liss, Inc.

Harnessing Big Data for Communicable Tropical and Sub-Tropical Disorders: Implications From a Systematic Review of the Literature.

PubMed

Gianfredi, Vincenza; Bragazzi, Nicola Luigi; Nucci, Daniele; Martini, Mariano; Rosselli, Roberto; Minelli, Liliana; Moretti, Massimo

2018-01-01

According to the World Health Organization (WHO), communicable tropical and sub-tropical diseases occur solely, or mainly in the tropics, thriving in hot, and humid conditions. Some of these disorders termed as neglected tropical diseases are particularly overlooked. Communicable tropical/sub-tropical diseases represent a diverse group of communicable disorders occurring in 149 countries, favored by tropical and sub-tropical conditions, affecting more than one billion people and imposing a dramatic societal and economic burden. A systematic review of the extant scholarly literature was carried out, searching in PubMed/MEDLINE and Scopus. The search string used included proper keywords, like big data, nontraditional data sources, social media, social networks, infodemiology, infoveillance, novel data streams (NDS), digital epidemiology, digital behavior, Google Trends, Twitter, Facebook, YouTube, Instagram, Pinterest, Ebola, Zika, dengue, Chikungunya, Chagas, and the other neglected tropical diseases. 47 original, observational studies were included in the current systematic review: 1 focused on Chikungunya, 6 on dengue, 19 on Ebola, 2 on Malaria, 1 on Mayaro virus, 2 on West Nile virus, and 16 on Zika. Fifteen were dedicated on developing and validating forecasting techniques for real-time monitoring of neglected tropical diseases, while the remaining studies investigated public reaction to infectious outbreaks. Most studies explored a single nontraditional data source, with Twitter being the most exploited tool (25 studies). Even though some studies have shown the feasibility of utilizing NDS as an effective tool for predicting epidemic outbreaks and disseminating accurate, high-quality information concerning neglected tropical diseases, some gaps should be properly underlined. Out of the 47 articles included, only 7 were focusing on neglected tropical diseases, while all the other covered communicable tropical/sub-tropical diseases, and the main determinant of this unbalanced coverage seems to be the media impact and resonance. Furthermore, efforts in integrating diverse NDS should be made. As such, taking into account these limitations, further research in the field is needed.

Lack of CCR5 modifies glial phenotypes and population of the nigral dopaminergic neurons, but not MPTP-induced dopaminergic neurodegeneration.

PubMed

Choi, Dong-Young; Lee, Myung Koo; Hong, Jin Tae

2013-01-01

Constitutive expression of C-C chemokine receptor (CCR) 5 has been detected in astrocytes, microglia and neurons, but its physiological roles in the central nervous system are obscure. The bidirectional interactions between neuron and glial cells through CCR5 and its ligands were thought to be crucial for maintaining normal neuronal activities. No study has described function of CCR5 in the dopaminergic neurodegeneration in Parkinson's disease. In order to examine effects of CCR5 on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopaminergic neurodegeneration, we employed CCR5 wild type (WT) and knockout (KO) mice. Immunostainings for tyrosine hydroxylase (TH) exhibited that CCR5 KO mice had lower number of TH-positive neurons even in the absence of MPTP. Difference in MPTP (15mg/kg×4 times, 2hr interval)-mediated loss of TH-positive neurons was subtle between CCR5 WT and KO mice, but there was larger dopamine depletion, behavioral impairments and microglial activation in CCR5 deficient mice. Intriguingly, CCR5 KO brains contained higher immunoreactivity for monoamine oxidase (MAO) B which was mainly localized within astrocytes. In agreement with upregulation of MAO B, concentration of MPP+ was higher in the substantia nigra and striatum of CCR5 KO mice after MPTP injection. We found remarkable activation of p38 MAPK in CCR5 deficient mice, which positively regulates MAO B expression. These results indicate that CCR5 deficiency modifies the nigrostriatal dopaminergic neuronal system and bidirectional interaction between neurons and glial cells via CCR5 might be important for dopaminergic neuronal survival. Copyright © 2012 Elsevier Inc. All rights reserved.

[CCR5, CCR2, apoe, p53, ITGB3 and HFE gene polymorphism in Western Siberia long-livers].

PubMed

Ivanoshchuk, D E; Mikhaĭlova, S V; Kulikov, I V; Maksimov, V N; Voevoda, M I; Romashchenko, A G

2012-01-01

In order to estimate the distribution of some polymorphisms for the CCR5, CCR2, apoE, p53, ITGB3, and HFE genes in Russian long-livers from Western Siberia, a sample of 271 individuals (range 90-105 years) was examined. It was demonstrated that carriage of the delta32 polymorphism for the CCR5 gene, V64/polymorphism for the CCR2 gene, e2/e3/e4 for the apoE gene, L33P for the ITGB3 gene, as well as H63D and S65C polymorphisms for the HFE gene does not influence on predisposition to the longevity; carriage of the 282 Y allele for the HFE gene negatively influences on the longevity; carriage of the heterozygous genotype for the R72P polymorphism for the p53 gene correlates with the longevity of elderly people.

Analysis of monocyte infiltration in MPTP mice reveals that microglial CX3CR1 protects against neurotoxic over-induction of monocyte-attracting CCL2 by astrocytes.

PubMed

Parillaud, Vincent R; Lornet, Guillaume; Monnet, Yann; Privat, Anne-Laure; Haddad, Andrei T; Brochard, Vanessa; Bekaert, Amaury; de Chanville, Camille Baudesson; Hirsch, Etienne C; Combadière, Christophe; Hunot, Stéphane; Lobsiger, Christian S

2017-03-21

Evidence from mice suggests that brain infiltrating immune cells contribute to neurodegeneration, and we previously identified a deleterious lymphocyte infiltration in Parkinson's disease mice. However, this remains controversial for monocytes, due to artifact-prone techniques used to distinguish them from microglia. Our aim was to reassess this open question, by taking advantage of the recent recognition that chemokine receptors CCR2 and CX3CR1 can differentiate between inflammatory monocytes and microglia, enabling to test whether CCR2 + monocytes infiltrate the brain during dopaminergic (DA) neurodegeneration and whether they contribute to neuronal death. This revealed unexpected insights into possible regulation of monocyte-attracting CCL2 induction. We used acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mice and assessed monocyte infiltration by combining laser microdissection-guided chemokine RNA profiling of the substantia nigra (SN) with immunohistochemistry and CCR2-GFP reporter mice. To determine contribution to neuronal loss, we used CCR2-deletion and CCL2-overexpression, to reduce and increase CCR2 + monocyte infiltration, and CX3CR1-deletion to assess a potential implication in CCL2 regulation. Nigral chemokine profiling revealed early CCL2/7/12-CCR2 axis induction, suggesting monocyte infiltration in MPTP mice. CCL2 protein showed early peak induction in nigral astrocytes, while CCR2-GFP mice revealed early but limited nigral monocyte infiltration. However, blocking infiltration by CCR2 deletion did not influence DA neuronal loss. In contrast, transgenic astrocytic CCL2 over-induction increased CCR2 + monocyte infiltration and DA neuronal loss in MPTP mice. Surprisingly, CCL2 over-induction was also detected in MPTP intoxicated CX3CR1-deleted mice, which are known to present increased DA neuronal loss. Importantly, CX3CR1/CCL2 double-deletion suggested that increased neurotoxicity was driven by astrocytic CCL2 over-induction. We show that CCR2 + monocytes infiltrate the affected CNS, but at the level observed in acute MPTP mice, this does not contribute to DA neuronal loss. In contrast, the underlying astrocytic CCL2 induction seemed to be tightly controled, as already moderate CCL2 over-induction led to increased neurotoxicity in MPTP mice, likely due to the increased CCR2 + monocyte infiltration. Importantly, we found evidence suggesting that during DA neurodegeneration, this control was mediated by microglial CX3CR1 signaling, which protects against such neurotoxic CCL2 over-induction by astrocytes, thus hinting at an endogenous mechanism to limit neurotoxic effects of the CCL2-CCR2 axis.

Generation and genetic engineering of human induced pluripotent stem cells using designed zinc finger nucleases.

PubMed

Ramalingam, Sivaprakash; London, Viktoriya; Kandavelou, Karthikeyan; Cebotaru, Liudmila; Guggino, William; Civin, Curt; Chandrasegaran, Srinivasan

2013-02-15

Zinc finger nucleases (ZFNs) have become powerful tools to deliver a targeted double-strand break at a pre-determined chromosomal locus in order to insert an exogenous transgene by homology-directed repair. ZFN-mediated gene targeting was used to generate both single-allele chemokine (C-C motif) receptor 5 (CCR5)-modified human induced pluripotent stem cells (hiPSCs) and biallele CCR5-modified hiPSCs from human lung fibroblasts (IMR90 cells) and human primary cord blood mononuclear cells (CBMNCs) by site-specific insertion of stem cell transcription factor genes flanked by LoxP sites into the endogenous CCR5 locus. The Oct4 and Sox2 reprogramming factors, in combination with valproic acid, induced reprogramming of human lung fibroblasts to form CCR5-modified hiPSCs, while 5 factors, Oct4/Sox2/Klf4/Lin28/Nanog, induced reprogramming of CBMNCs. Subsequent Cre recombinase treatment of the CCR5-modified IMR90 hiPSCs resulted in the removal of the Oct4 and Sox2 transgenes. Further genetic engineering of the single-allele CCR5-modified IMR90 hiPSCs was achieved by site-specific addition of the large CFTR transcription unit to the remaining CCR5 wild-type allele, using CCR5-specific ZFNs and a donor construct containing tdTomato and CFTR transgenes flanked by CCR5 homology arms. CFTR was expressed efficiently from the endogenous CCR5 locus of the CCR5-modified tdTomato/CFTR hiPSCs. These results suggest that it might be feasible to use ZFN-evoked strategies to (1) generate precisely targeted genetically well-defined patient-specific hiPSCs, and (2) then to reshape their function by targeted addition and expression of therapeutic genes from the CCR5 chromosomal locus for autologous cell-based transgene-correction therapy to treat various recessive monogenic human diseases in the future.

Particulate Air pollution mediated effects on insulin resistance in mice are independent of CCR2.

PubMed

Liu, Cuiqing; Xu, Xiaohua; Bai, Yuntao; Zhong, Jixin; Wang, Aixia; Sun, Lixian; Kong, Liya; Ying, Zhekang; Sun, Qinghua; Rajagopalan, Sanjay

2017-03-03

Chronic exposure to fine ambient particulate matter (PM 2.5 ) induces insulin resistance. CC-chemokine receptor 2 (CCR2) appears to be essential in diet-induced insulin resistance implicating an important role for systemic cellular inflammation in the process. We have previously suggested that CCR2 is important in PM 2.5 exposure-mediated inflammation leading to insulin resistance under high fat diet situation. The present study assessed the importance of CCR2 in PM 2.5 exposure-induced insulin resistance in the context of normal diet. C57BL/6 and CCR2 -/- mice were subjected to exposure to concentrated ambient PM 2.5 or filtered air for 6 months. In C57BL/6 mice, concentrated ambient PM 2.5 exposure induced whole-body insulin resistance, macrophage infiltration into the adipose tissue, and upregulation of phosphoenolpyruvate carboxykinase (PEPCK) in the liver. While CCR2 deficiency reduced adipose macrophage content in the PM 2.5 -exposed animals, it did not improve systemic insulin resistance. This lack of improvement in insulin resistance was paralleled by increased hepatic expression of genes in PEPCK and inflammation. CCR2 deletion failed to attenuate PM 2.5 exposure-induced insulin resistance in mice fed on normal diet. The present study indicates that PM 2.5 may dysregulate glucose metabolism directly without exerting proinflammatory effects.

CCR6(+) Th cell populations distinguish ACPA positive from ACPA negative rheumatoid arthritis.

PubMed

Paulissen, Sandra M J; van Hamburg, Jan Piet; Davelaar, Nadine; Vroman, Heleen; Hazes, Johanna M W; de Jong, Pascal H P; Lubberts, Erik

2015-11-30

Patients with rheumatoid arthritis (RA) can be separated into two major subpopulations based on the absence or presence of serum anti-citrullinated protein antibodies (ACPAs). The more severe disease course in ACPA(+) RA and differences in treatment outcome between these subpopulations suggest that ACPA(+) and ACPA(-) RA are different disease subsets. The identification of T-helper (Th) cells specifically recognizing citrullinated peptides, combined with the strong association between HLA-DRB1 and ACPA positivity, point toward a pathogenic role of Th cells in ACPA(+) RA. In this context we recently identified a potential pathogenic role for CCR6(+) Th cells in RA. Therefore, we examined whether Th cell population distributions differ by ACPA status. We performed a nested matched case-control study including 27 ACPA(+) and 27 ACPA(-) treatment-naive early RA patients matched for disease activity score in 44 joints, presence of rheumatoid factor, sex, age, duration of complaints and presence of erosions. CD4(+)CD45RO(+) (memory) Th cell distribution profiles from these patients were generated based on differential chemokine receptor expression and related with disease duration. ACPA status was not related to differences in total CD4(+) T cell or memory Th cell proportions. However, ACPA(+) patients had significantly higher proportions of Th cells expressing the chemokine receptors CCR6 and CXCR3. Similar proportions of CCR4(+) and CCR10(+) Th cells were found. Within the CCR6(+) cell population, four Th subpopulations were distinguished based on differential chemokine receptor expression: Th17 (CCR4(+)CCR10(-)), Th17.1 (CXCR3(+)), Th22 (CCR4(+)CCR10(+)) and CCR4/CXCR3 double-positive (DP) cells. In particular, higher proportions of Th22 (p = 0.02), Th17.1 (p = 0.03) and CCR4/CXCR3 DP (p = 0.01) cells were present in ACPA(+) patients. In contrast, ACPA status was not associated with differences in Th1 (CCR6(-)CXCR3(+); p = 0.90), Th2 (CCR6(-)CCR4(+); p = 0.27) and T-regulatory (CD25(hi)FOXP3(+); p = 0.06) cell proportions. Interestingly, CCR6(+) Th cells were inversely correlated with disease duration in ACPA(-) patients (R(2) = -0.35; p < 0.01) but not in ACPA(+) (R(2) < 0.01; p = 0.94) patients. These findings demonstrate that increased peripheral blood CCR6(+) Th cells proportions distinguish ACPA(+) RA from ACPA(-) RA. This suggests that CCR6(+) Th cells are involved in the differences in disease severity and treatment outcome between ACPA(+) and ACPA(-) RA.

Persistent Simian Immunodeficiency Virus Infection Drives Differentiation, Aberrant Accumulation, and Latent Infection of Germinal Center Follicular T Helper Cells

PubMed Central

Xu, Huanbin; Wang, Xiaolei; Malam, Naomi; Aye, Pyone P.; Alvarez, Xavier; Lackner, Andrew A.

2015-01-01

ABSTRACT CD4+ follicular T helper (Tfh) cells play a prominent role in humoral immune responses, but the mechanisms of their accumulation and infection in AIDS remain unclear. Here we found that germinal center (GC) Tfh cells, defined here as CXCR5+ PD-1HIGH CD4+ T cells, do not express the HIV coreceptor CCR5 yet serve as a latent reservoir in GCs. With disease progression, an expansion of GC Tfh cells is accompanied by increases in dysfunctional CD8+ T cells. In contrast, Tfh precursor (CXCR5− CD4+ T) cells in lymph nodes do express CCR5 and differentiate into GC Tfh cells following interleukin-6 (IL-6) and IL-21 stimulation, and viral DNA is detectable in fully differentiated GC Tfh cells ex vivo. This suggests that SIV-infected GC Tfh cells may be derived from Tfh precursor cell subsets that become infected in marginal zones and then migrate into GCs as fully mature GC Tfh cells that serve as persistent virus reservoirs. These findings suggest that viral persistence in lymph nodes drives compensatory differentiation, aberrant accumulation, and latent infection of GC Tfh cells, resulting in marked impairment of humoral immune responses. IMPORTANCE Generation of antibodies that can effectively eliminate viruses requires interactions of B cells with highly specialized T cells in GCs of lymphoid tissues called follicular T helper cells. Here we show that in simian immunodeficiency virus infection, these cells are initially infected in a precursor stage that leads to alterations in their homing, accumulation, and function that may be responsible for the inability of human immunodeficiency virus-infected patients to generate effective antibody responses. PMID:26608323

Persistent Simian Immunodeficiency Virus Infection Drives Differentiation, Aberrant Accumulation, and Latent Infection of Germinal Center Follicular T Helper Cells.

PubMed

Xu, Huanbin; Wang, Xiaolei; Malam, Naomi; Aye, Pyone P; Alvarez, Xavier; Lackner, Andrew A; Veazey, Ronald S

2016-02-01

CD4(+) follicular T helper (Tfh) cells play a prominent role in humoral immune responses, but the mechanisms of their accumulation and infection in AIDS remain unclear. Here we found that germinal center (GC) Tfh cells, defined here as CXCR5(+) PD-1(HIGH) CD4(+) T cells, do not express the HIV coreceptor CCR5 yet serve as a latent reservoir in GCs. With disease progression, an expansion of GC Tfh cells is accompanied by increases in dysfunctional CD8(+) T cells. In contrast, Tfh precursor (CXCR5(-) CD4(+) T) cells in lymph nodes do express CCR5 and differentiate into GC Tfh cells following interleukin-6 (IL-6) and IL-21 stimulation, and viral DNA is detectable in fully differentiated GC Tfh cells ex vivo. This suggests that SIV-infected GC Tfh cells may be derived from Tfh precursor cell subsets that become infected in marginal zones and then migrate into GCs as fully mature GC Tfh cells that serve as persistent virus reservoirs. These findings suggest that viral persistence in lymph nodes drives compensatory differentiation, aberrant accumulation, and latent infection of GC Tfh cells, resulting in marked impairment of humoral immune responses. Generation of antibodies that can effectively eliminate viruses requires interactions of B cells with highly specialized T cells in GCs of lymphoid tissues called follicular T helper cells. Here we show that in simian immunodeficiency virus infection, these cells are initially infected in a precursor stage that leads to alterations in their homing, accumulation, and function that may be responsible for the inability of human immunodeficiency virus-infected patients to generate effective antibody responses. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

A dual CCR2/CCR5 chemokine antagonist, BMS-813160? Evaluation of WO2011046916.

PubMed

Norman, Peter

2011-12-01

This application claims the dual CCR2/CCR5 antagonist (S)-1-[(1S,2R,4R)-4-isopropyl(methyl)amino)-2-propylcyclohexyl]-3-(6-(trifluoromethyl)quinazolin-4-ylamino)pyrrolidin-2-one, crystalline form N-1, a process for its preparation and therapeutic uses of it. The compound is shown to be a potent, well-absorbed, drug with a good duration of action. It shows better properties than similar compounds previously claimed by Bristol-Myers Squibb and appears to be a clinical development candidate.

Longitudinal Analysis of Cerebrospinal Fluid and Plasma HIV-1 Envelope Sequences Isolated From a Single Donor with HIV Asymptomatic Neurocognitive Impairment.

PubMed

Vázquez-Santiago, Fabián; García, Yashira; Rivera-Román, Ivelisse; Noel, Richard J; Wojna, Valerie; Meléndez, Loyda M; Rivera-Amill, Vanessa

Combined antiretroviral treatment (cART) has changed the clinical presentation of HIV-associated neurocognitive disorders (HAND) to that of the milder forms of the disease. Asymptomatic neurocognitive impairment (ANI) is now more prevalent and is associated with increased morbidity and mortality risk in HIV-1-infected people. HIV-1 envelope ( env ) genetic heterogeneity has been detected within the central nervous system (CNS) of individuals with ANI. Changes within env determine co-receptor use, cellular tropism, and neuropathogenesis. We hypothesize that compartmental changes are associated with HIV-1 env C2V4 during ANI and sought to analyze paired HIV-1 env sequences from plasma and cerebrospinal fluid (CSF) of a female subject undergoing long-term cART. Paired plasma and CSF samples were collected at 12-month intervals and HIV-1 env C2V4 was cloned and sequenced. Phylogenetic analysis of paired samples consistently showed genetic variants unique to the CSF. Phenotypic prediction showed CCR5 (R5) variants for all CSF-derived sequences and showed minor X4 variants (or dual-tropic) in the plasma at later time points. Viral compartmentalization was evident throughout the study, suggesting that the occurrence of distinctive env strains may contribute to the neuropathogenesis of HAND. Our study provides new insights about the genetic characteristics within the C2V4 of HIV-1 env that persist after long-term cART and during the course of persistent ANI.

Differential impact of transplantation on peripheral and tissue-associated viral reservoirs: Implications for HIV gene therapy

PubMed Central

Peterson, Christopher W.; Wang, Jianbin; Deleage, Claire; Reddy, Sowmya; Kaur, Jasbir; Polacino, Patricia; Reik, Andreas; Huang, Meei-Li; Holmes, Michael C.; Estes, Jacob D.

2018-01-01

Autologous transplantation and engraftment of HIV-resistant cells in sufficient numbers should recapitulate the functional cure of the Berlin Patient, with applicability to a greater number of infected individuals and with a superior safety profile. A robust preclinical model of suppressed HIV infection is critical in order to test such gene therapy-based cure strategies, both alone and in combination with other cure strategies. Here, we present a nonhuman primate (NHP) model of latent infection using simian/human immunodeficiency virus (SHIV) and combination antiretroviral therapy (cART) in pigtail macaques. We demonstrate that transplantation of CCR5 gene-edited hematopoietic stem/progenitor cells (HSPCs) persist in infected and suppressed animals, and that protected cells expand through virus-dependent positive selection. CCR5 gene-edited cells are readily detectable in tissues, namely those closely associated with viral reservoirs such as lymph nodes and gastrointestinal tract. Following autologous transplantation, tissue-associated SHIV DNA and RNA levels in suppressed animals are significantly reduced (p ≤ 0.05), relative to suppressed, untransplanted control animals. In contrast, the size of the peripheral reservoir, measured by QVOA, is variably impacted by transplantation. Our studies demonstrate that CCR5 gene editing is equally feasible in infected and uninfected animals, that edited cells persist, traffic to, and engraft in tissue reservoirs, and that this approach significantly reduces secondary lymphoid tissue viral reservoir size. Our robust NHP model of HIV gene therapy and viral persistence can be immediately applied to the investigation of combinatorial approaches that incorporate anti-HIV gene therapy, immune modulators, therapeutic vaccination, and latency reversing agents. PMID:29672640

Almendravirus: A Proposed New Genus of Rhabdoviruses Isolated from Mosquitoes in Tropical Regions of the Americas

PubMed Central

Contreras, Maria Angelica; Eastwood, Gillian; Guzman, Hilda; Popov, Vsevolod; Savit, Chelsea; Uribe, Sandra; Kramer, Laura D.; Wood, Thomas G.; Widen, Steven G.; Fish, Durland; Tesh, Robert B.; Vasilakis, Nikos; Walker, Peter J.

2017-01-01

The Rhabdoviridae is a diverse family of negative-sense single-stranded RNA viruses, many of which infect vertebrate hosts and are transmitted by hematophagous arthropods. Others appear to be arthropod specific, circulating only within arthropod populations. Herein, we report the isolation and characterization of three novel viruses from mosquitoes collected from the Americas. Coot Bay virus was isolated from Anopheles quadrimaculatus mosquitoes collected in the Everglades National Park, Florida; Rio Chico virus was isolated from Anopheles triannulatus mosquitoes collected in Panama; and Balsa virus was isolated from two pools of Culex erraticus mosquitoes collected in Colombia. Sequence analysis indicated that the viruses share a similar genome organization to Arboretum virus and Puerto Almendras virus that had previously been isolated from mosquitoes collected in Peru. Each genome features the five canonical rhabdovirus structural protein genes as well as a gene encoding a class 1A viroporin-like protein (U1) located between the G and L genes (3′-N-P-M-G-U1-L-5′). Phylogenetic analysis of complete L protein sequences indicated that all five viruses cluster in a unique clade that is relatively deeply rooted in the ancestry of animal rhabdoviruses. The failure of all viruses in this clade to grow in newborn mice or vertebrate cells in culture suggests that they may be poorly adapted to replication in vertebrates. PMID:27799634

The atypical chemokine receptor ACKR2 drives pulmonary fibrosis by tuning influx of CCR2+ and CCR5+ IFNγ-producing γδT cells in mice.

PubMed

Russo, Remo Castro; Savino, Benedetta; Mirolo, Massimiliano; Buracchi, Chiara; Germano, Giovanni; Anselmo, Achille; Zammataro, Luca; Pasqualini, Fabio; Mantovani, Alberto; Locati, Massimo; Teixeira, Mauro M

2018-02-22

Chemokines coordinate lung inflammation and fibrosis by acting on chemokine receptors expressed on leukocytes and other cell types. Atypical chemokine receptors (ACKRs) bind, internalize and degrade chemokines, tuning homeostasis and immune responses. ACKR2 recognizes and decreases levels of inflammatory CC chemokines. The role of ACKR2 in fibrogenesis is unknown. Investigate the role of ACKR2 in the context of pulmonary fibrosis. The effects of ACKR2 expression and deficiency during inflammation and fibrosis were analyzed using a bleomycin-model of fibrosis, ACKR2-deficient mice, bone marrow chimeras and antibody-mediated leukocyte depletion. ACKR2 was up-regulated acutely in response to bleomycin and normalized over time. ACKR2-/- mice showed reduced lethality and lung fibrosis. Bone marrow chimeras showed that lethality and fibrosis depended on ACKR2 expression in pulmonary resident (non-hematopoietic) cells but not on leukocytes. ACKR2-/- mice exhibited decreased expression of tissue remodeling genes, reduced leukocyte influx, pulmonary injury, and dysfunction. ACKR2-/- mice had early-increased levels of CCL5, CCL12, CCL17 and IFNγ, and increased number of CCR2+ and CCR5+ IFNγ-producing γδT cells in the airways counterbalanced by low Th17 lymphocyte influx. There was reduced accumulation of IFNγ-producing γδT cells in CCR2-/- and CCR5-/- mice. Moreover, depletion of γδT cells worsened the clinical symptoms induced by bleomycin and reversed the phenotype of ACKR2-/- mice exposed to bleomycin. ACKR2 controls the CC chemokine expression that drives the influx of CCR2+ and CCR5+ IFNγ-producing γδT cells tuning the Th17 response that mediate pulmonary fibrosis triggered by bleomycin instillation.

Induced pluripotent stem cell generation from a man carrying a complex chromosomal rearrangement as a genetic model for infertility studies

PubMed Central

Mouka, Aurélie; Izard, Vincent; Tachdjian, Gérard; Brisset, Sophie; Yates, Frank; Mayeur, Anne; Drévillon, Loïc; Jarray, Rafika; Leboulch, Philippe; Maouche-Chrétien, Leila; Tosca, Lucie

2017-01-01

Despite progress in human reproductive biology, the cause of male infertility often remains unknown, due to the lack of appropriate and convenient in vitro models of meiosis. Induced pluripotent stem cells (iPSCs) derived from the cells of infertile patients could provide a gold standard model for generating primordial germ cells and studying their development and the process of spermatogenesis. We report the characterization of a complex chromosomal rearrangement (CCR) in an azoospermic patient, and the successful generation of specific-iPSCs from PBMC-derived erythroblasts. The CCR was characterized by karyotype, fluorescence in situ hybridization and oligonucleotide-based array-comparative genomic hybridization. The CCR included five breakpoints and was caused by the inverted insertion of a chromosome 12 segment into the short arm of one chromosome 7 and a pericentric inversion of the structurally rearranged chromosome 12. Gene mapping of the breakpoints led to the identification of a candidate gene, SYCP3. Erythroblasts from the patient were reprogrammed with Sendai virus vectors to generate iPSCs. We assessed iPSC pluripotency by RT-PCR, immunofluorescence staining and teratoma induction. The generation of specific-iPSCs from patients with a CCR provides a valuable in vitro genetic model for studying the mechanisms by which chromosomal abnormalities alter meiosis and germ cell development. PMID:28045072

Host and Viral Factors in HIV-Mediated Bystander Apoptosis

PubMed Central

Garg, Himanshu; Joshi, Anjali

2017-01-01

Human immunodeficiency virus (HIV) infections lead to a progressive loss of CD4 T cells primarily via the process of apoptosis. With a limited number of infected cells and vastly disproportionate apoptosis in HIV infected patients, it is believed that apoptosis of uninfected bystander cells plays a significant role in this process. Disease progression in HIV infected individuals is highly variable suggesting that both host and viral factors may influence HIV mediated apoptosis. Amongst the viral factors, the role of Envelope (Env) glycoprotein in bystander apoptosis is well documented. Recent evidence on the variability in apoptosis induction by primary patient derived Envs underscores the role of Env glycoprotein in HIV disease. Amongst the host factors, the role of C-C Chemokine Receptor type 5 (CCR5), a coreceptor for HIV Env, is also becoming increasingly evident. Polymorphisms in the CCR5 gene and promoter affect CCR5 cell surface expression and correlate with both apoptosis and CD4 loss. Finally, chronic immune activation in HIV infections induces multiple defects in the immune system and has recently been shown to accelerate HIV Env mediated CD4 apoptosis. Consequently, those factors that affect CCR5 expression and/or immune activation in turn indirectly regulate HIV mediated apoptosis making this phenomenon both complex and multifactorial. This review explores the complex role of various host and viral factors in determining HIV mediated bystander apoptosis. PMID:28829402

[Prevalence study of the genetic markers associated with slow progression of human inmunodefiency virus type 1 in the Galician population (Northwest of Spain)].

PubMed

Rodríguez-Da Silva, Alfredo; Miralles, Celia; Ocampo, Antonio; Valverde, Diana

2017-02-01

The deletion in the CCR5 gene (CCR5Δ32), the HLA-B*27:05, and polymorphisms rs2395029 and rs9264942 have been associated with slower progression of HIV-1. An analysis was performed on 408 patients on follow-up. The analysis of viral load, CD4+ Tlymphocytes and other clinical variables since the diagnosis of the infection were collected. The prevalence of the genetic markers rs9264942, CCR5wt/Δ32, rs2395029, HLA-B*27:05 was 17.9%, 11.5%, 7.6%, and 6.4%, respectively. Of all the patients, 354 were classified as progressors and 46 as long-term non-progressors (LTNPs). Except for the HLA-B*27:05 allele, other genetic markers were associated with slower progression: CCR5wt/Δ32 (P=.011) and SNPs rs2395029 and rs9264942 (P<.0001), as well as their association (P<.0001). The prevalence of the HLA-B*57:01 allele was higher than described nationally. No association could be found between the HLA-B*27:05 allele and the presence of slower disease progression. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Neuronal apoptotic signaling pathways probed and intervened by synthetically and modularly modified (SMM) chemokines.

PubMed

Choi, Won-Tak; Kaul, Marcus; Kumar, Santosh; Wang, Jun; Kumar, I M Krishna; Dong, Chang-Zhi; An, Jing; Lipton, Stuart A; Huang, Ziwei

2007-03-09

As the main coreceptors for human immunodeficiency virus type 1 (HIV-1) entry, CXCR4 and CCR5 play important roles in HIV-associated dementia (HAD). HIV-1 glycoprotein gp120 contributes to HAD by causing neuronal damage and death, either directly by triggering apoptotic pathways or indirectly by stimulating glial cells to release neurotoxins. Here, to understand the mechanism of CXCR4 or CCR5 signaling in neuronal apoptosis associated with HAD, we have applied synthetically and modularly modified (SMM)-chemokine analogs derived from natural stromal cell-derived factor-1alpha or viral macrophage inflammatory protein-II as chemical probes of the mechanism(s) whereby these SMM-chemokines prevent or promote neuronal apoptosis. We show that inherently neurotoxic natural ligands of CXCR4, such as stromal cell-derived factor-1alpha or viral macrophage inflammatory protein-II, can be modified to protect neurons from apoptosis induced by CXCR4-preferring gp120(IIIB), and that the inhibition of CCR5 by antagonist SMM-chemokines, unlike neuroprotective CCR5 natural ligands, leads to neurotoxicity by activating a p38 mitogen-activated protein kinase (MAPK)-dependent pathway. Furthermore, we discover distinct signaling pathways activated by different chemokine ligands that are either natural agonists or synthetic antagonists, thus demonstrating a chemical biology strategy of using chemically engineered inhibitors of chemokine receptors to study the signaling mechanism of neuronal apoptosis and survival.

Arteriolar and Venular Remodeling Are Differentially Regulated by Bone Marrow-Derived Cell-Specific CX3CR1 and CCR2 Expression

PubMed Central

Meisner, Joshua K.; Song, Ji; Price, Richard J.

2012-01-01

The chemokine receptors CCR2 and CX3CR1 are critical for the recruitment of “inflammatory” and “resident” monocytes, respectively, subpopulations that differentially affect vascular remodeling in atherosclerosis. Here, we tested the hypothesis that bone marrow-derived cell (BMC)-specific CCR2 and CX3CR1 differentially control venular and arteriolar remodeling. Venular and arteriolar lumenal remodeling were observed by intravital microscopy in mice with either CCR2 or CX3CR1 deficient BMCs after implantation of a dorsal skinfold window chamber, a model in which arterioles and venules lumenally enlarge in wild-type (WT) mice. Arteriolar remodeling was abolished in mice with either CCR2 or CX3CR1-deficient BMCs. In contrast, the loss of CX3CR1 from BMCs, but not CCR2, significantly reduced small venule remodeling compared to WT controls. We conclude that microvascular remodeling is differentially regulated by BMC-expressed chemokine receptors. Both CCR2 and CX3CR1 regulate arteriole growth; however, only BMC-expressed CX3CR1 impacts small venule growth. These findings may provide a basis for additional investigations aimed at determining how patterns of monocyte subpopulation recruitment spatially influence microvascular remodeling. PMID:23029475

Antibodies Elicited by Multiple Envelope Glycoprotein Immunogens in Primates Neutralize Primary Human Immunodeficiency Viruses (HIV-1) Sensitized by CD4-Mimetic Compounds

PubMed Central

Madani, Navid; Princiotto, Amy M.; Easterhoff, David; Bradley, Todd; Luo, Kan; Williams, Wilton B.; Liao, Hua-Xin; Moody, M. Anthony; Phad, Ganesh E.; Vázquez Bernat, Néstor; Melillo, Bruno; Santra, Sampa; Smith, Amos B.; Karlsson Hedestam, Gunilla B.; Haynes, Barton

2016-01-01

ABSTRACT The human immunodeficiency virus (HIV-1) envelope glycoproteins (Env) mediate virus entry through a series of complex conformational changes triggered by binding to the receptors CD4 and CCR5/CXCR4. Broadly neutralizing antibodies that recognize conserved Env epitopes are thought to be an important component of a protective immune response. However, to date, HIV-1 Env immunogens that elicit broadly neutralizing antibodies have not been identified, creating hurdles for vaccine development. Small-molecule CD4-mimetic compounds engage the CD4-binding pocket on the gp120 exterior Env and induce Env conformations that are highly sensitive to neutralization by antibodies, including antibodies directed against the conserved Env region that interacts with CCR5/CXCR4. Here, we show that CD4-mimetic compounds sensitize primary HIV-1 to neutralization by antibodies that can be elicited in monkeys and humans within 6 months by several Env vaccine candidates, including gp120 monomers. Monoclonal antibodies directed against the gp120 V2 and V3 variable regions were isolated from the immunized monkeys and humans; these monoclonal antibodies neutralized a primary HIV-1 only when the virus was sensitized by a CD4-mimetic compound. Thus, in addition to their direct antiviral effect, CD4-mimetic compounds dramatically enhance the HIV-1-neutralizing activity of antibodies that can be elicited with currently available immunogens. Used as components of microbicides, the CD4-mimetic compounds might increase the protective efficacy of HIV-1 vaccines. IMPORTANCE Preventing HIV-1 transmission is a high priority for global health. Eliciting antibodies that can neutralize transmitted strains of HIV-1 is difficult, creating problems for the development of an effective vaccine. We found that small-molecule CD4-mimetic compounds sensitize HIV-1 to antibodies that can be elicited in vaccinated humans and monkeys. These results suggest an approach to prevent HIV-1 sexual transmission in which a virus-sensitizing microbicide is combined with a vaccine. PMID:26962221

Is the European spatial distribution of the HIV-1-resistant CCR5-Delta32 allele formed by a breakdown of the pathocenosis due to the historical Roman expansion?

PubMed

Faure, Eric; Royer-Carenzi, Manuela

2008-12-01

We studied the possible effects of the expansion of ancient Mediterranean civilizations during the five centuries before and after Christ on the European distribution of the mutant allele for the chemokine receptor gene CCR5 which has a 32-bp deletion (CCR5-Delta32). There is a strong evidence for the unitary origin of the CCR5-Delta32 mutation, this it is found principally in Europe and Western Asia, with generally a north-south downhill cline frequency. Homozygous carriers of this mutation show a resistance to HIV-1 infection and a slower progression towards AIDS. However, HIV has clearly emerged too recently to have been the selective force on CCR5. Our analyses showed strong negative correlations in Europe between the allele frequency and two historical parameters, i.e. the first colonization dates by the great ancient Mediterranean civilizations, and the distances from the Northern frontiers of the Roman Empire in its greatest expansion. Moreover, other studies have shown that the deletion frequencies in both German Bronze Age and Swedish Neolithic populations were similar to those found in the corresponding modern populations, and this deletion has been found in ancient DNA of around 7000 years ago, suggesting that in the past, the deletion frequency could have been relatively high in European populations. In addition, in West Nile virus pathogenesis, CCR5 plays an antimicrobial role showing that host genetic factors are highly pathogen-specific. Our results added to all these previous data suggest that the actual European allele frequency distribution might not be due to genes spreading, but to a negative selection resulting in the spread of pathogens principally during Roman expansion. Indeed, as gene flows from colonizers to European native populations were extremely low, the mutational changes might be associated with vulnerability to imported infections. To date, the nature of the parasites remains unknown; however, zoonoses could be incriminated.

Renal creatinine handling in very old patients with chronic renal disease.

PubMed

Musso, Carlos G; Michelángelo, Hernán; Vilas, Manuel; Martinez, Bernardo; Bonetto, Alberto; Jauregui, Ricardo; Algranati, Luis

2011-09-01

Renal creatinine handling is basically the result of its glomerular filtration and proximal tubular secretion. However, creatinine reabsorption has been documented in certain conditions, such as premature babies, newborns, and healthy elderly people. Additionally, it is known that there is an increase in the proportion of secreted creatinine in chronic renal disease. In this paper, we report our studies on the characteristic reabsorption pattern of creatinine in the elderly with chronic renal disease. We studied twenty-seven volunteers with chronic kidney disease, eleven of whom were young and the rest were very old (age > 75 years old). We measured creatinine clearance without (Ccr) and with cimetidine (CcrWC) and Ccr/CcrWC ratio from each volunteer, in timed urine samples. Then, Ccr, CcrWC, and Ccr/CcrWC ratio were compared between young and very old people in two chronic kidney disease subgroups: stages II-III and stages IV-V. Statistical analysis was performed applying a non-parametric test (Wilcoxon). We observed a tendency towards a lower Ccr/CcrWC ratio in the very old stage II-III group compared with the young one: 1 (0.96-1.26) (very old) vs 1.3 (1.1-1.5) (young), P = 0.09, on the contrary, there was no significant difference in Ccr/CcrWC ratio between very old and young person with stage IV-V CKD: 1.66 (1.41-2.21) (young) vs 1.77 (1.1-2.7) (young), P = NS. Creatinine secretion pattern in very old patients with advanced chronic renal disease is similar to that observed in young ones with similar level of CKD.

Role of Exonic Variation in Chemokine Receptor Genes on AIDS: CCRL2 F167Y Association with Pneumocystis Pneumonia

PubMed Central

An, Ping; Li, Rongling; Wang, Ji Ming; Yoshimura, Teizo; Takahashi, Munehisa; Samudralal, Ram; O'Brien, Stephen J.; Phair, John; Goedert, James J.; Kirk, Gregory D.; Troyer, Jennifer L.; Sezgin, Efe; Buchbinder, Susan P.; Donfield, Sharyne; Nelson, George W.; Winkler, Cheryl A.

2011-01-01

Chromosome 3p21–22 harbors two clusters of chemokine receptor genes, several of which serve as major or minor coreceptors of HIV-1. Although the genetic association of CCR5 and CCR2 variants with HIV-1 pathogenesis is well known, the role of variation in other nearby chemokine receptor genes remain unresolved. We genotyped exonic single nucleotide polymorphisms (SNPs) in chemokine receptor genes: CCR3, CCRL2, and CXCR6 (at 3p21) and CCR8 and CX3CR1 (at 3p22), the majority of which were non-synonymous. The individual SNPs were tested for their effects on disease progression and outcomes in five treatment-naïve HIV-1/AIDS natural history cohorts. In addition to the known CCR5 and CCR2 associations, significant associations were identified for CCR3, CCR8, and CCRL2 on progression to AIDS. A multivariate survival analysis pointed to a previously undetected association of a non-conservative amino acid change F167Y in CCRL2 with AIDS progression: 167F is associated with accelerated progression to AIDS (RH = 1.90, P = 0.002, corrected). Further analysis indicated that CCRL2-167F was specifically associated with more rapid development of pneumocystis pneumonia (PCP) (RH = 2.84, 95% CI 1.28–6.31) among four major AIDS–defining conditions. Considering the newly defined role of CCRL2 in lung dendritic cell trafficking, this atypical chemokine receptor may affect PCP through immune regulation and inducing inflammation. PMID:22046140

CCR2-V64I genetic polymorphism: a possible involvement in HER2+ breast cancer.

PubMed

Banin-Hirata, Bruna Karina; Losi-Guembarovski, Roberta; Oda, Julie Massayo Maeda; de Oliveira, Carlos Eduardo Coral; Campos, Clodoaldo Zago; Mazzuco, Tânia Longo; Borelli, Sueli Donizete; Ceribelli, Jesus Roberto; Watanabe, Maria Angelica Ehara

2016-05-01

Many tumor cells express chemokines and chemokine receptors, and these molecules can affect both tumor progression and anti-tumor immune response. Genetic polymorphisms of some chemokine receptors were found to be closely related to malignant tumors, especially in metastasis process, including breast cancer (BC). Considering this, it was investigated a possible role for CCR2-V64I (C-C chemokine receptor 2) and CCR5-Δ32 (C-C chemokine receptor 5) genetic variants in BC context. Patients were divided into subgroups according to immunohistochemical profile of estrogen (ER) and progesterone (PR) receptors and the human epidermal growth factor receptor 2 (HER2) overexpression. No significant associations were found in relation to susceptibility (CCR2-V64I: OR 1.32; 95 % CI 0.57-3.06; CCR5-∆32: OR 1.04; 95 % CI 0.60-1.81), clinical outcome (tumor size, lymph nodes commitment and/or distant metastasis, TNM staging and nuclear grade) or therapeutic response (recurrence and survival). However, it was found a significant correlation between CCR2-V64I allelic variant and HER2 immunohistochemical positive samples (p = 0.026). All in all, we demonstrate, for the first time, a positive correlation between CCR2 receptor gene polymorphism and a subgroup of BC related to poor prognosis, which deserves further investigation in larger samples for validation.

Polymorphisms in chemokine and receptor genes and gastric cancer risk and survival in a high risk Polish population

PubMed Central

Gawron, Andrew J.; Fought, Angela J.; Lissowska, Jolanta; Ye, Weimin; Zhang, Xiao; Chow, Wong-Ho; Freeman, Laura E. Beane; Hou, Lifang

2010-01-01

Objective To examine if genetic variations in chemokine receptor and ligand genes are associated with gastric cancer risk and survival. Methods The study included 298 cases and 417 controls from a population-based study of gastric cancer conducted in Warsaw, Poland in 1994–1996. We investigated seven single nucleotide polymorphisms in a chemokine ligand (CXCL12) and chemokine receptor (CCR2, CCR5, CX3CR1) genes and one frameshift deletion (CCR5) in blood leukocyte DNA in relation to gastric cancer risk and survival. Genotyping was conducted at the NCI Core Genotyping Facility. Odds ratios and 95% confidence intervals were computed using univariate and multivariate logistic regression models. Survival analysis was performed using Cox proportional hazards models. Results Gastric cancer risk was not associated with single chemokine polymorphisms. A CCR5 haplotype that contained the common alleles of IVS1+151 G>T (rs2734648), IVS2+80 C>T (rs1800024) and minor allele of IVS1+246 A>G (rs1799987) was associated with a borderline significantly increased risk (OR = 1.5, 95% CI: 1.0–2.2). For gastric cancer cases, there was a greater risk of death for carriers of the minor alleles of CCR2 Ex2+241 G>A (rs1799864) (HR = 1.5, 95% CI: 1.1–2.1) and CCR5 IVS2+80 C>T (rs1800024) (HR = 1.5, 95% CI: 1.1–2.1). Carriers of the CCR5 minor allele of IVS1+151 G>T (rs2734648) had a decreased risk of death compared to homozygote carriers of the common allele (HR = 0.8, 95% CI: 0.6–1.0). Conclusions Our findings do not support an association between gastric cancer risk and single chemokine genetic variation. The observed associations between cancer risk and a CCR5 haplotype and between survival and polymorphisms in CCR2 and CCR5 need replication in future studies. PMID:21091093

Improved Cognitive Performance and Reduced Monocyte Activation in Virally Suppressed Chronic HIV Following Dual CCR2 and CCR5 Antagonism.

PubMed

D'Antoni, Michelle L; Paul, Robert H; Mitchell, Brooks I; Kohorn, Lindsay; Fischer, Laurent; Lefebvre, Eric; Seyedkazemi, Star; Nakamoto, Beau K; Walker, Maegen; Kallianpur, Kalpana J; Ogata-Arakaki, Debra; Ndhlovu, Lishomwa C; Shikuma, Cecilia

2018-05-16

To evaluate changes in neuropsychological (NP) performance and in plasma and cell surface markers of peripheral monocyte activation/migration following treatment with cenicriviroc (CVC), a dual C-C chemokine receptor type 2 (CCR2) and type 5 (CCR5) antagonist, in treatment-experienced, HIV-infected individuals. Single-arm, 24-week, open-label clinical trial. HIV-infected individuals on antiretroviral therapy (ART) >1 year with plasma HIV RNA <50 copies/ml and below-normal cognitive performance [defined as age, gender and education-adjusted NP performance (NPZ) <-0.5 in a single cognitive domain or in global performance] were enrolled. Changes over 24 weeks were assessed for global and domain-specific NPZ scores, plasma markers of monocyte/macrophage activation [neopterin, soluble (s)CD14 and sCD163] quantified by ELISA, and CCR2 and CCR5 expression on monocytes and T cells measured by flow cytometry. Seventeen of 20 enrolled participants completed the study. Improvements over 24 weeks were observed in global NPZ [median change (Δ)=0.24; p=0.008], and in cognitive domains of attention (Δ0.23; p=0.011) and working memory (Δ0.44; p=0.017). Plasma levels of sCD163, sCD14, and neopterin decreased significantly (p's<0.01). CCR2 and CCR5 monocyte expression remained unchanged; however, CCR5 levels on CD4 and CD8 T cells and CCR2 expression on CD4 T cells increased (p's<0.01). CVC given over 24 weeks was associated with improved NP test performance and decreased plasma markers of monocyte immune activation in virally-suppressed, HIV-infected participants. These data potentially link changes in monocyte activation to cognitive performance. Further study of CVC for HIV cognitive impairment in a randomized controlled study is warranted.

Effect of C-C Chemokine Receptor 2 (CCR2) Knockout on Type-2 (Schistosomal Antigen-Elicited) Pulmonary Granuloma Formation

PubMed Central

Warmington, Kelly S.; Boring, Landin; Ruth, Jeffrey H.; Sonstein, Joanne; Hogaboam, Cory M.; Curtis, Jeffrey L.; Kunkel, Steven L.; Charo, Israel R.; Chensue, Stephen W.

1999-01-01

Monocyte chemotactic protein (MCP)-1 is postulated to play a role in cellular recruitment during inflammatory reactions. C-C chemokine receptor 2 (CCR2) is considered the major G-protein coupled receptor for MCP-1/JE. We reported that mice with knockout of the CCR2 gene display partially impaired type-1 granuloma formation. The present study similarly examined the effect of CCR2 deficiency on synchronously developing type-2 (Th2) cytokine-mediated lung granulomas elicited by embolization of beads coated with Ags of Schistosoma mansoni eggs. Systemically, blood monocytes were reduced by about half throughout the 8-day study period. At the local level, granuloma size and macrophage content were impaired during the early growth phase (days 1 to 2). By day 4, granuloma sizes were similar to controls. In granulomatous lungs, CCR2 knockout increased mRNA for CCR2 agonists, MCP-1, MCP-3, and MCP-5, but reduced IL-4 and IFNγ mRNA. The latter was possibly related to decreased CD4+ T cell recruitment. Regionally, draining lymph nodes showed panlymphoid hyperplasia with impaired production of IFNγ, IL-2, and IL-4, but not IL-5, IL-10, or IL-13. Analysis of procollagen gene expression indicated transient impairment of procollagen III transcripts on day 4 of granuloma formation. These findings indicate that agonists of CCR2 contribute to multiple facets of type-2 hypersensitivity granulomatous inflammation. PMID:10329593

Constitutive and Stress-induced Expression of CCL5 Machinery in Rodent Retina

PubMed Central

Duncan, D'Anne S.; McLaughlin, William M.; Vasilakes, Noah; Echevarria, Franklin D.; Formichella, Cathryn R.; Sappington, Rebecca M.

2017-01-01

Signaling by inflammatory cytokines and chemokines is associated with neurodegeneration in disease and injury. Here we examined expression of the β-chemokine CCL5 and its receptors in the mouse retina and evaluated its relevance in glaucoma, a common optic neuropathy associated with sensitivity to intraocular pressure (IOP). Using quantitative PCR, fluorescent in situ hybridization, immunohistochemistry and quantitative image analysis, we found CCL5 mRNA and protein was constitutively expressed in the inner retina and synaptic layers. CCL5 appeared to associate with Müller cells and RGCs as well as synaptic connections between horizontal cells and bipolar cells in the OPL and amacrine cells, bipolar cells and RGCs in the IPL. Although all three high-affinity receptors (CCR5, CCR3, CCR1) for CCL5 were expressed constitutively, CCR5 expression was significantly higher than CCR3, which was also markedly greater than CCR1. Localization patterns for constitutive CCR5, CCR3 and CCR1 expression differed, particularly with respect to expression in inner retinal neurons. Stress-related expression of CCL5 was primarily altered in aged DBA/2 mice with elevated IOP. In contrast, changes in expression and localization of both CCR3 and CCR5 were evident not only in aged DBA/2 mice, but also in age-matched control mice and young DBA/2 mice. These groups do not exhibit elevated IOP, but possess either the aging stress (control mice) or the genetic predisposition to glaucoma (DBA/2 mice). Together, these data indicate that CCL5 and its high-affinity receptors are constitutively expressed in murine retina and differentially induced by stressors associated with glaucomatous optic neuropathy. Localization patterns further indicate that CCL5 signaling may be relevant for modulation of synapses in both health and disease, particularly in the inner plexiform layer. PMID:28936366

Autologous Hematopoietic Stem Cells transplantation and genetic modification of CCR5 m303/m303 mutant patient for HIV/AIDS.

PubMed

Esmaeilzadeh, Abdolreza; Farshbaf, Alieh; Erfanmanesh, Maryam

2015-03-01

HIV and AIDS is one of the biggest challenges all over the world. There are an approximately 34 million people living with the virus, and a large number of them become infected each year. Although there are some antiviral drugs for HIV viral load reduction, they are not sufficient. There is no cure for AIDS. Nowadays natural resistance or immunity has absorbed attentions. Because in some HIV positive patients progression trend is slow or even they indicate resistance to AIDS. One of the most interesting approaches in this category is CCR5 gene. CCR5 is a main cc-chemokine co-receptor that facilitates HIV-1 entry to macrophage and CD4(+) T cells. To now, many polymorphisms have been known by CCR5 gene that produces a truncated protein with no function. So, HIV-1 could not entry to immune-cells and the body resistant to HIV/AIDS. Δ32/Δ32 and m303/m303 homozygotes are example of mutations that could create this resistance mechanism. There is a new treatment, such as Hematopoietic Stem Cell transplantation (HSCT) in Berlin and Boston patients for Δ32/Δ32 mutation. It could eliminate co-receptor antagonist and highly-active-anti retroviral therapy (HAART) drugs problems such as toxicity, low safety and side-effects. Now there, the aim of this hypothesis will be evaluation of a new mutation CCR5 m303/m303 as autologous HSCT. This novel hypothesis indicates that autologous HSCT for m303/m303 could be effective treatment for anyone HIV/AIDS affected patient worldwide. Copyright © 2015 Elsevier Ltd. All rights reserved.

CXC chemokine ligand 4 (CXCL4) down-regulates CC chemokine receptor expression on human monocytes.

PubMed

Schwartzkopff, Franziska; Petersen, Frank; Grimm, Tobias Alexander; Brandt, Ernst

2012-02-01

During acute inflammation, monocytes are essential in abolishing invading micro-organisms and encouraging wound healing. Recruitment by CC chemokines is an important step in targeting monocytes to the inflamed tissue. However, cell surface expression of the corresponding chemokine receptors is subject to regulation by various endogenous stimuli which so far have not been comprehensively identified. We report that the platelet-derived CXC chemokine ligand 4 (CXCL4), a known activator of human monocytes, induces down-regulation of CC chemokine receptors (CCR) 1, -2, and -5, resulting in drastic impairment of monocyte chemotactic migration towards cognate CC chemokine ligands (CCL) for these receptors. Interestingly, CXCL4-mediated down-regulation of CCR1, CCR2 and CCR5 was strongly dependent on the chemokine's ability to stimulate autocrine/paracrine release of TNF-α. In turn, TNF-α induced the secretion CCL3 and CCL4, two chemokines selective for CCR1 and CCR5, while the secretion of CCR2-ligand CCL2 was TNF-α-independent. Culture supernatants of CXCL4-stimulated monocytes as well as chemokine-enriched preparations thereof reproduced CXCL4-induced CCR down-regulation. In conclusion, CXCL4 may act as a selective regulator of monocyte migration by stimulating the release of autocrine, receptor-desensitizing chemokine ligands. Our results stress a co-ordinating role for CXCL4 in the cross-talk between platelets and monocytes during early inflammation.

Temporal dynamics of the primary human T cell response to yellow fever virus 17D as it matures from an effector- to a memory-type response.

PubMed

Blom, Kim; Braun, Monika; Ivarsson, Martin A; Gonzalez, Veronica D; Falconer, Karolin; Moll, Markus; Ljunggren, Hans-Gustaf; Michaëlsson, Jakob; Sandberg, Johan K

2013-03-01

The live attenuated yellow fever virus (YFV) 17D vaccine provides a good model to study immune responses to an acute viral infection in humans. We studied the temporal dynamics, composition, and character of the primary human T cell response to YFV. The acute YFV-specific effector CD8 T cell response was broad and complex; it was composed of dominant responses that persisted into the memory population, as well as of transient subdominant responses that were not detected at the memory stage. Furthermore, HLA-A2- and HLA-B7-restricted YFV epitope-specific effector cells predominantly displayed a CD45RA(-)CCR7(-)PD-1(+)CD27(high) phenotype, which transitioned into a CD45RA(+)CCR7(-)PD-1(-)CD27(low) memory population phenotype. The functional profile of the YFV-specific CD8 T cell response changed in composition as it matured from an effector- to a memory-type response, and it tended to become less polyfunctional during the course of this transition. Interestingly, activation of CD4 T cells, as well as FOXP3(+) T regulatory cells, in response to YFV vaccination preceded the kinetics of the CD8 T cell response. The present results contribute to our understanding of how immunodominance patterns develop, as well as the phenotypic and functional characteristics of the primary human T cell response to a viral infection as it evolves and matures into memory.

Editor's Highlight: CCR2 Regulates Inflammatory Cell Accumulation in the Lung and Tissue Injury following Ozone Exposure.

PubMed

Francis, Mary; Groves, Angela M; Sun, Richard; Cervelli, Jessica A; Choi, Hyejeong; Laskin, Jeffrey D; Laskin, Debra L

2017-02-01

Ozone-induced lung injury is associated with an accumulation of activated macrophages in the lung. Chemokine receptor CCR2 mediates the migration of inflammatory monocytes/macrophages to sites of tissue injury. It is also required for monocyte egress from the bone marrow. In the present studies, we analyzed the role of CCR2 in inflammatory cell trafficking to the lung in response to ozone. Treatment of mice with ozone (0.8 ppm, 3 h) resulted in increases in proinflammatory CCR2 + macrophages in the lung at 24 h, as well as proinflammatory CD11b  +  Ly6C Hi and iNOS +  macrophages at 24 and 48 h. Mannose receptor +  anti-inflammatory macrophages were also observed in the lung 24 and 48 h post-ozone. Loss of CCR2 was associated with reduced numbers of proinflammatory macrophages in the lung and decreased expression of the proinflammatory cytokines, IL-1β and TNFα. Decreases in anti-inflammatory CD11b  +  Ly6C Lo macrophages were also observed in lungs of CCR2 -/- mice treated with ozone, whereas mannose receptor +  macrophage accumulation was delayed; conversely, CX3CL1 and CX3CR1 were upregulated. Changes in lung macrophage subpopulations and inflammatory gene expression in CCR2 -/- mice were correlated with reduced ozone toxicity and oxidative stress, as measured by decreases in bronchoalveolar lavage protein content and reduced lung expression of heme-oxygenase-1, 4-hydroxynonenal and cytochrome b5. These data demonstrate that CCR2 plays a role in both pro- and anti-inflammatory macrophage accumulation in the lung following ozone exposure. The fact that ozone-induced lung injury and oxidative stress are reduced in CCR2 -/- mice suggests more prominent effects on proinflammatory macrophages. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Editor’s Highlight: CCR2 Regulates Inflammatory Cell Accumulation in the Lung and Tissue Injury following Ozone Exposure

PubMed Central

Francis, Mary; Groves, Angela M.; Sun, Richard; Cervelli, Jessica A.; Choi, Hyejeong; Laskin, Jeffrey D.; Laskin, Debra L.

2017-01-01

Ozone-induced lung injury is associated with an accumulation of activated macrophages in the lung. Chemokine receptor CCR2 mediates the migration of inflammatory monocytes/macrophages to sites of tissue injury. It is also required for monocyte egress from the bone marrow. In the present studies, we analyzed the role of CCR2 in inflammatory cell trafficking to the lung in response to ozone. Treatment of mice with ozone (0.8 ppm, 3 h) resulted in increases in proinflammatory CCR2+ macrophages in the lung at 24 h, as well as proinflammatory CD11b + Ly6CHi and iNOS+ macrophages at 24 and 48 h. Mannose receptor+ anti-inflammatory macrophages were also observed in the lung 24 and 48 h post-ozone. Loss of CCR2 was associated with reduced numbers of proinflammatory macrophages in the lung and decreased expression of the proinflammatory cytokines, IL-1β and TNFα. Decreases in anti-inflammatory CD11b + Ly6CLo macrophages were also observed in lungs of CCR2−/− mice treated with ozone, whereas mannose receptor+ macrophage accumulation was delayed; conversely, CX3CL1 and CX3CR1 were upregulated. Changes in lung macrophage subpopulations and inflammatory gene expression in CCR2−/− mice were correlated with reduced ozone toxicity and oxidative stress, as measured by decreases in bronchoalveolar lavage protein content and reduced lung expression of heme-oxygenase-1, 4-hydroxynonenal and cytochrome b5. These data demonstrate that CCR2 plays a role in both pro- and anti-inflammatory macrophage accumulation in the lung following ozone exposure. The fact that ozone-induced lung injury and oxidative stress are reduced in CCR2−/− mice suggests more prominent effects on proinflammatory macrophages. PMID:27837169

Membrane organization of virus and target cell plays a role in HIV entry.

PubMed

Dumas, Fabrice; Preira, Pascal; Salomé, Laurence

2014-12-01

The initial steps of the Human Immunodeficiency Virus (HIV) replication cycle play a crucial role that arbitrates viral tropism and infection efficiency. Before the release of its genome into the host cell cytoplasm, viruses operate a complex sequence of events that take place at the plasma membrane of the target cell. The first step is the binding of the HIV protein envelope (Env) to the cellular receptor CD4. This triggers conformational changes of the gp120 viral protein that allow its interaction with a co-receptor that can be either CCR5 or CXCR4, defining the tropism of the virus entering the cell. This sequential interaction finally drives the fusion of the viral and host cell membrane or to the endocytosis of the viruses. Here, we discuss how the membrane composition and organization of both the virus and the target cell can affect these steps and thus influence the capability of the viruses to infect cells. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

A chimeric measles virus with a lentiviral envelope replicates exclusively in CD4+/CCR5+ cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mourez, Thomas; APHP, GH Saint-Louis-Lariboisiere, Laboratoire de Bacteriologie-Virologie, F-75010 Paris; Universite Paris 7 Denis Diderot, F-75010 Paris

2011-10-25

We generated a replicating chimeric measles virus in which the hemagglutinin and fusion surface glycoproteins were replaced with the gp160 envelope glycoprotein of simian immunodeficiency virus (SIVmac239). Based on a previously cloned live-attenuated Schwarz vaccine strain of measles virus (MV), this chimera was rescued at high titers using reverse genetics in CD4+ target cells. Cytopathic effect consisted in the presence of large cell aggregates evolving to form syncytia, as observed during SIV infection. The morphology of the chimeric virus was identical to that of the parent MV particles. The presence of SIV gp160 as the only envelope protein on chimericmore » particles surface altered the cell tropism of the new virus from CD46+ to CD4+ cells. Used as an HIV candidate vaccine, this MV/SIVenv chimeric virus would mimic transient HIV-like infection, benefiting both from HIV-like tropism and the capacity of MV to replicate in dendritic cells, macrophages and lymphocytes.« less

Regional control of Drosophila gut stem cell proliferation: EGF establishes GSSC proliferative set point & controls emergence from quiescence.

PubMed

Strand, Marie; Micchelli, Craig A

2013-01-01

Adult stem cells vary widely in their rates of proliferation. Some stem cells are constitutively active, while others divide only in response to injury. The mechanism controlling this differential proliferative set point is not well understood. The anterior-posterior (A/P) axis of the adult Drosophila midgut has a segmental organization, displaying physiological compartmentalization and region-specific epithelia. These distinct midgut regions are maintained by defined stem cell populations with unique division schedules, providing an excellent experimental model with which to investigate this question. Here, we focus on the quiescent gastric stem cells (GSSCs) of the acidic copper cell region (CCR), which exhibit the greatest period of latency between divisions of all characterized gut stem cells, to define the molecular basis of differential stem cell activity. Our molecular genetic analysis demonstrates that the mitogenic EGF signaling pathway is a limiting factor controlling GSSC proliferation. We find that under baseline conditions, when GSSCs are largely quiescent, the lowest levels of EGF ligands in the midgut are found in the CCR. However, acute epithelial injury by enteric pathogens leads to an increase in EGF ligand expression in the CCR and rapid expansion of the GSSC lineage. Thus, the unique proliferative set points for gut stem cells residing in physiologically distinct compartments are governed by regional control of niche signals along the A/P axis.

CCR5 receptor antagonists in preclinical to phase II clinical development for treatment of HIV

PubMed Central

Kim, Michelle B.; Giesler, Kyle E.; Tahirovic, Yesim A.; Truax, Valarie M.; Liotta, Dennis C.; Wilson, Lawrence J.

2018-01-01

Introduction The chemokine receptor CCR5 has garnered significant attention in recent years as a target to treat HIV infection largely due to the approval and success of the drug Maraviroc. The side effects and inefficiencies with other first generation agents led to failed clinical trials, prompting the development of newer CCR5 antagonists. Areas covered This review aims to survey the current status of ‘next generation’ CCR5 antagonists in the preclinical pipeline with an emphasis on emerging agents for the treatment of HIV infection. These efforts have culminated in the identification of advanced second-generation agents to reach the clinic and the dual CCR5/CCR2 antagonist Cenicriviroc as the most advanced currently in phase II clinical studies. Expert opinion The clinical success of CCR5 inhibitors for treatment of HIV infection has rested largely on studies of Maraviroc and a second-generation dual CCR5/CCR2 antagonist Cenicriviroc. Although research efforts identified several promising preclinical candidates, these were dropped during early clinical studies. Despite patient access to Maraviroc, there is insufficient enthusiasm surrounding its use as front-line therapy for treatment of HIV. The non-HIV infection related development activities for Maraviroc and Cenicriviroc may help drive future interests. PMID:27791451

Aberrant in Vivo T Helper Type 2 Cell Response and Impaired Eosinophil Recruitment in Cc Chemokine Receptor 8 Knockout Mice

PubMed Central

Chensue, Stephen W.; Lukacs, Nicholas W.; Yang, Tong-Yuan; Shang, Xiaozhou; Frait, Kirsten A.; Kunkel, Steven L.; Kung, Ted; Wiekowski, Maria T.; Hedrick, Joseph A.; Cook, Donald N.; Zingoni, Alessandra; Narula, Satwant K.; Zlotnik, Albert; Barrat, Franck J.; O'Garra, Anne; Napolitano, Monica; Lira, Sergio A.

2001-01-01

Chemokine receptors transduce signals important for the function and trafficking of leukocytes. Recently, it has been shown that CC chemokine receptor (CCR)8 is selectively expressed by Th2 subsets, but its functional relevance is unclear. To address the biological role of CCR8, we generated CCR8 deficient (−/−) mice. Here we report defective T helper type 2 (Th2) immune responses in vivo in CCR8−/− mice in models of Schistosoma mansoni soluble egg antigen (SEA)-induced granuloma formation as well as ovalbumin (OVA)- and cockroach antigen (CRA)-induced allergic airway inflammation. In these mice, the response to SEA, OVA, and CRA showed impaired Th2 cytokine production that was associated with aberrant type 2 inflammation displaying a 50 to 80% reduction in eosinophils. In contrast, a prototypical Th1 immune response, elicited by Mycobacteria bovis purified protein derivative (PPD) was unaffected by CCR8 deficiency. Mechanistic analyses indicated that Th2 cells developed normally and that the reduction in eosinophil recruitment was likely due to systemic reduction in interleukin 5. These results indicate an important role for CCR8 in Th2 functional responses in vivo. PMID:11238588

Serotype-Specific Structural Differences in the Protease-Cofactor Complexes of the Dengue Virus Family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chandramouli, Sumana; Joseph, Jeremiah S.; Daudenarde, Sophie

With an estimated 40% of the world population at risk, dengue poses a significant threat to human health, especially in tropical and subtropical regions. Preventative and curative efforts, such as vaccine development and drug discovery, face additional challenges due to the occurrence of four antigenically distinct serotypes of the causative dengue virus (DEN1 to -4). Complex immune responses resulting from repeat assaults by the different serotypes necessitate simultaneous targeting of all forms of the virus. One of the promising targets for drug development is the highly conserved two-component viral protease NS2B-NS3, which plays an essential role in viral replication bymore » processing the viral precursor polyprotein into functional proteins. In this paper, we report the 2.1-{angstrom} crystal structure of the DEN1 NS2B hydrophilic core (residues 49 to 95) in complex with the NS3 protease domain (residues 1 to 186) carrying an internal deletion in the N terminus (residues 11 to 20). While the overall folds within the protease core are similar to those of DEN2 and DEN4 proteases, the conformation of the cofactor NS2B is dramatically different from those of other flaviviral apoprotease structures. The differences are especially apparent within its C-terminal region, implicated in substrate binding. The structure reveals for the first time serotype-specific structural elements in the dengue virus family, with the reported alternate conformation resulting from a unique metal-binding site within the DEN1 sequence. We also report the identification of a 10-residue stretch within NS3pro that separates the substrate-binding function from the catalytic turnover rate of the enzyme. Implications for broad-spectrum drug discovery are discussed.« less

ccrABEnt serine recombinase genes are widely distributed in the Enterococcus faecium and Enterococcus casseliflavus species groups and are expressed in E. faecium

PubMed Central

Bjørkeng, Eva Katrin; Tessema, Girum Tadesse; Lundblad, Eirik Wasmuth; Butaye, Patrick; Willems, Rob; Sollid, Johanna Ericsson; Sundsfjord, Arnfinn; Hegstad, Kristin

2010-01-01

The presence, distribution and expression of cassette chromosome recombinase (ccr) genes, which are homologous to the staphylococcal ccrAB genes and are designated ccrABEnt genes, were examined in enterococcal isolates (n=421) representing 13 different species. A total of 118 (28 %) isolates were positive for ccrABEnt genes by PCR, and a number of these were confirmed by Southern hybridization with a ccrAEnt probe (n=76) and partial DNA sequencing of ccrAEnt and ccrBEnt genes (n=38). ccrABEnt genes were present in Enterococcus faecium (58/216, 27 %), Enterococcus durans (31/38, 82 %), Enterococcus hirae (27/52, 50 %), Enterococcus casseliflavus (1/4, 25 %) and Enterococcus gallinarum (1/2, 50 %). In the eight other species tested, including Enterococcus faecalis (n=94), ccrABEnt genes were not found. Thirty-eight sequenced ccrABEnt genes from five different enterococcal species showed 94–100 % nucleotide sequence identity and linkage PCRs showed heterogeneity in the ccrABEnt flanking chromosomal genes. Expression analysis of ccrABEnt genes from the E. faecium DO strain showed constitutive expression as a bicistronic mRNA. The ccrABEnt mRNA levels were lower during log phase than stationary phase in relation to total mRNA. Multilocus sequence typing was performed on 39 isolates. ccrABEnt genes were detected in both hospital-related (10/29, 34 %) and non-hospital (4/10, 40 %) strains of E. faecium. Various sequence types were represented by both ccrABEnt positive and negative isolates, suggesting acquisition or loss of ccrABEnt in E. faecium. In summary, ccrABEnt genes, potentially involved in genome plasticity, are expressed in E. faecium and are widely distributed in the E. faecium and E. casseliflavus species groups. PMID:20817645

MBCP - Approach - Immunotherapy | Center for Cancer Research

Cancer.gov

Immunotherapy CCR investigators pioneered the use of the tuberculosis vaccine—Bacillus Calmette-Guerin (BCG)—in the treatment of bladder cancer. In cases where the tumor burden is not too high and direct contact can be made with the urothelium surface of the bladder, BCG application appears to elicit an immune response that attacks the tumor as well as the attenuated virus.

Innate immunity and HIV-1 infection.

PubMed

Lehner, T; Wang, Y; Whittall, T; Seidl, T

2011-04-01

HIV-1 is predominantly transmitted through mucosal tissues, targeting CD4(+)CCR5(+) T cells, 50% of which are destroyed within 2 weeks of infection. Conventional vaccination strategies have so far failed to prevent HIV-1 infection. Neither antibodies nor cytotoxic lymphocytes are capable of mounting a sufficiently rapid immune response to prevent early destruction of these cells. However, innate immunity is an early-response system, largely independent of prior encounter with a pathogen. Innate immunity can be classified into cellular, extracellular, and intracellular components, each of which is exemplified in this review by γδ T cells, CC chemokines, and APOBEC3G, respectively. First, γδ T cells are found predominantly in mucosal tissues and produce cytokines, CC chemokines, and antiviral factors. Second, the CC chemokines CCL-3, CCL-4, and CCL-5 can be upregulated by immunization of macaques with SIVgp120 and gag p27, and these can bind and downmodulate CCR5, thereby inhibiting HIV-1 entry into the host cells. Third, APOBEC3G is generated and maintained following rectal mucosal immunization in rhesus macaques for over 17 weeks, and the innate anti-SIV factor is generated by CD4(+)CD95(+)CCR7(-) effector memory T cells. Thus, innate anti-HIV-1 or SIV immunity can be linked with immune memory, mediated by CD4(+) T cells generating APOBEC3G. The multiple innate functions may mount an early anti-HIV-1 response and either prevent viral transmission or contain the virus until an effective adaptive immune response develops.

Green-synthesized silver nanoparticles as a novel control tool against dengue virus (DEN-2) and its primary vector Aedes aegypti

USDA-ARS?s Scientific Manuscript database

Dengue is an arthropod-borne viral infection mainly vectored through the bite of Aedes mosquitoes. Recently, its transmission has strongly increased in urban and semi-urban areas of tropical and sub-tropical regions worldwide, becoming a major international public health concern. There is no specifi...

CCL2 and CCR2 regulate pain-related behaviour and early gene expression in post-traumatic murine osteoarthritis but contribute little to chondropathy.

PubMed

Miotla Zarebska, J; Chanalaris, A; Driscoll, C; Burleigh, A; Miller, R E; Malfait, A M; Stott, B; Vincent, T L

2017-03-01

The role of inflammation in structural and symptomatic osteoarthritis (OA) remains unclear. One key mediator of inflammation is the chemokine CCL2, primarily responsible for attracting monocytes to sites of injury. We investigated the role of CCL2 and its receptor CCR2 in experimental OA. OA was induced in 10 weeks old male wild type (WT), Ccl2 -/- and Ccr2 -/- mice, by destabilisation of the medial meniscus (DMM). RNA was extracted from whole joints at 6 h and 7 days post-surgery and examined by reverse transcription polymerase chain reaction (RT-PCR). Gene expression changes between naïve and DMM-operated mice were compared. Chondropathy scores, from mice at 8, 12, 16 and 20 weeks post DMM were calculated using modified Osteoarthritis Research Society International (OARSI) grading systems. Changes in hind paw weight distribution, as a measure of pain, were assessed by Linton incapacitance. Absence of CCL2 strongly suppressed (>90%) selective inflammatory response genes in the joint 6 h post DMM, including arginase 1, prostaglandin synthase 2, nitric oxide synthase 2 and inhibin A. IL6, MMP3 and tissue inhibitor of metalloproteinase 1 were also significantly suppressed. Similar trends were also observed in the absence of CCR2. A lower average chondropathy score was observed in both Ccl2 -/- and Ccr2 -/- mice at 12, 16 and 20 weeks post DMM compared with WT mice, but this was only statistically significant at 20 weeks in Ccr2 -/- mice. Pain-related behaviour in Ccl2 -/- and Ccr2 -/- mice post DMM was delayed in onset. The CCL2/CCR2 axis plays an important role in the development of pain in murine OA, but contributes little to cartilage damage. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Evidence of CCR2-independent transmigration of Ly6C(hi) monocytes into the brain after permanent cerebral ischemia in mice.

PubMed

Chu, Hannah X; Kim, Hyun Ah; Lee, Seyoung; Broughton, Brad R S; Drummond, Grant R; Sobey, Christopher G

2016-04-15

Previously we showed that INCB3344, a CCR2 antagonist, inhibits transmigration of Ly6C(hi) monocytes into the brain after ischemia-reperfusion. Here we tested the effect of CCR2 inhibition during permanent cerebral ischemia. Mice were administered either vehicle (dimethyl sulfoxide/carboxymethylcellulose) or INCB3344 (30 or 100mg/kg IP) 1h before middle cerebral artery occlusion and at 2 and 6h after the initiation of ischemia. After 24h, we assessed functional outcome, infarct volume and quantified immune cells in blood and brain. The increase in circulating bone marrow-derived Ly6C(hi) monocytes, but not the infiltration of those cells into the brain, was blocked by the CCR2 antagonist. INCB3344 had no effect on either neurological deficit or infarct volume. Our data confirm that cerebral ischemia triggers a CCR2-dependent increase in circulating Ly6C(hi) monocytes, but suggest that in the absence of reperfusion these cells may transmigrate into the ischemic brain in a CCR2-independent manner. Copyright © 2016 Elsevier B.V. All rights reserved.

Bioinformatic analysis of neurotropic HIV envelope sequences identifies polymorphisms in the gp120 bridging sheet that increase macrophage-tropism through enhanced interactions with CCR5

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mefford, Megan E., E-mail: megan_mefford@hms.harvard.edu; Kunstman, Kevin, E-mail: kunstman@northwestern.edu; Wolinsky, Steven M., E-mail: s-wolinsky@northwestern.edu

Macrophages express low levels of the CD4 receptor compared to T-cells. Macrophage-tropic HIV strains replicating in brain of untreated patients with HIV-associated dementia (HAD) express Envs that are adapted to overcome this restriction through mechanisms that are poorly understood. Here, bioinformatic analysis of env sequence datasets together with functional studies identified polymorphisms in the β3 strand of the HIV gp120 bridging sheet that increase M-tropism. D197, which results in loss of an N-glycan located near the HIV Env trimer apex, was detected in brain in some HAD patients, while position 200 was estimated to be under positive selection. D197 andmore » T/V200 increased fusion and infection of cells expressing low CD4 by enhancing gp120 binding to CCR5. These results identify polymorphisms in the HIV gp120 bridging sheet that overcome the restriction to macrophage infection imposed by low CD4 through enhanced gp120–CCR5 interactions, thereby promoting infection of brain and other macrophage-rich tissues. - Highlights: • We analyze HIV Env sequences and identify amino acids in beta 3 of the gp120 bridging sheet that enhance macrophage tropism. • These amino acids at positions 197 and 200 are present in brain of some patients with HIV-associated dementia. • D197 results in loss of a glycan near the HIV Env trimer apex, which may increase exposure of V3. • These variants may promote infection of macrophages in the brain by enhancing gp120–CCR5 interactions.« less

Structure-kinetic relationships--an overlooked parameter in hit-to-lead optimization: a case of cyclopentylamines as chemokine receptor 2 antagonists.

PubMed

Vilums, Maris; Zweemer, Annelien J M; Yu, Zhiyi; de Vries, Henk; Hillger, Julia M; Wapenaar, Hannah; Bollen, Ilse A E; Barmare, Farhana; Gross, Raymond; Clemens, Jeremy; Krenitsky, Paul; Brussee, Johannes; Stamos, Dean; Saunders, John; Heitman, Laura H; Ijzerman, Adriaan P

2013-10-10

Preclinical models of inflammatory diseases (e.g., neuropathic pain, rheumatoid arthritis, and multiple sclerosis) have pointed to a critical role of the chemokine receptor 2 (CCR2) and chemokine ligand 2 (CCL2). However, one of the biggest problems of high-affinity inhibitors of CCR2 is their lack of efficacy in clinical trials. We report a new approach for the design of high-affinity and long-residence-time CCR2 antagonists. We developed a new competition association assay for CCR2, which allows us to investigate the relation of the structure of the ligand and its receptor residence time [i.e., structure-kinetic relationship (SKR)] next to a traditional structure-affinity relationship (SAR). By applying combined knowledge of SAR and SKR, we were able to re-evaluate the hit-to-lead process of cyclopentylamines as CCR2 antagonists. Affinity-based optimization yielded compound 1 with good binding (Ki = 6.8 nM) but very short residence time (2.4 min). However, when the optimization was also based on residence time, the hit-to-lead process yielded compound 22a, a new high-affinity CCR2 antagonist (3.6 nM), with a residence time of 135 min.

CCR2-dependent Gr1high monocytes promote kidney injury in shiga toxin-induced hemolytic uremic syndrome in mice.

PubMed

Pohl, Judith-Mira; Volke, Julia K; Thiebes, Stephanie; Brenzel, Alexandra; Fuchs, Kerstin; Beziere, Nicolas; Ehrlichmann, Walter; Pichler, Bernd J; Squire, Anthony; Gueler, Faikah; Engel, Daniel R

2018-06-01

The hemolytic uremic syndrome (HUS) is a life-threatening disease of the kidney that is induced by shiga toxin-producing E.coli. Major changes in the monocytic compartment and in CCR2-binding chemokines have been observed. However, the specific contribution of CCR2-dependent Gr1 high monocytes is unknown. To investigate the impact of these monocytes during HUS, we injected a combination of LPS and shiga toxin into mice. We observed an impaired kidney function and elevated levels of the CCR2-binding chemokine CCL2 after shiga toxin/LPS- injection, thus suggesting Gr1 high monocyte infiltration into the kidney. Indeed, the number of Gr1 high monocytes was strongly increased one day after HUS induction. Moreover, these cells expressed high levels of CD11b suggesting activation after tissue entry. Non-invasive PET-MR imaging revealed kidney injury mainly in the kidney cortex and this damage coincided with the detection of Gr1 high monocytes. Lack of Gr1 high monocytes in Ccr2-deficient animals reduced neutrophil gelatinase-associated lipocalin and blood urea nitrogen levels. Moreover, the survival of Ccr2-deficient animals was significantly improved. Conclusively, this study demonstrates that CCR2-dependent Gr1 high monocytes contribute to the kidney injury during HUS and targeting these cells is beneficial during this disease. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Helper-Dependent Properties of Friend Spleen Focus-Forming Virus: Effect of the Fv-1 Gene on the Late Stages in Virus Synthesis

PubMed Central

Eckner, Robert J.

1973-01-01

Co-infection of neonatal BALB/c mice with Friend virus (FV) complex (containing defective spleen focus-forming virus [SFFV] and endogenous N-tropic leukemia-inducing helper virus [LLV-F]) and B-tropic Tennant leukemia virus (TenLV) resulted in the inhibition of LLV-F by the Fv-1b gene and recovery of a TenLV pseudotype of SFFV, abbreviated SFFV(TenLV). The host range of this pseudotype was B-tropic, since SFFV(TenLV) was 10 to 100 times more infectious for B-type (Fv-1bb) than for N-type (Fv-1nn) mice. The similar patterns of neutralization of N-tropic and B-tropic SFFV by type-specific murine antisera suggested that the difference in infectivity between these two SFFV preparations did not reside in envelope determinants. Rather, helper control of SFFV's host range was only apparent and dependent upon the ability of associated virus to provide a helper function for late stages in SFFV synthesis. Early stages in SFFV's infectious cycle were shown to be helper independent. The Fv-1 gene did not act at the level of the cell membrane to effectively restrict SFFV infection, since SFFV-induced transformed cells could be detected in the absence of spleen focus formation and SFFV synthesis. Further, the generation of these transformed cells by SFFV followed a one-hit, dose-response pattern, suggesting that SFFV-induced cell transformation is helper independent. Finally, restriction of helper function by Fv-1 may be an intracellular event, because both SFFV and its associated LLV-F helper share common envelope determinants and presumably adsorb onto and penetrate target cells with equal efficiency. PMID:4127030

Characterizing Myeloid Cell Activation in NF1 Vasculopathy

DTIC Science & Technology

2017-07-01

stimulation of its receptor (CCR2) and the generation of reactive oxygen species, which are generated in excessive quantities by neurofibromin-deficient...macrophages via monocyte chemotactic peptide-1 (MCP-1) stimulation of its receptor (CCR2) and the generation of reactive oxygen species, which are...neurofibromatosis; stenosis; aneurysm; MCP-1; CCR2; reactive oxygen species; superoxide; macrophages; monocytes; arteries; cardiovascular disease Major

A dynamic spectrum of monocytes arising from the in situ reprogramming of CCR2+ monocytes at a site of sterile injury

PubMed Central

Dal-Secco, Daniela; Wang, Jing; Zeng, Zhutian; Kolaczkowska, Elzbieta; Wong, Connie H.Y.; Petri, Björn; Ransohoff, Richard M.; Charo, Israel F.

2015-01-01

Monocytes are recruited from the blood to sites of inflammation, where they contribute to wound healing and tissue repair. There are at least two subsets of monocytes: classical or proinflammatory (CCR2hiCX3CR1low) and nonclassical, patrolling, or alternative (CCR2lowCX3CR1hi) monocytes. Using spinning-disk confocal intravital microscopy and mice with fluorescent reporters for each of these subsets, we were able to track the dynamic spectrum of monocytes that enter a site of sterile hepatic injury in vivo. We observed that the CCR2hiCX3CR1low monocytes were recruited early and persisted for at least 48 h, forming a ringlike structure around the injured area. These monocytes transitioned, in situ, from CCR2hiCx3CR1low to CX3CR1hiCCR2low within the ringlike structure and then entered the injury site. This phenotypic conversion was essential for optimal repair. These results demonstrate a local, cytokine driven reprogramming of classic, proinflammatory monocytes into nonclassical or alternative monocytes to facilitate proper wound-healing. PMID:25800956

Downregulation of mouse CCR3 by lentiviral shRNA inhibits proliferation and induces apoptosis of mouse eosinophils.

PubMed

Zhu, Xin-Hua; Liao, Bing; Xu, Yi; Liu, Ke; Huang, Yun; Huang, Quan-Long; Liu, Yue-Hui

2017-02-01

RNA interference has been considered as an effective gene silencing method in basic and preclinical investigations. The aims of the present study were to construct a lentiviral vector expressing a short hairpin RNA (shRNA) targeting the murine CC chemokine receptor 3 (mCCR3), and to investigate its effects on the proliferation and apoptosis of mouse eosinophils. A recombinant lentiviral vector expressing four fragments of mouse CCR3 shRNA (pLVX‑mCCR3‑1+2+3+4‑shRNA) was constructed using subcloning techniques. This novel lentivirus was then packaged into 293T cells by co‑transduction with plasmids, including Baculo p35, pCMV R8.2 and VSV. The interference effects of the vector were verified using polymerase chain reaction (PCR) and western blot analyses. The effects of the interference on the proliferation and apoptosis of mouse eosinophils were investigated using 3‑(4,5‑dimethylthiazol‑2‑yl)‑5‑(3‑carboxymethoxyphenyl)‑2‑(4‑sulfophenyl)‑2H‑tetrazolium and terminal deoxynucleotidyl transferase dUTP nick end labeling methods, respectively. The results of the PCR and western blot analyses confirmed that the novel recombinant vector, pLVX‑mCCR3‑1+2+3+4‑shRNA, had high efficiency in inhibiting the mRNA and protein expression levels of mCCR3 in mouse eosinophils. The downregulation of mCCR3 significantly inhibited proliferation of the eosinophils. Furthermore, the present study found that the downregulation of mCCR3 significantly promoted apoptosis of the eosinophils. Therefore, the downregulation of mCCR3 led to the inhibition of proliferation and induction of apoptosis in mouse eosinophils. The predominant characteristics of allergic rhinitis are eosinophil infiltration and release of inflammatory mediators, which appear in a variety of clinical manifestations. The results of the present study indicate that mCCR3 silencing may serve as a putative approach for the treatment of allergic rhinitis.

Multimodality vaccination against clade C SHIV: partial protection against mucosal challenges with a heterologous tier 2 virus.

PubMed

Lakhashe, Samir K; Byrareddy, Siddappa N; Zhou, Mingkui; Bachler, Barbara C; Hemashettar, Girish; Hu, Shiu-Lok; Villinger, Francois; Else, James G; Stock, Shannon; Lee, Sandra J; Vargas-Inchaustegui, Diego A; Cofano, Egidio Brocca; Robert-Guroff, Marjorie; Johnson, Welkin E; Polonis, Victoria R; Forthal, Donald N; Loret, Erwann P; Rasmussen, Robert A; Ruprecht, Ruth M

2014-11-12

We sought to test whether vaccine-induced immune responses could protect rhesus macaques (RMs) against upfront heterologous challenges with an R5 simian-human immunodeficiency virus, SHIV-2873Nip. This SHIV strain exhibits many properties of transmitted HIV-1, such as tier 2 phenotype (relatively difficult to neutralize), exclusive CCR5 tropism, and gradual disease progression in infected RMs. Since no human AIDS vaccine recipient is likely to encounter an HIV-1 strain that exactly matches the immunogens, we immunized the RMs with recombinant Env proteins heterologous to the challenge virus. For induction of immune responses against Gag, Tat, and Nef, we explored a strategy of immunization with overlapping synthetic peptides (OSP). The immune responses against Gag and Tat were finally boosted with recombinant proteins. The vaccinees and a group of ten control animals were given five low-dose intrarectal (i.r.) challenges with SHIV-2873Nip. All controls and seven out of eight vaccinees became systemically infected; there was no significant difference in viremia levels of vaccinees vs. controls. Prevention of viremia was observed in one vaccinee which showed strong boosting of virus-specific cellular immunity during virus exposures. The protected animal showed no challenge virus-specific neutralizing antibodies in the TZM-bl or A3R5 cell-based assays and had low-level ADCC activity after the virus exposures. Microarray data strongly supported a role for cellular immunity in the protected animal. Our study represents a case of protection against heterologous tier 2 SHIV-C by vaccine-induced, virus-specific cellular immune responses. Copyright © 2014 Elsevier Ltd. All rights reserved.

Rapamycin causes down-regulation of CCR5 and accumulation of anti-HIV β-chemokines: An approach to suppress R5 strains of HIV-1

PubMed Central

Heredia, A.; Amoroso, A.; Davis, C.; Le, N.; Reardon, E.; Dominique, J. K.; Klingebiel, E.; Gallo, R. C.; Redfield, R. R.

2003-01-01

Propagation of R5 strains of HIV-1 on CD4 lymphocytes and macrophages requires expression of the CCR5 coreceptor on the cell surface. Individuals lacking CCR5 (CCR5Δ32 homozygous genotype) are phenotypically normal and resistant to infection with HIV-1. CCR5 expression on lymphocytes depends on signaling through the IL-2 receptor. By FACS analysis we demonstrate that rapamycin (RAPA), a drug that disrupts IL-2 receptor signaling, reduces CCR5 surface expression on T cells at concentrations as low as 1 nM. In addition, lower concentrations of RAPA (0.01 nM) were sufficient to reduce CCR5 surface expression on maturing monocytes. PCR analysis on peripheral blood mononuclear cells (PBMCs) showed that RAPA interfered with CCR5 expression at the transcriptional level. Reduced expression of CCR5 on PBMCs cultured in the presence of RAPA was associated with increased extracellular levels of macrophage inflammatory protein (MIP)-1α and MIP-1β. In infectivity assays, RAPA suppressed the replication of R5 strains of HIV-1 both in PBMC and macrophage cultures. In total PBMC cultures, RAPA-mediated inhibition of CCR5-using strains of HIV-1 occurred at 0.01 nM, a concentration of drug that is ∼103 times lower than therapeutic through levels of drug in renal transplant recipients. In addition, RAPA enhanced the antiviral activity of the CCR5 antagonist TAK-779. These results suggest that low concentrations of RAPA may have a role in both the treatment and prevention of HIV-1 infection. PMID:12915736

Transfection of Fv-1 permissive and restrictive mouse cells with integrated DNA of murine leukemia viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hsu, I.C.; Yang, W.K.; Tennant, R.W.

1978-03-01

Whole-cell DNA preparations isolated from SC-1 cells chronically infected with N- or B-tropic murine leukemia viruses (MuLV) were tested for infectious activity in an Fv-I/sup n/ (NIH-3T3) and two Fv-I/sup b/ (C57BL/6 and SV-A31) cell cultures. Efficiency of transfection for all DNAs was better in the NIH-3T3 cells than in C57BL/6 or SV-A31 cells; and an (N-tropic MuLV)SC-1 cell DNA preparation was slightly more infectious than a (B-tropic MuLV)SC-1 cell DNA preparation in all three cell cultures, regardless of their Fv-1 genotypes. Progeny viruses from the transfection showed N- or B-tropism corresponding to that of the parent viruses produced bymore » the infected SC-1 cells that were used for the DNA preparation. DNA dose-response studies in NIH-3T3 cells revealed a one-hit mechanism for both the (B-tropic MuLV)SC-1 cell DNA and the (N-tropic MuLV)SC-1 cell DNA preparation. These results demonstrate that, in contrast to virion infection, transfection of N- and B-tropic MuLV with DNA preparations from chronically infected cells is not affected by the Fv-1 gene.« less

Transfection of Fv-1 permissive and restrictive mouse cells with integrated DNA of murine leukemia viruses (host range restriction/Fv-1 gene/DNA transfection)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hsu, I.C.; Yang, W.K.; Tennant, R.W.

1978-03-01

Whole-cell DNA preparations isolated from SC-1 cells chronically infected with N- or B-tropic murine leukemia viruses (MuLV) were tested for infectious activity in an Fv-1/sup n/ (NIH-3T3) and two Fv-1/sup b/ (C57BL/6 and SV-A31) cell cultures. Efficiency of transfection for all DNAs was better in the NIH-3T3 cells than in C57BL/6 or SV-A31 cells; and an (N-tropic MuLV)SC-1 cell DNA preparation was slightly more infectious than a (B-tropic MuLV)SC-1 cell DNA preparation in all three cell cultures, regardless of their Fv-1 geonotypes. Progeny viruses from the transfection showed N- or B-tropism corresponding to that of the parent viruses produced bymore » the infected SC-I cells that were used for the DNA preparation. DNA dose-response studies in NIH-3T3 cells revealed a one-hit mechanism for both the (B-tropic MuLV)SC-1 cell DNA and the (N-tropic MuLV)SC-1 cell DNA preparation. These results demonstrate that, in contrast to virion infection, transfection of N- and B-tropic MuLV with DNA preparations from chronically infected cells is not affected by the Fv-1 gene.« less

Improving Dengue Virus Capture Rates in Humans and Vectors in Kamphaeng Phet Province, Thailand, Using an Enhanced Spatiotemporal Surveillance Strategy

DTIC Science & Technology

2015-05-18

THOMAS AND OTHERS ENHANCED SURVEILLANCE FOR DENGUE Improving Dengue Virus Capture Rates in Humans and Vectors in Kamphaeng Phet Province...of Medical Sciences, Bangkok, Thailand. Abstract. Dengue is of public health importance in tropical and sub-tropical regions. Dengue virus (DENV...with confirmed dengue (initiates) and associated cluster individuals (associates) with entomologic sampling. A total of 438 associates were enrolled

An Anti-HIV-1 V3 Loop Antibody Fully Protects Cross-Clade and Elicits T-Cell Immunity in Macaques Mucosally Challenged with an R5 Clade C SHIV

PubMed Central

Lakhashe, Samir K.; Humbert, Michael; Sholukh, Anton; Hemashettar, Girish; Wong, Yin Ling; Yoon, John K.; Wang, Wendy; Novembre, Francis J.; Villinger, Francois; Ibegbu, Chris; Patel, Kalpana; Corti, Davide; Agatic, Gloria; Vanzetta, Fabrizia; Bianchi, Siro; Heeney, Jonathan L.; Sallusto, Federica; Lanzavecchia, Antonio; Ruprecht, Ruth M.

2011-01-01

Neutralizing antibodies have been shown to protect macaques against SHIV challenge. However, genetically diverse HIV-1 clades have evolved, and a key question left unanswered is whether neutralizing antibodies can confer cross-clade protection in vivo. The novel human monoclonal antibody HGN194 was isolated from an individual infected with an HIV-1 clade AG recombinant circulating recombinant form (CRF). HGN194 targets an epitope in the third hypervariable loop (V3) of HIV-1 gp120 and neutralizes a range of relatively neutralization-sensitive and resistant viruses. We evaluated the potential of HGN194 to protect infant rhesus monkeys against a SHIV encoding a primary CCR5-tropic HIV-1 clade C envelope. After high-dose mucosal challenge, all untreated controls became highly viremic while all HGN194-treated animals (50 mg/kg) were completely protected. When HGN194 was given at 1 mg/kg, one out of two monkeys remained aviremic, whereas the other had delayed, lower peak viremia. Interestingly, all protected monkeys given high-dose HGN194 developed Gag-specific proliferative responses of both CD4+ and CD8+ T cells. To test whether generation of the latter involved cryptic infection, we ablated CD8+ cells after HGN194 clearance. No viremia was detected in any protected monkeys, thus ruling out virus reservoirs. Thus, induction of CD8 T-cell immunity may have resulted from transient “Hit and Run” infection or cross priming via Ag-Ab-mediated cross-presentation. Together, our data identified the HGN194 epitope as protective and provide proof-of-concept that this anti-V3 loop mAb can prevent infection with sterilizing immunity after challenge with virus of a different clade, implying that V3 is a potential vaccine target. PMID:21483815

A NEW COPPER (II)-IMIDAZOLE DERIVATIVE EFFECTIVELY INHIBITS REPLICATION OF DENV-2 IN VERO CELL

PubMed Central

Sucipto, Teguh Hari; Churrotin, Siti; Setyawati, Harsasi; Martak, Fahimah; Mulyatno, Kris Cahyo; Amarullah, Ilham Harlan; Kotaki, Tomohiro; Kameoka, Masanori; Yotopranoto, Subagyo; Soegijanto, and Soegeng

2018-01-01

Background: Dengue is a kind of infectious disease that was distributed in the tropical and sub-tropical areas. To date, there is no clinically approved dengue vaccine or antiviral for humans, even though there have been great efforts towards this end. Therefore, finding the effective compound against dengue virus (DENV) replication is very important. Among the complex compounds, copper(II)-imidazole derivatives are of interest because of their biological and medicinal benefits. Materials and Methods: In the present study, antiviral activity of [Cu(2,4,5-triphenylimidazole)2]n, was evaluated against different stages of dengue virus type 2 (DENV-2) replication in Vero cell using focus forming unit reduction assay and quantitative ELISA. Results: [Cu(2,4,5-triphenylimidazole)2]n inhibited DENV-2 replication in Vero cells with IC50 = 2.3 μg/ml and SI= 19.42 when cells were treated 2 days after virus infection, whereas its CC50 for cytotoxicity to Vero cells was 44.174 μg/ml. Conclusion: The compound has high anti-DENV2 activity, less toxicity, and a high possibility to be considered a drug candidate. PMID:29619441

Increased loss of CCR5+ CD45RA- CD4+ T cells in CD8+ lymphocyte-depleted Simian immunodeficiency virus-infected rhesus monkeys.

PubMed

Veazey, Ronald S; Acierno, Paula M; McEvers, Kimberly J; Baumeister, Susanne H C; Foster, Gabriel J; Rett, Melisa D; Newberg, Michael H; Kuroda, Marcelo J; Williams, Kenneth; Kim, Eun-Young; Wolinsky, Steven M; Rieber, E Peter; Piatak, Michael; Lifson, Jeffrey D; Montefiori, David C; Brown, Charles R; Hirsch, Vanessa M; Schmitz, Jörn E

2008-06-01

Previously we have shown that CD8(+) T cells are critical for containment of simian immunodeficiency virus (SIV) viremia and that rapid and profound depletion of CD4(+) T cells occurs in the intestinal tract of acutely infected macaques. To determine the impact of SIV-specific CD8(+) T-cell responses on the magnitude of the CD4(+) T-cell depletion, we investigated the effect of CD8(+) lymphocyte depletion during primary SIV infection on CD4(+) T-cell subsets and function in peripheral blood, lymph nodes, and intestinal tissues. In peripheral blood, CD8(+) lymphocyte-depletion changed the dynamics of CD4(+) T-cell loss, resulting in a more pronounced loss 2 weeks after infection, followed by a temporal rebound approximately 2 months after infection, when absolute numbers of CD4(+) T cells were restored to baseline levels. These CD4(+) T cells showed a markedly skewed phenotype, however, as there were decreased levels of memory cells in CD8(+) lymphocyte-depleted macaques compared to controls. In intestinal tissues and lymph nodes, we observed a significantly higher loss of CCR5(+) CD45RA(-) CD4(+) T cells in CD8(+) lymphocyte-depleted macaques than in controls, suggesting that these SIV-targeted CD4(+) T cells were eliminated more efficiently in CD8(+) lymphocyte-depleted animals. Also, CD8(+) lymphocyte depletion significantly affected the ability to generate SIV Gag-specific CD4(+) T-cell responses and neutralizing antibodies. These results reemphasize that SIV-specific CD8(+) T-cell responses are absolutely critical to initiate at least partial control of SIV infection.

Genetic interactions in the spontaneous production of endogenous murine leukemia virus in low leukemic mouse strains

PubMed Central

1982-01-01

The spontaneous expression of ecotropic murine leukemia virus (MuLV) in spleen cells of BALB/c, C57BL/6 (B6), and derivative mice was examined as a function of age. The patterns of spontaneous virus induction in vivo correlate with the patterns of virus induction in vitro, which result from the action of two loci, Inc-l and Inb-l (7). Whereas mice carrying Inc-l or Inb-l have similar phenotypes in vitro, they have significantly different phenotypes in vivo. Mice of the Inb-l+/+ genotype, e.g., B6, rarely expressed MuLV, and the titer of MuLV recovered from rare MuLV-positive mice of this genotype was usually low. Mice of the Inc-l+/+ genotype, e.g., BALB/c, expressed low amounts of MuLV early in life, however, from 6-12 mo of age approximately one- half of the Inc-l+/+ mice expressed virus, frequently of high titer. Equal numbers of N-tropic and B-tropic MuLV were recovered from Inb-l+ mice, but predominantly N-tropic MuLV was recovered from Inc-l+ mice. Strains that carry dominant (+) alleles at both Inc-l and Inb-l show higher titers of MuLV earlier in life than strains that carry only Inc- l or Inb-l. The presence of dominant alleles at both loci results in the appearance of predominantly N-tropic virus early in life. These results demonstrate that the principal determinants of spontaneous virus expression in these low leukemic strains of mice are the In loci or genes linked to them. A further inference that can be drawn from these studies is that the appearance of B-tropic virus is by no means a random process but rather results from predictable patterns of MuLV expression and alteration. PMID:6284854

CD44+ Cancer Stem-Like Cells in EBV-Associated Nasopharyngeal Carcinoma

PubMed Central

Lun, Samantha Wei-Man; Cheung, Siu Tim; Cheung, Phyllis Fung Yi; To, Ka-Fai; Woo, John Kong-Sang; Choy, Kwong-Wai; Chow, Chit; Cheung, Chartia Ching-Mei; Chung, Grace Tin-Yun; Cheng, Alice Suk-Hang; Ko, Chun-Wai; Tsao, Sai-Wah; Busson, Pierre; Ng, Margaret Heung-Ling; Lo, Kwok-Wai

2012-01-01

Nasopharyngeal carcinoma (NPC) is a unique EBV-associated epithelial malignancy, showing highly invasive and metastatic phenotype. Despite increasing evidence demonstrating the critical role of cancer stem-like cells (CSCs) in the maintenance and progression of tumors in a variety of malignancies, the existence and properties of CSC in EBV-associated NPC are largely unknown. Our study aims to elucidate the presence and role of CSCs in the pathogenesis of this malignant disease. Sphere-forming cells were isolated from an EBV-positive NPC cell line C666-1 and its tumor-initiating properties were confirmed by in vitro and in vivo assays. In these spheroids, up-regulation of multiple stem cell markers were found. By flow cytometry, we demonstrated that both CD44 and SOX2 were overexpressed in a majority of sphere-forming C666-1 cells. The CD44+SOX2+ cells was detected in a minor population in EBV-positive xenografts and primary tumors and considered as potential CSC in NPC. Notably, the isolated CD44+ NPC cells were resistant to chemotherapeutic agents and with higher spheroid formation efficiency, showing CSC properties. On the other hand, microarray analysis has revealed a number of differentially expressed genes involved in transcription regulation (e.g. FOXN4, GLI1), immune response (CCR7, IL8) and transmembrane transport (e.g. ABCC3, ABCC11) in the spheroids. Among these genes, increased expression of CCR7 in CD44+ CSCs was confirmed in NPC xenografts and primary tumors. Importantly, blocking of CCR7 abolished the sphere-forming ability of C666-1 in vitro. Expression of CCR7 was associated with recurrent disease and distant metastasis. The current study defined the specific properties of a CSC subpopulation in EBV-associated NPC. Our findings provided new insights into developing effective therapies targeting on CSCs, thereby potentiating treatment efficacy for NPC patients. PMID:23285037

Clinical and Virological Descriptive Study in the 2011 Outbreak of Dengue in the Amazonas, Brazil

PubMed Central

Martins, Valquiria do Carmo Alves; Bastos, Michele de Souza; Ramasawmy, Rajendranath; de Figueiredo, Regina Pinto; Gimaque, João Bosco Lima; Braga, Wornei Silva Miranda; Nogueira, Mauricio Lacerda; Nozawa, Sergio; Naveca, Felipe Gomes; Figueiredo, Luiz Tadeu Moraes; Mourão, Maria Paula Gomes

2014-01-01

Background Dengue is a vector-borne disease in the tropical and subtropical region of the world and is transmitted by the mosquito Aedes aegypti. In the state of Amazonas, Brazil during the 2011 outbreak of dengue all the four Dengue virus (DENV) serotypes circulating simultaneously were observed. The aim of the study was to describe the clinical epidemiology of dengue in Manaus, the capital city of the state of the Amazonas, where all the four DENV serotypes were co-circulating simultaneously. Methodology Patients with acute febrile illness during the 2011 outbreak of dengue, enrolled at the Fundação de Medicina Tropical Dr. Heitor Viera Dourado (FMT-HVD), a referral centre for tropical and infectious diseases in Manaus, were invited to participate in a clinical and virological descriptive study. Sera from 677 patients were analyzed by RT-nested-PCRs for flaviviruses (DENV 1–4, Saint Louis encephalitis virus-SLEV, Bussuquara virus-BSQV and Ilheus virus-ILHV), alphavirus (Mayaro virus-MAYV) and orthobunyavirus (Oropouche virus-OROV). Principal Findings Only dengue viruses were detected in 260 patients (38.4%). Thirteen patients were co-infected with more than one DENV serotype and six (46.1%) of them had a more severe clinical presentation of the disease. Nucleotide sequencing showed that DENV-1 belonged to genotype V, DENV-2 to the Asian/American genotype, DENV-3 to genotype III and DENV-4 to genotype II. Conclusions Co-infection with more than one DENV serotype was observed. This finding should be warning signs to health authorities in situations of the large dispersal of serotypes that are occurring in the world. PMID:24978469

Investigation on the phytosanitary status of major ornamental hibiscus species in Italy to assess virus infection

USDA-ARS?s Scientific Manuscript database

The genus Hibiscus (family Malvaceae) includes about 250 species that vary from annual to perennial herbs, and shrubs to small trees that are native to tropical, sub-tropical and temperate climates. A study in 2010-2011 examined viruses associated with symptoms observed on hibiscus plants in Italy....

Tropical arthritogenic alphaviruses.

PubMed

Mejía, Carla-Ruth; López-Vélez, Rogelio

Tropical alphaviruses have special tropism for bone and joint tissue. Patients can develop chronic rheumatic disorders similar to rheumatoid arthritis and ankylosing spondylitis. The prototype is Chikungunya virus, although other lesser known viruses in our environment such as Sindbis, Ross River, Mayaro, O'nyong nyong and Barmah Forest viruses have the potential to be sped through vectors and cause chronic rheumatic disease. International population movements have increased the numbers of patients diagnosed with these tropical viruses in areas in which they are not endemic. Since they can leave persistent symptoms and affect the quality of life of the patients, it is important that we be aware of them. Changes in ecosystems have favored the expansion of competent mosquitoes, making fears of local transmission in southern Europe a reality. The objective of this review is to provide a clinical approach to the different arthritogenic tropical alphaviruses, especially those in which chronic rheumatic disease is more frequent. Copyright © 2017 Elsevier España, S.L.U. and Sociedad Española de Reumatología y Colegio Mexicano de Reumatología. All rights reserved.

Longitudinal Analysis of Cerebrospinal Fluid and Plasma HIV-1 Envelope Sequences Isolated From a Single Donor with HIV Asymptomatic Neurocognitive Impairment

PubMed Central

Vázquez-Santiago, Fabián; García, Yashira; Rivera-Román, Ivelisse; Noel, Richard J.; Wojna, Valerie; Meléndez, Loyda M.; Rivera-Amill, Vanessa

2015-01-01

Objective Combined antiretroviral treatment (cART) has changed the clinical presentation of HIV-associated neurocognitive disorders (HAND) to that of the milder forms of the disease. Asymptomatic neurocognitive impairment (ANI) is now more prevalent and is associated with increased morbidity and mortality risk in HIV-1–infected people. HIV-1 envelope (env) genetic heterogeneity has been detected within the central nervous system (CNS) of individuals with ANI. Changes within env determine co-receptor use, cellular tropism, and neuropathogenesis. We hypothesize that compartmental changes are associated with HIV-1 env C2V4 during ANI and sought to analyze paired HIV-1 env sequences from plasma and cerebrospinal fluid (CSF) of a female subject undergoing long-term cART. Methods Paired plasma and CSF samples were collected at 12-month intervals and HIV-1 env C2V4 was cloned and sequenced. Results Phylogenetic analysis of paired samples consistently showed genetic variants unique to the CSF. Phenotypic prediction showed CCR5 (R5) variants for all CSF-derived sequences and showed minor X4 variants (or dual-tropic) in the plasma at later time points. Viral compartmentalization was evident throughout the study, suggesting that the occurrence of distinctive env strains may contribute to the neuropathogenesis of HAND. Conclusions Our study provides new insights about the genetic characteristics within the C2V4 of HIV-1 env that persist after long-term cART and during the course of persistent ANI. PMID:26167513

Eotaxin-1 promotes prostate cancer cell invasion via activation of the CCR3-ERK pathway and upregulation of MMP-3 expression.

PubMed

Zhu, Feng; Liu, Pei; Li, Jun; Zhang, Yan

2014-05-01

Chemokines have been reported to play crucial roles in tumor progression. Eotaxin-1 (CCL11), a member of the CC chemokine family, is elevated in many types of human cancer. Here, to reveal the molecular mechanisms of eotaxin-1 in prostate cancer cell invasion, the expression of eotaxin-1 receptors [CC chemokine receptor (CCR)2, CCR3 and CCR5] were silenced by small interfering RNA (siRNA). The ERK pathway was inhibited by the specific MEK inhibitor U0126. The role of eotaxin-1 and the CCR3-ERK pathway in prostate cancer cell invasion was assessed by invasion and migration assays. MMP-3 expression was detected by real-time PCR and ELISA assay. The results demonstrated that eotaxin-1 promoted the invasion and migration of DU-145 cells, and increased ERK1/2 activation and MMP-3 expression. Knockdown of CCR3 inhibited the invasion and migration of prostate cancer cells, and attenuated the eotaxin-1-induced ERK1/2 activation and MMP-3 expression. Furthermore, inactivation of the ERK pathway suppressed the eotaxin‑1-promoted invasion and migration, and decreased MMP-3 expression in the prostate cancer cells. Together, the present study suggests that eotaxin-1 increases MMP-3 expression via the CCR3-ERK pathway, thereby promoting prostate cancer cell invasion and migration. Thus, therapies that block eotaxin-1 and CCR3 may be effective interventions for prostate cancer.

CCL22-specific Antibodies Reveal That Engagement of Two Distinct Binding Domains on CCL22 Is Required for CCR4-mediated Function.

PubMed

Santulli-Marotto, Sandra; Wheeler, John; Lacy, Eilyn R; Boakye, Ken; Luongo, Jennifer; Wu, Sheng-Jiun; Ryan, Mary

2015-12-01

CCL22 inactivation in vivo occurs by cleavage at the N-terminus; however, it is unclear whether this encompasses the entire site of CCR4 interaction. CCL17 also binds CCR4 and its function requires binding via two discrete binding sites. Using monoclonal antibodies (MAbs), we report that there are two separate sites on CCL22 that are required for CCR4-mediated function. The CCL22-specific antibodies bind with affinities of 632 ± 297 pM (MC2B7) and 308 ± 43 pM (MAB4391) and neither exhibited detectable binding to CCL17. Both antibodies are comparable in their ability to inhibit CCL22-mediated calcium mobilization; however, competition binding studies demonstrate that MC2B7 and MAB4391 bind to distinct epitopes on CCL22. Both antibodies inhibit function through CCR4, which is demonstrated by loss of β-arrestin recruitment in a reporter cell line. In both assays, blocking either site independently abolished CCL22 function, suggesting that concurrent engagement of both sites with CCR4 is necessary for function. This is the first demonstration that CCL22 has two distinct binding sites that are required for CCR4 function. These antibodies are valuable tools for better understanding the interaction and function of CCL22 and CCR4 and will potentially help further understanding of the differential outcomes of CCL17 and CCL22 interaction with CCR4.

An Improved X-Band Maser System for Deep Space Network Applications

NASA Astrophysics Data System (ADS)

Britcliffe, M.; Hanson, T.; Fernandez, J.

2000-01-01

An 8450-MHz (X-band) maser system utilizing a commercial Gifford--McMahon (GM) closed-cycle cryocooler (CCR) was designed, fabricated, and demonstrated. The CCR system was used to cool a maser operating at 8450 MHz. The prototype GM CCR system meets or exceeds all Deep Space Network requirements for maser performance. The two-stage GM CCR operates at 4.2 K; for comparison, the DSN's current three-stage cryocooler, which uses a Joule--Thompson cooling stage in addition to GM cooling, operates at 4.5 K. The new CCR withstands heat loads of 1.5 W at 4.2 K as compared to 1 W at 4.5 K for the existing DSN cryocooler used for cooling masers. The measured noise temperature, T_e, of the maser used for these tests is defined at the ambient connection to the antenna feed system. The T_e measured 5.0 K at a CCR temperature of 4.5 K, about 1.5 K higher than the noise temperature of a typical DSN Block II-A X-band traveling-wave maser (TWM). Reducing the temperature of the CCR significantly lowers the maser noise temperature and increases maser gain and bandwidth. The new GM CCR gives future maser systems significant operational advantages, including reduced maintenance time and logistics requirements. The results of a demonstration of this new system are presented. Advantages of using a GM-cooled maser and the effects of the reduced CCR temperature on maser performance are discussed.

Chemokine and Chemokine Receptor Profiles in Metastatic Salivary Adenoid Cystic Carcinoma.

PubMed

Mays, Ashley C; Feng, Xin; Browne, James D; Sullivan, Christopher A

2016-08-01

To characterize the chemokine pattern in metastatic salivary adenoid cystic carcinoma (SACC). Real-time polymerase chain reaction (RT-PCR) was used to compare chemokine and chemokine receptor gene expression in two SACC cell lines: SACC-83 and SACC-LM (lung metastasis). Chemokines and receptor genes were then screened and their expression pattern characterized in human tissue samples of non-recurrent SACC and recurrent SACC with perineural invasion. Expression of chemokine receptors C5AR1, CCR1, CCR3, CCR6, CCR7, CCR9, CCR10, CXCR4, CXCR6, CXCR7, CCRL1 and CCRL2 were higher in SACC-83 compared to SACC-LM. CCRL1, CCBP2, CMKLR1, XCR1 and CXCR2 and 6 chemokine genes (CCL13, CCL27, CXCL14, CMTM1, CMTM2, CKLF) were more highly expressed in tissues of patients without tumor recurrence/perineural invasion compared to those with tumor recurrence. CCRL1 (receptor), CCL27, CMTM1, CMTM2, and CKLF (chemokine) genes were more highly expressed in SACC-83 and human tissues of patients without tumor recurrence/perineural invasion. CCRL1, CCL27, CMTM1, CMTM2 and CKLF may play important roles in the development of tumor metastases in SACC. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Novel CCR3 Antagonists Are Effective Mono- and Combination Inhibitors of Choroidal Neovascular Growth and Vascular Permeability.

PubMed

Nagai, Nori; Ju, Meihua; Izumi-Nagai, Kanako; Robbie, Scott J; Bainbridge, James W; Gale, David C; Pierre, Esaie; Krauss, Achim H P; Adamson, Peter; Shima, David T; Ng, Yin-Shan

2015-09-01

Choroidal neovascularization (CNV) is a defining feature of wet age-related macular degeneration. We examined the functional role of CCR3 in the development of CNV in mice and primates. CCR3 was associated with spontaneous CNV lesions in the newly described JR5558 mice, whereas CCR3 ligands localized to CNV-associated macrophages and the retinal pigment epithelium/choroid complex. Intravitreal injection of neutralizing antibodies against vascular endothelial growth factor receptor 2, CCR3, CC chemokine ligand 11/eotaxin-1, and CC chemokine ligand 24/eotaxin-2 all reduced CNV area and lesion number in these mice. Systemic administration of the CCR3 antagonists GW766994X and GW782415X reduced spontaneous CNV in JR5558 mice and laser-induced CNV in mouse and primate models in a dose-dependent fashion. Combination treatment with antivascular endothelial growth factor receptor 2 antibody and GW766994X yielded additive reductions in CNV area and hyperpermeability in mice. Interestingly, topical GW766994X and intravitreal anti-CCR3 antibody yielded strong systemic effects, reducing CNV in the untreated, contralateral eye. Contrarily, ocular administration of GW782415X in primates failed to substantially elevate plasma drug levels or to reduce the development of grade IV CNV lesions. These findings suggest that CCR3 signaling may be an attractive therapeutic target for CNV, utilizing a pathway that is at least partly distinct from that of vascular endothelial growth factor receptor. The findings also demonstrate that systemic exposure to CCR3 antagonists may be crucial for CNV-targeted activity. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Clinicopathological significance of chemokine receptor (CCR1, CCR3, CCR4, CCR5, CCR7 and CXCR4) expression in head and neck squamous cell carcinomas.

PubMed

González-Arriagada, Wilfredo A; Lozano-Burgos, Carlos; Zúñiga-Moreta, Rodrigo; González-Díaz, Paulina; Coletta, Ricardo D

2018-05-24

Head and neck squamous cell carcinoma shows high prevalence of lymph node metastasis at diagnosis, and despite the advances in treatment, the overall 5-year survival is still under 50%. Chemokine receptors have a role in the development and progression of cancer, but their effect in head and neck carcinoma remains poorly characterised. This study aimed to assess the prognostic value of CCR1, CCR3, CCR4, CCR5, CCR7 and CXCR4 in head and neck squamous cell carcinomas. Immunohistochemical expression of chemokine receptors was evaluated in a retrospective cohort of 76 cases of head and neck squamous cell carcinoma. Clinicopathological associations were analysed using the chi-square test, survival curves were analysed according to the Kaplan-Meier method, and the Cox proportional hazard model was applied for multivariate survival analysis. The chemokine receptors were highly expressed in primary carcinomas, except for CCR1 and CCR3. Significant associations were detected, including the associations between CCR5 expression and lymph node metastasis (N stage, P = .03), advanced clinical stage (P = .003), poor differentiation of tumours (P = .05) and recurrence (P = .01). The high expression of CCR5 was also associated with shortened disease-free survival (HR: 2.85, 95% CI: 1.09-8.14, P = .05), but the association did not withstand the Cox multivariate survival analysis. At univariate analysis, high expression of CCR7 was associated with disease-free survival and low levels of CXCR4 were significantly associated with both disease-specific and disease-free survival. These findings show that chemokine receptors may have an important role in head and neck squamous cell carcinoma progression, regional lymph node metastasis and recurrence. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Gene editing of CCR5 in autologous CD4 T cells of persons infected with HIV.

PubMed

Tebas, Pablo; Stein, David; Tang, Winson W; Frank, Ian; Wang, Shelley Q; Lee, Gary; Spratt, S Kaye; Surosky, Richard T; Giedlin, Martin A; Nichol, Geoff; Holmes, Michael C; Gregory, Philip D; Ando, Dale G; Kalos, Michael; Collman, Ronald G; Binder-Scholl, Gwendolyn; Plesa, Gabriela; Hwang, Wei-Ting; Levine, Bruce L; June, Carl H

2014-03-06

CCR5 is the major coreceptor for human immunodeficiency virus (HIV). We investigated whether site-specific modification of the gene ("gene editing")--in this case, the infusion of autologous CD4 T cells in which the CCR5 gene was rendered permanently dysfunctional by a zinc-finger nuclease (ZFN)--is safe. We enrolled 12 patients in an open-label, nonrandomized, uncontrolled study of a single dose of ZFN-modified autologous CD4 T cells. The patients had chronic aviremic HIV infection while they were receiving highly active antiretroviral therapy. Six of them underwent an interruption in antiretroviral treatment 4 weeks after the infusion of 10 billion autologous CD4 T cells, 11 to 28% of which were genetically modified with the ZFN. The primary outcome was safety as assessed by treatment-related adverse events. Secondary outcomes included measures of immune reconstitution and HIV resistance. One serious adverse event was associated with infusion of the ZFN-modified autologous CD4 T cells and was attributed to a transfusion reaction. The median CD4 T-cell count was 1517 per cubic millimeter at week 1, a significant increase from the preinfusion count of 448 per cubic millimeter (P<0.001). The median concentration of CCR5-modified CD4 T cells at 1 week was 250 cells per cubic millimeter. This constituted 8.8% of circulating peripheral-blood mononuclear cells and 13.9% of circulating CD4 T cells. Modified cells had an estimated mean half-life of 48 weeks. During treatment interruption and the resultant viremia, the decline in circulating CCR5-modified cells (-1.81 cells per day) was significantly less than the decline in unmodified cells (-7.25 cells per day) (P=0.02). HIV RNA became undetectable in one of four patients who could be evaluated. The blood level of HIV DNA decreased in most patients. CCR5-modified autologous CD4 T-cell infusions are safe within the limits of this study. (Funded by the National Institute of Allergy and Infectious Diseases and others; ClinicalTrials.gov number, NCT00842634.).

Matrix metalloproteinase-9 is up-regulated by CCL21/CCR7 interaction via extracellular signal-regulated kinase-1/2 signaling and is involved in CCL21-driven B-cell chronic lymphocytic leukemia cell invasion and migration.

PubMed

Redondo-Muñoz, Javier; José Terol, María; García-Marco, José A; García-Pardo, Angeles

2008-01-01

B-cell chronic lymphocytic leukemia (B-CLL) progression is frequently accompanied by clinical lymphadenopathy, and the CCL21 chemokine may play an important role in this process. Indeed, CCR7 (the CCL21 receptor), as well as matrix metalloproteinase-9 (MMP-9), are overexpressed in infiltrating B-CLL cells. We have studied whether MMP-9 is regulated by CCL21 and participates in CCL21-dependent migration. CCL21 significantly increased B-CLL MMP-9 production, measured by gelatin zymography. This was inhibited by blocking extracellular signal-regulated kinase-1/2 (ERK1/2) activity or by cell transfection with CCR7-siRNA. Accordingly, CCL21/CCR7 interaction activated the ERK1/2/c-Fos pathway and increased MMP-9 mRNA. CCL21-driven B-CLL cell migration through Matrigel or human umbilical vein endothelial cells (HUVEC) was blocked by anti-CCR7 antibodies, CCR7-siRNA transfection, or the ERK1/2 inhibitor U0126, as well as by anti-MMP-9 antibodies or tissue inhibitor of metalloproteinase 1 (TIMP-1). These results strongly suggest that MMP-9 is involved in B-CLL nodal infiltration and expand the roles of MMP-9 and CCR7 in B-CLL progression. Both molecules could thus constitute therapeutic targets for this disease.

Isolation and evolutionary analysis of Australasian topotype of bluetongue virus serotype 4 from India.

PubMed

Reddy, Y V; Susmitha, B; Patil, S; Krishnajyothi, Y; Putty, K; Ramakrishna, K V; Sunitha, G; Devi, B V; Kavitha, K; Deepthi, B; Krovvidi, S; Reddy, Y N; Reddy, G H; Singh, K P; Maan, N S; Hemadri, D; Maan, S; Mertens, P P; Hegde, N R; Rao, P P

2018-04-01

Bluetongue (BT) is a Culicoides-borne disease caused by several serotypes of bluetongue virus (BTV). Similar to other insect-borne viral diseases, distribution of BT is limited to distribution of Culicoides species competent to transmit BTV. In the tropics, vector activity is almost year long, and hence, the disease is endemic, with the circulation of several serotypes of BTV, whereas in temperate areas, seasonal incursions of a limited number of serotypes of BTV from neighbouring tropical areas are observed. Although BTV is endemic in all the three major tropical regions (parts of Africa, America and Asia) of the world, the distribution of serotypes is not alike. Apart from serological diversity, geography-based diversity of BTV genome has been observed, and this is the basis for proposal of topotypes. However, evolution of these topotypes is not well understood. In this study, we report the isolation and characterization of several BTV-4 isolates from India. These isolates are distinct from BTV-4 isolates from other geographical regions. Analysis of available BTV seg-2 sequences indicated that the Australasian BTV-4 diverged from African viruses around 3,500 years ago, whereas the American viruses diverged relatively recently (1,684 CE). Unlike Australasia and America, BTV-4 strains of the Mediterranean area evolved through several independent incursions. We speculate that independent evolution of BTV in different geographical areas over long periods of time might have led to the diversity observed in the current virus population. © 2017 Blackwell Verlag GmbH.

Effects of neferine on CCL5 and CCR5 expression in SCG of type 2 diabetic rats.

PubMed

Li, Guilin; Xu, Hong; Zhu, Shuanghua; Xu, Wenyuan; Qin, Shulan; Liu, Shuangmei; Tu, Guihua; Peng, Haiying; Qiu, Shuyi; Yu, Shicheng; Zhu, Qicheng; Fan, Bo; Zheng, Chaoran; Li, Guodong; Liang, Shangdong

2013-01-01

Chemokines and their receptors have the key role in inflammatory responses. The phenomenon of low grade inflammation is associated with the development of type 2 diabetes. Postprandial hyperglycemia increases the systemic inflammatory responses, which promotes the development of type 2 diabetic associating autonomic nervous injuries or cardiovascular disease. Neferine is a bisbenzylisoquinline alkaloid isolated from a Chinese medicinal herb. The objectives of this study will examine the CCL5 and CCR5 expression in the superior cervical ganglion (SCG) of type 2 diabetic rats. The effects of neferine on the expression of CCL5 and CCR5 mRNA and protein in the superior cervical ganglion (SCG) of type 2 diabetic rats will also be observed. The studies showed that in type 2 diabetic rats, body weight, blood pressure, heart rates, fasting blood glucose, insulin, total cholesterol and triglyceride were enhanced and high density lipoprotein was decreased, and CCL5 and CCR5 expression levels in the SCG of type 2 diabetic rats were up-regulated. In type 2 diabetic rats treated with neferine, body weight, blood pressure, fasting blood glucose, insulin, total cholesterol and triglyceride were decreased and high density lipoprotein was increased. The elevated expressions of CCL5 and CCR5 in SCG were decreased after type 2 diabetic rats treated with neferine. The motor nerve conduction velocity (MNCV) in diabetic rats treated with neferine group showed a significantly increment in comparison with that in type 2 diabetic group. Neferine can decrease the expression of CCL5 and CCR5 in the SCG and reduce the SCG neuronal signaling mediated by CCL5 and CCR5 in regulating diabetic cardiovascular autonomic complications. Copyright © 2012 Elsevier Inc. All rights reserved.

Human intestinal pro-inflammatory CD11chighCCR2+CX3CR1+ macrophages, but not their tolerogenic CD11c-CCR2-CX3CR1- counterparts, are expanded in inflammatory bowel disease.

PubMed

Bernardo, D; Marin, A C; Fernández-Tomé, S; Montalban-Arques, A; Carrasco, A; Tristán, E; Ortega-Moreno, L; Mora-Gutiérrez, I; Díaz-Guerra, A; Caminero-Fernández, R; Miranda, P; Casals, F; Caldas, M; Jiménez, M; Casabona, S; De la Morena, F; Esteve, M; Santander, C; Chaparro, M; Gisbert, J P

2018-05-09

Although macrophages (Mϕ) maintain intestinal immune homoeostasis, there is not much available information about their subset composition, phenotype and function in the human setting. Human intestinal Mϕ (CD45 + HLA-DR + CD14 + CD64 + ) can be divided into subsets based on the expression of CD11c, CCR2 and CX3CR1. Monocyte-like cells can be identified as CD11c high CCR2 + CX3CR1 + cells, a phenotype also shared by circulating CD14 + monocytes. On the contrary, their Mϕ-like tissue-resident counterparts display a CD11c - CCR2 - CX3CR1 - phenotype. CD11c high monocyte-like cells produced IL-1β, both in resting conditions and after LPS stimulation, while CD11c - Mϕ-like cells produced IL-10. CD11c high pro-inflammatory monocyte-like cells, but not the others, were increased in the inflamed colon from patients with inflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis. Tolerogenic IL-10-producing CD11c - Mϕ-like cells were generated from monocytes following mucosal conditioning. Finally, the colonic mucosa recruited circulating CD14 + monocytes in a CCR2-dependent manner, being such capacity expanded in IBD. Mϕ subsets represent, therefore, transition stages from newly arrived pro-inflammatory monocyte-like cells (CD11c high CCR2 + CX3CR1 + ) into tolerogenic tissue-resident (CD11c - CCR2 - CX3CR1 - ) Mϕ-like cells as reflected by the mucosal capacity to recruit circulating monocytes and induce CD11c - Mϕ. The process is nevertheless dysregulated in IBD, where there is an increased migration and accumulation of pro-inflammatory CD11c high monocyte-like cells.

Association of chemokine receptor gene (CCR2-CCR5) haplotypes with acquisition and control of HIV-1 infection in Zambians

PubMed Central

2011-01-01

Background Polymorphisms in chemokine (C-C motif) receptors 2 and 5 genes (CCR2 and CCR5) have been associated with HIV-1 infection and disease progression. We investigated the impact of CCR2-CCR5 haplotypes on HIV-1 viral load (VL) and heterosexual transmission in an African cohort. Between 1995 and 2006, cohabiting Zambian couples discordant for HIV-1 (index seropositive and HIV-1 exposed seronegative {HESN}) were monitored prospectively to determine the role of host genetic factors in HIV-1 control and heterosexual transmission. Genotyping for eight CCR2 and CCR5 variants resolved nine previously recognized haplotypes. By regression and survival analytic techniques, controlling for non-genetic factors, we estimated the effects of these haplotypic variants on a) index partner VL, b) seroconverter VL, c) HIV-1 transmission by index partners, d) HIV-1 acquisition by HESN partners. Results Among 567 couples, 240 virologically linked transmission events had occurred through 2006. HHF*2 homozygosity was associated with significantly lower VL in seroconverters (mean beta = -0.58, log10 P = 0.027) and the HHD/HHE diplotype was associated with significantly higher VL in the seroconverters (mean beta = 0.54, log10 P = 0.014) adjusted for age and gender in multivariable model. HHD/HHE was associated with more rapid acquisition of infection by the HESNs (HR = 2.0, 95% CI = 1.20-3.43, P = 0.008), after adjustments for index partner VL and the presence of genital ulcer or inflammation in either partner in Cox multivariable models. The HHD/HHE effect was stronger in exposed females (HR = 2.1, 95% CI = 1.14-3.95, P = 0.018). Conclusions Among Zambian discordant couples, HIV-1 coreceptor gene haplotypes and diplotypes appear to modulate HIV-1 VL in seroconverters and alter the rate of HIV-1 acquisition by HESNs. These associations replicate or resemble findings reported in other African and European populations. PMID:21429204

CCL3L1-CCR5 genotype improves the assessment of AIDS Risk in HIV-1-infected individuals.

PubMed

Kulkarni, Hemant; Agan, Brian K; Marconi, Vincent C; O'Connell, Robert J; Camargo, Jose F; He, Weijing; Delmar, Judith; Phelps, Kenneth R; Crawford, George; Clark, Robert A; Dolan, Matthew J; Ahuja, Sunil K

2008-09-08

Whether vexing clinical decision-making dilemmas can be partly addressed by recent advances in genomics is unclear. For example, when to initiate highly active antiretroviral therapy (HAART) during HIV-1 infection remains a clinical dilemma. This decision relies heavily on assessing AIDS risk based on the CD4+ T cell count and plasma viral load. However, the trajectories of these two laboratory markers are influenced, in part, by polymorphisms in CCR5, the major HIV coreceptor, and the gene copy number of CCL3L1, a potent CCR5 ligand and HIV-suppressive chemokine. Therefore, we determined whether accounting for both genetic and laboratory markers provided an improved means of assessing AIDS risk. In a prospective, single-site, ethnically-mixed cohort of 1,132 HIV-positive subjects, we determined the AIDS risk conveyed by the laboratory and genetic markers separately and in combination. Subjects were assigned to a low, moderate or high genetic risk group (GRG) based on variations in CCL3L1 and CCR5. The predictive value of the CCL3L1-CCR5 GRGs, as estimated by likelihood ratios, was equivalent to that of the laboratory markers. GRG status also predicted AIDS development when the laboratory markers conveyed a contrary risk. Additionally, in two separate and large groups of HIV+ subjects from a natural history cohort, the results from additive risk-scoring systems and classification and regression tree (CART) analysis revealed that the laboratory and CCL3L1-CCR5 genetic markers together provided more prognostic information than either marker alone. Furthermore, GRGs independently predicted the time interval from seroconversion to CD4+ cell count thresholds used to guide HAART initiation. The combination of the laboratory and genetic markers captures a broader spectrum of AIDS risk than either marker alone. By tracking a unique aspect of AIDS risk distinct from that captured by the laboratory parameters, CCL3L1-CCR5 genotypes may have utility in HIV clinical management. These findings illustrate how genomic information might be applied to achieve practical benefits of personalized medicine.

Genetic Polymorphism at CCL5 Is Associated With Protection in Chagas’ Heart Disease: Antagonistic Participation of CCR1+ and CCR5+ Cells in Chronic Chagasic Cardiomyopathy

PubMed Central

Batista, Angelica Martins; Alvarado-Arnez, Lucia Elena; Alves, Silvia Marinho; Melo, Gloria; Pereira, Isabela Resende; Ruivo, Leonardo Alexandre de Souza; da Silva, Andrea Alice; Gibaldi, Daniel; da Silva, Thayse do E. S. Protásio; de Lorena, Virginia Maria Barros; de Melo, Adriene Siqueira; de Araújo Soares, Ana Karine; Barros, Michelle da Silva; Costa, Vláudia Maria Assis; Cardoso, Cynthia C.; Pacheco, Antonio G.; Carrazzone, Cristina; Oliveira, Wilson; Moraes, Milton Ozório; Lannes-Vieira, Joseli

2018-01-01

Chronic cardiomyopathy is the main clinical manifestation of Chagas disease (CD), a disease caused by Trypanosoma cruzi infection. A hallmark of chronic chagasic cardiomyopathy (CCC) is a fibrogenic inflammation mainly composed of CD8+ and CD4+ T cells and macrophages. CC-chemokine ligands and receptors have been proposed to drive cell migration toward the heart tissue of CD patients. Single nucleotide polymorphisms (SNPs) in CC-chemokine ligand and receptor genes may determine protein expression. Herein, we evaluated the association of SNPs in the CC-chemokines CCL2 (rs1024611) and CCL5 (rs2107538, rs2280788) and the CCL5/RANTES receptors CCR1 (rs3181077, rs1491961, rs3136672) and CCR5 (rs1799987) with risk and progression toward CCC. We performed a cross-sectional association study of 406 seropositive patients from endemic areas for CD in the State of Pernambuco, Northeast Brazil. The patients were classified as non-cardiopathic (A, n = 110) or cardiopathic (mild, B1, n = 163; severe, C, n = 133). Serum levels of CCL5 and CCL2/MCP-1 were elevated in CD patients but were neither associated with risk/severity of CCC nor with SNP genotypes. After logistic regression analysis with adjustment for the covariates gender and ethnicity, CCL5 −403 (rs2107538) CT heterozygotes (OR = 0.5, P-value = 0.04) and T carriers (OR = 0.5, P-value = 0.01) were associated with protection against CCC. To gain insight into the participation of the CCL5–CCR5/CCR1 axis in CCC, mice were infected with the Colombian T. cruzi strain. Increased CCL5 concentrations were detected in cardiac tissue. In spleen, frequencies of CCR1+ CD8+ T cells and CD14+ macrophages were decreased, while frequencies of CCR5+ cells were increased. Importantly, CCR1+CD14+ macrophages were mainly IL-10+, while CCR5+ cells were mostly TNF+. CCR5-deficient infected mice presented reduced TNF concentrations and injury in heart tissue. Selective blockade of CCR1 (Met-RANTES therapy) in infected Ccr5−/− mice supported a protective role for CCR1 in CCC. Furthermore, parasite antigen stimulation of CD patient blood cells increased the frequency of CCR1+CD8+ T cells and CCL5 production. Collectively, our data support that a genetic variant of CCL5 and CCR1+ cells confer protection against Chagas heart disease, identifying the CCL5-CCR1 axis as a target for immunostimulation. PMID:29696014

Genetic Polymorphism at CCL5 Is Associated With Protection in Chagas' Heart Disease: Antagonistic Participation of CCR1+ and CCR5+ Cells in Chronic Chagasic Cardiomyopathy.

PubMed

Batista, Angelica Martins; Alvarado-Arnez, Lucia Elena; Alves, Silvia Marinho; Melo, Gloria; Pereira, Isabela Resende; Ruivo, Leonardo Alexandre de Souza; da Silva, Andrea Alice; Gibaldi, Daniel; da Silva, Thayse do E S Protásio; de Lorena, Virginia Maria Barros; de Melo, Adriene Siqueira; de Araújo Soares, Ana Karine; Barros, Michelle da Silva; Costa, Vláudia Maria Assis; Cardoso, Cynthia C; Pacheco, Antonio G; Carrazzone, Cristina; Oliveira, Wilson; Moraes, Milton Ozório; Lannes-Vieira, Joseli

2018-01-01

Chronic cardiomyopathy is the main clinical manifestation of Chagas disease (CD), a disease caused by Trypanosoma cruzi infection. A hallmark of chronic chagasic cardiomyopathy (CCC) is a fibrogenic inflammation mainly composed of CD8 + and CD4 + T cells and macrophages. CC-chemokine ligands and receptors have been proposed to drive cell migration toward the heart tissue of CD patients. Single nucleotide polymorphisms (SNPs) in CC-chemokine ligand and receptor genes may determine protein expression. Herein, we evaluated the association of SNPs in the CC-chemokines CCL2 (rs1024611) and CCL5 (rs2107538, rs2280788) and the CCL5/RANTES receptors CCR1 (rs3181077, rs1491961, rs3136672) and CCR5 (rs1799987) with risk and progression toward CCC. We performed a cross-sectional association study of 406 seropositive patients from endemic areas for CD in the State of Pernambuco, Northeast Brazil. The patients were classified as non-cardiopathic (A, n  = 110) or cardiopathic (mild, B1, n  = 163; severe, C, n  = 133). Serum levels of CCL5 and CCL2/MCP-1 were elevated in CD patients but were neither associated with risk/severity of CCC nor with SNP genotypes. After logistic regression analysis with adjustment for the covariates gender and ethnicity, CCL5 -403 (rs2107538) CT heterozygotes (OR = 0.5, P -value = 0.04) and T carriers (OR = 0.5, P -value = 0.01) were associated with protection against CCC. To gain insight into the participation of the CCL5-CCR5/CCR1 axis in CCC, mice were infected with the Colombian T. cruzi strain. Increased CCL5 concentrations were detected in cardiac tissue. In spleen, frequencies of CCR1 + CD8 + T cells and CD14 + macrophages were decreased, while frequencies of CCR5 + cells were increased. Importantly, CCR1 + CD14 + macrophages were mainly IL-10 + , while CCR5 + cells were mostly TNF + . CCR5-deficient infected mice presented reduced TNF concentrations and injury in heart tissue. Selective blockade of CCR1 (Met-RANTES therapy) in infected Ccr5 -/- mice supported a protective role for CCR1 in CCC. Furthermore, parasite antigen stimulation of CD patient blood cells increased the frequency of CCR1 + CD8 + T cells and CCL5 production. Collectively, our data support that a genetic variant of CCL5 and CCR1 + cells confer protection against Chagas heart disease, identifying the CCL5-CCR1 axis as a target for immunostimulation.

The virus–receptor interaction in the replication of feline immunodeficiency virus (FIV)☆

PubMed Central

Willett, Brian J; Hosie, Margaret J

2013-01-01

The feline and human immunodeficiency viruses (FIV and HIV) target helper T cells selectively, and in doing so they induce a profound immune dysfunction. The primary determinant of HIV cell tropism is the expression pattern of the primary viral receptor CD4 and co-receptor(s), such as CXCR4 and CCR5. FIV employs a distinct strategy to target helper T cells; a high affinity interaction with CD134 (OX40) is followed by binding of the virus to its sole co-receptor, CXCR4. Recent studies have demonstrated that the way in which FIV interacts with its primary receptor, CD134, alters as infection progresses, changing the cell tropism of the virus. This review examines the contribution of the virus–receptor interaction to replication in vivo as well as the significance of these findings to the development of vaccines and therapeutics. PMID:23992667

Investigation on the phytosanitary status of major ornamental hibiscus species in Italy to assess virus infection

USDA-ARS?s Scientific Manuscript database

The genus Hibiscus (family Malvaceae) includes more than 250 species that vary from annual to perennial herbs, and shrubs to small trees that are native to tropical, sub-tropical and temperate climates. A study in 2010-2011 examined viruses associated with symptoms observed on hibiscus plants in It...

Prediction of HIV-1 coreceptor usage (tropism) by sequence analysis using a genotypic approach.

PubMed

Sierra, Saleta; Kaiser, Rolf; Lübke, Nadine; Thielen, Alexander; Schuelter, Eugen; Heger, Eva; Däumer, Martin; Reuter, Stefan; Esser, Stefan; Fätkenheuer, Gerd; Pfister, Herbert; Oette, Mark; Lengauer, Thomas

2011-12-01

Maraviroc (MVC) is the first licensed antiretroviral drug from the class of coreceptor antagonists. It binds to the host coreceptor CCR5, which is used by the majority of HIV strains in order to infect the human immune cells (Fig. 1). Other HIV isolates use a different coreceptor, the CXCR4. Which receptor is used, is determined in the virus by the Env protein (Fig. 2). Depending on the coreceptor used, the viruses are classified as R5 or X4, respectively. MVC binds to the CCR5 receptor inhibiting the entry of R5 viruses into the target cell. During the course of disease, X4 viruses may emerge and outgrow the R5 viruses. Determination of coreceptor usage (also called tropism) is therefore mandatory prior to administration of MVC, as demanded by EMA and FDA. The studies for MVC efficiency MOTIVATE, MERIT and 1029 have been performed with the Trofile assay from Monogram, San Francisco, U.S.A. This is a high quality assay based on sophisticated recombinant tests. The acceptance for this test for daily routine is rather low outside of the U.S.A., since the European physicians rather tend to work with decentralized expert laboratories, which also provide concomitant resistance testing. These laboratories have undergone several quality assurance evaluations, the last one being presented in 2011. For several years now, we have performed tropism determinations based on sequence analysis from the HIV env-V3 gene region (V3). This region carries enough information to perform a reliable prediction. The genotypic determination of coreceptor usage presents advantages such as: shorter turnover time (equivalent to resistance testing), lower costs, possibility to adapt the results to the patients' needs and possibility of analysing clinical samples with very low or even undetectable viral load (VL), particularly since the number of samples analysed with VL < 1000 copies/μl roughly increased in the last years (Fig. 3). The main steps for tropism testing (Fig. 4) demonstrated in this video: Collection of a blood sample Isolation of the HIV RNA from the plasma and/or HIV proviral DNA from blood mononuclear cells Amplification of the env region Amplification of the V3 region Sequence reaction of the V3 amplicon Purification of the sequencing samples Sequencing the purified samples Sequence editing Sequencing data interpretation and tropism prediction.

CXCR3/CCR5 pathways in metastatic melanoma patients treated with adoptive therapy and interleukin-2

PubMed Central

Bedognetti, D; Spivey, T L; Zhao, Y; Uccellini, L; Tomei, S; Dudley, M E; Ascierto, M L; De Giorgi, V; Liu, Q; Delogu, L G; Sommariva, M; Sertoli, M R; Simon, R; Wang, E; Rosenberg, S A; Marincola, F M

2013-01-01

Background: Adoptive therapy with tumour-infiltrating lymphocytes (TILs) induces durable complete responses (CR) in ∼20% of patients with metastatic melanoma. The recruitment of T cells through CXCR3/CCR5 chemokine ligands is critical for immune-mediated rejection. We postulated that polymorphisms and/or expression of CXCR3/CCR5 in TILs and the expression of their ligands in tumour influence the migration of TILs to tumours and tumour regression. Methods: Tumour-infiltrating lymphocytes from 142 metastatic melanoma patients enrolled in adoptive therapy trials were genotyped for CXCR3 rs2280964 and CCR5-Δ32 deletion, which encodes a protein not expressed on the cell surface. Expression of CXCR3/CCR5 in TILs and CXCR3/CCR5 and ligand genes in 113 available parental tumours was also assessed. Tumour-infiltrating lymphocyte data were validated by flow cytometry (N=50). Results: The full gene expression/polymorphism model, which includes CXCR3 and CCR5 expression data, CCR5-Δ32 polymorphism data and their interaction, was significantly associated with both CR and overall response (OR; P=0.0009, and P=0.007, respectively). More in detail, the predicted underexpression of both CXCR3 and CCR5 according to gene expression and polymorphism data (protein prediction model, PPM) was associated with response to therapy (odds ratio=6.16 and 2.32, for CR and OR, respectively). Flow cytometric analysis confirmed the PPM. Coordinate upregulation of CXCL9, CXCL10, CXCL11, and CCL5 in pretreatment tumour biopsies was associated with OR. Conclusion: Coordinate overexpression of CXCL9, CXCL10, CXCL11, and CCL5 in pretreatment tumours was associated with responsiveness to treatment. Conversely, CCR5-Δ32 polymorphism and CXCR3/CCR5 underexpression influence downregulation of the corresponding receptors in TILs and were associated with likelihood and degree of response. PMID:24129241

Creatinine reabsorption by the aged kidney.

PubMed

Musso, Carlos G; Michelángelo, Hernán; Vilas, Manuel; Reynaldi, Juliana; Martinez, Bernardo; Algranati, Luis; Macías Núñez, Juan F

2009-01-01

The handling of renal creatinine in human beings has classically been described as the result of two particular physiological processes: glomerular filtration and proximal tubular secretion. However, there are particular physiological situations in which tubular creatinine reabsorption has been documented, such as in the case of healthy newborns and premature babies. We performed a prospective study in order to evaluate if there is tubular creatinine reabsorption in healthy elderly people. We studied prospectively nine healthy volunteers, four of them young (20-33 years old) and the remaining five, old (65-73 years old). Since creatinine is secreted in the proximal tubules, and its secretion can be completely blocked by cimetidine administration, a creatinine clearance with cimetidine reliably represents the glomerular filtration rate. Therefore, if the ratio creatinine clearance (Ccr)/creatinine clearance with cimetidine (CcrWC) is higher than one, this would indicate net creatinine secretion, whereas a ratio lower than one would indicate a net renal creatinine tubular reabsorption; a ratio equal to one indicates creatinine filtration. Finally, the Ccr, CcrWC, and Ccr/CcrWC ratios were compared between the young and old group. Mann-Whitney and Wilcoxon tests were used. As expected, creatinine clearance in the elderly was significantly lower than in the young [Ccr: 74.4 ml/min (47.9-100.9) (old) vs. 153.8 ml/min (108.3-199.2) (young), p = 0.014]. Similarly, the creatinine clearance with cimetidine (CcrWC) was significantly lower in the elderly compared to the young [CcrWC: 81.8 ml/min (69.2-94.5) (old) vs. 122.5 ml/min (82.6-162.4) (young), p = 0.028]. The ratio of Ccr/CcrWC was 0.9 in the elderly vs. 1.26 in the young (p = 0.014), indicating net creatinine reabsorption in the elderly and net creatinine secretion in the young. Our findings indicate that there seems to be a net reabsorption of creatinine in the renal tubules of healthy old persons.

Haplotypes in CCR5-CCR2, CCL3 and CCL5 are associated with natural resistance to HIV-1 infection in a Colombian cohort.

PubMed

Vega, Jorge A; Villegas-Ospina, Simón; Aguilar-Jiménez, Wbeimar; Rugeles, María T; Bedoya, Gabriel; Zapata, Wildeman

2017-06-01

Variants in genes encoding for HIV-1 co-receptors and their natural ligands have been individually associated to natural resistance to HIV-1 infection. However, the simultaneous presence of these variants has been poorly studied. To evaluate the association of single and multilocus haplotypes in genes coding for the viral co-receptors CCR5 and CCR2, and their ligands CCL3 and CCL5, with resistance or susceptibility to HIV-1 infection. Nine variants in CCR5-CCR2, two SNPs in CCL3 and two in CCL5 were genotyped by PCR-RFLP in 35 seropositive (cases) and 49 HIV-1-exposed seronegative Colombian individuals (controls). Haplotypes were inferred using the Arlequin software, and their frequency in individual or combined loci was compared between cases and controls by the chi-square test. A p' value ;0.05 after Bonferroni correction was considered significant. Homozygosis of the human haplogroup (HH) E was absent in controls and frequent in cases, showing a tendency to susceptibility. The haplotypes C-C and T-T in CCL3 were associated with susceptibility (p'=0.016) and resistance (p';0.0001) to HIV-1 infection, respectively. Finally, in multilocus analysis, the haplotype combinations formed by HHC in CCR5-CCR2, T-T in CCL3 and G-C in CCL5 were associated with resistance (p'=0.006). Our results suggest that specific combinations of variants in genes from the same signaling pathway can define an HIV-1 resistant phenotype. Despite our small sample size, our statistically significant associations suggest strong effects; however, these results should be further validated in larger cohorts.

Isolation, molecular characterization, and phylogenetic analysis of encephalomyocarditis virus from South China tigers in China.

PubMed

Liu, Huimin; Yan, Qi; Zhao, Bo; Luo, Jing; Wang, Chengmin; Du, Yingchun; Yan, Jing; He, Hongxuan

2013-10-01

Although encephalomyocarditis virus (EMCV) can infect many host species and cause myocarditis and sudden death in many species, little is known about EMCV infection in tigers. A virus was isolated from organs of dead South China tigers with sudden death in southern China. The production of cytopathic effect on BHK cells, and the results of PCR, electron microscopy (EM), and whole genome sequencing indicated that the pathogen was EMCV, the strain was named FJ13. Other pathogenic agents were excluded as possible pathogenic agents. Phylogenetic analyses of the whole genome, ORF (open reading frame) and CCR (capsid coding region) using the neighbour-joining method revealed that EMCV isolates cluster into two groups (group 1 and 2) with two sub-clusters within group 1 (group 1a and 1b), and FJ13 belongs to group 1a. Animal experiment showed that the isolated strain FJ13 could cause clinical symptoms and pathological changes. The results of this study indicated that FJ13 caused myocarditis of tigers and provided new epidemiologic data on EMCV in China. Copyright © 2013 Elsevier B.V. All rights reserved.

Determination of HIV-1 co-receptor usage.

PubMed

Cavarelli, Mariangela; Scarlatti, Gabriella

2014-01-01

Human immunodeficiency virus type I (HIV-1) infects target cells through interaction with the CD4 molecule and chemokine receptors, mainly the β-chemokine receptor 5 (CCR5) and the α-chemokine receptor 4 (CXCR4). Viral isolates can be phenotypically classified based on the co-receptor they utilize to infect target cells. In this chapter, methods to determine the co-receptor usage of HIV-1 variants are described.

Herpes Simplex Virus Type 1 Infects Enteric Neurons and Triggers Gut Dysfunction via Macrophage Recruitment.

PubMed

Brun, Paola; Qesari, Marsela; Marconi, Peggy C; Kotsafti, Andromachi; Porzionato, Andrea; Macchi, Veronica; Schwendener, Reto A; Scarpa, Marco; Giron, Maria C; Palù, Giorgio; Calistri, Arianna; Castagliuolo, Ignazio

2018-01-01

Herpes Simplex Virus type 1 (HSV-1), a neurotropic pathogen widespread in human population, infects the enteric nervous system (ENS) in humans and rodents and causes intestinal neuromuscular dysfunction in rats. Although infiltration of inflammatory cells in the myenteric plexus and neurodegeneration of enteric nerves are common features of patients suffering from functional intestinal disorders, the proof of a pathogenic link with HSV-1 is still unsettled mainly because the underlying mechanisms are largely unknown. In this study we demonstrated that following intragastrical administration HSV-1 infects neurons within the myenteric plexus resulting in functional and structural alterations of the ENS. By infecting mice with HSV-1 replication-defective strain we revealed that gastrointestinal neuromuscular anomalies were however independent of viral replication. Indeed, enteric neurons exposed to UV-inactivated HSV-1 produced monocyte chemoattractant protein-1 (MCP-1/CCL2) to recruit activated macrophages in the longitudinal muscle myenteric plexus. Infiltrating macrophages produced reactive oxygen and nitrogen species and directly harmed enteric neurons resulting in gastrointestinal dysmotility. In HSV-1 infected mice intestinal neuromuscular dysfunctions were ameliorated by in vivo administration of (i) liposomes containing dichloromethylene bisphosphonic acid (clodronate) to deplete tissue macrophages, (ii) CCR2 chemokine receptor antagonist RS504393 to block the CCL2/CCR2 pathway, (iii) Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) and AR-C 102222 to quench production of nitrogen reactive species produced via iNOS. Overall these data demonstrate that HSV-1 infection makes enteric neurons recruit macrophages via production of a specific chemoattractant factor. The resulting inflammatory reaction is mandatory for intestinal dysmotility. These findings provide insights into the neuro-immune communication that occurs in the ENS following HSV-1 infection and allow recognition of an original pathophysiologic mechanism underlying gastrointestinal diseases as well as identification of novel therapeutic targets.

CCL19/CCR7 contributes to the pathogenesis of endometriosis via PI3K/Akt pathway by regulating the proliferation and invasion of ESCs.

PubMed

Diao, Ruiying; Wei, Weixia; Zhao, Jinghui; Tian, Fuying; Cai, Xueyong; Duan, Yong-Gang

2017-11-01

The level of CCL19 increased in the peritoneal fluid of women with endometriosis, but the precise mechanism of CCL19/CCR7 in the pathogenesis of endometriosis remains unknown. ELISA and immunohistochemistry were performed to analyze CCL19/CCR7 expressions in peritoneal fluid and endometrium from women with endometriosis (n = 38) and controls (n = 32). Cell proliferation and transwell invasion assays were applied to detect proliferation and invasion of human endometrial stromal cells (ESCs). Expressions of Bcl2, MMP2, MMP9, and p-AKT/AKT were analyzed by Western blot. Peritoneal fluid concentration of CCL19 in patients with endometriosis was higher than that in controls. Those patients with moderate/severe endometriosis had significantly higher peritoneal fluid concentrations of CCL19 compared to those with minimal/mild endometriosis. Higher CCL19 and CCR7 were found in the endometrium with endometriosis compared to control. CCL19 significantly enhanced ESC proliferation and invasion through CCR7 via activating PI3K/Akt signal pathways. CCL19/CCR7 interaction significantly enhanced phosphorylation of Akt, Bcl2, MMP2, and MMP9 in ESCs. These data indicate CCL19/CCR7 contributes to proliferation and invasion of ESCs, which are conducive to the pathogenesis of endometriosis through activating PI3K/Akt pathway. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

48 CFR 52.204-7 - Central Contractor Registration.

Code of Federal Regulations, 2011 CFR

2011-10-01

... (CCR) database means the primary Government repository for Contractor information required for the...) for the same concern. Registered in the CCR database means that— (1) The Contractor has entered all mandatory information, including the DUNS number or the DUNS+4 number, into the CCR database; and (2) The...

CCR6 Defines Memory B Cell Precursors in Mouse and Human Germinal Centers, Revealing Light-Zone Location and Predominant Low Antigen Affinity.

PubMed

Suan, Dan; Kräutler, Nike J; Maag, Jesper L V; Butt, Danyal; Bourne, Katherine; Hermes, Jana R; Avery, Danielle T; Young, Clara; Statham, Aaron; Elliott, Michael; Dinger, Marcel E; Basten, Antony; Tangye, Stuart G; Brink, Robert

2017-12-19

Memory B cells (MBCs) and plasma cells (PCs) constitute the two cellular outputs of germinal center (GC) responses that together facilitate long-term humoral immunity. Although expression of the transcription factor BLIMP-1 identifies cells undergoing PC differentiation, no such marker exists for cells committed to the MBC lineage. Here, we report that the chemokine receptor CCR6 uniquely marks MBC precursors in both mouse and human GCs. CCR6 + GC B cells were highly enriched within the GC light zone (LZ), were the most quiescent of all GC B cells, exhibited a cell-surface phenotype and gene expression signature indicative of an MBC transition, and possessed the augmented response characteristics of MBCs. MBC precursors within the GC LZ predominantly possessed a low affinity for antigen but also included cells from within the high-affinity pool. These data indicate a fundamental dichotomy between the processes that drive MBC and PC differentiation during GC responses. Copyright © 2017 Elsevier Inc. All rights reserved.

Interactions Between HIV-Infected Monocytes and the Extracellular Matrix: HIV-Infected Monocytes Secrete Neutral Metalloproteases That Degrade Basement Membrane Protein Matrices,

DTIC Science & Technology

1992-08-01

p2 4 antigen (Ag) by flow "cytometry. All cells were recovered from Matrigel (Col- 20• laborative Biomedical Products) after treatment with a bac...of HIV-infected mono- tropic virus originally isolated and passaged in monocytes, at a multiplicity of infection of 0.05 infectious virus/target cell

Genome-Wide Mutagenesis of Dengue Virus Reveals Plasticity of the NS1 Protein and Enables Generation of Infectious Tagged Reporter Viruses

PubMed Central

Johnson, Stephen M.; Eltahla, Auda A.; Aloi, Maria; Aloia, Amanda L.; McDevitt, Christopher A.; Bull, Rowena A.

2017-01-01

ABSTRACT Dengue virus (DENV) is a major global pathogen that causes significant morbidity and mortality in tropical and subtropical areas worldwide. An improved understanding of the regions within the DENV genome and its encoded proteins that are required for the virus replication cycle will expedite the development of urgently required therapeutics and vaccines. We subjected an infectious DENV genome to unbiased insertional mutagenesis and used next-generation sequencing to identify sites that tolerate 15-nucleotide insertions during the virus replication cycle in hepatic cell culture. This revealed that the regions within capsid, NS1, and the 3′ untranslated region were the most tolerant of insertions. In contrast, prM- and NS2A-encoding regions were largely intolerant of insertions. Notably, the multifunctional NS1 protein readily tolerated insertions in regions within the Wing, connector, and β-ladder domains with minimal effects on viral RNA replication and infectious virus production. Using this information, we generated infectious reporter viruses, including a variant encoding the APEX2 electron microscopy tag in NS1 that uniquely enabled high-resolution imaging of its localization to the surface and interior of viral replication vesicles. In addition, we generated a tagged virus bearing an mScarlet fluorescent protein insertion in NS1 that, despite an impact on fitness, enabled live cell imaging of NS1 localization and traffic in infected cells. Overall, this genome-wide profile of DENV genome flexibility may be further dissected and exploited in reporter virus generation and antiviral strategies. IMPORTANCE Regions of genetic flexibility in viral genomes can be exploited in the generation of reporter virus tools and should arguably be avoided in antiviral drug and vaccine design. Here, we subjected the DENV genome to high-throughput insertional mutagenesis to identify regions of genetic flexibility and enable tagged reporter virus generation. In particular, the viral NS1 protein displayed remarkable tolerance of small insertions. This genetic flexibility enabled generation of several novel NS1-tagged reporter viruses, including an APEX2-tagged virus that we used in high-resolution imaging of NS1 localization in infected cells by electron microscopy. For the first time, this analysis revealed the localization of NS1 within viral replication factories known as “vesicle packets” (VPs), in addition to its acknowledged localization to the luminal surface of these VPs. Together, this genetic profile of DENV may be further refined and exploited in the identification of antiviral targets and the generation of reporter virus tools. PMID:28956770

Gas6 Promotes Inflammatory (CCR2hiCX3CR1lo) Monocyte Recruitment in Venous Thrombosis.

PubMed

Laurance, Sandrine; Bertin, François-René; Ebrahimian, Talin; Kassim, Yusra; Rys, Ryan N; Lehoux, Stéphanie; Lemarié, Catherine A; Blostein, Mark D

2017-07-01

Coagulation and inflammation are inter-related. Gas6 (growth arrest-specific 6) promotes venous thrombosis and participates to inflammation through endothelial-innate immune cell interactions. Innate immune cells can provide the initiating stimulus for venous thrombus development. We hypothesize that Gas6 promotes monocyte recruitment during venous thrombosis. Deep venous thrombosis was induced in wild-type and Gas6-deficient (-/-) mice using 5% FeCl 3 and flow reduction in the inferior vena cava. Total monocyte depletion was achieved by injection of clodronate before deep venous thrombosis. Inflammatory monocytes were depleted using an anti-C-C chemokine receptor type 2 (CCR2) antibody. Similarly, injection of an anti-chemokine ligand 2 (CCL2) antibody induced CCL2 depletion. Flow cytometry and immunofluorescence were used to characterize the monocytes recruited to the thrombus. In vivo, absence of Gas6 was associated with a reduction of monocyte recruitment in both deep venous thrombosis models. Global monocyte depletion by clodronate leads to smaller thrombi in wild-type mice. Compared with wild type, the thrombi from Gas6 -/- mice contain less inflammatory (CCR2 hi CX 3 CR1 lo ) monocytes, consistent with a Gas6-dependent recruitment of this monocyte subset. Correspondingly, selective depletion of CCR2 hi CX 3 CR1 lo monocytes reduced the formation of venous thrombi in wild-type mice demonstrating a predominant role of the inflammatory monocytes in thrombosis. In vitro, the expression of both CCR2 and CCL2 were Gas6 dependent in monocytes and endothelial cells, respectively, impacting monocyte migration. Moreover, Gas6-dependent CCL2 expression and monocyte migration were mediated via JNK (c-Jun N-terminal kinase). This study demonstrates that Gas6 specifically promotes the recruitment of inflammatory CCR2 hi CX 3 CR1 lo monocytes through the regulation of both CCR2 and CCL2 during deep venous thrombosis. © 2017 American Heart Association, Inc.

AhR activation increases IL-2 production by alloreactive CD4+ T cells initiating the differentiation of mucosal-homing Tim3+ Lag3+ Tr1 cells.

PubMed

Ehrlich, Allison K; Pennington, Jamie M; Tilton, Susan; Wang, Xisheng; Marshall, Nikki B; Rohlman, Diana; Funatake, Castle; Punj, Sumit; O'Donnell, Edmond; Yu, Zhen; Kolluri, Siva K; Kerkvliet, Nancy I

2017-11-01

Activation of the aryl hydrocarbon receptor (AhR) by immunosuppressive ligands promotes the development of regulatory T (Treg) cells. Although AhR-induced Foxp3 + Treg cells have been well studied, much less is known about the development and fate of AhR-induced Type 1 Treg (AhR-Tr1) cells. In the current study, we identified the unique transcriptional and functional changes in murine CD4 + T cells that accompany the differentiation of AhR-Tr1 cells during the CD4 + T-cell-dependent phase of an allospecific cytotoxic T lymphocyte (allo-CTL) response. AhR activation increased the expression of genes involved in T-cell activation, immune regulation and chemotaxis, as well as a global downregulation of genes involved in cell cycling.  Increased IL-2 production was responsible for the early AhR-Tr1 activation phenotype previously characterized as CD25 + CTLA4 + GITR + on day 2. The AhR-Tr1 phenotype was further defined by the coexpression of the immunoregulatory receptors Lag3 and Tim3 and non-overlapping expression of CCR4 and CCR9. Consistent with the increased expression of CCR9, real-time imaging showed enhanced migration of AhR-Tr1 cells to the lamina propria of the small intestine and colon. The discovery of mucosal imprinting of AhR-Tr1 cells provides an additional mechanism by which therapeutic AhR ligands can control immunopathology. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Isolation of Madre de Dios Virus (Orthobunyavirus; Bunyaviridae), an Oropouche Virus Species Reassortant, from a Monkey in Venezuela.

PubMed

Navarro, Juan-Carlos; Giambalvo, Dileyvic; Hernandez, Rosa; Auguste, Albert J; Tesh, Robert B; Weaver, Scott C; Montañez, Humberto; Liria, Jonathan; Lima, Anderson; Travassos da Rosa, Jorge Fernando Soares; da Silva, Sandro P; Vasconcelos, Janaina M; Oliveira, Rodrigo; Vianez, João L S G; Nunes, Marcio R T

2016-08-03

Oropouche virus (OROV), genus Orthobunyavirus, family Bunyaviridae, is an important cause of human illness in tropical South America. Herein, we report the isolation, complete genome sequence, genetic characterization, and phylogenetic analysis of an OROV species reassortant, Madre de Dios virus (MDDV), obtained from a sick monkey (Cebus olivaceus Schomburgk) collected in a forest near Atapirire, a small rural village located in Anzoategui State, Venezuela. MDDV is one of a growing number of naturally occurring OROV species reassortants isolated in South America and was known previously only from southern Peru. © The American Society of Tropical Medicine and Hygiene.

Enteric viruses in a mangrove lagoon, survival and shellfish incidence

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lopez de Cardona, I.; Bermudez, M.; Billmire, E.

Mangrove oysters (Crassostrea rhizophorae) were screened for enteric viruses. For 18 months oysters were collected from Cano Boqueron, a tropical mangrove lagoon on the southwest coast of Puerto Rico. This popular tourist resort has two primary sewage treatment plants which service 158 single family cabanas. In spite of the heavy seasonal input of sewage to Cano Boqueron and high densities of fecal coliform bacteria, enteric viruses were not detected in shellfish meat. Because no viruses were detected in the oysters, a virus survival study was performed. Poliovirus type 1 was placed in diffusion chambers in situ at two sites inmore » Cano Boqueron. More than 95% of the poliovirus inactivation occurred within 24 h. Virus inactivation was significantly different by site, indicating different inactivation rates within the lagoon. Chamber studies done simultaneously with Escherichia coli did not reveal differences between sites. It is suggested that the sewage effluent had an antiviral effect in the absence of an antibacterial effect. This study demonstrates the importance for establishing microbial contamination standards for shellfish growing waters in the tropics based upon in situ studies with tropical species, e.g. mangrove oyster.« less

Colonic eosinophilic inflammation in experimental colitis is mediated by Ly6Chigh CCR2+ inflammatory monocyte/macrophage-derived CCL11

PubMed Central

Waddell, Amanda; Ahrens, Richard; Steinbrecher, Kris; Donovan, Burke; Rothenberg, Marc E.; Munitz, Ariel; Hogan, Simon P.

2011-01-01

Recent genome-wide association studies of pediatric IBD have implicated the 17q12 loci, which contains the eosinophil specific chemokine gene CCL11, with early-onset IBD susceptibility. In the present study, we employed a murine model of experimental colitis to define the molecular pathways that regulate CCL11 expression in the chronic intestinal inflammation and pathophysiology of experimental colitis. Bone marrow chimera experiments showed that hematopoietic cell-derived CCL11 is sufficient for CCL11-mediated colonic eosinophilic inflammation. We show that DSS treatment promotes the recruitment of F4/80+CD11b+CCR2+Ly6Chigh inflammatory monocytes into the colon. F4/80+CD11b+CCR2+Ly6Chigh monocytes express CCL11, and their recruitment positively correlated with colonic eosinophilic inflammation. Phenotypic analysis of purified Ly6Chigh intestinal inflammatory MΦs revealed that these cells express both M1- and M2-associated genes, including Il6, Ccl4 and Cxcl2, and Arg1, Chi3l3, Ccl11 and IL-10, respectively. Attenuation of DSS-induced F4/80+CD11b+CCR2+Ly6Chigh monocyte recruitment to the colon in CCR2−/− mice was associated with decreased colonic CCL11 expression, eosinophilic inflammation and DSS-induced histopathology. These studies identify a mechanism for DSS-induced colonic eosinophilia mediated by Ly6ChighCCR2+ inflammatory monocyte/MΦ-derived CCL11. PMID:21498668

Isoleucine/leucine2 is essential for chemoattractant activity of beta-defensin Defb14 through chemokine receptor 6.

PubMed

Tyrrell, Christine; De Cecco, Martin; Reynolds, Natalie L; Kilanowski, Fiona; Campopiano, Dominic; Barran, Perdita; Macmillan, Derek; Dorin, Julia R

2010-03-01

Beta-defensins are both antimicrobial and able to chemoattract various immune cells including immature dendritic cells and CD4 T cells through CCR6. They are short, cationic peptides with a highly conserved six-cysteine motif. It has been shown that only the fifth cysteine is critical for chemoattraction of cells expressing CCR6. In order to identify other residues essential for functional interaction with CCR6 we used a library of peptide deletion derivatives based on Defb14. Loss of the initial two amino acids from the Defb14-1C(V) derivative destroys its ability to chemoattract cells expressing CCR6. As the second amino acid is an evolutionarily conserved leucine, we make full-length Defb14-1C(V) peptides with substitution of the leucine(2) for glycine (L2G), lysine (L2K) or isoleucine (L2I). Defb14-1C(V) L2G and L2K and are unable to chemoattract CCR6 expressing cells but the semi-conservative change L2I has activity. By circular dichroism spectroscopy we can see no evidence for a significant change in secondary structure as a consequence of these substitutions and so cannot attribute loss of chemotactic activity with disruption of the N-terminal helix. We conclude that isoleucine/leucine in the N-terminal alpha-helix region of this beta-defensin is essential for CCR6-mediated chemotaxis. Copyright 2009 Elsevier Ltd. All rights reserved.

Clinical Applications of Genome Editing to HIV Cure.

PubMed

Wang, Cathy X; Cannon, Paula M

2016-12-01

Despite significant advances in HIV drug treatment regimens, which grant near-normal life expectancies to infected individuals who have good virological control, HIV infection itself remains incurable. In recent years, novel gene- and cell-based therapies have gained increasing attention due to their potential to provide a functional or even sterilizing cure for HIV infection with a one-shot treatment. A functional cure would keep the infection in check and prevent progression to AIDS, while a sterilizing cure would eradicate all HIV viruses from the patient. Genome editing is the most precise form of gene therapy, able to achieve permanent genetic disruption, modification, or insertion at a predesignated genetic locus. The most well-studied candidate for anti-HIV genome editing is CCR5, an essential coreceptor for the majority of HIV strains, and the lack of which confers HIV resistance in naturally occurring homozygous individuals. Genetic disruption of CCR5 to treat HIV has undergone clinical testing, with seven completed or ongoing trials in T cells and hematopoietic stem and progenitor cells, and has shown promising safety and potential efficacy profiles. Here we summarize clinical findings of CCR5 editing for HIV therapy, as well as other genome editing-based approaches under pre-clinical development. The anticipated development of more sophisticated genome editing technologies should continue to benefit HIV cure efforts.

The Structural Changes of Tropical Cyclones Upon Interaction with Vertical Wind Shear

NASA Technical Reports Server (NTRS)

Ritchie, Elizabeth A.

2003-01-01

The Fourth Convection and Moisture Experiment (CAMEX-4) provided a unique opportunity to observe the distributions and document the roles of important atmospheric factors that impact the development of the core asymmetries and core structural changes of tropical cyclones embedded in vertical wind shear. The state-of-the-art instruments flown on the NASA DC-8 and ER-2, in addition to those on the NOAA aircraft, provided a unique set of observations that documented the core structure throughout the depth of the tropical cyclone. These data have been used to conduct a combined observational and modeling study using a state-of-the-art, high- resolution mesoscale model to examine the role of the environmental vertical wind shear in producing tropical cyclone core asymmetries, and the effects on the structure and intensity of tropical cyclones.The scientific objectives of this study were to obtain in situ measurements that would allow documentation of the physical mechanisms that influence the development of the asymmetric convection and its effect on the core structure of the tropical cyclone.

Increased expression of chemokine receptor CCR3 and its ligands in ulcerative colitis: the role of colonic epithelial cells in in vitro studies.

PubMed

Manousou, P; Kolios, G; Valatas, V; Drygiannakis, I; Bourikas, L; Pyrovolaki, K; Koutroubakis, I; Papadaki, H A; Kouroumalis, E

2010-11-01

Human colonic epithelial cells express T helper type 1 (Th1)-associated chemoattractants, yet little is known about the production of Th2-associated chemoattractants. CCL11/eotaxin-1, CCL24/eotaxin-2 and CCL26/eotaxin-3 are known to attract CCR3-expressing, Th2-polarized lymphocytes. We studied constitutive and inflammation-induced expression and production of CCR3 together with its ligands in the colon and peripheral blood of patients with inflammatory bowel disease (IBD) by flow cytometry, reverse transcription–polymerase chain reaction (RT–PCR) and enzyme-linked immunosorbent assay (ELISA). We further defined the regulated expression of these chemokines by RT–PCR and ELISA using cultured human epithelial cell lines. A higher fraction of peripheral T lymphocytes were found to be positive for CCR3 in patients with ulcerative colitis (UC) compared to Crohn’s disease (CD), while almost no CCR3(+) T cells were found in normal controls (NC). Similarly, higher and more frequent expression of CCR3 was observed in colonic biopsies from patients with UC, regardless of the disease activity, when compared to CD or NCs. Serum CCL11/eotaxin-1 was increased significantly in UC (306 ± 87 pg/ml) and less so in CD (257 ± 43 pg/ml), whereas CCL24/eotaxin-2, and CCL26/eotaxin-3 were increased only in UC. Colonic expression of the three chemokines was minimal in NCs but high in inflammatory bowel diseases (especially UC) and was independent of disease activity. Th2, and to a lesser extent Th1, cytokines were able to induce expression and production of all three eotaxins from colonic epithelial cells in culture. CCR3 and ligands over-expression would appear to be a characteristic of UC. The production of CCR3 ligands by human colonic epithelial cells suggests further that epithelium can play a role in modulating pathological T cell-mediated mucosal inflammation.

Evidence for viral virulence as a predominant factor limiting human immunodeficiency virus vaccine efficacy.

PubMed

Mooij, P; Bogers, W M; Oostermeijer, H; Koornstra, W; Ten Haaft, P J; Verstrepen, B E; Van Der Auwera, G; Heeney, J L

2000-05-01

Current strategies in human immunodeficiency virus type 1 (HIV-1) vaccine development are often based on the production of different vaccine antigens according to particular genetic clades of HIV-1 variants. To determine if virus virulence or genetic distance had a greater impact on HIV-1 vaccine efficacy, we designed a series of heterologous chimeric simian/human immunodeficiency virus (SHIV) challenge experiments in HIV-1 subunit-vaccinated rhesus macaques. Of a total of 22 animals, 10 nonimmunized animals served as controls; the remainder were vaccinated with the CCR5 binding envelope of HIV-1(W6.1D). In the first study, heterologous challenge included two nonpathogenic SHIV chimeras encoding the envelopes of the divergent clade B HIV-1(han2) and HIV-1(sf13) strains. In the second study, all immunized animals were rechallenged with SHIV(89. 6p), a virus closely related to the vaccine strain but highly virulent. Protection from either of the divergent SHIV(sf13) or SHIV(han2) challenges was demonstrated in the majority of the vaccinated animals. In contrast, upon challenge with the more related but virulent SHIV(89.6p), protection was achieved in only one of the previously protected vaccinees. A secondary but beneficial effect of immunization on virus load and CD4(+) T-cell counts was observed despite failure to protect from infection. In addition to revealing different levels of protective immunity, these results suggest the importance of developing vaccine strategies capable of protecting from particularly virulent variants of HIV-1.

Zika and Spondweni Viruses: Historic Evidence of Misidentification, Misdiagnosis and Serious Clinical Disease Manifestations

DTIC Science & Technology

2016-10-01

in 37 Nigeria in 1952 (strain Chuku) (2). Cross-reactivity using the neutralization test led to the 38 misidentification of the SPONV Chuku strain...509-20. 149 2. Macnamara FN. Zika virus: a report on three cases of human infection during an 150 epidemic of jaundice in Nigeria . Transactions of...two serological surveys made in South Western Nigeria . Transactions 165 of the Royal Society of Tropical Medicine and Hygiene. 1959;53(2):202-12

Differential impact of lytic viruses on prokaryotic morphopopulations in a tropical estuarine system (Cochin estuary, India).

PubMed

Jasna, Vijayan; Pradeep Ram, Angia Sriram; Parvathi, Ammini; Sime-Ngando, Telesphore

2018-01-01

Our understanding on the importance of viral lysis in the functioning of tropical estuarine ecosystem is limited. This study examines viral infection of prokaryotes and subsequent lysis of cells belonging to different morphotypes across a salinity gradient in monsoon driven estuarine ecosystem (Cochin estuary, India). High standing stock of viruses and prokaryotes accompanied by lytic infection rates in the euryhaline/mesohaline region of the estuary suggests salinity to have an influential role in driving interactions between prokaryotes and viruses. High prokaryotic mortality rates, up to 42% of prokaryote population in the pre-monsoon season is further substantiated by a high virus to prokaryote ratio (VPR), suggesting that maintenance of a high number of viruses is dependent on the most active fraction of bacterioplankton. Although myoviruses were the dominant viral morphotype (mean = 43%) throughout the study period, there was significant variation among prokaryotic morphotypes susceptible to viral infection. Among them, the viral infected short rod prokaryote morphotype with lower burst estimates (mean = 18 viruses prokaryote-1) was dominant (35%) in the dry seasons whereas a substantial increase in cocci forms (30%) infected by viruses with high burst size (mean = 31 viruses prokaryote-1) was evident during the monsoon season. Such preferential infections of prokaryotic morphopopulations with respect to seasons can have a strong and variable impact on the carbon and energy flow in this tropical ecosystem.

Differential impact of lytic viruses on prokaryotic morphopopulations in a tropical estuarine system (Cochin estuary, India)

PubMed Central

Jasna, Vijayan; Pradeep Ram, Angia Sriram; Sime-Ngando, Telesphore

2018-01-01

Our understanding on the importance of viral lysis in the functioning of tropical estuarine ecosystem is limited. This study examines viral infection of prokaryotes and subsequent lysis of cells belonging to different morphotypes across a salinity gradient in monsoon driven estuarine ecosystem (Cochin estuary, India). High standing stock of viruses and prokaryotes accompanied by lytic infection rates in the euryhaline/mesohaline region of the estuary suggests salinity to have an influential role in driving interactions between prokaryotes and viruses. High prokaryotic mortality rates, up to 42% of prokaryote population in the pre-monsoon season is further substantiated by a high virus to prokaryote ratio (VPR), suggesting that maintenance of a high number of viruses is dependent on the most active fraction of bacterioplankton. Although myoviruses were the dominant viral morphotype (mean = 43%) throughout the study period, there was significant variation among prokaryotic morphotypes susceptible to viral infection. Among them, the viral infected short rod prokaryote morphotype with lower burst estimates (mean = 18 viruses prokaryote-1) was dominant (35%) in the dry seasons whereas a substantial increase in cocci forms (30%) infected by viruses with high burst size (mean = 31 viruses prokaryote-1) was evident during the monsoon season. Such preferential infections of prokaryotic morphopopulations with respect to seasons can have a strong and variable impact on the carbon and energy flow in this tropical ecosystem. PMID:29534102

Zoonotic encephalitides caused by arboviruses: transmission and epidemiology of alphaviruses and flaviviruses

PubMed Central

Balasuriya, Udeni B. R.; Lee, Chong-kyo

2014-01-01

In this review, we mainly focus on zoonotic encephalitides caused by arthropod-borne viruses (arboviruses) of the families Flaviviridae (genus Flavivirus) and Togaviridae (genus Alphavirus) that are important in both humans and domestic animals. Specifically, we will focus on alphaviruses (Eastern equine encephalitis virus, Western equine encephalitis virus, Venezuelan equine encephalitis virus) and flaviviruses (Japanese encephalitis virus and West Nile virus). Most of these viruses were originally found in tropical regions such as Africa and South America or in some regions in Asia. However, they have dispersed widely and currently cause diseases around the world. Global warming, increasing urbanization and population size in tropical regions, faster transportation and rapid spread of arthropod vectors contribute in continuous spreading of arboviruses into new geographic areas causing reemerging or resurging diseases. Most of the reemerging arboviruses also have emerged as zoonotic disease agents and created major public health issues and disease epidemics. PMID:24427764

Zoonotic encephalitides caused by arboviruses: transmission and epidemiology of alphaviruses and flaviviruses.

PubMed

Go, Yun Young; Balasuriya, Udeni B R; Lee, Chong-Kyo

2014-01-01

In this review, we mainly focus on zoonotic encephalitides caused by arthropod-borne viruses (arboviruses) of the families Flaviviridae (genus Flavivirus) and Togaviridae (genus Alphavirus) that are important in both humans and domestic animals. Specifically, we will focus on alphaviruses (Eastern equine encephalitis virus, Western equine encephalitis virus, Venezuelan equine encephalitis virus) and flaviviruses (Japanese encephalitis virus and West Nile virus). Most of these viruses were originally found in tropical regions such as Africa and South America or in some regions in Asia. However, they have dispersed widely and currently cause diseases around the world. Global warming, increasing urbanization and population size in tropical regions, faster transportation and rapid spread of arthropod vectors contribute in continuous spreading of arboviruses into new geographic areas causing reemerging or resurging diseases. Most of the reemerging arboviruses also have emerged as zoonotic disease agents and created major public health issues and disease epidemics.

Relevance of CCL3/CCR5 axis in oral carcinogenesis.

PubMed

da Silva, Janine Mayra; Moreira Dos Santos, Tálita Pollyanna; Sobral, Lays Martin; Queiroz-Junior, Celso Martins; Rachid, Milene Alvarenga; Proudfoot, Amanda E I; Garlet, Gustavo Pompermaier; Batista, Aline Carvalho; Teixeira, Mauro Martins; Leopoldino, Andréia Machado; Russo, Remo Castro; Silva, Tarcília Aparecida

2017-08-01

The chemokine CCL3 is a chemotactic cytokine crucial for inflammatory cell recruitment in homeostatic and pathological conditions. CCL3 might stimulate cancer progression by promoting leukocyte accumulation, angiogenesis and tumour growth. The expression of CCL3 and its receptors CCR1 and CCR5 was demonstrated in oral squamous cell carcinoma (OSCC), but their role was not defined. Here, the functions of CCL3 were assessed using a model of chemically induced tongue carcinogenesis with 4-nitroquinoline-1-oxide (4NQO). Lineages of OSCC were used to analyse the effects of CCL3 in vitro . The 4NQO-induced lesions exhibited increased expression of CCL3, CCR1 and CCR5. CCL3 -/- and CCR5 -/- mice presented reduced incidence of tongue tumours compared to wild-type (WT) and CCR1 -/- mice. Consistently, attenuated cytomorphological atypia and reduced cell proliferation were observed in lesions of CCL3 -/- and CCR5 -/- mice. OSCC from CCL3 -/- mice exhibited lower infiltration of eosinophils and reduced expression of Egf, Fgf1, Tgf-β1, Vegfa, Vegfb, Itga-4, Vtn, Mmp-1a, Mmp-2 and Mmp-9 than WT mice. In vitro , CCL3 induced invasion and production of CCL5, IL-6, MMP -2, -8, -9. Blockage of CCL3 in vitro using α-CCL3 or Evasin-1 (a CCL3-binding protein) impaired tumour cell invasion. In conclusion, CCL3/CCR5 axis has pro-tumourigenic effects in oral carcinogenesis. The induction of inflammatory and angiogenic pathways and eosinophils recruitment appear to be the underlying mechanism explaining these effects. These data reveal potential protective effects of CCL3 blockade in oral cancer.

Genetic modification of mesenchymal stem cells overexpressing CCR1 increases cell viability, migration, engraftment, and capillary density in the injured myocardium.

PubMed

Huang, Jing; Zhang, Zhiping; Guo, Jian; Ni, Aiguo; Deb, Arjun; Zhang, Lunan; Mirotsou, Maria; Pratt, Richard E; Dzau, Victor J

2010-06-11

Although mesenchymal stem cell (MSC) transplantation has been shown to promote cardiac repair in acute myocardial injury in vivo, its overall restorative capacity appears to be restricted mainly because of poor cell viability and low engraftment in the ischemic myocardium. Specific chemokines are upregulated in the infarcted myocardium. However the expression levels of the corresponding chemokine receptors (eg, CCR1, CXCR2) in MSCs are very low. We hypothesized that this discordance may account for the poor MSC engraftment and survival. To determine whether overexpression of CCR1 or CXCR2 chemokine receptors in MSCs augments their cell survival, migration and engraftment after injection in the infarcted myocardium. Overexpression of CCR1, but not CXCR2, dramatically increased chemokine-induced murine MSC migration and protected MSC from apoptosis in vitro. Moreover, when MSCs were injected intramyocardially one hour after coronary artery ligation, CCR1-MSCs accumulated in the infarcted myocardium at significantly higher levels than control-MSCs or CXCR2-MSCs 3 days postmyocardial infarction (MI). CCR1-MSC-injected hearts exhibited a significant reduction in infarct size, reduced cardiomyocytes apoptosis and increased capillary density in injured myocardium 3 days after MI. Furthermore, intramyocardial injection of CCR1-MSCs prevented cardiac remodeling and restored cardiac function 4 weeks after MI. Our results demonstrate the in vitro and in vivo salutary effects of genetic modification of stem cells. Specifically, overexpression of chemokine receptor enhances the migration, survival and engraftment of MSCs, and may provide a new therapeutic strategy for the injured myocardium.

The Chemokine Receptor CCR1 Is Constitutively Active, Which Leads to G Protein-independent, β-Arrestin-mediated Internalization*

PubMed Central

Gilliland, C. Taylor; Salanga, Catherina L.; Kawamura, Tetsuya; Trejo, JoAnn; Handel, Tracy M.

2013-01-01

Activation of G protein-coupled receptors by their associated ligands has been extensively studied, and increasing structural information about the molecular mechanisms underlying ligand-dependent receptor activation is beginning to emerge with the recent expansion in GPCR crystal structures. However, some GPCRs are also able to adopt active conformations in the absence of agonist binding that result in the initiation of signal transduction and receptor down-modulation. In this report, we show that the CC-type chemokine receptor 1 (CCR1) exhibits significant constitutive activity leading to a variety of cellular responses. CCR1 expression is sufficient to induce inhibition of cAMP formation, increased F-actin content, and basal migration of human and murine leukocytes. The constitutive activity leads to basal phosphorylation of the receptor, recruitment of β-arrestin-2, and subsequent receptor internalization. CCR1 concurrently engages Gαi and β-arrestin-2 in a multiprotein complex, which may be accommodated by homo-oligomerization or receptor clustering. The data suggest the presence of two functional states for CCR1; whereas receptor coupled to Gαi functions as a canonical GPCR, albeit with high constitutive activity, the CCR1·β-arrestin-2 complex is required for G protein-independent constitutive receptor internalization. The pertussis toxin-insensitive uptake of chemokine by the receptor suggests that the CCR1·β-arrestin-2 complex may be related to a potential scavenging function of the receptor, which may be important for maintenance of chemokine gradients and receptor responsiveness in complex fields of chemokines during inflammation. PMID:24056371

Suppressive effects of a novel CC chemokine receptor 4 antagonist on Th2 cell trafficking in ligand- and antigen-induced mouse models.

PubMed

Komiya, Takaki; Sugiyama, Tetsuya; Takeda, Kazuhiko; Watanabe, Noriki; Imai, Masamichi; Kokubo, Masaya; Tokuda, Natsuko; Ochiai, Hiroshi; Habashita, Hiromu; Shibayama, Shiro

2013-11-15

CC chemokine receptor 4 (CCR4) has been implicated as a preferential marker for T helper type 2 (Th2) cells, and is believed to be involved in the pathology of allergic diseases by controlling Th2 cell trafficking into inflamed tissues. The objective of the study was to characterize the pharmacological properties of E0001-163, a novel CCR4 antagonist. E0001-163 was tested in both in vitro chemotaxis assays as well as in vivo mouse models of CCR4 ligand-induced air pouch and antigen-induced airway inflammation by utilizing in vitro-polarized Th2 cells. In vitro, E0001-163 inhibited migratory response of human Th2-polarized cells to CCL22, a CCR4 ligand, with an IC50 value of 11.9 nM. E0001-163 significantly suppressed CCL22-induced Th2 cell trafficking into mouse air pouch in a dose-dependent manner at doses of 3 and 10mg/kg, suggesting that E0001-163 has an inhibitory effect on CCR4-mediated T cell trafficking in vivo. In addition, E0001-163 partially decreased Th2 cell trafficking and the level of IL-4 in the lungs in Th2-tansferred and ovalbumin (OVA)-challenged mice. T cell trafficking involves multiple chemokine receptors both in acute and chronic phases, and our findings suggest that CCR4, together with other chemokine receptors, may be involved in Th2 cell trafficking under disease conditions. © 2013 Elsevier B.V. All rights reserved.

MBCP - Approach - Immunotherapy | Center for Cancer Research

Cancer.gov

Immunotherapy CCR investigators pioneered the use of the tuberculosis vaccine—Bacillus Calmette-Guerin (BCG)—in the treatment of bladder cancer. In cases where the tumor burden is not too high and direct contact can be made with the urothelium surface of the bladder, BCG application appears to elicit an immune response that attacks the tumor as well as the attenuated virus. Ongoing clinical trials focusing on enhancing the patient’s immune system are listed below.

CCL20 and β-defensin-2 induce arrest of human Th17 cells on inflamed endothelium in vitro under flow conditions.

PubMed

Ghannam, Soufiane; Dejou, Cécile; Pedretti, Nathalie; Giot, Jean-Philipe; Dorgham, Karim; Boukhaddaoui, Hassan; Deleuze, Virginie; Bernard, François-Xavier; Jorgensen, Christian; Yssel, Hans; Pène, Jérôme

2011-02-01

CCR6 is a chemokine receptor that is expressed at the cell surface of Th17 cells, an IL-17- and IL-22-secreting population of CD4(+) T cells with antipathogenic, as well as inflammatory, properties. In the current study, we have determined the involvement of CCR6 in human Th17 lymphocyte migration toward inflamed tissue by analyzing the capacity of its ligands to induce arrest of these cells onto inflamed endothelium in vitro under flow conditions. We show that polarized, in situ-differentiated, skin-derived Th17 clones activated via the TCR-CD3 complex produce CCL20 in addition to IL-17 and IL-22. The latter cytokines induce, in a synergic fashion, the production of human β-defensin (hBD)-2, but neither hBD-1 nor hBD-3, by epidermal keratinocytes. Both CCL20 and hBD-2 are capable of inducing the arrest of Th17 cells, but not Th1 or Th2 cells, on HUVEC in an CD54-dependent manner that is CCR6 specific and independent from the expression of CXCR4, reported to be an alternative receptor for hBD-2. In addition, Ag-specific activation induces a transient loss of CCR6 expression, both at the transcriptional and protein level, which occurs with slow kinetics and is not due to endogenous CCL20-mediated internalization of CCR6. Together, these results indicate that Ag-specific activation will initially contribute to CCR6-mediated Th17 cell trafficking toward and sequestration in inflamed tissue, but that it eventually results in a transitory state of nonresponsiveness to further stimulation of these cells with CCR6 ligands, thus permitting their subsequent migration out of the inflamed site.

Experience with Hong Kong influenza in tropical areas*

PubMed Central

Buescher, E. L.; Smith, T. J.; Zachary, I. H.

1969-01-01

Following the introduction of A2/Hong Kong/68 influenza virus into 2 different susceptible populations residing in Thailand and the Panama Canal Zone, epidemic disease occurred within approximately 1 month. The establishment and transmission of the virus, and the disease it caused, were studied in detail. The rates of progression and extent of overt epidemic disease and the proportions of overt to subclinical infection in the 2 areas were different. In Korat Royal Thai Air Force Base, only 8% of US servicemen became ill, although 13% of them were infected. The epidemic progressed slowly, so that no more than 1.5% of the population were ill at any one time during the 3 months of its recognized presence. In contrast, in the Canal Zone the clinical attack rate approached 50%, and subclinical infection occurred in 5% or less of the population during a 6-week period when absenteeism from schools and work was quite obvious. Factors such as immunization status of the populations and virulence of virus seemed not to be primarily responsible for the differences. Rather, the latter appear to be related to differences in environmental circumstances, with crowding contributing to the higher disease incidence in the Canal Zone. The data show that this variant A2 virus may produce different epidemiological patterns of disease in tropical areas, similar to those observed in temperate zones. PMID:5309443

Experience with Hong Kong influenza in tropical areas.

PubMed

Buescher, E L; Smith, T J; Zachary, I H

1969-01-01

Following the introduction of A2/Hong Kong/68 influenza virus into 2 different susceptible populations residing in Thailand and the Panama Canal Zone, epidemic disease occurred within approximately 1 month. The establishment and transmission of the virus, and the disease it caused, were studied in detail. The rates of progression and extent of overt epidemic disease and the proportions of overt to subclinical infection in the 2 areas were different. In Korat Royal Thai Air Force Base, only 8% of US servicemen became ill, although 13% of them were infected. The epidemic progressed slowly, so that no more than 1.5% of the population were ill at any one time during the 3 months of its recognized presence. In contrast, in the Canal Zone the clinical attack rate approached 50%, and subclinical infection occurred in 5% or less of the population during a 6-week period when absenteeism from schools and work was quite obvious. Factors such as immunization status of the populations and virulence of virus seemed not to be primarily responsible for the differences. Rather, the latter appear to be related to differences in environmental circumstances, with crowding contributing to the higher disease incidence in the Canal Zone.The data show that this variant A2 virus may produce different epidemiological patterns of disease in tropical areas, similar to those observed in temperate zones.

Molecular cloning and expression analysis of the canine chemokine receptor CCR9.

PubMed

Maeda, Shingo; Ohno, Koichi; Tsukamoto, Atsushi; Nakashima, Ko; Fukushima, Kenjiro; Goto-Koshino, Yuko; Fujino, Yasuhito; Tsujimoto, Hajime

2012-01-15

The chemokine receptor CCR9, which interacts with the thymus-expressed chemokine TECK/CCL25, contributes to the localization of lymphocytes to the small intestine, and is implicated in the development of human inflammatory bowel disease (IBD); however, their role in canine IBD is unknown. The objective of this study was to isolate cDNA encoding CCR9 and to investigate CCR9 expression in normal canine tissues and lymphoid cell lines. The complete open reading frame contained 1104 bp, encoding 367 amino acids, with 85% and 81% identity to human and mouse homologs, respectively. CCR9 mRNA was detected in all tissues investigated with the highest expression level in the small intestine. CCR9 mRNA was also expressed in GL-1, a canine B cell leukemia cell line, but not in CLBL-1, a canine B cell lymphoma cell line. Immunoblot and flow cytometry analyses of these cell lines using an anti-human CCR9 monoclonal antibody revealed that CCR9 protein expression was detected only in GL-1, indicating the cross-reactivity of the antibody. Using the antibody, flow cytometry showed that the proportions of CCR9(+) cells were small (mean, 4.88%; SD, 2.15%) in the normal canine PBMCs. This study will be useful in understanding canine intestinal immunity and the immunopathogenesis of canine IBD. Copyright © 2011 Elsevier B.V. All rights reserved.

Viremic HIV Infected Individuals with High CD4 T Cells and Functional Envelope Proteins Show Anti-gp41 Antibodies with Unique Specificity and Function

PubMed Central

Curriu, Marta; Fausther-Bovendo, Hughes; Pernas, María; Massanella, Marta; Carrillo, Jorge; Cabrera, Cecilia; López-Galíndez, Cecilio; Clotet, Bonaventura; Debré, Patrice; Vieillard, Vincent; Blanco, Julià

2012-01-01

Background CD4 T-cell decay is variable among HIV-infected individuals. In exceptional cases, CD4 T-cell counts remain stable despite high plasma viremia. HIV envelope glycoprotein (Env) properties, namely tropism, fusion or the ability to induce the NK ligand NKp44L, or host factors that modulate Env cytopathic mechanisms may be modified in such situation. Methods We identified untreated HIV-infected individuals showing non-cytopathic replication (VL>10,000 copies/mL and CD4 T-cell decay<50 cells/µL/year, Viremic Non Progressors, VNP) or rapid progression (CD4 T-cells<350 cells/µL within three years post-infection, RP). We isolated full-length Env clones and analyzed their functions (tropism, fusion activity and capacity to induce NKp44L expression on CD4 cells). Anti-Env humoral responses were also analyzed. Results Env clones isolated from VNP or RP individuals showed no major phenotypic differences. The percentage of functional clones was similar in both groups. All clones tested were CCR5-tropic and showed comparable expression and fusogenic activity. Moreover, no differences were observed in their capacity to induce NKp44L expression on CD4 T cells from healthy donors through the 3S epitope of gp41. In contrast, anti- Env antibodies showed clear functional differences: plasma from VNPs had significantly higher capacity than RPs to block NKp44L induction by autologous viruses. Consistently, CD4 T-cells isolated from VNPs showed undetectable NKp44L expression and specific antibodies against a variable region flanking the highly conserved 3S epitope were identified in plasma samples from these patients. Conversely, despite continuous antigen stimulation, VNPs were unable to mount a broad neutralizing response against HIV. Conclusions Env functions (fusion and induction of NKp44L) were similar in viremic patients with slow or rapid progression to AIDS. However, differences in humoral responses against gp41 epitopes nearby 3S sequence may contribute to the lack of CD4 T cell decay in VNPs by blocking the induction of NKp44L by gp41. PMID:22312424

Characterization and analysis of CCR and CAD gene families at the whole-genome level for lignin synthesis of stone cells in pear (Pyrus bretschneideri) fruit

PubMed Central

Cheng, Xi; Li, Manli; Li, Dahui; Zhang, Jinyun; Jin, Qing; Sheng, Lingling; Lin, Yi

2017-01-01

ABSTRACT The content of stone cells has significant effects on the flavour and quality of pear fruit. Previous research suggested that lignin deposition is closely related to stone cell formation. In the lignin biosynthetic pathway, cinnamoyl-CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD), dehydrogenase/reductase family members, catalyse the last two steps in monolignol synthesis. However, there is little knowledge of the characteristics of the CCR and CAD families in pear and their involvement in lignin synthesis of stone cells. In this study, 31 CCRs and 26 CADs were identified in the pear genome. Phylogenetic trees for CCRs and CADs were constructed; key amino acid residues were analysed, and three-dimensional structures were predicted. Using quantitative real-time polymerase chain reaction (qRT-PCR), PbCAD2, PbCCR1, -2 and -3 were identified as participating in lignin synthesis of stone cells in pear fruit. Subcellular localization analysis showed that the expressed proteins (PbCAD2, PbCCR1, -2 and -3) are found in the cytoplasm or at the cell membrane. These results reveal the evolutionary features of the CCR and CAD families in pear as well as the genes responsible for regulation of lignin synthesis and stone cell development in pear fruit. PMID:29141952

Postdoctoral Fellow | Center for Cancer Research

Cancer.gov

The Neuro-Oncology Branch (NOB), Center for Cancer Research (CCR), National Cancer Institute (NCI) of the National Institutes of Health (NIH) is seeking outstanding postdoctoral candidates interested in studying metabolic and cell signaling pathways in the context of brain cancers through construction of computational models amenable to formal computational analysis and simulation. The ability to closely collaborate with the modern metabolomics center developed at CCR provides a unique opportunity for a postdoctoral candidate with a strong theoretical background and interest in demonstrating the incredible potential of computational approaches to solve problems from scientific disciplines and improve lives. The candidate will be given the opportunity to both construct data-driven models, as well as biologically validate the models by demonstrating the ability to predict the effects of altering tumor metabolism in laboratory and clinical settings.

Identification of men with low-risk biopsy-confirmed prostate cancer as candidates for active surveillance.

PubMed

Lin, Daniel W; Crawford, E David; Keane, Thomas; Evans, Brent; Reid, Julia; Rajamani, Saradha; Brown, Krystal; Gutin, Alexander; Tward, Jonathan; Scardino, Peter; Brawer, Michael; Stone, Steven; Cuzick, Jack

2018-06-01

A combined clinical cell-cycle risk (CCR) score that incorporates prognostic molecular and clinical information has been recently developed and validated to improve prostate cancer mortality (PCM) risk stratification over clinical features alone. As clinical features are currently used to select men for active surveillance (AS), we developed and validated a CCR score threshold to improve the identification of men with low-risk disease who are appropriate for AS. The score threshold was selected based on the 90th percentile of CCR scores among men who might typically be considered for AS based on NCCN low/favorable-intermediate risk criteria (CCR = 0.8). The threshold was validated using 10-year PCM in an unselected, conservatively managed cohort and in the subset of the same cohort after excluding men with high-risk features. The clinical effect was evaluated in a contemporary clinical cohort. In the unselected validation cohort, men with CCR scores below the threshold had a predicted mean 10-year PCM of 2.7%, and the threshold significantly dichotomized low- and high-risk disease (P = 1.2 × 10 -5 ). After excluding high-risk men from the validation cohort, men with CCR scores below the threshold had a predicted mean 10-year PCM of 2.3%, and the threshold significantly dichotomized low- and high-risk disease (P = 0.020). There were no prostate cancer-specific deaths in men with CCR scores below the threshold in either analysis. The proportion of men in the clinical testing cohort identified as candidates for AS was substantially higher using the threshold (68.8%) compared to clinicopathologic features alone (42.6%), while mean 10-year predicted PCM risks remained essentially identical (1.9% vs. 2.0%, respectively). The CCR score threshold appropriately dichotomized patients into low- and high-risk groups for 10-year PCM, and may enable more appropriate selection of patients for AS. Copyright © 2018 Elsevier Inc. All rights reserved.

The chemokines CCR1 and CCRL2 have a role in colorectal cancer liver metastasis.

PubMed

Akram, Israa G; Georges, Rania; Hielscher, Thomas; Adwan, Hassan; Berger, Martin R

2016-02-01

C-C chemokine receptor type 1 (CCR1) and chemokine C-C motif receptor-like 2 (CCRL2) have not yet been sufficiently investigated for their role in colorectal cancer (CRC). Here, we investigated their expression in rat and human CRC samples, their modulation of expression in a rat liver metastasis model, as well as the effects on cellular properties resulting from their knockdown. One rat and five human colorectal cancer cell lines were used. CC531 rat colorectal cells were injected via the portal vein into rats and re-isolated from rat livers after defined periods. Following mRNA isolation, the gene expression was investigated by microarray. In addition, all cell lines were screened for mRNA expression of CCR1 and CCRL2 by reverse transcription polymerase chain reaction (RT-PCR). Cell lines with detectable expression were used for knockdown experiments; and the respective influence was determined on the cells' proliferation, scratch closure, and colony formation. Finally, specimens from the primaries of 50 patients with CRC were monitored by quantitative RT-PCR for CCR1 and CCRL2 expression levels. The microarray studies showed peak increases of CCR1 and CCRL2 in the early phase of liver colonization. Knockdown was sufficient at mRNA but only moderate at protein levels and resulted in modest but significant inhibition of proliferation (p < 0.05), scratch closure, and colony formation (p < 0.05). All human CRC samples were positive for CCR1 and CCRL2 and showed a significant pairwise correlation (p < 0.0004), but there was no correlation with tumor stage or age of patients. In summary, the data point to an important role of CCR1 and CCRL2 under conditions of organ colonization and both chemokine receptors qualify as targets of treatment during early colorectal cancer liver metastasis.

Diversity of staphylococcal cassette chromosome mec elements in nosocomial multiresistant Staphylococcus haemolyticus isolates.

PubMed

Szczuka, Ewa; Krajewska, Magdalena; Lijewska, Dagmara; Bosacka, Karolina; Kaznowski, Adam

2016-11-01

Staphylococcus haemolyticus is the second, most frequently isolated coagulase-negative staphyloccus (CoNS) from patients with hospital-acquired infections, and it is usually resistant to methicillin and other semisynthetic penicillins. The purpose of this study was to characterize staphylococcal cassette chromosome mec (SCCmec) elements and assess the in-vitro activity of antibiotics against 60 S. haemolyticus strains recovered from hospitalized patients. All these strains expressed methicillin resistance and carried a mecA gene. Moreover, all strains possessed a multiresistant phenotype, i.e., exhibited resistance to more than three classes of antibiotics. Eleven strains (18 %) harbored the SCCmec type V, containing ccrC and mec complex C. Three isolates harboring the ccrC gene did not contain a known mec complex. One strain positive for mec complex C was not typeable for ccr. This suggests that ccrC and mec complex C may exist autonomously. Only four strains carried mec complex B, whereas none of the S. haemolyticus harboured mec complex A. A new combination, which is mec complex B-ccrAB ship , was found in S. haemolitycus. The ccrAB ship was also identified in two strains of S. haemolitycus in which the mec gene complex was not identified. The results of the present study indicate that in S. haemolyticus the mec gene complex and the ccr genes are highly divergent. However, ccr sequence analysis does not allow the identification of a new allotype, based on a cut-off value of 85 % identity. The ccr genes in the S. haemolitycus strain showed ≥96 % sequence identity to the ccrAB2 genes.

Detecting the emergence of novel, zoonotic viruses pathogenic to humans.

PubMed

Rosenberg, Ronald

2015-03-01

RNA viruses, with their high potential for mutation and epidemic spread, are the most common class of pathogens found as new causes of human illness. Despite great advances made in diagnostic technology since the 1950s, the annual rate at which novel virulent viruses have been found has remained at 2-3. Most emerging viruses are zoonoses; they have jumped from mammal or bird hosts to humans. An analysis of virus discovery indicates that the small number of novel viruses discovered annually is an artifact of inadequate surveillance in tropical and subtropical countries, where even established endemic pathogens are often misdiagnosed. Many of the emerging viruses of the future are already infecting humans but remain to be uncovered by a strategy of disease surveillance in selected populations.

Expression of chemokine receptor CCR7 is a negative prognostic factor for patients with gastric cancer: a meta-analysis.

PubMed

Du, Peizhun; Liu, Yongchao; Ren, Hong; Zhao, Jing; Zhang, Xiaodan; Patel, Rajan; Hu, Chenen; Gan, Jun; Huang, Guangjian

2017-03-01

The prognostic significance of CC chemokine receptor type 7 (CCR7) for survival of patients with gastric cancer remains controversial. To investigate the impacts of CCR7 on clinicopathological findings and survival outcome in gastric cancer, we performed a meta-analysis. A comprehensive search in PubMed, Embase, the Cochrane Library, and the CNKI database (1966 to November 2015) was undertaken for relevant studies. The relative risk and hazard ratios with their 95 % confidence intervals were used as measures to investigate the correlation between CCR7 expression and clinicopathological findings and overall survival rate. Sensitivity analysis was conducted to assess the stability of outcomes. Fifteen eligible studies comprising 1697 participants were included in our analysis. The pooled relative risks indicated CCR7 expression was significantly associated with deeper tumor invasion [0.61, 95 % confidence interval (CI) 0.45-0.84, p = 0.003], advanced stage (0.47, 95 % CI 0.32-0.69, p < 0.001), vascular invasion (2.12, 95 % CI 1.20-3.73, p = 0.009), lymph node metastasis (2.00, 95 % CI 1.48-2.70, p < 0.001), and lymphatic invasion (1.98, 95 % CI 1.43-2.72, p < 0.001) but not with age, tumor size, and histological type. The pooling of hazard ratios showed a significant relationship between positive CCR7 expression and worse 5-year overall survival rate (0.46, 95 % CI 0.31-0.70, p < 0.001). Our meta-analysis indicated high CCR7 expression is likely to be a negative clinicopathological prognostic factor for patients with gastric cancer and to predict a worse long-term survival outcome.

Efficient Single Tobamoviral Vector-Based Bioproduction of Broadly Neutralizing Anti-HIV-1 Monoclonal Antibody VRC01 in Nicotiana benthamiana Plants and Utility of VRC01 in Combination Microbicides

PubMed Central

Hamorsky, Krystal Teasley; Grooms-Williams, Tiffany W.; Husk, Adam S.; Bennett, Lauren J.; Palmer, Kenneth E.

2013-01-01

Broadly neutralizing monoclonal antibodies (bnMAbs) may offer powerful tools for HIV-1 preexposure prophylaxis, such as topical microbicides. However, this option is hampered due to expensive MAb biomanufacturing based on mammalian cell culture. To address this issue, we developed a new production system for bnMAb VRC01 in Nicotiana benthamiana plants using a tobamovirus replicon vector. Unlike conventional two-vector-based expression, this system was designed to overexpress full-length IgG1 from a single polypeptide by means of kex2p-like enzyme recognition sites introduced between the heavy and light chains. An enzyme-linked immunosorbent assay (ELISA) revealed that gp120-binding VRC01 IgG1 was maximally accumulated on 5 to 7 days following vector inoculation, yielding ∼150 mg of the bnMAb per kg of fresh leaf material. The plant-made VRC01 (VRC01p) was efficiently purified by protein A affinity followed by hydrophobic-interaction chromatography. ELISA, surface plasmon resonance, and an HIV-1 neutralization assay demonstrated that VRC01p has gp120-binding affinity and HIV-1-neutralization capacity virtually identical to the human-cell-produced counterpart. To advance VRC01p's use in topical microbicides, we analyzed combinations of the bnMAb with other microbicide candidates holding distinct antiviral mechanisms in an HIV-1 neutralization assay. VRC01p exhibited clear synergy with the antiviral lectin griffithsin, the CCR5 antagonist maraviroc, and the reverse transcriptase inhibitor tenofovir in multiple CCR5-tropic HIV-1 strains from clades A, B, and C. In summary, VRC01p is amenable to robust, rapid, and large-scale production and may be developed as an active component in combination microbicides with other anti-HIV agents such as antiviral lectins, CCR5 antagonists, and reverse transcriptase inhibitors. PMID:23403432

Improving GEOS-5 seven day forecast skill by assimilation of quality controlled AIRS temperature profiles

NASA Astrophysics Data System (ADS)

Susskind, J.; Rosenberg, R. I.

2016-12-01

The GEOS-5 Data Assimilation System (DAS) generates a global analysis every six hours by combining the previous six hour forecast for that time period with contemporaneous observations. These observations include in-situ observations as well as those taken by satellite borne instruments, such as AIRS/AMSU on EOS Aqua and CrIS/ATMS on S-NPP. Operational data assimilation methodology assimilates observed channel radiances Ri for IR sounding instruments such as AIRS and CrIS, but only for those channels i in a given scene whose radiances are thought to be unaffected by clouds. A limitation of this approach is that radiances in most tropospheric sounding channels are affected by clouds under partial cloud cover conditions, which occurs most of the time. The AIRS Science Team Version-6 retrieval algorithm generates cloud cleared radiances (CCR's) for each channel in a given scene, which represent the radiances AIRS would have observed if the scene were cloud free, and then uses them to determine quality controlled (QC'd) temperature profiles T(p) under all cloud conditions. There are potential advantages to assimilate either AIRS QC'd CCR's or QC'd T(p) instead of Ri in that the spatial coverage of observations is greater under partial cloud cover. We tested these two alternate data assimilation approaches by running three parallel data assimilation experiments over different time periods using GEOS-5. Experiment 1 assimilated all observations as done operationally, Experiment 2 assimilated QC'd values of AIRS CCRs in place of AIRS radiances, and Experiment 3 assimilated QC'd values of T(p) in place of observed radiances. Assimilation of QC'd AIRS T(p) resulted in significant improvement in seven day forecast skill compared to assimilation of CCR's or assimilation of observed radiances, especially in the Southern Hemisphere Extra-tropics.

Expression and function of CCL2/CCR2 in rat micturition reflexes and somatic sensitivity with urinary bladder inflammation

PubMed Central

Arms, Lauren; Girard, Beatrice M.; Malley, Susan E.

2013-01-01

Chemokines are proinflammatory mediators of the immune response, and there is growing evidence for chemokine/receptor signaling involvement in pronociception. Bladder pain syndrome (BPS)/interstitial cystitis (IC) is a chronic pain syndrome characterized by pain, pressure, or discomfort perceived to be bladder-related with at least one urinary symptom. We have explored the expression and functional roles of CCL2 (monocyte chemoattractant protein-1) and its high-affinity receptor, CCR2, in micturition reflex function and somatic sensitivity in rats with urinary bladder inflammation induced by cyclophosphamide (CYP) treatment of varying duration (4 h, 48 h, chronic). Real-time quantitative RT-PCR, ELISAs, and immunohistochemistry demonstrated significant (P ≤ 0.01) increases in CCL2 and CCR2 expression in the urothelium and in Fast Blue-labeled bladder afferent neurons in lumbosacral dorsal root ganglia with CYP-induced cystitis. Intravesical infusion of RS504393 (5 μM), a specific CCR2 antagonist, reduced voiding frequency and increased bladder capacity and void volume in rats with CYP-induced cystitis (4 h), as determined with open outlet, conscious cystometry. In addition, CCR2 blockade, at the level of the urinary bladder, reduced referred somatic sensitivity of the hindpaw and pelvic region in rats with CYP treatment, as determined with von Frey filament testing. We provide evidence of functional roles for CCL2/CCR2 signaling at the level of the urinary bladder in reducing voiding frequency and somatic sensitivity following CYP-induced cystitis (4 h). These studies suggest that chemokines/receptors may be novel targets with therapeutic potential in the context of urinary bladder inflammation. PMID:23594826

The proportionality of global warming to cumulative carbon emissions.

PubMed

Matthews, H Damon; Gillett, Nathan P; Stott, Peter A; Zickfeld, Kirsten

2009-06-11

The global temperature response to increasing atmospheric CO(2) is often quantified by metrics such as equilibrium climate sensitivity and transient climate response. These approaches, however, do not account for carbon cycle feedbacks and therefore do not fully represent the net response of the Earth system to anthropogenic CO(2) emissions. Climate-carbon modelling experiments have shown that: (1) the warming per unit CO(2) emitted does not depend on the background CO(2) concentration; (2) the total allowable emissions for climate stabilization do not depend on the timing of those emissions; and (3) the temperature response to a pulse of CO(2) is approximately constant on timescales of decades to centuries. Here we generalize these results and show that the carbon-climate response (CCR), defined as the ratio of temperature change to cumulative carbon emissions, is approximately independent of both the atmospheric CO(2) concentration and its rate of change on these timescales. From observational constraints, we estimate CCR to be in the range 1.0-2.1 degrees C per trillion tonnes of carbon (Tt C) emitted (5th to 95th percentiles), consistent with twenty-first-century CCR values simulated by climate-carbon models. Uncertainty in land-use CO(2) emissions and aerosol forcing, however, means that higher observationally constrained values cannot be excluded. The CCR, when evaluated from climate-carbon models under idealized conditions, represents a simple yet robust metric for comparing models, which aggregates both climate feedbacks and carbon cycle feedbacks. CCR is also likely to be a useful concept for climate change mitigation and policy; by combining the uncertainties associated with climate sensitivity, carbon sinks and climate-carbon feedbacks into a single quantity, the CCR allows CO(2)-induced global mean temperature change to be inferred directly from cumulative carbon emissions.

Characterization of influenza activity based on virological surveillance of influenza-like illness in tropical Singapore, 2010-2014.

PubMed

Ang, Li Wei; Tien, Wee Siong; Lin, Raymond Tzer-Pin; Cui, Lin; Cutter, Jeffery; James, Lyn; Goh, Kee Tai

2016-12-01

Singapore is situated in the tropics where the seasonality of influenza is not as well defined as that of temperate countries. We examined the circulation of influenza viruses in the community in terms of the characteristics of influenza activity. We reviewed laboratory-confirmed virological data collected between 2010 and 2014 under the national influenza surveillance programme. Influenza activity was measured by the proportion of specimens from outpatients with influenza-like illness tested positive for influenza virus based on 4-weekly moving interval. Seasonal epidemics occurred around the end of previous year or the beginning and middle of the year. Increases in influenza positivity were more pronounced when there was a change in the predominant circulating influenza virus type/subtype to influenza A(H3N2). Influenza epidemics lasted about 12 weeks on average, with longer duration when there was a change in the predominant influenza type/subtype and especially when it was associated with influenza A(H3N2). Continuous influenza surveillance is important as it could provide early warning of imminent surges in virus transmission, and allow for timely implementation of public health prevention and control interventions to minimize influenza-associated disease burden. J. Med. Virol. 88:2069-2077, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Performance of the Cockcroft-Gault and MDRD equations in adult Nigerians with chronic kidney disease.

PubMed

Agaba, Emmanuel I; Wigwe, Chinyere M; Agaba, Patricia A; Tzamaloukas, Antonios H

2009-01-01

Estimation of the glomerular filtration rate (GFR) is required in the assessment of patients with chronic kidney disease (CKD) in order to provide information regarding the functional status of the kidneys. Current guidelines advocate the use of prediction equations, such as the Cockcroft-Gault (CG) formula and the Modification of Diet in Renal Disease (MDRD) study-derived equations, over clearance of endogenous creatinine (Ccr) in achieving this aim. We were interested in knowing the accuracy of these equations in predicting the GFR in adult Nigerians with CKD. We conducted a review of records of patients who were evaluated for CKD at the Nephrology Clinic of the Jos University Teaching Hospital between 2001 and 2003. We compared the CG and MDRD equations against the Ccr in predicting the GFR in 130 patients (88 males and 42 females) with CKD. The means +/- standard deviation (SD) for the measured and predicted GFR by the CG and MDRD equations were similar (17.6 +/- 25.8 ml/min, 19.9 +/- 24.0 ml/min and 21.5 +/- 28.2 ml/min, respectively; analysis of variance [ANOVA], F = 0.68, P = 0.5). The mean difference between CG and Ccr was -2.2 +/- 14.8 ml/min, with discordance at Ccr values >25 ml/min. The mean difference between MDRD and Ccr was -3.9 +/- 18.1 ml/min, with discordance at Ccr values >40 ml/min. The CG and MDRD equations provide reliable alternatives to measured Ccr in the estimation of the GFR in Nigerian patients with CKD.

Expansion of somatically reverted memory CD8+ T cells in patients with X-linked lymphoproliferative disease caused by selective pressure from Epstein-Barr virus

PubMed Central

Low, Carol; Bell, Andrew I.; Abbott, Rachel J.M.; Phan, Tri Giang; Riminton, D. Sean; Choo, Sharon; Smart, Joanne M.; Lougaris, Vassilios; Giliani, Silvia; Buckley, Rebecca H.; Grimbacher, Bodo; Alvaro, Frank; Klion, Amy D.; Nichols, Kim E.; Adelstein, Stephen; Rickinson, Alan B.

2012-01-01

Patients with the primary immunodeficiency X-linked lymphoproliferative disease (XLP), which is caused by mutations in SH2D1A, are highly susceptible to Epstein-Barr virus (EBV) infection. Nonetheless, some XLP patients demonstrate less severe clinical manifestations after primary infection. SH2D1A encodes the adaptor molecule SLAM-associated protein (SAP), which is expressed in T and natural killer cells and is required for cytotoxicity against B cells, the reservoir for EBV. It is not known why the clinical presentation of XLP is so variable. In this study, we report for the first time the occurrence of somatic reversion in XLP. Reverted SAP-expressing cells resided exclusively within the CD8+ T cell subset, displayed a CD45RA−CCR7− effector memory phenotype, and were maintained at a stable level over time. Importantly, revertant CD8+ SAP+ T cells, but not SAP− cells, proliferated in response to EBV and killed EBV-infected B cells. As somatic reversion correlated with EBV infection, we propose that the virus exerts a selective pressure on the reverted cells, resulting in their expansion in vivo and host protection against ongoing infection. PMID:22493517

Gene expression analysis of whole blood RNA from pigs infected with low and high pathogenic African swine fever viruses

DOE PAGES

Jaing, Crystal; Rowland, Raymond R. R.; Allen, Jonathan E.; ...

2017-08-31

African swine fever virus (ASFV) is a macrophage-tropic virus responsible for ASF, a transboundary disease that threatens swine production world-wide. Since there are no vaccines available to control ASF after an outbreak, obtaining an understanding of the virus-host interaction is important for developing new intervention strategies. In this study, a whole transcriptomic RNA-Seq method was used to characterize differentially expressed genes in pigs infected with a low pathogenic ASFV isolate, OUR T88/3 (OURT), or the highly pathogenic Georgia 2007/1 (GRG). After infection, pigs infected with OURT showed no or few clinical signs; whereas, GRG produced clinical signs consistent with acutemore » ASF. RNA-Seq detected the expression of ASFV genes from the whole blood of the GRG, but not the OURT pigs, consistent with the pathotypes of these strains and the replication of GRG in circulating monocytes. Even though GRG and OURT possess different pathogenic properties, there was significant overlap in the most upregulated host genes. A small number of differentially expressed microRNAs were also detected in GRG and OURT pigs. These data confirm previous studies describing the response of macrophages and lymphocytes to ASFV infection, as well as reveal unique gene pathways upregulated in response to infection with GRG.« less

Identification of a disease complex involving a novel monopartite begomovirus with beta- and alphasatellites associated with okra leaf curl disease in Oman.

PubMed

Akhtar, Sohail; Khan, Akhtar J; Singh, Achuit S; Briddon, Rob W

2014-05-01

Okra leaf curl disease (OLCD) is an important viral disease of okra in tropical and subtropical areas. The disease is caused by begomovirus-satellite complexes. A begomovirus and associated betasatellite and alphasatellite were identified in symptomatic okra plants from Barka, in the Al-Batinah region of Oman. Analysis of the begomovirus sequences showed them to represent a new begomovirus most closely related to cotton leaf curl Gezira virus (CLCuGeV), a begomovirus of African origin. The sequences showed less than 85 % nucleotide sequence identity to CLCuGeV isolates. The name okra leaf curl Oman virus (OLCOMV) is proposed for the new virus. Further analysis revealed that the OLCOMV is a recombinant begomovirus that evolved by the recombination of CLCuGeV isolates with tomato yellow leaf curl virus-Oman (TYLCV-OM). An alpha- and a betasatellite were also identified from the same plant sample, which were also unique when compared to sequences available in the databases. However, although the betasatellite appeared to be of African origin, the alphasatellite was most closely related to alphasatellites originating from South Asia. This is the first report of a begomovirus-satellite complex infecting okra in Oman.

Gene expression analysis of whole blood RNA from pigs infected with low and high pathogenic African swine fever viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaing, Crystal; Rowland, Raymond R. R.; Allen, Jonathan E.

African swine fever virus (ASFV) is a macrophage-tropic virus responsible for ASF, a transboundary disease that threatens swine production world-wide. Since there are no vaccines available to control ASF after an outbreak, obtaining an understanding of the virus-host interaction is important for developing new intervention strategies. In this study, a whole transcriptomic RNA-Seq method was used to characterize differentially expressed genes in pigs infected with a low pathogenic ASFV isolate, OUR T88/3 (OURT), or the highly pathogenic Georgia 2007/1 (GRG). After infection, pigs infected with OURT showed no or few clinical signs; whereas, GRG produced clinical signs consistent with acutemore » ASF. RNA-Seq detected the expression of ASFV genes from the whole blood of the GRG, but not the OURT pigs, consistent with the pathotypes of these strains and the replication of GRG in circulating monocytes. Even though GRG and OURT possess different pathogenic properties, there was significant overlap in the most upregulated host genes. A small number of differentially expressed microRNAs were also detected in GRG and OURT pigs. These data confirm previous studies describing the response of macrophages and lymphocytes to ASFV infection, as well as reveal unique gene pathways upregulated in response to infection with GRG.« less

Comparative study of enteric viruses, coliphages and indicator bacteria for evaluating water quality in a tropical high-altitude system.

PubMed

Espinosa, Ana C; Arias, Carlos F; Sánchez-Colón, Salvador; Mazari-Hiriart, Marisa

2009-10-27

Bacteria used as indicators for pathogenic microorganisms in water are not considered adequate as enteric virus indicators. Surface water from a tropical high-altitude system located in Mexico City that receives rainwater, treated and non-treated wastewater used for irrigation, and groundwater used for drinking, was studied. The presence of enterovirus, rotavirus, astrovirus, coliphage, coliform bacteria, and enterococci was determined during annual cycles in 2001 and 2002. Enteric viruses in concentrated water samples were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). Coliphages were detected using the double agar layer method. Bacteria analyses of the water samples were carried out by membrane filtration. The presence of viruses and bacteria in the water used for irrigation showed no relationship between current bacterial indicator detection and viral presence. Coliphages showed strong association with indicator bacteria and enterovirus, but weak association with other enteric viruses. Enterovirus and rotavirus showed significant seasonal differences in water used for irrigation, although this was not clear for astrovirus. Coliphages proved to be adequate faecal pollution indicators for the irrigation water studied. Viral presence in this tropical high-altitude system showed a similar trend to data previously reported for temperate zones.

The innate immune response to lower respiratory tract E. Coli infection and the role of the CCL2-CCR2 axis in neonatal mice.

PubMed

McGrath-Morrow, Sharon A; Ndeh, Roland; Collaco, Joseph M; Poupore, Amy K; Dikeman, Dustin; Zhong, Qiong; Singer, Benjamin D; D'Alessio, Franco; Scott, Alan

2017-09-01

Neonates have greater morbidity/mortality from lower respiratory tract infections (LRTI) compared to older children. Lack of conditioning of the pulmonary immune system due to limited environmental exposures and/or infectious challenges likely contributes to the increase susceptibility in the neonate. In this study, we sought to gain insights into the nature and dynamics of the neonatal pulmonary immune response to LRTI using a murine model. Wildtype (WT) and Ccr2 -/- C57BL/6 neonatal and juvenile mice received E. coli or PBS by direct pharyngeal aspiration. Flow cytometry was used to measure immune cell dynamics and identify cytokine-producing cells. Real-time PCR and ELISA were used to measure cytokine/chemokine expression. Innate immune cell recruitment in response to E. coli-induced LRTI was delayed in the neonatal lung compared to juvenile lung. Lung clearance of bacteria was also significantly delayed in the neonate. Ccr2 -/- neonates, which lack an intact CCL2-CCR2 axis, had higher mortality after E. coli challenged than Ccr2 +/+ neonates. A greater percentage of CD8 + T cells and monocytes from WT neonates challenged with E. coli produced TNF compared to controls. The pulmonary immune response to E. coli-induced LRTI differed significantly between neonatal and juvenile mice. Neonates were more susceptible to increasing doses of E. coli and exhibited greater mortality than juveniles. In the absence of an intact CCL2-CCR2 axis, susceptibility to LRTI-induced mortality was further increased in neonatal mice. Taken together these findings underscore the importance of age-related differences in the innate immune response to LRTI during early stages of postnatal life. Copyright © 2017 Elsevier Ltd. All rights reserved.

Simulation of high frequency nitrous oxide emissions from irrigated sub-tropical soils using DAYCENT

USDA-ARS?s Scientific Manuscript database

A unique high temporal frequency dataset from an irrigated cotton-wheat rotation was used to test the agroecosystem model DayCent to simulate daily N2O emissions from sub-tropical vertisols under different irrigation intensities. DayCent was able to simulate the effect of different irrigation intens...

Structural and Biochemical Characterization of Cinnamoyl-CoA Reductases1

PubMed Central

Walker, Alexander M.

2017-01-01

Cinnamoyl-coenzyme A reductase (CCR) catalyzes the reduction of hydroxycinnamoyl-coenzyme A (CoA) esters using NADPH to produce hydroxycinnamyl aldehyde precursors in lignin synthesis. The catalytic mechanism and substrate specificity of cinnamoyl-CoA reductases from sorghum (Sorghum bicolor), a strategic plant for bioenergy production, were deduced from crystal structures, site-directed mutagenesis, and kinetic and thermodynamic analyses. Although SbCCR1 displayed higher affinity for caffeoyl-CoA or p-coumaroyl-CoA than for feruloyl-CoA, the enzyme showed significantly higher activity for the latter substrate. Through molecular docking and comparisons between the crystal structures of the Vitis vinifera dihydroflavonol reductase and SbCCR1, residues threonine-154 and tyrosine-310 were pinpointed as being involved in binding CoA-conjugated phenylpropanoids. Threonine-154 of SbCCR1 and other CCRs likely confers strong substrate specificity for feruloyl-CoA over other cinnamoyl-CoA thioesters, and the T154Y mutation in SbCCR1 led to broader substrate specificity and faster turnover. Through data mining using our structural and biochemical information, four additional putative CCR genes were discovered from sorghum genomic data. One of these, SbCCR2, displayed greater activity toward p-coumaroyl-CoA than did SbCCR1, which could imply a role in the synthesis of defense-related lignin. Taken together, these findings provide knowledge about critical residues and substrate preference among CCRs and provide, to our knowledge, the first three-dimensional structure information for a CCR from a monocot species. PMID:27956488

Optimized dosing of a CCR2 antagonist for amplification of vaccine immunity.

PubMed

Mitchell, Leah A; Hansen, Ryan J; Beaupre, Adam J; Gustafson, Daniel L; Dow, Steven W

2013-02-01

We have recently discovered that inflammatory monocytes recruited to lymph nodes in response to vaccine-induced inflammation can function as potent negative regulators of both humoral and cell-mediated immune responses to vaccination. Monocyte depletion or migration blockade can significantly amplify both antibody titers and cellular immune responses to vaccination with several different antigens in mouse models. Thus, we hypothesized that the use of small molecule CCR2 inhibitors to block monocyte migration into lymph nodes may represent a broadly effective means of amplifying vaccine immunity. To address this question, the role of CCR2 in monocyte recruitment to vaccine draining lymph nodes was initially explored in CCR2-/- mice. Next, a small molecule antagonist of CCR2 (RS102895) was evaluated in mouse vaccination models. Initial studies revealed that a single intraperitoneal dose of RS102895 failed to effectively block monocyte recruitment following vaccination. Pharmacokinetic analysis of RS102895 revealed a short half-life (approximately 1h), and suggested that a multi-dose treatment regimen would be more effective. We found that administration of RS102895 every 6 h resulted in consistent plasma levels of 20 ng/ml or greater, which effectively blocked monocyte migration to lymph nodes following vaccination. Moreover, administration of RS102895 with concurrent vaccination markedly enhanced vaccine responses following immunization against the influenza antigen HA1. We concluded that administration of small molecule CCR2 antagonists such as RS102895 in the immediate post-vaccine period could be used as a novel means of significantly enhancing vaccine immunity. Copyright © 2012 Elsevier B.V. All rights reserved.

Multicohort Genomewide Association Study Reveals a New Signal of Protection Against HIV-1 Acquisition

PubMed Central

Limou, Sophie; Delaneau, Olivier; van Manen, Daniëlle; An, Ping; Sezgin, Efe; Le Clerc, Sigrid; Coulonges, Cédric; Troyer, Jennifer L.; Veldink, Jan H.; van den Berg, Leonard H.; Spadoni, Jean-Louis; Taing, Lieng; Labib, Taoufik; Montes, Matthieu; Delfraissy, Jean-François; Schachter, François; O’Brien, Stephen J.; Buchbinder, Susan; van Natta, Mark L.; Jabs, Douglas A.; Froguel, Philippe; Schuitemaker, Hanneke; Winkler, Cheryl A.

2012-01-01

Background. To date, only mutations in CCR5 have been shown to confer resistance to human immunodeficiency virus type 1 (HIV-1) infection, and these explain only a small fraction of the observed variability in HIV susceptibility. Methods. We performed a meta-analysis between 2 independent European genomewide association studies, each comparing HIV-1 seropositive cases with normal population controls known to be HIV uninfected, to identify single-nucleotide polymorphisms (SNPs) associated with the HIV-1 acquisition phenotype. SNPs exhibiting P < 10−5 in this first stage underwent second-stage analysis in 2 independent US cohorts of European descent. Results. After the first stage, a single highly significant association was revealed for the chromosome 8 rs6996198 with HIV-1 acquisition and was replicated in both second-stage cohorts. Across the 4 groups, the rs6996198-T allele was consistently associated with a significant reduced risk of HIV-1 infection, and the global meta-analysis reached genomewide significance: Pcombined = 7.76 × 10−8. Conclusions. We provide strong evidence of association for a common variant with HIV-1 acquisition in populations of European ancestry. This protective signal against HIV-1 infection is the first identified outside the CCR5 nexus. First clues point to a potential functional role for a nearby candidate gene, CYP7B1, but this locus warrants further investigation. PMID:22362864

Finding a Vulnerable Spot in HIV’s Armor by Investigating the Structure of HIV | Center for Cancer Research

Cancer.gov

The Human Immunodeficiency Virus (HIV) infects and eventually kills CD4-expressing T cells, which are essential for the immune system to function appropriately. Loss of significant numbers of T cells leads to Acquired Immunodeficiency Syndrome (AIDS), a disease that kills over two million people around the world every year. HIV infection depends on two proteins expressed on the virus surface: gp41, which sits in the virus membrane, and gp120, which sits on top of gp41. Three copies, or trimers, of each gp41/gp120 pair make up the envelope glycoprotein, Env. Env coats the virus surface and interacts with its receptor, CD4, and a co-receptor, either CCR5 or CXCR4, on the T cell. Binding to the receptors is thought to cause a structural reorganization of Env, which exposes a fusion peptide that inserts into the T cell membrane and actually forces the virus and host membranes together, initiating an infection. However, the structural details of this process are lacking.

Ebola Virus Persistence in Semen Ex Vivo.

PubMed

Fischer, Robert J; Judson, Seth; Miazgowicz, Kerri; Bushmaker, Trent; Munster, Vincent J

2016-02-01

On March 20, 2015, a case of Ebola virus disease was identified in Liberia that most likely was transmitted through sexual contact. We assessed the efficiency of detecting Ebola virus in semen samples by molecular diagnostics and the stability of Ebola virus in ex vivo semen under simulated tropical conditions.

A Critical Role for Monocytes/Macrophages During Intestinal Inflammation-associated Lymphangiogenesis

PubMed Central

Becker, Felix; Kurmaeva, Elvira; Gavins, Felicity N. E.; Stevenson, Emily V.; Navratil, Aaron R.; Jin, Long; Tsunoda, Ikuo; Orr, A. Wayne; Alexander, Jonathan S.; Ostanin, Dmitry V.

2016-01-01

Background Inflammation-associated lymphangiogenesis (IAL) is frequently observed in inflammatory bowel diseases. IAL is believed to limit inflammation by enhancing fluid and immune cell clearance. Although monocytes/macrophages (MΦ) are known to contribute to intestinal pathology in inflammatory bowel disease, their role in intestinal IAL has never been studied mechanistically. We investigated contributions of monocytes/MΦ to the development of intestinal inflammation and IAL. Methods Because inflammatory monocytes express CC chemokine receptor 2 (CCR2), we used CCR2 diphtheria toxin receptor transgenic (CCR2.DTR) mice, in which monocytes can be depleted by diphtheria toxin injection, and CCR2−/− mice, which have reduced circulating monocytes. Acute or chronic colitis was induced by dextran sodium sulfate or adoptive transfer of CD4+CD45RBhigh T cells, respectively. Intestinal inflammation was assessed by flow cytometry, immunofluorescence, disease activity, and histopathology, whereas IAL was assessed by lymphatic vessel morphology and density. Results We demonstrated that intestinal MΦ expressed vascular endothelial growth factor-C/D. In acute colitis, monocyte-depleted mice were protected from intestinal injury and showed reduced IAL, which was reversed after transfer of wild-type monocytes into CCR2−/− mice. In chronic colitis, CCR2 deficiency did not attenuate inflammation but reduced IAL. Conclusions We propose a dual role of MΦ in (1) promoting acute inflammation and (2) contributing to IAL. Our data suggest that intestinal inflammation and IAL could occur independently, because IAL was reduced in the absence of monocytes/MΦ, even when inflammation was present. Future inflammatory bowel disease therapies might exploit promotion of IAL and suppression of MΦ independently, to restore lymphatic clearance and reduce inflammation. PMID:26950310

CCR6+ Th cell distribution differentiates systemic lupus erythematosus patients based on anti-dsDNA antibody status.

PubMed

Zhong, Wei; Jiang, Zhenyu; Wu, Jiang; Jiang, Yanfang; Zhao, Ling

2018-01-01

Systemic lupus erythematosus (SLE) disease has been shown to be associated with the generation of multiple auto-antibodies. Among these, anti-dsDNA antibodies (anti-DNAs) are specific and play a pathogenic role in SLE. Indeed, anti-DNA + SLE patients display a worse disease course. The generation of these pathogenic anti-DNAs has been attributed to the interaction between aberrant T helper (Th) cells and autoimmune B cells. Thus, in this study we have investigated whether CCR6 + Th cells have the ability to differentiate SLE patients based on anti-DNA status, and if their distribution has any correlation with disease activity. We recruited 25 anti-DNA + and 25 anti-DNA - treatment-naive onset SLE patients, matched for various clinical characteristics in our nested matched case-control study. CCR6 + Th cells and their additional subsets were analyzed in each patient by flow cytometry. Anti-DNA + SLE patients specifically had a higher percentage of Th cells expressing CCR6 and CXCR3. Further analysis of CCR6 + Th cell subsets showed that anti-DNA + SLE patients had elevated proportions of Th9, Th17, Th17.1 and CCR4/CXCR3 double-negative (DN) cells. However, the proportions of CCR6 - Th subsets, including Th1 and Th2 cells, did not show any association with anti-DNA status. Finally, we identified a correlation between CCR6 + Th subsets and clinical indicators, specifically in anti-DNA + SLE patients. Our data indicated that CCR6 + Th cells and their subsets were elevated and correlated with disease activity in anti-DNA + SLE patients. We speculated that CCR6 + Th cells may contribute to distinct disease severity in anti-DNA + SLE patients.

The simultaneous repression of CCR and CAD, two enzymes of the lignin biosynthetic pathway, results in sterility and dwarfism in Arabidopsis thaliana.

PubMed

Thévenin, Johanne; Pollet, Brigitte; Letarnec, Bruno; Saulnier, Luc; Gissot, Lionel; Maia-Grondard, Alessandra; Lapierre, Catherine; Jouanin, Lise

2011-01-01

Cinnamoyl CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) catalyze the last steps of monolignol biosynthesis. In Arabidopsis, one CCR gene (CCR1, At1g15950) and two CAD genes (CAD C At3g19450 and CAD D At4g34230) are involved in this pathway. A triple cad c cad d ccr1 mutant, named ccc, was obtained. This mutant displays a severe dwarf phenotype and male sterility. The lignin content in ccc mature stems is reduced to 50% of the wild-type level. In addition, stem lignin structure is severely affected, as shown by the dramatic enrichment in resistant inter-unit bonds and incorporation into the polymer of monolignol precursors such as coniferaldehyde, sinapaldehyde, and ferulic acid. Male sterility is due to the lack of lignification in the anther endothecium, which causes the failure of anther dehiscence and of pollen release. The ccc hypolignified stems accumulate higher amounts of flavonol glycosides, sinapoyl malate and feruloyl malate, which suggests a redirection of the phenolic pathway. Therefore, the absence of CAD and CCR, key enzymes of the monolignol pathway, has more severe consequences on the phenotype than the individual absence of each of them. Induction of another CCR (CCR2, At1g80820) and another CAD (CAD1, At4g39330) does not compensate the absence of the main CCR and CAD activities. This lack of CCR and CAD activities not only impacts lignification, but also severely affects the development of the plants. These consequences must be carefully considered when trying to reduce the lignin content of plants in order to facilitate the lignocellulose-to-bioethanol conversion process.

CCR8 Signaling Influences Toll-Like Receptor 4 Responses in Human Macrophages in Inflammatory Diseases ▿

PubMed Central

Kvist Reimer, Martina; Brange, Charlotte; Rosendahl, Alexander

2011-01-01

CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients. PMID:21976223

CCR8 signaling influences Toll-like receptor 4 responses in human macrophages in inflammatory diseases.

PubMed

Reimer, Martina Kvist; Brange, Charlotte; Rosendahl, Alexander

2011-12-01

CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients.

Modeled Forecasts of Dengue Fever in San Juan, PR Using NASA Satellite Enhanced Weather Forecasts

NASA Technical Reports Server (NTRS)

Morin, Cory; Quattrochi, Dale; Zavodsky, Bradley; Case, Jonathan

2015-01-01

Dengue virus is transmitted between humans and mosquitoes of the genus Aedes and causes approximately 96 million cases of disease (dengue fever) each year (Bhatet al. 2013). Symptoms of dengue fever include fever, headache, nausea, vomiting, and eye, muscle and joint pain (CDC). More sever manifestations such as abdominal pain, bleeding from nose and gums, vomiting of blood, and clammy skin occur in rare cases of dengue hemorrhagic fever (CDC). Dengue fever occurs throughout tropical and sub-tropical regions worldwide, however, the geographical range and size of epidemics is increasing. Weather and climate are drivers of dengue virus transmission dynamics (Morin et al. 2013) by affecting mosquito proliferation and the virus extrinsic incubation period (i.e. required time for the virus to replicate and disseminate within the mosquito before it can retransmit the virus).

Use of cardiocerebral resuscitation or AHA/ERC 2005 Guidelines is associated with improved survival from out-of-hospital cardiac arrest: a systematic review and meta-analysis.

PubMed

Salmen, Marcus; Ewy, Gordon A; Sasson, Comilla

2012-01-01

To determine whether the use of cardiocerebral resuscitation (CCR) or AHA/ERC 2005 Resuscitation Guidelines improved patient outcomes from out-of-hospital cardiac arrest (OHCA) compared to older guidelines. Systematic review and meta-analysis. MEDLINE, EMBASE, Web of Science and the Cochrane Library databases. We also hand-searched study references and consulted experts. Design: randomised controlled trials and observational studies. OHCA patients, age >17 years. 'Control' protocol versus 'Study' protocol. 'Control' protocol defined as AHA/ERC 2000 Guidelines for cardiopulmonary resuscitation (CPR). 'Study' protocol defined as AHA/ERC 2005 Guidelines for CPR, or a CCR protocol. Survival to hospital discharge. High-quality or medium-quality studies, as measured by the Newcastle Ottawa Scale using predefined categories. Twelve observational studies met inclusion criteria. All the three studies using CCR demonstrated significantly improved survival compared to use of AHA 2000 Guidelines, as did five of the nine studies using AHA/ERC 2005 Guidelines. Pooled data demonstrate that use of a CCR protocol has an unadjusted OR of 2.26 (95% CI 1.64 to 3.12) for survival to hospital discharge among all cardiac arrest patients. Among witnessed ventricular fibrillation/ventricular tachycardia (VF/VT) patients, CCR increased survival by an OR of 2.98 (95% CI 1.92 to 4.62). Studies using AHA/ERC 2005 Guidelines showed an overall trend towards increased survival, but significant heterogeneity existed among these studies. We demonstrate an association with improved survival from OHCA when CCR protocols or AHA/ERC 2005 Guidelines are compared to use of older guidelines. In the subgroup of patients with witnessed VF/VT, there was a threefold increase in OHCA survival when CCR was used. CCR appears to be a promising resuscitation protocol for Emergency Medical Services providers in increasing survival from OHCA. Future research will need to be conducted to directly compare AHA/ERC 2010 Guidelines with the CCR approach.

Role of human pulmonary fibroblast-derived MCP-1 in cell activation and migration in experimental silicosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Xueting; Fang, Shencun; Liu, Haijun

Background: Silicosis is a systemic disease caused by inhaling silicon dioxide (SiO{sub 2}). Phagocytosis of SiO{sub 2} in the lung initiates an inflammatory cascade that results in fibroblast proliferation and migration and subsequent fibrosis. Clinical evidence indicates that the activation of alveolar macrophages by SiO{sub 2} produces rapid and sustained inflammation that is characterized by the generation of monocyte chemotactic protein 1 (MCP-1), which induces fibrosis. Pulmonary fibroblast-derived MCP-1 may play a critical role in fibroblast proliferation and migration. Methods and results: Experiments using primary cultured adult human pulmonary fibroblasts (HPF-a) demonstrated the following results: 1) SiO{sub 2} treatment resultedmore » in the rapid and sustained induction of MCP-1 as well as the elevation of the CC chemokine receptor type 2 (CCR2) protein levels; 2) pretreatment of HPF-a with RS-102895, a specific CCR2 inhibitor, abolished the SiO{sub 2}-induced increase in cell activation and migration in both 2D and 3D culture systems; and 3) RNA interference targeting CCR2 prevented the SiO{sub 2}-induced increase in cell migration. Conclusion: These data demonstrated that the up-regulation of pulmonary fibroblast-derived MCP-1 is involved in pulmonary fibroblast migration induced by SiO{sub 2}. CCR2 was also up-regulated in response to SiO{sub 2}, and this up-regulation facilitated the effect of MCP-1 on fibroblasts. Our study deciphered the link between fibroblast-derived MCP-1 and SiO{sub 2}-induced cell migration. This finding provides novel insight into the potential of MCP-1 in the development of novel therapeutic strategies for silicosis. - Highlights: • Role of pulmonary fibroblast-derived MCP-1 in experimental silicosis was studied. • SiO{sub 2} induced MCP-1 release from cultured human pulmonary fibroblast (HPF-a). • SiO{sub 2} directly activated HPF-a via the MCP-1/CCR2 pathway. • SiO{sub 2} increased HPF-a migration in both 2D and 3D model via the MCP-1/CCR2 pathway. • RNA-i of MCP-1/CCR2 decreased HPF-a activation and migration induced by SiO{sub 2}.« less

Vaccination against mosquito borne viral infections: current status.

PubMed

Wiwanitkit, Viroj

2007-12-01

Mosquito borne infectious diseases are among important group of diseases worldwide. Vaccination is available for some tropical mosquito-borne diseases, especially for Japanese encephalitis virus infection and yellow fever. There are also several attempts to develop new vaccines for the other mosquito-borne diseases such as malaria, dengue infection and West Nile virus infection. In this article, the author reviews the issues on vaccination of some important tropical mosquito borne infectious diseases.

Prenatal exposure to ethanol stimulates hypothalamic CCR2 chemokine receptor system: Possible relation to increased density of orexigenic peptide neurons and ethanol drinking in adolescent offspring

PubMed Central

Chang, G.-Q.; Karatayev, O.; Leibowitz, S. F.

2015-01-01

Clinical and animal studies indicate that maternal consumption of ethanol during pregnancy increases alcohol drinking in the offspring. Possible underlying mechanisms may involve orexigenic peptides, which are stimulated by prenatal ethanol exposure and themselves promote drinking. Building on evidence that ethanol stimulates neuroimmune factors such as the chemokine CCL2 that in adult rats is shown to colocalize with the orexigenic peptide, melanin-concentrating hormone (MCH) in the lateral hypothalamus (LH), the present study sought to investigate the possibility that CCL2 or its receptor CCR2 in LH are stimulated by prenatal ethanol exposure, perhaps specifically within MCH neurons. Our paradigm of intraoral administration of ethanol to pregnant rats, at low-to-moderate doses (1 or 3 g/kg/day) during peak hypothalamic neurogenesis, caused in adolescent male offspring two-fold increase in drinking of and preference for ethanol and reinstatement of ethanol drinking in a two-bottle choice paradigm under an intermittent access schedule. This effect of prenatal ethanol exposure was associated with an increased expression of MCH and density of MCH+ neurons in LH of preadolescent offspring. Whereas CCL2+ cells at this age were low in density and unaffected by ethanol, CCR2+ cells were dense in LH and increased by prenatal ethanol, with a large percentage (83–87%) identified as neurons and found to colocalize MCH. Prenatal ethanol also stimulated the genesis of CCR2+ and MCH+ neurons in the embryo, which co-labeled the proliferation marker, BrdU. Ethanol also increased the genesis and density of neurons that co-expressed CCR2 and MCH in LH, with triple-labeled CCR2+/MCH+/BrdU+ neurons that were absent in control rats accounting for 35% of newly generated neurons in ethanol-exposed rats. With both the chemokine and MCH systems believed to promote ethanol consumption, this greater density of CCR2+/MCH+ neurons in the LH of preadolescent rats suggests that these systems function together in promoting alcohol drinking during adolescence. PMID:26365610

Comprehensive cardiac rehabilitation for secondary prevention after transient ischemic attack or mild stroke: I: feasibility and risk factors.

PubMed

Prior, Peter L; Hachinski, Vladimir; Unsworth, Karen; Chan, Richard; Mytka, Sharon; O'Callaghan, Christina; Suskin, Neville

2011-11-01

Comprehensive cardiac rehabilitation (CCR), which integrates structured lifestyle interventions and medications, reduces morbidity and mortality among cardiac patients. CCR has not typically been used with cerebrovascular populations, despite important commonalities with heart patients. We tested feasibility and effectiveness of 6-month outpatient CCR for secondary prevention after transient ischemic attack or mild, nondisabling stroke. This article presents risk factors. A future article will discuss psychological outcomes. Consecutive consenting subjects having sustained a transient ischemic attack or mild, nondisabling stroke within the previous 12 months (mean, 11.5 weeks; event-to-CCR entry) with ≥1 vascular risk factor, were recruited from a stroke prevention clinic providing usual care. We measured 6-month CCR outcomes following a prospective cohort design. Of 110 subjects recruited from January 2005 to April 2006, 100 subjects (mean age, 64.9 years; 46 women) entered and 80 subjects completed CCR. We obtained favorable, significant intake-to-exit changes in: aerobic capacity (+31.4%; P<0.001), total cholesterol (-0.30 mmol/L; P=0.008), total cholesterol/high-density lipoprotein (-11.6%; P<0.001), triglycerides (-0.27 mmol/L; P=0.003), waist circumference (-2.44 cm; P<0.001), body mass index (-0.53 kg/m(2); P=0.003), and body weight (-1.43 kg; P=0.001). Low-density lipoprotein (-0.24 mmol/L), high-density lipoprotein (+0.06 mmol/L), systolic (-3.21 mm Hg) and diastolic (-2.34 mm Hg) blood pressure changed favorably, but nonsignificantly. A significant shift toward nonsmoking occurred (P=0.008). Compared with intake, 11 more individuals (25.6% increase) finished CCR in the lowest-mortality risk category of the Duke Treadmill Score (P<0.001). CCR is feasible and effective for secondary prevention after transient ischemic attack or mild, nondisabling stroke, offering a promising model for vascular protection across chronic disease entities. We know of no similar previous investigation, and are now conducting a randomized trial.

Ross River virus and Barmah Forest virus infections: a review of history, ecology, and predictive models, with implications for tropical northern Australia.

PubMed

Jacups, Susan P; Whelan, Peter I; Currie, Bart J

2008-04-01

The purpose of the present article is to present a review of the Ross River virus (RRV) and Barmah Forest virus (BFV) literature in relation to potential implications for future disease in tropical northern Australia. Ross River virus infection is the most common and most widespread arboviral disease in Australia, with an average of 4,800 national notifications annually. Of recent concern is the sudden rise in BFV infections; the 2005-2006 summer marked the largest BFV epidemic on record in Australia, with 1,895 notifications. Although not life-threatening, infection with either virus can cause arthritis, myalgia, and fatigue for 6 months or longer, resulting in substantial morbidity and economic impact. The geographic distribution of mosquito species and their seasonal activity is determined in large part by temperature and rainfall. Predictive models can be useful tools in providing early warning systems for epidemics of RRV and BFV infection. Various models have been developed to predict RRV outbreaks, but these appear to be mostly only regionally valid, being dependent on local ecological factors. Difficulties have arisen in developing useful models for the tropical northern parts of Australia, and to date no models have been developed for the Northern Territory. Only one model has been developed for predicting BFV infections using climate and tide variables. It is predicted that the exacerbation of current greenhouse conditions will result in longer periods of high mosquito activity in the tropical regions where RRV and BFV are already common. In addition, the endemic locations may expand further within temperate regions, and epidemics may become more frequent in those areas. Further development of predictive models should benefit public health planning by providing early warning systems of RRV and BFV infection outbreaks in different geographical locations.

Polymorphisms of large effect explain the majority of the host genetic contribution to variation of HIV-1 virus load

PubMed Central

Coulonges, Cedric; Bartha, István; Lenz, Tobias L.; Deutsch, Aaron J.; Bashirova, Arman; Buchbinder, Susan; Carrington, Mary N.; Cossarizza, Andrea; Dalmau, Judith; De Luca, Andrea; Goedert, James J.; Gurdasani, Deepti; Haas, David W.; Herbeck, Joshua T.; Johnson, Eric O.; Kirk, Gregory D.; Lambotte, Olivier; Luo, Ma; Mallal, Simon; van Manen, Daniëlle; Martinez-Picado, Javier; Meyer, Laurence; Miro, José M.; Mullins, James I.; Obel, Niels; Poli, Guido; Sandhu, Manjinder S.; Schuitemaker, Hanneke; Shea, Patrick R.; Theodorou, Ioannis; Walker, Bruce D.; Weintrob, Amy C.; Winkler, Cheryl A.; Wolinsky, Steven M.; Raychaudhuri, Soumya; Goldstein, David B.; Telenti, Amalio; de Bakker, Paul I. W.; Zagury, Jean-François; Fellay, Jacques

2015-01-01

Previous genome-wide association studies (GWAS) of HIV-1–infected populations have been underpowered to detect common variants with moderate impact on disease outcome and have not assessed the phenotypic variance explained by genome-wide additive effects. By combining the majority of available genome-wide genotyping data in HIV-infected populations, we tested for association between ∼8 million variants and viral load (HIV RNA copies per milliliter of plasma) in 6,315 individuals of European ancestry. The strongest signal of association was observed in the HLA class I region that was fully explained by independent effects mapping to five variable amino acid positions in the peptide binding grooves of the HLA-B and HLA-A proteins. We observed a second genome-wide significant association signal in the chemokine (C-C motif) receptor (CCR) gene cluster on chromosome 3. Conditional analysis showed that this signal could not be fully attributed to the known protective CCR5Δ32 allele and the risk P1 haplotype, suggesting further causal variants in this region. Heritability analysis demonstrated that common human genetic variation—mostly in the HLA and CCR5 regions—explains 25% of the variability in viral load. This study suggests that analyses in non-European populations and of variant classes not assessed by GWAS should be priorities for the field going forward. PMID:26553974

The distribution of HIV DNA and RNA in cell subsets differs in gut and blood of HIV-positive patients on ART: implications for viral persistence.

PubMed

Yukl, Steven A; Shergill, Amandeep K; Ho, Terence; Killian, Maudi; Girling, Valerie; Epling, Lorrie; Li, Peilin; Wong, Lisa K; Crouch, Pierre; Deeks, Steven G; Havlir, Diane V; McQuaid, Kenneth; Sinclair, Elizabeth; Wong, Joseph K

2013-10-15

Even with optimal antiretroviral therapy, human immunodeficiency virus (HIV) persists in plasma, blood cells, and tissues. To develop new therapies, it is essential to know what cell types harbor residual HIV. We measured levels of HIV DNA, RNA, and RNA/DNA ratios in sorted subsets of CD4+ T cells (CCR7+, transitional memory, and effector memory) and non-CD4+ T leukocytes from blood, ileum, and rectum of 8 ART-suppressed HIV-positive subjects. Levels of HIV DNA/million cells in CCR7+ and effector memory cells were higher in the ileum than blood. When normalized by cell frequencies, most HIV DNA and RNA in the blood were found in CCR7+ cells, whereas in both gut sites, most HIV DNA and RNA were found in effector memory cells. HIV DNA and RNA were observed in non-CD4+ T leukocytes at low levels, particularly in gut tissues. Compared to the blood, the ileum had higher levels of HIV DNA and RNA in both CD4+ T cells and non-CD4+ T leukocytes, whereas the rectum had higher HIV DNA levels in both cell types but lower RNA levels in CD4+ T cells. Future studies should determine whether different mechanisms allow HIV to persist in these distinct reservoirs, and the degree to which different therapies can affect each reservoir.

Bats Worldwide Carry Hepatitis E Virus-Related Viruses That Form a Putative Novel Genus within the Family Hepeviridae

PubMed Central

Drexler, Jan Felix; Seelen, Annika; Corman, Victor Max; Fumie Tateno, Adriana; Cottontail, Veronika; Melim Zerbinati, Rodrigo; Gloza-Rausch, Florian; Klose, Stefan M.; Adu-Sarkodie, Yaw; Oppong, Samuel K.; Kalko, Elisabeth K. V.; Osterman, Andreas; Rasche, Andrea; Adam, Alexander; Müller, Marcel A.; Ulrich, Rainer G.; Leroy, Eric M.; Lukashev, Alexander N.

2012-01-01

Hepatitis E virus (HEV) is one of the most common causes of acute hepatitis in tropical and temperate climates. Tropical genotypes 1 and 2 are associated with food-borne and waterborne transmission. Zoonotic reservoirs (mainly pigs, wild boar, and deer) are considered for genotypes 3 and 4, which exist in temperate climates. In view of the association of several zoonotic viruses with bats, we analyzed 3,869 bat specimens from 85 different species and from five continents for hepevirus RNA. HEVs were detected in African, Central American, and European bats, forming a novel phylogenetic clade in the family Hepeviridae. Bat hepeviruses were highly diversified and comparable to human HEV in sequence variation. No evidence for the transmission of bat hepeviruses to humans was found in over 90,000 human blood donations and individual patient sera. Full-genome analysis of one representative virus confirmed formal classification within the family Hepeviridae. Sequence- and distance-based taxonomic evaluations suggested that bat hepeviruses constitute a distinct genus within the family Hepeviridae and that at least three other genera comprising human, rodent, and avian hepeviruses can be designated. This may imply that hepeviruses invaded mammalian hosts nonrecently and underwent speciation according to their host restrictions. Human HEV-related viruses in farmed and peridomestic animals might represent secondary acquisitions of human viruses, rather than animal precursors causally involved in the evolution of human HEV. PMID:22696648

Salinity Drives the Virioplankton Abundance but Not Production in Tropical Coastal Lagoons.

PubMed

Junger, Pedro C; Amado, André M; Paranhos, Rodolfo; Cabral, Anderson S; Jacques, Saulo M S; Farjalla, Vinicius F

2018-01-01

Viruses are the most abundant components of microbial food webs and play important ecological and biogeochemical roles in aquatic ecosystems. Virioplankton is regulated by several environmental factors, such as salinity, turbidity, and humic substances. However, most of the studies aimed to investigate virioplankton regulation were conducted in temperate systems combining a limited range of environmental variables. In this study, virus abundance and production were determined and their relation to bacterial and limnological variables was assessed in 20 neighboring shallow tropical coastal lagoons that present wide environmental gradients of turbidity (2.32-571 NTU), water color (1.82-92.49 m -1 ), dissolved organic carbon (0.71-16.7 mM), salinity (0.13-332.1‰), and chlorophyll-a (0.28 to 134.5 μg L -1 ). Virus abundance varied from 0.37 × 10 8 to 117 × 10 8 virus-like-particle (VLP) mL -1 , with the highest values observed in highly salty aquatic systems. Salinity and heterotrophic bacterial abundance were the main variables positively driving viral abundances in these lagoons. We suggest that, with increased salinity, there is a decrease in the protozoan control on bacterial populations and lower bacterial diversity (higher encounter rates with virus specific hosts), both factors positively affecting virus abundance. Virus production varied from 0.68 × 10 7 to 56.5 × 10 7 VLP mL -1 h -1 and was regulated by bacterial production and total phosphorus, but it was not directly affected by salinity. The uncoupling between virus abundance and virus production supports that the hypothesis that the lack of grazing pressure on viral and bacterial populations is an important mechanism causing virus abundance to escalate with increasing salt concentrations.

Subsurface Ice Detection via Low Frequency Surface Electromagnetic Method

NASA Astrophysics Data System (ADS)

Stillman, D. E.; Grimm, R. E.; Mcginnis, R. N.

2014-12-01

The geophysical detection of ice in the Cryosphere is typically conducted by measuring the absence of water. These interpretations can become non-unique in dry soils or in clay- and silt-rich soils that contain significant quantities of unfrozen water. Extensive laboratory measurements of electrical properties were made on permafrost samples as a function of frequency, temperature, and water content. These laboratory measurements show that the amount of ice can be uniquely obtained by measuring a frequency dependence of the electrical properties over a large frequency range (20 kHz - 10 Hz). In addition, the electrical properties of permafrost are temperature dependent, which can allow for an estimate of subsurface temperature. In order to test this approach in the field, we performed field surveys at four locations in Alaska. We used three low frequency electromagnetic methods: Spectral Induced Polarization (SIP: 20 kHz - 10 Hz), Capacively Coupled Resistivity (CCR: OhmMapper - 16.5 kHz), and DC Resistivity (Syscal ~ 8 Hz). At the Cold Regions Research and Engineering Laboratory permafrost tunnel near Fox, AK, we used SIP to measure the average ice concentration of 80 v% and determined the temperature to be -3±1°C by matching survey results to lab data. SIP data acquisition is very slow; therefore, at three sites near Tok, AK, we used CCR to perform reconnaissance of the area. Then SIP and DC resistivity were performed at anomalous areas. The three survey types give very similar absolute resistivity values. We found that while SIP gives the most quantitative results, the frequency dependence from the CCR and DC resistivity surveys is all that are needed to determine ice content in permafrost.

Delineating CD4 dependency of HIV-1: Adaptation to infect low level CD4 expressing target cells widens cellular tropism but severely impacts on envelope functionality

PubMed Central

Beauparlant, David; Rusert, Peter; Magnus, Carsten; Weber, Jacqueline; Uhr, Therese; Clapham, Paul R.; Metzner, Karin J.

2017-01-01

A hallmark of HIV-1 infection is the continuously declining number of the virus’ predominant target cells, activated CD4+ T cells. With diminishing CD4+ T cell levels, the capacity to utilize alternate cell types and receptors, including cells that express low CD4 receptor levels such as macrophages, thus becomes crucial. To explore evolutionary paths that allow HIV-1 to acquire a wider host cell range by infecting cells with lower CD4 levels, we dissected the evolution of the envelope-CD4 interaction under in vitro culture conditions that mimicked the decline of CD4high target cells, using a prototypic subtype B, R5-tropic strain. Adaptation to CD4low targets proved to severely alter envelope functions including trimer opening as indicated by a higher affinity to CD4 and loss in shielding against neutralizing antibodies. We observed a strikingly decreased infectivity on CD4high target cells, but sustained infectivity on CD4low targets, including macrophages. Intriguingly, the adaptation to CD4low targets altered the kinetic of the entry process, leading to rapid CD4 engagement and an extended transition time between CD4 and CCR5 binding during entry. This phenotype was also observed for certain central nervous system (CNS) derived macrophage-tropic viruses, highlighting that the functional perturbation we defined upon in vitro adaptation to CD4low targets occurs in vivo. Collectively, our findings suggest that CD4low adapted envelopes may exhibit severe deficiencies in entry fitness and shielding early in their evolution. Considering this, adaptation to CD4low targets may preferentially occur in a sheltered and immune-privileged environment such as the CNS to allow fitness restoring compensatory mutations to occur. PMID:28264054

Analysis of the Spanish CCR5-∆32 inventory of cord blood units: lower cell counts in homozygous donors.

PubMed

Enrich, Emma; Vidal, Francisco; Sánchez-Gordo, Francisco; Gómez-Zumaquero, Juan M; Balas, Antonio; Rudilla, Francesc; Barea, Luisa; Castro, Ana; Larrea, Luis; Perez-Vaquero, Miguel A; Prat, Isidro; Querol, Sergio; Garrido, Gregorio; Matesanz, Rafael; Carreras, Enric; Duarte, Rafael F

2018-02-06

The possibility to use CCR5-∆32 umbilical cord blood to cure HIV infection in patients in need of a hematopoietic transplant has been suggested. The less stringent HLA compatibility needed in this type of transplant facilitates the search of a suitable donor having the CCR5-∆32 mutation. To achieve an inventory of CCR5-∆32 cord blood units, the 20,236 best cell quality units of the Spanish Registry were genotyped. Furthermore, their CD34 + and total nucleated cells counts, blood type, gender, HLA and donor's geographical and ancestral origin were analyzed. The results showed 130 (0.64%) units homozygous for the deletion, 2,646 (13.08%) heterozygous and 17,460 (86.28%) did not present the mutation. Interestingly, a significant lower amount of CD34 + cells was found in the CCR5-∆32 homozygous units. In addition, a significant association was found among donor's ancestral origin and the mutation, with a higher percentage of CCR5-∆32 units with a European ancestry. In summary, identification of a relatively high number of CCR5-∆32 units is feasible and will facilitate the development of clinical trials for HIV cure in patients requiring hematopoietic transplantation. Further studies are required to understand the significance of lower cell counts within the CCR5-∆32 homozygous group and its clinical impact.

Phylogenetic reconstruction of dengue virus type 2 in Colombia

PubMed Central

2012-01-01

Background Dengue fever is perhaps the most important viral re-emergent disease especially in tropical and sub-tropical countries, affecting about 50 million people around the world yearly. In Colombia, dengue virus was first detected in 1971 and still remains as a major public health issue. Although four viral serotypes have been recurrently identified, dengue virus type 2 (DENV-2) has been involved in the most important outbreaks during the last 20 years, including 2010 when the fatality rate highly increased. As there are no major studies reviewing virus origin and genotype distribution in this country, the present study attempts to reconstruct the phylogenetic history of DENV-2 using a sequence analysis from a 224 bp PCR-amplified product corresponding to the carboxyl terminus of the envelope (E) gene from 48 Colombian isolates. Results As expected, the oldest isolates belonged to the American genotype (subtype V), but the strains collected since 1990 represent the American/Asian genotype (subtype IIIb) as previously reported in different American countries. Interestingly, the introduction of this genotype coincides with the first report of dengue hemorrhagic fever in Colombia at the end of 1989 and the increase of cases during the next years. Conclusion After replacement of the American genotype, several lineages of American/Asian subtype have rapidly spread all over the country evolving in new clades. Nevertheless, the direct association of these new variants in the raise of lethality rate observed during the last outbreak has to be demonstrated. PMID:22405440

Phylogeography of Influenza A(H3N2) Virus in Peru, 2010-2012.

PubMed

Pollett, Simon; Nelson, Martha I; Kasper, Matthew; Tinoco, Yeny; Simons, Mark; Romero, Candice; Silva, Marita; Lin, Xudong; Halpin, Rebecca A; Fedorova, Nadia; Stockwell, Timothy B; Wentworth, David; Holmes, Edward C; Bausch, Daniel G

2015-08-01

It remains unclear whether lineages of influenza A(H3N2) virus can persist in the tropics and seed temperate areas. We used viral gene sequence data sampled from Peru to test this source-sink model for a Latin American country. Viruses were obtained during 2010-2012 from influenza surveillance cohorts in Cusco, Tumbes, Puerto Maldonado, and Lima. Specimens positive for influenza A(H3N2) virus were randomly selected and underwent hemagglutinin sequencing and phylogeographic analyses. Analysis of 389 hemagglutinin sequences from Peru and 2,192 global sequences demonstrated interseasonal extinction of Peruvian lineages. Extensive mixing occurred with global clades, but some spatial structure was observed at all sites; this structure was weakest in Lima and Puerto Maldonado, indicating that these locations may experience greater viral traffic. The broad diversity and co-circulation of many simultaneous lineages of H3N2 virus in Peru suggests that this country should not be overlooked as a potential source for novel pandemic strains.

Phylogeography of Influenza A(H3N2) Virus in Peru, 2010–2012

PubMed Central

Nelson, Martha I.; Kasper, Matthew; Tinoco, Yeny; Simons, Mark; Romero, Candice; Silva, Marita; Lin, Xudong; Halpin, Rebecca A.; Fedorova, Nadia; Stockwell, Timothy B.; Wentworth, David; Holmes, Edward C.; Bausch, Daniel G.

2015-01-01

It remains unclear whether lineages of influenza A(H3N2) virus can persist in the tropics and seed temperate areas. We used viral gene sequence data sampled from Peru to test this source–sink model for a Latin American country. Viruses were obtained during 2010–2012 from influenza surveillance cohorts in Cusco, Tumbes, Puerto Maldonado, and Lima. Specimens positive for influenza A(H3N2) virus were randomly selected and underwent hemagglutinin sequencing and phylogeographic analyses. Analysis of 389 hemagglutinin sequences from Peru and 2,192 global sequences demonstrated interseasonal extinction of Peruvian lineages. Extensive mixing occurred with global clades, but some spatial structure was observed at all sites; this structure was weakest in Lima and Puerto Maldonado, indicating that these locations may experience greater viral traffic. The broad diversity and co-circulation of many simultaneous lineages of H3N2 virus in Peru suggests that this country should not be overlooked as a potential source for novel pandemic strains. PMID:26196599

Itaya virus, a Novel Orthobunyavirus Associated with Human Febrile Illness, Peru.

PubMed

Hontz, Robert D; Guevara, Carolina; Halsey, Eric S; Silvas, Jesus; Santiago, Felix W; Widen, Steven G; Wood, Thomas G; Casanova, Wilma; Vasilakis, Nikos; Watts, Douglas M; Kochel, Tadeusz J; Ebihara, Hideki; Aguilar, Patricia V

2015-05-01

Our genetic analyses of uncharacterized bunyaviruses isolated in Peru identified a possible reassortant virus containing small and large gene segment sequences closely related to the Caraparu virus and a medium gene segment sequence potentially derived from an unidentified group C orthobunyavirus. Neutralization tests confirmed serologic distinction among the newly identified virus and the prototype and Caraparu strains. This virus, named Itaya, was isolated in 1999 and 2006 from febrile patients in the cities of Iquitos and Yurimaguas in Peru. The geographic distance between the 2 cases suggests that the Itaya virus could be widely distributed throughout the Amazon basin in northeastern Peru. Identification of a new Orthobunyavirus species that causes febrile disease in humans reinforces the need to expand viral disease surveillance in tropical regions of South America.

Itaya virus, a Novel Orthobunyavirus Associated with Human Febrile Illness, Peru

PubMed Central

Hontz, Robert D.; Guevara, Carolina; Halsey, Eric S.; Silvas, Jesus; Santiago, Felix W.; Widen, Steven G.; Wood, Thomas G.; Casanova, Wilma; Vasilakis, Nikos; Watts, Douglas M.; Kochel, Tadeusz J.; Ebihara, Hideki

2015-01-01

Our genetic analyses of uncharacterized bunyaviruses isolated in Peru identified a possible reassortant virus containing small and large gene segment sequences closely related to the Caraparu virus and a medium gene segment sequence potentially derived from an unidentified group C orthobunyavirus. Neutralization tests confirmed serologic distinction among the newly identified virus and the prototype and Caraparu strains. This virus, named Itaya, was isolated in 1999 and 2006 from febrile patients in the cities of Iquitos and Yurimaguas in Peru. The geographic distance between the 2 cases suggests that the Itaya virus could be widely distributed throughout the Amazon basin in northeastern Peru. Identification of a new Orthobunyavirus species that causes febrile disease in humans reinforces the need to expand viral disease surveillance in tropical regions of South America. PMID:25898901

Altered expression of glial markers, chemokines, and opioid receptors in the spinal cord of type 2 diabetic monkeys.

PubMed

Kiguchi, Norikazu; Ding, Huiping; Peters, Christopher M; Kock, Nancy D; Kishioka, Shiroh; Cline, J Mark; Wagner, Janice D; Ko, Mei-Chuan

2017-01-01

Neuroinflammation is a pathological condition that underlies diabetes and affects sensory processing. Given the high prevalence of pain in diabetic patients and crosstalk between chemokines and opioids, it is pivotal to know whether neuroinflammation-associated mediators are dysregulated in the central nervous system of diabetic primates. Therefore, the aim of this study was to investigate whether mRNA expression levels of glial markers, chemokines, and opioid receptors are altered in the spinal cord and thalamus of naturally occurring type 2 diabetic monkeys (n=7) compared with age-matched non-diabetic monkeys (n=6). By using RT-qPCR, we found that mRNA expression levels of both GFAP and IBA1 were up-regulated in the spinal dorsal horn (SDH) of diabetic monkeys compared with non-diabetic monkeys. Among all chemokines, expression levels of three chemokine ligand-receptor systems, i.e., CCL2-CCR2, CCL3-CCR1/5, and CCL4-CCR5, were up-regulated in the SDH of diabetic monkeys. Moreover, in the SDH, seven additional chemokine receptors, i.e., CCR4, CCR6, CCR8, CCR10, CXCR3, CXCR5, and CXCR6, were also up-regulated in diabetic monkeys. In contrast, expression levels of MOP, KOP, and DOP, but not NOP receptors, were down-regulated in the SDH of diabetic monkeys, and the thalamus had fewer changes in the glial markers, chemokines and opioids. These findings indicate that neuroinflammation, manifested as glial activation and simultaneous up-regulation of multiple chemokine ligands and receptors, seems to be permanent in type 2 diabetic monkeys. As chemokines and opioids are important pain modulators, this first-in-primate study provides a translational bridge for determining the functional efficacy of spinal drugs targeting their signaling cascades. Copyright © 2016 Elsevier B.V. All rights reserved.

The genomes of four novel begomoviruses and a new Sida micrantha mosaic virus strain from Bolivian weeds.

PubMed

Wyant, Patrícia Soares; Gotthardt, Diether; Schäfer, Benjamin; Krenz, Björn; Jeske, Holger

2011-02-01

Begomovirus is the largest genus within the family Geminiviridae and includes economically important plant DNA viruses infecting a broad range of plant species and causing devastating crop diseases, mainly in subtropical and tropical countries. Besides cultivated plants, many weeds act as virus reservoirs. Eight begomovirus isolates from Bolivian weeds were examined using rolling-circle amplification (RCA) and restriction fragment length polymorphism (RFLP). An efficient, novel cloning strategy using limited Sau3A digestion to obtain tandem-repeat inserts allowed the sequencing of the complete genomes. The viruses were classified by phylogenetic analysis as typical bipartite New World begomoviruses. Four of them represented distinct new virus species, for which the names Solanum mosaic Bolivia virus, Sida mosaic Bolivia virus 1, Sida mosaic Bolivia virus 2, and Abutilon mosaic Bolivia virus are proposed. Three were variants of a new strain of Sida micrantha mosaic virus (SimMV), SimMV-rho[BoVi07], SimMV-rho[Bo:CF1:07] and SimMV-rho[Bo:CF2:07], and one was a new variant of a previously described SimMV, SimMV-MGS2:07-Bo.

Tropism-independent protection of macaques against vaginal transmission of three SHIVs by the HIV-1 fusion inhibitor T-1249.

PubMed

Veazey, Ronald S; Ketas, Thomas A; Klasse, Per Johan; Davison, Donna K; Singletary, Morgan; Green, Linda C; Greenberg, Michael L; Moore, John P

2008-07-29

We have assessed the potential of the fusion inhibitory peptide T-1249 for development as a vaginal microbicide to prevent HIV-1 sexual transmission. When formulated as a simple gel, T-1249 provided dose-dependent protection to macaques against high-dose challenge with three different SHIVs that used either CCR5 or CXCR4 for infection (the R5 virus SHIV-162P3, the X4 virus SHIV-KU1 and the R5X4 virus SHIV-89.6P), and it also protected against SIVmac251 (R5). Protection of half of the test animals was estimated by interpolation to occur at T-1249 concentrations of approximately 40-130 muM, whereas complete protection was observed at 0.1-2 mM. In vitro, T-1249 had substantial breadth of activity against HIV-1 strains from multiple genetic subtypes and in a coreceptor-independent manner. Thus, at 1 muM in a peripheral blood mononuclear cell-based replication assay, T-1249 inhibited all 29 R5 viruses, all 12 X4 viruses and all 7 R5X4 viruses in the test panel, irrespective of their genetic subtype. Combining lower concentrations of T-1249 with other entry inhibitors (CMPD-167, BMS-C, or AMD3465) increased the proportion of test viruses that could be blocked. In the PhenoSense assay, T-1249 was active against 636 different HIV-1 Env-pseudotyped viruses of varying tropism and derived from clinical samples, with IC(50) values typically clustered in a 10-fold range approximately 10 nM. Overall, these results support the concept of using T-1249 as a component of an entry inhibitor-based combination microbicide to prevent the sexual transmission of diverse HIV-1 variants.