Targeting Virus-host Interactions of HIV Replication.

Science.gov (United States)

Weydert, Caroline; De Rijck, Jan; Christ, Frauke; Debyser, Zeger

2016-01-01

Cellular proteins that are hijacked by HIV in order to complete its replication cycle, form attractive new targets for antiretroviral therapy. In particular, the protein-protein interactions between these cellular proteins (cofactors) and viral proteins are of great interest to develop new therapies. Research efforts have led to the validation of different cofactors and some successes in therapeutic applications. Maraviroc, the first cofactor inhibitor approved for human medicinal use, provided a proof of concept. Furthermore, compounds developed as Integrase-LEDGF/p75 interaction inhibitors (LEDGINs) have advanced to early clinical trials. Other compounds targeting cofactors and cofactor-viral protein interactions are currently under development. Likewise, interactions between cellular restriction factors and their counteracting HIV protein might serve as interesting targets in order to impair HIV replication. In this respect, compounds targeting the Vif-APOBEC3G interaction have been described. In this review, we focus on compounds targeting the Integrase- LEDGF/p75 interaction, the Tat-P-TEFb interaction and the Vif-APOBEC3G interaction. Additionally we give an overview of currently discovered compounds presumably targeting cellular cofactor-HIV protein interactions.

Modeling HIV-1 intracellular replication: two simulation approaches

NARCIS (Netherlands)

Zarrabi, N.; Mancini, E.; Tay, J.; Shahand, S.; Sloot, P.M.A.

2010-01-01

Many mathematical and computational models have been developed to investigate the complexity of HIV dynamics, immune response and drug therapy. However, there are not many models which consider the dynamics of virus intracellular replication at a single level. We propose a model of HIV intracellular

Developing strategies for HIV-1 eradication

Science.gov (United States)

Durand, Christine M.; Blankson, Joel N.; Siliciano, Robert F.

2014-01-01

Highly active antiretroviral therapy (HAART) suppresses HIV-1 replication, transforming the outlook for infected patients. However, reservoirs of replication-competent forms of the virus persist during HAART, and when treatment is stopped, high rates of HIV-1 replication return. Recent insights into HIV-1 latency, as well as a report that HIV-1 infection was eradicated in one individual, have renewed interest in finding a cure for HIV-1 infection. Strategies for HIV-1 eradication include gene therapy and hematopoietic stem cell transplantation, stimulating host immunity to control HIV-1 replication, and targeting latent HIV-1 in resting memory CD4+ T cells. Future efforts should aim to provide better understanding of how to reconstitute the CD4+ T cell compartment with genetically engineered cells, exert immune control over HIV-1 replication, and identify and eliminate all viral reservoirs. PMID:22867874

Hili Inhibits HIV Replication in Activated T Cells.

Science.gov (United States)

Peterlin, B Matija; Liu, Pingyang; Wang, Xiaoyun; Cary, Daniele; Shao, Wei; Leoz, Marie; Hong, Tian; Pan, Tao; Fujinaga, Koh

2017-06-01

P-element-induced wimpy-like (Piwil) proteins restrict the replication of mobile genetic elements in the germ line. They are also expressed in many transformed cell lines. In this study, we discovered that the human Piwil 2 (Hili) protein can also inhibit HIV replication, especially in activated CD4 + T cells that are the preferred target cells for this virus in the infected host. Although resting cells did not express Hili, its expression was rapidly induced following T cell activation. In these cells and transformed cell lines, depletion of Hili increased levels of viral proteins and new viral particles. Further studies revealed that Hili binds to tRNA. Some of the tRNAs represent rare tRNA species, whose codons are overrepresented in the viral genome. Targeting tRNA Arg (UCU) with an antisense oligonucleotide replicated effects of Hili and also inhibited HIV replication. Finally, Hili also inhibited the retrotransposition of the endogenous intracysternal A particle (IAP) by a similar mechanism. Thus, Hili joins a list of host proteins that inhibit the replication of HIV and other mobile genetic elements. IMPORTANCE Piwil proteins inhibit the movement of mobile genetic elements in the germ line. In their absence, sperm does not form and male mice are sterile. This inhibition is thought to occur via small Piwi-interacting RNAs (piRNAs). However, in some species and in human somatic cells, Piwil proteins bind primarily to tRNA. In this report, we demonstrate that human Piwil proteins, especially Hili, not only bind to select tRNA species, including rare tRNAs, but also inhibit HIV replication. Importantly, T cell activation induces the expression of Hili in CD4 + T cells. Since Hili also inhibited the movement of an endogenous retrovirus (IAP), our finding shed new light on this intracellular resistance to exogenous and endogenous retroviruses as well as other mobile genetic elements. Copyright © 2017 American Society for Microbiology.

[Applying the Modified Delphi Technique to Develop the Role of HIV Case Managers and Essential Nursing Competencies in HIV Care].

Science.gov (United States)

Ko, Nai-Ying; Hsieh, Chia-Yin; Chen, Yen-Chin; Tsai, Chen-Hsi; Liu, Hsiao-Ying; Liu, Li-Fang

2015-08-01

Since 2005, the Taiwan Centers for Disease Control (Taiwan CDC) initiated an HIV case management program in AIDS-designated hospitals to provide integrative services and risk-reduction counseling for HIV-infected individuals. In light of the increasingly complex and highly specialized nature of clinical care, expanding and improving competency-based professional education is important to enhance the quality of HIV/AIDS care. The aim of this study was to develop the essential competency framework for HIV care for HIV case managers in Taiwan. We reviewed essential competencies of HIV care from Canada, the United Kingdom, and several African countries and devised descriptions of the roles of case managers and of the associated core competencies for HIV care in Taiwan. The modified Delphi technique was used to evaluate the draft framework of these roles and core competencies. A total of 15 HIV care experts were invited to join the expert panel to review and rank the draft framework. The final framework consisted of 7 roles and 27 competencies for HIV case managers. In Round 1, only 3 items did not receive consensus approval from the experts. After modification based on opinions of the experts, 7 roles and 27 competencies received 97.06% consensus approval in Round 2 and were organized into the final framework for HIV case managers. These roles and associated core competencies were: HIV Care Expert (9 competencies), Communicator (1 competency), Collaborator (4 competencies), Navigator (2 competencies), Manager (4 competencies), Advocate (2 competencies), and Professional (5 competencies). The authors developed an essential competency framework for HIV care using the consensus of a multidisciplinary expert panel. Curriculum developers and advanced nurses and practitioners may use this framework to support developments and to ensure a high quality of HIV care.

Modulation of HIV replication in monocyte derived macrophages (MDM) by steroid hormones.

Science.gov (United States)

Devadas, Krishnakumar; Biswas, Santanu; Ragupathy, Viswanath; Lee, Sherwin; Dayton, Andrew; Hewlett, Indira

2018-01-01

Significant sex specific differences in the progression of HIV/AIDS have been reported. Several studies have implicated steroid hormones in regulating host factor expression and modulating HIV transmission and replication. However, the exact mechanism exerted by steroid hormones estrogen and progesterone in the regulation of HIV-1 replication is still unclear. Results from the current study indicated a dose dependent down regulation of HIV-1 replication in monocyte derived macrophages pre-treated with high concentrations of estrogen or progesterone. To elucidate the molecular mechanisms associated with the down regulation of HIV-1 replication by estrogen and progesterone we used PCR arrays to analyze the expression profile of host genes involved in antiviral responses. Several chemokines, cytokines, transcription factors, interferon stimulated genes and genes involved in type-1 interferon signaling were down regulated in cells infected with HIV-1 pre-treated with high concentrations of estrogen or progesterone compared to untreated HIV-1 infected cells or HIV-1 infected cells treated with low concentrations of estrogen or progesterone. The down regulation of CXCL9, CXCL10 and CXCL11 chemokines and IL-1β, IL-6 cytokines in response to high concentrations of estrogen and progesterone pre-treatment in HIV-1 infected cells was confirmed at the protein level by quantitating chemokine and cytokine concentrations in the culture supernatant. These results demonstrate that a potent anti-inflammatory response is mediated by pre-treatment with high concentrations of estrogen and progesterone. Thus, our study suggests a strong correlation between the down-modulation of anti-viral and pro-inflammatory responses mediated by estrogen and progesterone pre-treatment and the down regulation of HIV-1 replication. These findings may be relevant to clinical observations of sex specific differences in patient populations and point to the need for further investigation.

Insulated hsp70B' promoter: stringent heat-inducible activity in replication-deficient, but not replication-competent adenoviruses.

Science.gov (United States)

Rohmer, Stanimira; Mainka, Astrid; Knippertz, Ilka; Hesse, Andrea; Nettelbeck, Dirk M

2008-04-01

Key to the realization of gene therapy is the development of efficient and targeted gene transfer vectors. Therapeutic gene transfer by replication-deficient or more recently by conditionally replication-competent/oncolytic adenoviruses has shown much promise. For specific applications, however, it will be advantageous to provide vectors that allow for external control of gene expression. The efficient cellular heat shock system in combination with available technology for focused and controlled hyperthermia suggests heat-regulated transcription control as a promising tool for this purpose. We investigated the feasibility of a short fragment of the human hsp70B' promoter, with and without upstream insulator elements, for the regulation of transgene expression by replication-deficient or oncolytic adenoviruses. Two novel adenoviral vectors with an insulated hsp70B' promoter were developed and showed stringent heat-inducible gene expression with induction ratios up to 8000-fold. In contrast, regulation of gene expression from the hsp70B' promoter without insulation was suboptimal. In replication-competent/oncolytic adenoviruses regulation of the hsp70B' promoter was lost specifically during late replication in permissive cells and could not be restored by the insulators. We developed novel adenovirus gene transfer vectors that feature improved and stringent regulation of transgene expression from the hsp70B' promoter using promoter insulation. These vectors have potential for gene therapy applications that benefit from external modulation of therapeutic gene expression or for combination therapy with hyperthermia. Furthermore, our study reveals that vector replication can deregulate inserted cellular promoters, an observation which is of relevance for the development of replication-competent/oncolytic gene transfer vectors. (c) 2008 John Wiley & Sons, Ltd.

Dimensions of Poverty and Health Outcomes Among People Living with HIV Infection: Limited Resources and Competing Needs.

Science.gov (United States)

Kalichman, Seth C; Hernandez, Dominica; Kegler, Christopher; Cherry, Chauncey; Kalichman, Moira O; Grebler, Tamar

2015-08-01

HIV infection is concentrated in populations living in poverty. We examined the overlapping and independent effects of multiple poverty indicators on HIV-related health status. Because substance use can create competing survival needs when resources are limited, we also sought to objectively measure expenditures on food relative to alcohol and tobacco products. To achieve these aims, 459 men and 212 women living with HIV infection in Atlanta, GA completed measures of socio-demographic and heath characteristics as well as multiple indicators of poverty including housing stability, transportation, food insecurity, and substance use. Participants were given a $30 grocery gift card for their participation and we collected receipts which were coded for alcohol (beer, wine, liquors) and tobacco purchases. Results showed that participants with unsuppressed HIV replication were significantly more likely to experience multiple indicators of poverty. In addition, one in four participants purchased alcohol or tobacco products with their gift cards, with as much as one-fourth of money spent on these products. A multivariable logistic regression model showed that food insecurity was independently associated with unsuppressed HIV, and purchasing alcohol or tobacco products did not moderate this association. Results confirm previous research to show the primacy of food insecurity in relation to HIV-related health outcomes. Competing survival needs, including addictive substances, should be addressed in programs that aim to alleviate poverty to enhance the health and well-being of people with HIV infection.

Selective serotonin reuptake inhibitor suppression of HIV infectivity and replication.

Science.gov (United States)

Benton, Tami; Lynch, Kevin; Dubé, Benoit; Gettes, David R; Tustin, Nancy B; Ping Lai, Jian; Metzger, David S; Blume, Joshua; Douglas, Steven D; Evans, Dwight L

2010-11-01

To test the hypothesis that the selective serotonin reuptake inhibitor (SSRI) citalopram would down-regulate human immunodeficiency virus (HIV) infectivity and that the greatest effects would be seen in people with depression. Depression is a risk factor for morbidity and mortality in HIV/acquired immune deficiency syndrome. Serotonin (5-HT) neurotransmission has been implicated in the pathobiology of depression, and pharmacologic therapies for depression target this system. The 5-HT transporter and 5-HT receptors are widely distributed throughout the central nervous and immune systems. Depression has been associated with suppression of natural killer cells and CD8(+) lymphocytes, key regulators of HIV infection. Ex vivo models for acute and chronic HIV infection were used to study the effects of citalopram on HIV viral infection and replication in 48 depressed and nondepressed women. For both the acute and chronic infection models, HIV reverse transcriptase activity was measured in the citalopram treatment condition and the control condition. The SSRI significantly down-regulated the reverse transcriptase response in both the acute and chronic infection models. Specifically, citalopram significantly decreased the acute HIV infectivity of macrophages. Citalopram also significantly decreased HIV viral replication in the latently infected T-cell line and in the latently infected macrophage cell line. There was no difference in down-regulation by depression status. These studies suggest that an SSRI enhances natural killer/CD8 noncytolytic HIV suppression in HIV/acquired immune deficiency syndrome and decreases HIV viral infectivity of macrophages, ex vivo, suggesting the need for in vivo studies to determine a potential role for agents targeting serotonin in the host defense against HIV.

Constitutively Active MAVS Inhibits HIV-1 Replication via Type I Interferon Secretion and Induction of HIV-1 Restriction Factors.

Directory of Open Access Journals (Sweden)

Sachin Gupta

Full Text Available Type I interferon is known to inhibit HIV-1 replication through the induction of interferon stimulated genes (ISG, including a number of HIV-1 restriction factors. To better understand interferon-mediated HIV-1 restriction, we constructed a constitutively active form of the RIG-I adapter protein MAVS. Constitutive MAVS was generated by fusion of full length MAVS to a truncated form of the Epstein Barr virus protein LMP1 (ΔLMP1. Supernatant from ΔLMP1-MAVS-transfected 293T cells contained high levels of type I interferons and inhibited HIV replication in both TZM-bl and primary human CD4+ T cells. Supernatant from ΔLMP1-MAVS-transfected 293T cells also inhibited replication of VSV-G pseudotyped single cycle SIV in TZM-bl cells, suggesting restriction was post-entry and common to both HIV and SIV. Gene array analysis of ΔLMP1-MAVS-transfected 293T cells and trans-activated CD4+ T cells showed significant upregulation of ISG, including previously characterized HIV restriction factors Viperin, Tetherin, MxB, and ISG56. Interferon blockade studies implicated interferon-beta in this response. In addition to direct viral inhibition, ΔLMP1-MAVS markedly enhanced secretion of IFN-β and IL-12p70 by dendritic cells and the activation and maturation of dendritic cells. Based on this immunostimulatory activity, an adenoviral vector (Ad5 expressing ΔLMP1-MAVS was tested as a molecular adjuvant in an HIV vaccine mouse model. Ad5-Gag antigen combined with Ad5-ΔLMP1-MAVS enhanced control of vaccinia-gag replication in a mouse challenge model, with 4/5 animals showing undetectable virus following challenge. Overall, ΔLMP1-MAVS is a promising reagent to inhibit HIV-1 replication in infected tissues and enhance vaccine-mediated immune responses, while avoiding toxicity associated with systemic type I interferon administration.

Nursing accounting competencies related to HIV in a Papua New Guinea context.

Science.gov (United States)

Brown, Alistair M

2013-01-01

Nursing administration is an important part of the campaign to eliminate HIV across Papua New Guinea (PNG). This paper considers the critical importance of developing nursing leadership in effective accounting competencies in relation to HIV projects in PNG. The results of the study's textual analysis of audit reports of the Auditor General of PNG revealed a failure on the part of PNG's main health agencies involved with its national HIV program to provide competent financial reporting. In light of these results, this study shows how improving accounting and other financial competencies among nursing leaders would benefit the implementation of the PNG HIV national strategy. The findings of this study have implications not only for the internal control of HIV nursing competencies but also for nursing leadership related to HIV issues in a developing-country context. Copyright © 2013 Association of Nurses in AIDS Care. Published by Elsevier Inc. All rights reserved.

Artificial 64-Residue HIV-1 Enhancer-Binding Peptide Is a Potent Inhibitor of Viral Replication in HIV-1-Infected Cells.

Science.gov (United States)

Oufir, Mouhssin; Bisset, Leslie R; Hoffmann, Stefan R K; Xue, Gongda; Klauser, Stephan; Bergamaschi, Bianca; Gervaix, Alain; Böni, Jürg; Schüpbach, Jörg; Gutte, Bernd

2011-01-01

An artificial HIV-1 enhancer-binding peptide was extended by nine consecutive arginine residues at the C-terminus and by the nuclear localization signal of SV40 large T antigen at the N-terminus. The resulting synthetic 64-residue peptide was found to bind to the two enhancers of the HIV-1 long terminal repeat, cross the plasma membrane and the nuclear envelope of human cells, and suppress the HIV-1 enhancer-controlled expression of a green fluorescent protein reporter gene. Moreover, HIV-1 replication is inhibited by this peptide in HIV-1-infected CEM-GFP cells as revealed by HIV-1 p24 ELISA and real-time RT-PCR of HIV-1 RNA. Rapid uptake of this intracellular stable and inhibitory peptide into the cells implies that this peptide may have the potential to attenuate HIV-1 replication in vivo.

Artificial 64-Residue HIV-1 Enhancer-Binding Peptide Is a Potent Inhibitor of Viral Replication in HIV-1-Infected Cells

Directory of Open Access Journals (Sweden)

Mouhssin Oufir

2011-01-01

Full Text Available An artificial HIV-1 enhancer-binding peptide was extended by nine consecutive arginine residues at the C-terminus and by the nuclear localization signal of SV40 large T antigen at the N-terminus. The resulting synthetic 64-residue peptide was found to bind to the two enhancers of the HIV-1 long terminal repeat, cross the plasma membrane and the nuclear envelope of human cells, and suppress the HIV-1 enhancer-controlled expression of a green fluorescent protein reporter gene. Moreover, HIV-1 replication is inhibited by this peptide in HIV-1-infected CEM-GFP cells as revealed by HIV-1 p24 ELISA and real-time RT-PCR of HIV-1 RNA. Rapid uptake of this intracellular stable and inhibitory peptide into the cells implies that this peptide may have the potential to attenuate HIV-1 replication in vivo.

A reporter system for replication-competent gammaretroviruses: the inGluc-MLV-DERSE assay

Science.gov (United States)

Aloia, Amanda L.; Duffy, Lisa; Pak, Vladimir; Lee, KyeongEun; Sanchez-Martinez, Silvia; Derse, David; Heidecker, Gisela; Cornetta, Kenneth; Rein, Alan

2012-01-01

While novel retroviral vectors for use in gene-therapy products are reducing the potential for formation of replication-competent retrovirus (RCR), it remains crucial to screen products for RCR for both research and clinical purposes. For clinical grade gammaretrovirus-based vectors, RCR screening is achieved by an extended S+L− or marker rescue assay, while standard methods for replication-competent lentivirus detection are still in development. In this report, we describe a rapid and sensitive method for replication-competent gammaretrovirus detection. We used this assay to detect three members of the gammaretrovirus family and compared the sensitivity of our assay with well-established methods for retrovirus detection, including the extended S+L− assay. Results presented here demonstrate that this assay should be useful for gene-therapy product testing. PMID:22402321

Cytotoxic effects of replication-competent adenoviruses on human esophageal carcinoma are enhanced by forced p53 expression

International Nuclear Information System (INIS)

Yang, Shan; Kawamura, Kiyoko; Okamoto, Shinya; Yamauchi, Suguru; Shingyoji, Masato; Sekine, Ikuo; Kobayashi, Hiroshi; Tada, Yuji; Tatsumi, Koichiro; Hiroshima, Kenzo; Shimada, Hideaki; Tagawa, Masatoshi

2015-01-01

Improvement of transduction and augmentation of cytotoxicity are crucial for adenoviruses (Ad)-mediated gene therapy for cancer. Down-regulated expression of type 5 Ad (Ad5) receptors on human tumors hampered Ad-mediated transduction. Furthermore, a role of the p53 pathways in cytotoxicity mediated by replication-competent Ad remained uncharacterized. We constructed replication-competent Ad5 of which the E1 region genes were activated by a transcriptional regulatory region of the midkine or the survivin gene, which is expressed preferentially in human tumors. We also prepared replication-competent Ad5 which were regulated by the same region but had a fiber-knob region derived from serotype 35 (AdF35). We examined the cytotoxicity of these Ad and a possible combinatory use of the replication-competent AdF35 and Ad5 expressing the wild-type p53 gene (Ad5/p53) in esophageal carcinoma cells. Expression levels of molecules involved in cell death, anti-tumor effects in vivo and production of viral progenies were also investigated. Replication-competent AdF35 in general achieved greater cytotoxic effects to esophageal carcinoma cells than the corresponding replication-competent Ad5. Infection with the AdF35 induced cleavages of caspases and increased sub-G1 fractions, but did not activate the autophagy pathway. Transduction with Ad5/p53 in combination with the replication-competent AdF35 further enhanced the cytotoxicity in a synergistic manner. We also demonstrated the combinatory effects in an animal model. Transduction with Ad5/p53 however suppressed production of replication-competent AdF35 progenies, but the combination augmented Ad5/p53-mediated p53 expression levels and the downstream pathways. Combination of replication-competent AdF35 and Ad5/p53 achieved synergistic cytotoxicity due to enhanced p53-mediated apoptotic pathways. The online version of this article (doi:10.1186/s12885-015-1482-8) contains supplementary material, which is available to authorized

Improved innate and adaptive immunostimulation by genetically modified HIV-1 protein expressing NYVAC vectors.

Directory of Open Access Journals (Sweden)

Esther D Quakkelaar

Full Text Available Attenuated poxviruses are safe and capable of expressing foreign antigens. Poxviruses are applied in veterinary vaccination and explored as candidate vaccines for humans. However, poxviruses express multiple genes encoding proteins that interfere with components of the innate and adaptive immune response. This manuscript describes two strategies aimed to improve the immunogenicity of the highly attenuated, host-range restricted poxvirus NYVAC: deletion of the viral gene encoding type-I interferon-binding protein and development of attenuated replication-competent NYVAC. We evaluated these newly generated NYVAC mutants, encoding HIV-1 env, gag, pol and nef, for their ability to stimulate HIV-specific CD8 T-cell responses in vitro from blood mononuclear cells of HIV-infected subjects. The new vectors were evaluated and compared to the parental NYVAC vector in dendritic cells (DCs, RNA expression arrays, HIV gag expression and cross-presentation assays in vitro. Deletion of type-I interferon-binding protein enhanced expression of interferon and interferon-induced genes in DCs, and increased maturation of infected DCs. Restoration of replication competence induced activation of pathways involving antigen processing and presentation. Also, replication-competent NYVAC showed increased Gag expression in infected cells, permitting enhanced cross-presentation to HIV-specific CD8 T cells and proliferation of HIV-specific memory CD8 T-cells in vitro. The recombinant NYVAC combining both modifications induced interferon-induced genes and genes involved in antigen processing and presentation, as well as increased Gag expression. This combined replication-competent NYVAC is a promising candidate for the next generation of HIV vaccines.

HIV-1 Replication and the Cellular Eukaryotic Translation Apparatus

Directory of Open Access Journals (Sweden)

Santiago Guerrero

2015-01-01

Full Text Available Eukaryotic translation is a complex process composed of three main steps: initiation, elongation, and termination. During infections by RNA- and DNA-viruses, the eukaryotic translation machinery is used to assure optimal viral protein synthesis. Human immunodeficiency virus type I (HIV-1 uses several non-canonical pathways to translate its own proteins, such as leaky scanning, frameshifting, shunt, and cap-independent mechanisms. Moreover, HIV-1 modulates the host translation machinery by targeting key translation factors and overcomes different cellular obstacles that affect protein translation. In this review, we describe how HIV-1 proteins target several components of the eukaryotic translation machinery, which consequently improves viral translation and replication.

Astrocyte Apoptosis and HIV Replication Are Modulated in Host Cells Coinfected with Trypanosoma cruzi

Directory of Open Access Journals (Sweden)

Javier M. Urquiza

2017-08-01

Full Text Available The protozoan Trypanosoma cruzi is the etiological agent of Chagas disease. In immunosuppressed individuals, as it occurs in the coinfection with human immunodeficiency virus (HIV, the central nervous system may be affected. In this regard, reactivation of Chagas disease is severe and often lethal, and it accounts for meningoencephalitis. Astrocytes play a crucial role in the environment maintenance of healthy neurons; however, they can host HIV and T. cruzi. In this report, human astrocytes were infected in vitro with both genetically modified-pathogens to express alternative fluorophore. As evidenced by fluorescence microscopy and flow cytometry, HIV and T. cruzi coexist in the same astrocyte, likely favoring reciprocal interactions. In this context, lower rates of cell death were observed in both T. cruzi monoinfected-astrocytes and HIV-T. cruzi coinfection in comparison with those infected only with HIV. The level of HIV replication is significantly diminished under T. cruzi coinfection, but without affecting the infectivity of the HIV progeny. This interference with viral replication appears to be related to the T. cruzi multiplication rate or its increased intracellular presence but does not require their intracellular cohabitation or infected cell-to-cell contact. Among several Th1/Th2/Th17 profile-related cytokines, only IL-6 was overexpressed in HIV-T. cruzi coinfection exhibiting its cytoprotective role. This study demonstrates that T. cruzi and HIV are able to coinfect astrocytes thus altering viral replication and apoptosis.

The Alphabet Soup of HIV Reservoir Markers.

Science.gov (United States)

Sharaf, Radwa R; Li, Jonathan Z

2017-04-01

Despite the success of antiretroviral therapy in suppressing HIV, life-long therapy is required to avoid HIV reactivation from long-lived viral reservoirs. Currently, there is intense interest in searching for therapeutic interventions that can purge the viral reservoir to achieve complete remission in HIV patients off antiretroviral therapy. The evaluation of such interventions relies on our ability to accurately and precisely measure the true size of the viral reservoir. In this review, we assess the most commonly used HIV reservoir assays, as a clear understanding of the strengths and weaknesses of each is vital for the accurate interpretation of results and for the development of improved assays. The quantification of intracellular or plasma HIV RNA or DNA levels remains the most commonly used tests for the characterization of the viral reservoir. While cost-effective and high-throughput, these assays are not able to differentiate between replication-competent or defective fractions or quantify the number of infected cells. Viral outgrowth assays provide a lower bound for the fraction of cells that can produce infectious virus, but these assays are laborious, expensive and substantially underestimate the potential reservoir of replication-competent provirus. Newer assays are now available that seek to overcome some of these problems, including full-length proviral sequencing, inducible HIV RNA assays, ultrasensitive p24 assays and murine adoptive transfer techniques. The development and evaluation of strategies for HIV remission rely upon our ability to accurately and precisely quantify the size of the remaining viral reservoir. At this time, all current HIV reservoir assays have drawbacks such that combinations of assays are generally needed to gain a more comprehensive view of the viral reservoir. The development of novel, rapid, high-throughput assays that can sensitively quantify the levels of the replication-competent HIV reservoir is still needed.

Suppression of HIV replication by CD8+regulatory T-cells in elite controllers

Directory of Open Access Journals (Sweden)

Wei eLu

2016-04-01

Full Text Available We previously demonstrated in the Chinese macaque model that an oral vaccine made of inactivated SIV and lactobacillus plantarum induced CD8+regulatory T-cells which suppressed the activation of SIV+CD4+T-cells, prevented SIV replication and protected macaques from SIV challenges.Here ,we sought whether a similar population of CD8+T-regs would induce the suppression of HIV replication in elite controllers (ECs, a small population (3‰ of HIV-infected patients with undetectable HIV replication. For that purpose, we investigated the in vitro antiviral activity of fresh CD8+T-cells on HIV-infected CD4+T-cells taken from 10 ECs. The 10 ECs had a classical genomic profile: all of them carried the KIR3DL1 gene and nine carried at least one allele of HLA-B:Bw4-80Ile ( i.e. with an isoleucine residue at position 80. In the nine HLA-B:Bw4-80Ile positive patients, we demonstrated a strong viral suppression byKIR3DL1-expressing CD8+T-cells that required cell-to-cell contact to switch off the activation signals in infected CD4+T-cells. KIR3DL1-expressing CD8+T-cells withdrawal and KIR3DL1 neutralization by a specific anti-KIR antibody inhibited the suppression of viral replication. Our findings provide the first evidence for an instrumental role of KIR-expressing CD8+ regulatory T- cells in the natural control of HIV-1 infection.

Human cellular restriction factors that target HIV-1 replication

Directory of Open Access Journals (Sweden)

Jeang Kuan-Teh

2009-09-01

Full Text Available Abstract Recent findings have highlighted roles played by innate cellular factors in restricting intracellular viral replication. In this review, we discuss in brief the activities of apolipoprotein B mRNA-editing enzyme 3G (APOBEC3G, bone marrow stromal cell antigen 2 (BST-2, cyclophilin A, tripartite motif protein 5 alpha (Trim5α, and cellular microRNAs as examples of host restriction factors that target HIV-1. We point to countermeasures encoded by HIV-1 for moderating the potency of these cellular restriction functions.

Development of a replication-competent lentivirus assay for dendritic cell-targeting lentiviral vectors

Directory of Open Access Journals (Sweden)

Daniel C Farley

Full Text Available It is a current regulatory requirement to demonstrate absence of detectable replication-competent lentivirus (RCL in lentiviral vector products prior to use in clinical trials. Immune Design previously described an HIV-1-based integration-deficient lentiviral vector for use in cancer immunotherapy (VP02. VP02 is enveloped with E1001, a modified Sindbis virus glycoprotein which targets dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN expressed on dendritic cells in vivo. Vector enveloped with E1001 does not transduce T-cell lines used in standard HIV-1-based RCL assays, making current RCL testing formats unsuitable for testing VP02. We therefore developed a novel assay to test for RCL in clinical lots of VP02. This assay, which utilizes a murine leukemia positive control virus and a 293F cell line expressing the E1001 receptor DC-SIGN, meets a series of evaluation criteria defined in collaboration with US regulatory authorities and demonstrates the ability of the assay format to amplify and detect a hypothetical RCL derived from VP02 vector components. This assay was qualified and used to test six independent GMP production lots of VP02, in which no RCL was detected. We propose that the evaluation criteria used to rationally design this novel method should be considered when developing an RCL assay for any lentiviral vector.

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

Science.gov (United States)

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.

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

Directory of Open Access Journals (Sweden)

Giacaman Rodrigo A

2008-07-01

Full Text Available Abstract Background 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. Results 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-NL4-3Δenv-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. Conclusion 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.

Casp8p41: The Protean Mediator of Death in CD4 T-cells that Replicate HIV

Directory of Open Access Journals (Sweden)

Rahul Sampath

2016-01-01

Full Text Available HIV cure is now the focus of intense research after Timothy Ray Brown (the Berlin patient set the precedent of being the first and only person cured. A major barrier to achieving this goal on a meaningful scale is an elimination of the latent reservoir, which is thought to comprise CD4-positive cells that harbor integrated, replication-competent HIV provirus. These cells do not express viral proteins, are indistinguishable from uninfected CD4 cells, and are thought to be responsible for HIV viral rebound–-that occurs within weeks of combination anti retroviral therapy (cART interruption. Modalities to engineer transcriptional stimulation (reactivation of this dormant integrated HIV provirus, leading to expression of cytotoxic viral proteins, are thought to be a specific way to eradicate the latently infected CD4 pool and are becoming increasingly relevant in the era of HIV cure. HIV protease is one such protein produced after HIV reactivation that cleaves procaspase-8 to generate a novel protein Casp8p41. Casp8p41 then binds to the BH3 domain of BAK, leading to BAK oligomerization, mitochondrial depolarization, and apoptosis. In central memory T cells (TCMs from HIV-infected patients, an elevated Bcl-2/procaspase-8 ratio was observed, and Casp8p41 binding to Bcl-2 was associated with a lack of reactivation-induced cell death. This was reversed by priming cells with a specific Bcl-2 antagonist prior to reactivation, resulting in increased cell death and decreased HIV DNA in a Casp8p41-dependent pathway. This review describes the biology, clinical relevance, and implications of Casp8p41 for a potential cure.

Anti-HIV Antibody Responses and the HIV Reservoir Size during Antiretroviral Therapy.

Directory of Open Access Journals (Sweden)

Sulggi A Lee

Full Text Available A major challenge to HIV eradication strategies is the lack of an accurate measurement of the total burden of replication-competent HIV (the "reservoir". We assessed the association of anti-HIV antibody responses and the estimated size of the reservoir during antiretroviral therapy (ART.We evaluated anti-HIV antibody profiles using luciferase immunoprecipitation systems (LIPS assay in relation to several blood-based HIV reservoir measures: total and 2-LTR DNA (rtPCR or droplet digital PCR; integrated DNA (Alu PCR; unspliced RNA (rtPCR, multiply-spliced RNA (TILDA, residual plasma HIV RNA (single copy PCR, and replication-competent virus (outgrowth assay. We also assessed total HIV DNA and RNA in gut-associated lymphoid tissue (rtPCR. Spearman correlations and linear regressions were performed using log-transformed blood- or tissue-based reservoir measurements as predictors and log-transformed antibody levels as outcome variables.Among 51 chronically HIV-infected ART-suppressed participants (median age = 57, nadir CD4+ count = 196 cells/mm3, ART duration = 9 years, the most statistically significant associations were between antibody responses to integrase and HIV RNA in gut-associated lymphoid tissue (1.17 fold-increase per two-fold RNA increase, P = 0.004 and between antibody responses to matrix and integrated HIV DNA in resting CD4+ T cells (0.35 fold-decrease per two-fold DNA increase, P = 0.003. However, these associations were not statistically significant after a stringent Bonferroni-adjustment of P<0.00045. Multivariate models including age and duration of ART did not markedly alter results.Our findings suggest that anti-HIV antibody responses may reflect the size of the HIV reservoir during chronic treated HIV disease, possibly via antigen recognition in reservoir sites. Larger, prospective studies are needed to validate the utility of antibody levels as a measure of the total body burden of HIV during treatment.

Increased T cell trafficking as adjunct therapy for HIV-1

OpenAIRE

Fryer, HR; Wolinsky, SM; McLean, AR

2018-01-01

Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical mode...

Transcriptional profiling reveals molecular signatures associated with HIV permissiveness in Th1Th17 cells and identifies Peroxisome Proliferator-Activated Receptor Gamma as an intrinsic negative regulator of viral replication

Science.gov (United States)

2013-01-01

Background We previously demonstrated that primary Th1Th17 cells are highly permissive to HIV-1, whereas Th1 cells are relatively resistant. Molecular mechanisms underlying these differences remain unknown. Results Exposure to replication competent and single-round VSV-G pseudotyped HIV strains provide evidence that superior HIV replication in Th1Th17 vs. Th1 cells was regulated by mechanisms located at entry and post-entry levels. Genome-wide transcriptional profiling identified transcripts upregulated (n = 264) and downregulated (n = 235) in Th1Th17 vs. Th1 cells (p-value Th17 (nuclear receptors, trafficking, p38/MAPK, NF-κB, p53/Ras, IL-23) vs. Th1 cells (proteasome, interferon α/β). Differentially expressed genes were classified into biological categories using Gene Ontology. Th1Th17 cells expressed typical Th17 markers (IL-17A/F, IL-22, CCL20, RORC, IL-26, IL-23R, CCR6) and transcripts functionally linked to regulating cell trafficking (CEACAM1, MCAM), activation (CD28, CD40LG, TNFSF13B, TNFSF25, PTPN13, MAP3K4, LTB, CTSH), transcription (PPARγ, RUNX1, ATF5, ARNTL), apoptosis (FASLG), and HIV infection (CXCR6, FURIN). Differential expression of CXCR6, PPARγ, ARNTL, PTPN13, MAP3K4, CTSH, SERPINB6, PTK2, and ISG20 was validated by RT-PCR, flow cytometry and/or confocal microscopy. The nuclear receptor PPARγ was preferentially expressed by Th1Th17 cells. PPARγ RNA interference significantly increased HIV replication at levels post-entry and prior HIV-DNA integration. Finally, the activation of PPARγ pathway via the agonist Rosiglitazone induced the nuclear translocation of PPARγ and a robust inhibition of viral replication. Conclusions Thus, transcriptional profiling in Th1Th17 vs. Th1 cells demonstrated that HIV permissiveness is associated with a superior state of cellular activation and limited antiviral properties and identified PPARγ as an intrinsic negative regulator of viral replication. Therefore, triggering PPARγ pathway via non

Human-Phosphate-Binding-Protein inhibits HIV-1 gene transcription and replication

Directory of Open Access Journals (Sweden)

Candolfi Ermanno

2011-07-01

Full Text Available Abstract The Human Phosphate-Binding protein (HPBP is a serendipitously discovered lipoprotein that binds phosphate with high affinity. HPBP belongs to the DING protein family, involved in various biological processes like cell cycle regulation. We report that HPBP inhibits HIV-1 gene transcription and replication in T cell line, primary peripherical blood lymphocytes and primary macrophages. We show that HPBP is efficient in naïve and HIV-1 AZT-resistant strains. Our results revealed HPBP as a new and potent anti HIV molecule that inhibits transcription of the virus, which has not yet been targeted by HAART and therefore opens new strategies in the treatment of HIV infection.

Subtype-Specific Differences in Gag-Protease-Driven Replication Capacity Are Consistent with Intersubtype Differences in HIV-1 Disease Progression.

Science.gov (United States)

Kiguoya, Marion W; Mann, Jaclyn K; Chopera, Denis; Gounder, Kamini; Lee, Guinevere Q; Hunt, Peter W; Martin, Jeffrey N; Ball, T Blake; Kimani, Joshua; Brumme, Zabrina L; Brockman, Mark A; Ndung'u, Thumbi

2017-07-01

There are marked differences in the spread and prevalence of HIV-1 subtypes worldwide, and differences in clinical progression have been reported. However, the biological reasons underlying these differences are unknown. Gag-protease is essential for HIV-1 replication, and Gag-protease-driven replication capacity has previously been correlated with disease progression. We show that Gag-protease replication capacity correlates significantly with that of whole isolates ( r = 0.51; P = 0.04), indicating that Gag-protease is a significant contributor to viral replication capacity. Furthermore, we investigated subtype-specific differences in Gag-protease-driven replication capacity using large well-characterized cohorts in Africa and the Americas. Patient-derived Gag-protease sequences were inserted into an HIV-1 NL4-3 backbone, and the replication capacities of the resulting recombinant viruses were measured in an HIV-1-inducible reporter T cell line by flow cytometry. Recombinant viruses expressing subtype C Gag-proteases exhibited substantially lower replication capacities than those expressing subtype B Gag-proteases ( P identified Gag residues 483 and 484, located within the Alix-binding motif involved in virus budding, as major contributors to subtype-specific replicative differences. In East African cohorts, we observed a hierarchy of Gag-protease-driven replication capacities, i.e., subtypes A/C differences in disease progression. We thus hypothesize that the lower Gag-protease-driven replication capacity of subtypes A and C slows disease progression in individuals infected with these subtypes, which in turn leads to greater opportunity for transmission and thus increased prevalence of these subtypes. IMPORTANCE HIV-1 subtypes are unevenly distributed globally, and there are reported differences in their rates of disease progression and epidemic spread. The biological determinants underlying these differences have not been fully elucidated. Here, we show that

HIV-1 replication through hHR23A-mediated interaction of Vpr with 26S proteasome.

Directory of Open Access Journals (Sweden)

Ge Li

2010-06-01

Full Text Available HIV-1 Vpr is a virion-associated protein. Its activities link to viral pathogenesis and disease progression of HIV-infected patients. In vitro, Vpr moderately activates HIV-1 replication in proliferating T cells, but it is required for efficient viral infection and replication in vivo in non-dividing cells such as macrophages. How exactly Vpr contributes to viral replication remains elusive. We show here that Vpr stimulates HIV-1 replication at least in part through its interaction with hHR23A, a protein that binds to 19S subunit of the 26S proteasome and shuttles ubiquitinated proteins to the proteasome for degradation. The Vpr-proteasome interaction was initially discovered in fission yeast, where Vpr was shown to associate with Mts4 and Mts2, two 19S-associated proteins. The interaction of Vpr with the 19S subunit of the proteasome was further confirmed in mammalian cells where Vpr associates with the mammalian orthologues of fission yeast Mts4 and S5a. Consistently, depletion of hHR23A interrupts interaction of Vpr with proteasome in mammalian cells. Furthermore, Vpr promotes hHR23A-mediated protein-ubiquitination, and down-regulation of hHR23A using RNAi significantly reduced viral replication in non-proliferating MAGI-CCR5 cells and primary macrophages. These findings suggest that Vpr-proteasome interaction might counteract certain host restriction factor(s to stimulate viral replication in non-dividing cells.

HIV-1 replication in cell lines harboring INI1/hSNF5 mutations

Directory of Open Access Journals (Sweden)

Wu Xuhong

2006-08-01

Full Text Available Abstract Background INI1/hSNF5 is a cellular protein that directly interacts with HIV-1 integrase (IN. It is specifically incorporated into HIV-1 virions. A dominant negative mutant derived from INI1 inhibits HIV-1 replication. Recent studies indicate that INI1 is associated with pre-integration and reverse transcription complexes that are formed upon viral entry into the target cells. INI1 also is a tumor suppressor, biallelically deleted/mutated in malignant rhabdoid tumors. We have utilized cell lines derived from the rhabdoid tumors, MON and STA-WT1, that harbor either null or truncating mutations of INI1 respectively, to assess the effect of INI1 on HIV-1 replication. Results We found that while HIV-1 virions produced in 293T cells efficiently transduced MON and STA-WT1 cells, HIV-1 particle production was severely reduced in both of these cells. Reintroduction of INI1 into MON and STA-WT1 significantly enhanced the particle production in both cell lines. HIV-1 particles produced in MON cells were reduced for infectivity, while those produced in STA-WT1 were not. Further analysis indicated the presence of INI1 in those virions produced from STA-WT1 but not from those produced from MON cells. HIV-1 produced in MON cells were defective for synthesis of early and late reverse transcription products in the target cells. Furthermore, virions produced in MON cells were defective for exogenous reverse transcriptase activity carried out using exogenous template, primer and substrate. Conclusion Our results suggest that INI1-deficient cells exhibit reduced particle production that can be partly enhanced by re-introduction of INI1. Infectivity of HIV-1 produced in some but not all INI1 defective cells, is affected and this defect may correlate to the lack of INI1 and/or some other proteins in these virions. The block in early events of virion produced from MON cells appears to be at the stage of reverse transcription. These studies suggest that

HIV-1 replication in cell lines harboring INI1/hSNF5 mutations.

Science.gov (United States)

Sorin, Masha; Yung, Eric; Wu, Xuhong; Kalpana, Ganjam V

2006-08-31

INI1/hSNF5 is a cellular protein that directly interacts with HIV-1 integrase (IN). It is specifically incorporated into HIV-1 virions. A dominant negative mutant derived from INI1 inhibits HIV-1 replication. Recent studies indicate that INI1 is associated with pre-integration and reverse transcription complexes that are formed upon viral entry into the target cells. INI1 also is a tumor suppressor, biallelically deleted/mutated in malignant rhabdoid tumors. We have utilized cell lines derived from the rhabdoid tumors, MON and STA-WT1, that harbor either null or truncating mutations of INI1 respectively, to assess the effect of INI1 on HIV-1 replication. We found that while HIV-1 virions produced in 293T cells efficiently transduced MON and STA-WT1 cells, HIV-1 particle production was severely reduced in both of these cells. Reintroduction of INI1 into MON and STA-WT1 significantly enhanced the particle production in both cell lines. HIV-1 particles produced in MON cells were reduced for infectivity, while those produced in STA-WT1 were not. Further analysis indicated the presence of INI1 in those virions produced from STA-WT1 but not from those produced from MON cells. HIV-1 produced in MON cells were defective for synthesis of early and late reverse transcription products in the target cells. Furthermore, virions produced in MON cells were defective for exogenous reverse transcriptase activity carried out using exogenous template, primer and substrate. Our results suggest that INI1-deficient cells exhibit reduced particle production that can be partly enhanced by re-introduction of INI1. Infectivity of HIV-1 produced in some but not all INI1 defective cells, is affected and this defect may correlate to the lack of INI1 and/or some other proteins in these virions. The block in early events of virion produced from MON cells appears to be at the stage of reverse transcription. These studies suggest that presence of INI1 or some other host factor in virions and

Potent inhibition of HIV-1 replication by a Tat mutant.

Directory of Open Access Journals (Sweden)

Luke W Meredith

Full Text Available Herein we describe a mutant of the two-exon HIV-1 Tat protein, termed Nullbasic, that potently inhibits multiple steps of the HIV-1 replication cycle. Nullbasic was created by replacing the entire arginine-rich basic domain of wild type Tat with glycine/alanine residues. Like similarly mutated one-exon Tat mutants, Nullbasic exhibited transdominant negative effects on Tat-dependent transactivation. However, unlike previously reported mutants, we discovered that Nullbasic also strongly suppressed the expression of unspliced and singly-spliced viral mRNA, an activity likely caused by redistribution and thus functional inhibition of HIV-1 Rev. Furthermore, HIV-1 virion particles produced by cells expressing Nullbasic had severely reduced infectivity, a defect attributable to a reduced ability of the virions to undergo reverse transcription. Combination of these inhibitory effects on transactivation, Rev-dependent mRNA transport and reverse transcription meant that permissive cells constitutively expressing Nullbasic were highly resistant to a spreading infection by HIV-1. Nullbasic and its activities thus provide potential insights into the development of potent antiviral therapeutics that target multiple stages of HIV-1 infection.

Ruxolitinib and Tofacitinib Are Potent and Selective Inhibitors of HIV-1 Replication and Virus Reactivation In Vitro

Science.gov (United States)

Gavegnano, Christina; Detorio, Mervi; Montero, Catherine; Bosque, Alberto; Planelles, Vicente

2014-01-01

The JAK-STAT pathway is activated in both macrophages and lymphocytes upon human immunodeficiency virus type 1 (HIV-1) infection and thus represents an attractive cellular target to achieve HIV suppression and reduced inflammation, which may impact virus sanctuaries. Ruxolitinib and tofacitinib are JAK1/2 inhibitors that are FDA approved for rheumatoid arthritis and myelofibrosis, respectively, but their therapeutic application for treatment of HIV infection was unexplored. Both drugs demonstrated submicromolar inhibition of infection with HIV-1, HIV-2, and a simian-human immunodeficiency virus, RT-SHIV, across primary human or rhesus macaque lymphocytes and macrophages, with no apparent significant cytotoxicity at 2 to 3 logs above the median effective antiviral concentration. Combination of tofacitinib and ruxolitinib increased the efficacy by 53- to 161-fold versus that observed for monotherapy, respectively, and each drug applied alone to primary human lymphocytes displayed similar efficacy against HIV-1 containing various polymerase substitutions. Both drugs inhibited virus replication in lymphocytes stimulated with phytohemagglutinin (PHA) plus interleukin-2 (IL-2), but not PHA alone, and inhibited reactivation of latent HIV-1 at low-micromolar concentrations across the J-Lat T cell latency model and in primary human central memory lymphocytes. Thus, targeted inhibition of JAK provided a selective, potent, and novel mechanism to inhibit HIV-1 replication in lymphocytes and macrophages, replication of drug-resistant HIV-1, and reactivation of latent HIV-1 and has the potential to reset the immunologic milieu in HIV-infected individuals. PMID:24419350

Vaginal Lactobacillus Inhibits HIV-1 Replication in Human Tissues Ex Vivo

Directory of Open Access Journals (Sweden)

Rogers A. Ñahui Palomino

2017-05-01

Full Text Available Lactobacillus species, which dominate vaginal microbiota of healthy reproductive-age women, lower the risks of sexually transmitted infections, including the risk of human immunodeficiency virus (HIV acquisition. The exact mechanisms of this protection remain to be understood. Here, we investigated these mechanisms in the context of human cervico-vaginal and lymphoid tissues ex vivo. We found that all six Lactobacillus strains tested in these systems significantly suppressed HIV type-1 (HIV-1 infection. We identified at least three factors that mediated this suppression: (i Acidification of the medium. The pH of the undiluted medium conditioned by lactobacilli was between 3.8 and 4.6. Acidification of the culture medium with hydrochloric acid (HCl to this pH in control experiments was sufficient to abrogate HIV-1 replication. However, the pH of the Lactobacillus-conditioned medium (CM diluted fivefold, which reached ∼6.9, was also suppressive for HIV-1 infection, while in control experiments HIV-1 infection was not abrogated when the pH of the medium was brought to 6.9 through the use of HCl. This suggested the existence of other factors responsible for HIV-1 inhibition by lactobacilli. (ii Lactic acid. There was a correlation between the concentration of lactic acid in the Lactobacillus-CM and its ability to suppress HIV-1 infection in human tissues ex vivo. Addition of lactic acid isomers D and L to tissue culture medium at the concentration that corresponded to their amount released by lactobacilli resulted in HIV-1 inhibition. Isomer L was produced in higher quantities than isomer D and was mostly responsible for HIV-1 inhibition. These results indicate that lactic acid, in particular its L-isomer, inhibits HIV-1 independently of lowering of the pH. (iii Virucidal effect. Incubation of HIV-1 in Lactobacillus-CM significantly suppressed viral infectivity for human tissues ex vivo. Finally, lactobacilli adsorb HIV-1, serving as a sink

Cellular specificity of HIV-1 replication can be controlled by LTR sequences

International Nuclear Information System (INIS)

Reed-Inderbitzin, Edward; Maury, Wendy

2003-01-01

Two well-established determinants of retroviral tropism are envelope sequences that regulate entry and LTR sequences that can regulate viral expression in a cell-specific manner. Studies with human immunodeficiency virus-1 (HIV-1) have demonstrated that tropism of this virus maps primarily to variable envelope sequences. Studies have demonstrated that T cell and macrophage-specific transcription factor binding motifs exist in the upstream region of the LTR U3; however, the ability of the core enhancer/promoter proximal elements (two NF-κB and three Sp1 sites) to function well in macrophages and T cells have led many to conclude that HIV LTR sequences are not primary determinants of HIV tropism. To determine if cellular specificity could be imparted to HIV by the core enhancer elements, the enhancer/promoter proximal region of the HIV LTR was substituted with motifs that control gene expression in a myeloid-specific manner. The enhancer region from equine infectious anemia virus (EIAV) when substituted for the HIV enhancer/promoter proximal region was found to drive expression in a macrophage-specific manner and was responsive to HIV Tat. The addition of a 5' methylation-dependent binding site (MDBP) and a promoter proximal Sp1 motif increased expression without altering cellular specificity. Spacing between the promoter proximal region and the TATA box was also found to influence LTR activity. Infectivity studies using chimeric LTRs within the context of a dual-tropic infectious molecular clone established that these LTRs directed HIV replication and production of infectious virions in macrophages but not primary T cells or T cell lines. This investigation demonstrates that cellular specificity can be imparted onto HIV-1 replication at the level of viral transcription and not entry

An integrated chemical biology approach reveals the mechanism of action of HIV replication inhibitors.

Science.gov (United States)

Pagano, Nicholas; Teriete, Peter; Mattmann, Margrith E; Yang, Li; Snyder, Beth A; Cai, Zhaohui; Heil, Marintha L; Cosford, Nicholas D P

2017-12-01

Continuous flow (microfluidic) chemistry was employed to prepare a small focused library of dihydropyrimidinone (DHPM) derivatives. Compounds in this class have been reported to exhibit activity against the human immunodeficiency virus (HIV), but their molecular target had not been identified. We tested the initial set of DHPMs in phenotypic assays providing a hit (1i) that inhibited the replication of the human immunodeficiency virus HIV in cells. Flow chemistry-driven optimization of 1i led to the identification of HIV replication inhibitors such as 1l with cellular potency comparable with the clinical drug nevirapine (NVP). Mechanism of action (MOA) studies using cellular and biochemical assays coupled with 3D fingerprinting and in silico modeling demonstrated that these drug-like probe compounds exert their effects by inhibiting the viral reverse transcriptase polymerase (RT). This led to the design and synthesis of the novel DHPM 1at that inhibits the replication of drug resistant strains of HIV. Our work demonstrates that combining flow chemistry-driven analogue refinement with phenotypic assays, in silico modeling and MOA studies is a highly effective strategy for hit-to-lead optimization applicable to the discovery of future therapeutic agents. Copyright © 2017. Published by Elsevier Ltd.

Effects of integration and replication on transcription of the HIV-1 long terminal repeat

NARCIS (Netherlands)

Jeang, K. T.; Berkhout, B.; Dropulic, B.

1993-01-01

The activity of a promoter is influenced by chromosomal and cell cycle/replication context. We analyzed the influences of integration and replication on transcription of the human immunodeficiency virus (HIV)-1 long terminal repeat (LTR). We found that one requirement for Tat trans-activated

On the Road to HIV/AIDS Competence in the Household: Building a Health-Enabling Environment for People Living with HIV/AIDS

Directory of Open Access Journals (Sweden)

Caroline Masquillier

2015-03-01

Full Text Available When aiming to provide chronic disease care within the context of human resource shortages, we should not only consider the responsibility of the individual person living with HIV/AIDS (PLWHA but also the capacity of the social environment to actively encourage a lifestyle that fosters health. In this social environment, extensive efforts are thus required to increase HIV/AIDS knowledge, reduce stigma, stimulate HIV testing, improve health care-seeking behavior, and encourage safe sexual practices—described in the literature as the need for AIDS competence. In accordance with socio-ecological theory, one cannot restrict the research focus to communities, as AIDS competence studies should also incorporate the intermediate household level. In responding to this research need, the aim of this article is to conceptualize an “HIV/AIDS competent household” based on qualitative interviews and focus group discussions conducted in a township on the outskirts of Cape Town, South Africa. Our results show that a household’s supportive response to disclosure allows a patient to live openly as HIV positive in the household concerned. This may mark the start of the road to HIV/AIDS competence in the household, meaning the PLWHA receives sustainable support throughout the care continuum and positive living becomes the norm for the PLWHA and his or her household. A feedback loop might also be created in which other household members are encouraged to be tested and to disclose their status, which is an important step towards a sustainable response to HIV/AIDS-related challenges. Despite the fact that this road to HIV/AIDS competence at the household level is fragile and prone to various barriers, this article shows that the household has the potential to be a health-enabling environment for PLWHA.

A stable latent reservoir for HIV-1 in resting CD4+ T lymphocytes in infected children

Science.gov (United States)

Persaud, Deborah; Pierson, Theodore; Ruff, Christian; Finzi, Diana; Chadwick, Karen R.; Margolick, Joseph B.; Ruff, Andrea; Hutton, Nancy; Ray, Stuart; Siliciano, Robert F.

2000-01-01

HIV-1 persists in a latent state in resting CD4+ T lymphocytes of infected adults despite prolonged highly active antiretroviral therapy (HAART). To determine whether a latent reservoir for HIV-1 exists in infected children, we performed a quantitative viral culture assay on highly purified resting CD4+ T cells from 21 children with perinatally acquired infection. Replication-competent HIV-1 was recovered from all 18 children from whom sufficient cells were obtained. The frequency of latently infected resting CD4+ T cells directly correlated with plasma virus levels, suggesting that in children with ongoing viral replication, most latently infected cells are in the labile preintegration state of latency. However, in each of 7 children who had suppression of viral replication to undetectable levels for 1–3 years on HAART, latent replication-competent HIV-1 persisted with little decay, owing to a stable reservoir of infected cells in the postintegration stage of latency. Drug-resistance mutations generated by previous nonsuppressive regimens persisted in this compartment despite more than 1 year of fully suppressive HAART, rendering untenable the idea of recycling drugs that were part of failed regimens. Thus the latent reservoir for HIV-1 in resting CD4+ T cells will be a major obstacle to HIV-1 eradication in children. PMID:10749578

Understanding HIV infection for the design of a therapeutic vaccine. Part II: Vaccination strategies for HIV.

Science.gov (United States)

de Goede, A L; Vulto, A G; Osterhaus, A D M E; Gruters, R A

2015-05-01

HIV infection leads to a gradual loss CD4(+) T lymphocytes comprising immune competence and progression to AIDS. Effective treatment with combined antiretroviral drugs (cART) decreases viral load below detectable levels but is not able to eliminate the virus from the body. The success of cART is frustrated by the requirement of expensive lifelong adherence, accumulating drug toxicities and chronic immune activation resulting in increased risk of several non-AIDS disorders, even when viral replication is suppressed. Therefore, there is a strong need for therapeutic strategies as an alternative to cART. Immunotherapy, or therapeutic vaccination, aims to increase existing immune responses against HIV or induce de novo immune responses. These immune responses should provide a functional cure by controlling viral replication and preventing disease progression in the absence of cART. The key difficulty in the development of an HIV vaccine is our ignorance of the immune responses that control of viral replication, and thus how these responses can be elicited and how they can be monitored. Part one of this review provides an extensive overview of the (patho-) physiology of HIV infection. It describes the structure and replication cycle of HIV, the epidemiology and pathogenesis of HIV infection and the innate and adaptive immune responses against HIV. Part two of this review discusses therapeutic options for HIV. Prevention modalities and antiretroviral therapy are briefly touched upon, after which an extensive overview on vaccination strategies for HIV is provided, including the choice of immunogens and delivery strategies. Copyright © 2014. Published by Elsevier Masson SAS.

Increased T cell trafficking as adjunct therapy for HIV-1

Science.gov (United States)

Wolinsky, Steven M.; McLean, Angela R.

2018-01-01

Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical model to illustrate a new approach to eliminating the part of the reservoir attributable to persistent replication in drug sanctuaries. Reducing the residency time of CD4 T cells in drug sanctuaries renders ongoing replication unsustainable in those sanctuaries. We hypothesize that, in combination with antiretroviral drugs, a strategy to orchestrate CD4 T cell trafficking could contribute to a functional cure for HIV-1 infection. PMID:29499057

Nef does not contribute to replication differences between R5 pre-AIDS and AIDS HIV-1 clones from patient ACH142

Directory of Open Access Journals (Sweden)

Rekosh David

2008-05-01

Full Text Available Abstract AIDS-associated, CCR5-tropic (R5 HIV-1 clones, isolated from a patient that never developed CXCR4-tropic HIV-1, replicate to a greater extent and cause greater cytopathic effects than R5 HIV-1 clones isolated before the onset of AIDS. Previously, we showed that HIV-1 Env substantially contributed to the enhanced replication of an AIDS clone. In order to determine if Nef makes a similar contribution, we cloned and phenotypically analyzed nef genes from a series of patient ACH142 derived R5 HIV-1 clones. The AIDS-associated Nef contains a series of residues found in Nef proteins from progressors 1. In contrast to other reports 123, this AIDS-associated Nef downmodulated MHC-I to a greater extent and CD4 less than pre-AIDS Nef proteins. Additionally, all Nef proteins enhanced infectivity similarly in a single round of replication. Combined with our previous study, these data show that evolution of the HIV-1 env gene, but not the nef gene, within patient ACH142 significantly contributed to the enhanced replication and cytopathic effects of the AIDS-associated R5 HIV-1 clone.

Genome-wide association study identifies single nucleotide polymorphism in DYRK1A associated with replication of HIV-1 in monocyte-derived macrophages.

Directory of Open Access Journals (Sweden)

Sebastiaan M Bol

2011-02-01

Full Text Available HIV-1 infected macrophages play an important role in rendering resting T cells permissive for infection, in spreading HIV-1 to T cells, and in the pathogenesis of AIDS dementia. During highly active anti-retroviral treatment (HAART, macrophages keep producing virus because tissue penetration of antiretrovirals is suboptimal and the efficacy of some is reduced. Thus, to cure HIV-1 infection with antiretrovirals we will also need to efficiently inhibit viral replication in macrophages. The majority of the current drugs block the action of viral enzymes, whereas there is an abundance of yet unidentified host factors that could be targeted. We here present results from a genome-wide association study identifying novel genetic polymorphisms that affect in vitro HIV-1 replication in macrophages.Monocyte-derived macrophages from 393 blood donors were infected with HIV-1 and viral replication was determined using Gag p24 antigen levels. Genomic DNA from individuals with macrophages that had relatively low (n = 96 or high (n = 96 p24 production was used for SNP genotyping with the Illumina 610 Quad beadchip. A total of 494,656 SNPs that passed quality control were tested for association with HIV-1 replication in macrophages, using linear regression. We found a strong association between in vitro HIV-1 replication in monocyte-derived macrophages and SNP rs12483205 in DYRK1A (p = 2.16 × 10(-5. While the association was not genome-wide significant (p<1 × 10(-7, we could replicate this association using monocyte-derived macrophages from an independent group of 31 individuals (p = 0.0034. Combined analysis of the initial and replication cohort increased the strength of the association (p = 4.84 × 10(-6. In addition, we found this SNP to be associated with HIV-1 disease progression in vivo in two independent cohort studies (p = 0.035 and p = 0.0048.These findings suggest that the kinase DYRK1A is involved in the replication of HIV-1, in vitro in macrophages

N-terminally truncated POM121C inhibits HIV-1 replication.

Directory of Open Access Journals (Sweden)

Hideki Saito

Full Text Available Recent studies have identified host cell factors that regulate early stages of HIV-1 infection including viral cDNA synthesis and orientation of the HIV-1 capsid (CA core toward the nuclear envelope, but it remains unclear how viral DNA is imported through the nuclear pore and guided to the host chromosomal DNA. Here, we demonstrate that N-terminally truncated POM121C, a component of the nuclear pore complex, blocks HIV-1 infection. This truncated protein is predominantly localized in the cytoplasm, does not bind to CA, does not affect viral cDNA synthesis, reduces the formation of 2-LTR and diminished the amount of integrated proviral DNA. Studies with an HIV-1-murine leukemia virus (MLV chimeric virus carrying the MLV-derived Gag revealed that Gag is a determinant of this inhibition. Intriguingly, mutational studies have revealed that the blockade by N-terminally-truncated POM121C is closely linked to its binding to importin-β/karyopherin subunit beta 1 (KPNB1. These results indicate that N-terminally-truncated POM121C inhibits HIV-1 infection after completion of reverse transcription and before integration, and suggest an important role for KPNB1 in HIV-1 replication.

Understanding HIV infection for the design of a therapeutic vaccine. Part I: Epidemiology and pathogenesis of HIV infection.

Science.gov (United States)

de Goede, A L; Vulto, A G; Osterhaus, A D M E; Gruters, R A

2015-03-01

HIV infection leads to a gradual loss CD4+ T lymphocytes comprising immune competence and progression to AIDS. Effective treatment with combined antiretroviral drugs (cART) decreases viral load below detectable levels but is not able to eliminate the virus from the body. The success of cART is frustrated by the requirement of expensive life-long adherence, accumulating drug toxicities and chronic immune activation resulting in increased risk of several non-AIDS disorders, even when viral replication is suppressed. Therefore there is a strong need for therapeutic strategies as an alternative to cART. Immunotherapy, or therapeutic vaccination, aims to increase existing immune responses against HIV or induce de novo immune responses. These immune responses should provide a functional cure by controlling viral replication and preventing disease progression in the absence of cART. The key difficulty in the development of an HIV vaccine is our ignorance of the immune responses that control of viral replication, and thus how these responses can be elicited and how they can be monitored. Part one of this review provides an extensive overview of the (patho-) physiology of HIV infection. It describes the structure and replication cycle of HIV, the epidemiology and pathogenesis of HIV infection and the innate and adaptive immune responses against HIV. Part two of this review discusses therapeutic options for HIV. Prevention modalities and antiretroviral therapy are briefly touched upon, after which an extensive overview on vaccination strategies for HIV is provided, including the choice of immunogens and delivery strategies. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Reduced evolutionary rates in HIV-1 reveal extensive latency periods among replicating lineages.

Science.gov (United States)

Immonen, Taina T; Leitner, Thomas

2014-10-16

HIV-1 can persist for the duration of a patient's life due in part to its ability to hide from the immune system, and from antiretroviral drugs, in long-lived latent reservoirs. Latent forms of HIV-1 may also be disproportionally involved in transmission. Thus, it is important to detect and quantify latency in the HIV-1 life cycle. We developed a novel molecular clock-based phylogenetic tool to investigate the prevalence of HIV-1 lineages that have experienced latency. The method removes alternative sources that may affect evolutionary rates, such as hypermutation, recombination, and selection, to reveal the contribution of generation-time effects caused by latency. Our method was able to recover latent lineages with high specificity and sensitivity, and low false discovery rates, even on relatively short branches on simulated phylogenies. Applying the tool to HIV-1 sequences from 26 patients, we show that the majority of phylogenetic lineages have been affected by generation-time effects in every patient type, whether untreated, elite controller, or under effective or failing treatment. Furthermore, we discovered extensive effects of latency in sequence data (gag, pol, and env) from reservoirs as well as in the replicating plasma population. To better understand our phylogenetic findings, we developed a dynamic model of virus-host interactions to investigate the proportion of lineages in the actively replicating population that have ever been latent. Assuming neutral evolution, our dynamic modeling showed that under most parameter conditions, it is possible for a few activated latent viruses to propagate so that in time, most HIV-1 lineages will have been latent at some time in their past. These results suggest that cycling in and out of latency plays a major role in the evolution of HIV-1. Thus, no aspect of HIV-1 evolution can be fully understood without considering latency - including treatment, drug resistance, immune evasion, transmission, and pathogenesis.

Genome-Wide Association Study Identifies Single Nucleotide Polymorphism in DYRK1A Associated with Replication of HIV-1 in Monocyte-Derived Macrophages

Science.gov (United States)

Bol, Sebastiaan M.; Moerland, Perry D.; Limou, Sophie; van Remmerden, Yvonne; Coulonges, Cédric; van Manen, Daniëlle; Herbeck, Joshua T.; Fellay, Jacques; Sieberer, Margit; Sietzema, Jantine G.; van 't Slot, Ruben; Martinson, Jeremy; Zagury, Jean-François; Schuitemaker, Hanneke; van 't Wout, Angélique B.

2011-01-01

Background HIV-1 infected macrophages play an important role in rendering resting T cells permissive for infection, in spreading HIV-1 to T cells, and in the pathogenesis of AIDS dementia. During highly active anti-retroviral treatment (HAART), macrophages keep producing virus because tissue penetration of antiretrovirals is suboptimal and the efficacy of some is reduced. Thus, to cure HIV-1 infection with antiretrovirals we will also need to efficiently inhibit viral replication in macrophages. The majority of the current drugs block the action of viral enzymes, whereas there is an abundance of yet unidentified host factors that could be targeted. We here present results from a genome-wide association study identifying novel genetic polymorphisms that affect in vitro HIV-1 replication in macrophages. Methodology/Principal Findings Monocyte-derived macrophages from 393 blood donors were infected with HIV-1 and viral replication was determined using Gag p24 antigen levels. Genomic DNA from individuals with macrophages that had relatively low (n = 96) or high (n = 96) p24 production was used for SNP genotyping with the Illumina 610 Quad beadchip. A total of 494,656 SNPs that passed quality control were tested for association with HIV-1 replication in macrophages, using linear regression. We found a strong association between in vitro HIV-1 replication in monocyte-derived macrophages and SNP rs12483205 in DYRK1A (p = 2.16×10−5). While the association was not genome-wide significant (p<1×10−7), we could replicate this association using monocyte-derived macrophages from an independent group of 31 individuals (p = 0.0034). Combined analysis of the initial and replication cohort increased the strength of the association (p = 4.84×10−6). In addition, we found this SNP to be associated with HIV-1 disease progression in vivo in two independent cohort studies (p = 0.035 and p = 0.0048). Conclusions/Significance These findings suggest that

Human prostate supports more efficient replication of HIV-1 R5 than X4 strains ex vivo

Directory of Open Access Journals (Sweden)

Denis Hélène

2008-12-01

Full Text Available Abstract Background In order to determine whether human prostate can be productively infected by HIV-1 strains with different tropism, and thus represent a potential source of HIV in semen, an organotypic culture of prostate from men undergoing prostatic adenomectomy for benign prostate hypertrophy (BPH was developed. The presence of potential HIV target cells in prostate tissues was investigated using immunohistochemistry. The infection of prostate explants following exposures with HIV-1 R5, R5X4 and X4 strains was analyzed through the measure of RT activity in culture supernatants, the quantification of HIV DNA in the explants and the detection of HIV RNA+ cells in situ. Results The overall prostate characteristics were retained for 21/2 weeks in culture. Numerous potential HIV-1 target cells were detected in the prostate stroma. Whilst HIV-1 R5SF162 strain consistently productively infected prostatic T lymphocytes and macrophages, the prototypic X4IIIB strain and a primary R5X4 strain showed less efficient replication in this organ. Conclusion The BPH prostate is a site of HIV-1 R5 replication that could contribute virus to semen. A limited spreading of HIV-1 X4 and R5X4 in this organ could participate to the preferential sexual transmission of HIV-1 R5 strains.

Effects of Cigarette Smoke Condensate on Oxidative Stress, Apoptotic Cell Death, and HIV Replication in Human Monocytic Cells.

Directory of Open Access Journals (Sweden)

Pss Rao

Full Text Available While cigarette smoking is prevalent amongst HIV-infected patients, the effects of cigarette smoke constituents in cells of myeloid lineage are poorly known. Recently, we have shown that nicotine induces oxidative stress through cytochrome P450 (CYP 2A6-mediated pathway in U937 monocytic cells. The present study was designed to examine the effect of cigarette smoke condensate (CSC, which contains majority of tobacco constituents, on oxidative stress, cytotoxicity, expression of CYP1A1, and/or HIV-1 replication in HIV-infected (U1 and uninfected U937 cells. The effects of CSC on induction of CYP1 enzymes in HIV-infected primary macrophages were also analyzed. The results showed that the CSC-mediated increase in production of reactive oxygen species (ROS in U937 cells is dose- and time-dependent. Moreover, CSC treatment was found to induce cytotoxicity in U937 cells through the apoptotic pathway via activation of caspase-3. Importantly, pretreatment with vitamin C blocked the CSC-mediated production of ROS and induction of caspase-3 activity. In U1 cells, acute treatment of CSC increased ROS production at 6H (>2-fold and both ROS (>2 fold and HIV-1 replication (>3-fold after chronic treatment. The CSC mediated effects were associated with robust induction in the expression of CYP1A1 mRNA upon acute CSC treatment of U937 and U1 cells (>20-fold, and upon chronic CSC treatment to U1 cells (>30-fold. In addition, the CYP1A1 induction in U937 cells was mediated through the aromatic hydrocarbon receptor pathway. Lastly, CSC, which is known to increase viral replication in primary macrophages, was also found to induce CYP1 enzymes in HIV-infected primary macrophages. While mRNA levels of both CYP1A1 and CYP1B1 were elevated following CSC treatment, only CYP1B1 protein levels were increased in HIV-infected primary macrophages. In conclusion, these results suggest a possible association between oxidative stress, CYP1 expression, and viral replication in

Ebselen, a Small-Molecule Capsid Inhibitor of HIV-1 Replication.

Science.gov (United States)

Thenin-Houssier, Suzie; de Vera, Ian Mitchelle S; Pedro-Rosa, Laura; Brady, Angela; Richard, Audrey; Konnick, Briana; Opp, Silvana; Buffone, Cindy; Fuhrmann, Jakob; Kota, Smitha; Billack, Blase; Pietka-Ottlik, Magdalena; Tellinghuisen, Timothy; Choe, Hyeryun; Spicer, Timothy; Scampavia, Louis; Diaz-Griffero, Felipe; Kojetin, Douglas J; Valente, Susana T

2016-04-01

The human immunodeficiency virus type 1 (HIV-1) capsid plays crucial roles in HIV-1 replication and thus represents an excellent drug target. We developed a high-throughput screening method based on a time-resolved fluorescence resonance energy transfer (HTS-TR-FRET) assay, using the C-terminal domain (CTD) of HIV-1 capsid to identify inhibitors of capsid dimerization. This assay was used to screen a library of pharmacologically active compounds, composed of 1,280in vivo-active drugs, and identified ebselen [2-phenyl-1,2-benzisoselenazol-3(2H)-one], an organoselenium compound, as an inhibitor of HIV-1 capsid CTD dimerization. Nuclear magnetic resonance (NMR) spectroscopic analysis confirmed the direct interaction of ebselen with the HIV-1 capsid CTD and dimer dissociation when ebselen is in 2-fold molar excess. Electrospray ionization mass spectrometry revealed that ebselen covalently binds the HIV-1 capsid CTD, likely via a selenylsulfide linkage with Cys198 and Cys218. This compound presents anti-HIV activity in single and multiple rounds of infection in permissive cell lines as well as in primary peripheral blood mononuclear cells. Ebselen inhibits early viral postentry events of the HIV-1 life cycle by impairing the incoming capsid uncoating process. This compound also blocks infection of other retroviruses, such as Moloney murine leukemia virus and simian immunodeficiency virus, but displays no inhibitory activity against hepatitis C and influenza viruses. This study reports the use of TR-FRET screening to successfully identify a novel capsid inhibitor, ebselen, validating HIV-1 capsid as a promising target for drug development. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

HIV/AIDS Competent Households: Interaction between a Health-Enabling Environment and Community-Based Treatment Adherence Support for People Living with HIV/AIDS in South Africa.

Science.gov (United States)

Masquillier, Caroline; Wouters, Edwin; Mortelmans, Dimitri; van Wyk, Brian; Hausler, Harry; Van Damme, Wim

2016-01-01

In the context of severe human resource shortages in HIV care, task-shifting and especially community-based support are increasingly being cited as potential means of providing durable care to chronic HIV patients. Socio-ecological theory clearly stipulates that-in all social interventions-the interrelatedness and interdependency between individuals and their immediate social contexts should be taken into account. People living with HIV/AIDS (PLWHA) seldom live in isolation, yet community-based interventions for supporting chronic HIV patients have largely ignored the social contexts in which they are implemented. Research is thus required to investigate such community-based support within its context. The aim of this study is to address this research gap by examining the way in which HIV/AIDS competence in the household hampers or facilitates community-based treatment adherence support. The data was analyzed carefully in accordance with the Grounded Theory procedures, using Nvivo 10. More specifically, we analyzed field notes from participatory observations conducted during 48 community-based treatment adherence support sessions in townships on the outskirts of Cape Town, transcripts of 32 audio-recorded in-depth interviews with PLWHA and transcripts of 4 focus group discussions with 36 community health workers (CHWs). Despite the fact that the CHWs try to present themselves as not being openly associated with HIV/AIDS services, results show that the presence of a CHW is often seen as a marker of the disease. Depending on the HIV/AIDS competence in the household, this association can challenge the patient's hybrid identity management and his/her attempt to regulate the interference of the household in the disease management. The results deepen our understanding of how the degree of HIV/AIDS competence present in a PLWHA's household affects the manner in which the CHW can perform his or her job and the associated benefits for the patient and his/her household

HIV/AIDS Competent Households: Interaction between a Health-Enabling Environment and Community-Based Treatment Adherence Support for People Living with HIV/AIDS in South Africa.

Directory of Open Access Journals (Sweden)

Caroline Masquillier

Full Text Available In the context of severe human resource shortages in HIV care, task-shifting and especially community-based support are increasingly being cited as potential means of providing durable care to chronic HIV patients. Socio-ecological theory clearly stipulates that-in all social interventions-the interrelatedness and interdependency between individuals and their immediate social contexts should be taken into account. People living with HIV/AIDS (PLWHA seldom live in isolation, yet community-based interventions for supporting chronic HIV patients have largely ignored the social contexts in which they are implemented. Research is thus required to investigate such community-based support within its context. The aim of this study is to address this research gap by examining the way in which HIV/AIDS competence in the household hampers or facilitates community-based treatment adherence support. The data was analyzed carefully in accordance with the Grounded Theory procedures, using Nvivo 10. More specifically, we analyzed field notes from participatory observations conducted during 48 community-based treatment adherence support sessions in townships on the outskirts of Cape Town, transcripts of 32 audio-recorded in-depth interviews with PLWHA and transcripts of 4 focus group discussions with 36 community health workers (CHWs. Despite the fact that the CHWs try to present themselves as not being openly associated with HIV/AIDS services, results show that the presence of a CHW is often seen as a marker of the disease. Depending on the HIV/AIDS competence in the household, this association can challenge the patient's hybrid identity management and his/her attempt to regulate the interference of the household in the disease management. The results deepen our understanding of how the degree of HIV/AIDS competence present in a PLWHA's household affects the manner in which the CHW can perform his or her job and the associated benefits for the patient and his

Low-level HIV-1 replication and the dynamics of the resting CD4+ T cell reservoir for HIV-1 in the setting of HAART

Directory of Open Access Journals (Sweden)

Wilke Claus O

2008-01-01

Full Text Available Abstract Background In the setting of highly active antiretroviral therapy (HAART, plasma levels of human immunodeficiency type-1 (HIV-1 rapidly decay to below the limit of detection of standard clinical assays. However, reactivation of remaining latently infected memory CD4+ T cells is a source of continued virus production, forcing patients to remain on HAART despite clinically undetectable viral loads. Unfortunately, the latent reservoir decays slowly, with a half-life of up to 44 months, making it the major known obstacle to the eradication of HIV-1 infection. However, the mechanism underlying the long half-life of the latent reservoir is unknown. The most likely potential mechanisms are low-level viral replication and the intrinsic stability of latently infected cells. Methods Here we use a mathematical model of T cell dynamics in the setting of HIV-1 infection to probe the decay characteristics of the latent reservoir upon initiation of HAART. We compare the behavior of this model to patient derived data in order to gain insight into the role of low-level viral replication in the setting of HAART. Results By comparing the behavior of our model to patient derived data, we find that the viral dynamics observed in patients on HAART could be consistent with low-level viral replication but that this replication would not significantly affect the decay rate of the latent reservoir. Rather than low-level replication, the intrinsic stability of latently infected cells and the rate at which they are reactivated primarily determine the observed reservoir decay rate according to the predictions of our model. Conclusion The intrinsic stability of the latent reservoir has important implications for efforts to eradicate HIV-1 infection and suggests that intensified HAART would not accelerate the decay of the latent reservoir.

Low-level HIV-1 replication and the dynamics of the resting CD4+ T cell reservoir for HIV-1 in the setting of HAART

Science.gov (United States)

Sedaghat, Ahmad R; Siliciano, Robert F; Wilke, Claus O

2008-01-01

Background In the setting of highly active antiretroviral therapy (HAART), plasma levels of human immunodeficiency type-1 (HIV-1) rapidly decay to below the limit of detection of standard clinical assays. However, reactivation of remaining latently infected memory CD4+ T cells is a source of continued virus production, forcing patients to remain on HAART despite clinically undetectable viral loads. Unfortunately, the latent reservoir decays slowly, with a half-life of up to 44 months, making it the major known obstacle to the eradication of HIV-1 infection. However, the mechanism underlying the long half-life of the latent reservoir is unknown. The most likely potential mechanisms are low-level viral replication and the intrinsic stability of latently infected cells. Methods Here we use a mathematical model of T cell dynamics in the setting of HIV-1 infection to probe the decay characteristics of the latent reservoir upon initiation of HAART. We compare the behavior of this model to patient derived data in order to gain insight into the role of low-level viral replication in the setting of HAART. Results By comparing the behavior of our model to patient derived data, we find that the viral dynamics observed in patients on HAART could be consistent with low-level viral replication but that this replication would not significantly affect the decay rate of the latent reservoir. Rather than low-level replication, the intrinsic stability of latently infected cells and the rate at which they are reactivated primarily determine the observed reservoir decay rate according to the predictions of our model. Conclusion The intrinsic stability of the latent reservoir has important implications for efforts to eradicate HIV-1 infection and suggests that intensified HAART would not accelerate the decay of the latent reservoir. PMID:18171475

Novel Latency Reversal Agents for HIV-1 Cure.

Science.gov (United States)

Spivak, Adam M; Planelles, Vicente

2018-01-29

Antiretroviral therapy (ART) has rendered HIV-1 infection a treatable illness; however, ART is not curative owing to the persistence of replication-competent, latent proviruses in long-lived resting T cells. Strategies that target these latently infected cells and allow immune recognition and clearance of this reservoir will be necessary to eradicate HIV-1 in infected individuals. This review describes current pharmacologic approaches to reactivate the latent reservoir so that infected cells can be recognized and targeted, with the ultimate goal of achieving an HIV-1 cure.

Effects of root, shoot, leaf and seed extracts of seven Artemisia species on HIV-1 replication and CD4 expression

Directory of Open Access Journals (Sweden)

Hassan Mohabatkar

2015-12-01

Full Text Available Objective: To investigate the effects of flower, leaf, shoot and root extracts of seven Artemisia species on peripheral blood mononuclear cells (PBMCs toxicity and HIV-1 replication. Methods: The studied Artemisia species were Artemisia absinthium, Artemisia khorasanica, Artemisia deserti, Artemisia fragrans, Artemisia aucheri, Artemisia sieberi and Artemisia vulgaris. The activity of these plant extracts on HIV-1 replication and CD4 expression was performed by HIV-1 p24 antigen kit and flow cytometry respectively. Results: The results demonstrated that flower extracts of all species increased PBMCs number more than shoot, leaf and root extracts. However, the frequency of CD4 expression in PBMC was not increased in the presence of all flower extracts. The flower extracts of all species had inhibitory effect on HIV-1 replication. Conclusions: In conclusion, the results demonstrated that flower extracts of Artemisia species are good candidates for further studies as anticancer agents.

Circulating Cxcr5-Expressing Cd8+T-Cells are Major Producers of Il-21 and Associate with Limited Hiv Replication.

Science.gov (United States)

Perdomo-Celis, Federico; Taborda, Natalia A; Rugeles, Maria T

2018-04-10

Despite advances made with the highly active anti-retroviral therapy (HAART) in the control of the human immunodeficiency virus 1 (HIV) infection, a cure has not been achieved due to the persistence of viral reservoirs. The major HIV reservoirs remain in the lymphoid follicles due to, among other factors, the partial absence of CD8T-cells in these structures. Recently, lymphoid follicle-confined and circulating CD8T-cells expressing the C-X-C chemokine receptor type 5 (CXCR5) were described, possessing antiviral mechanisms which could help to control HIV replication. and methods: By flow cytometry, we characterized the phenotype and function of circulating CXCR5-expressing CD8T-cells in HIV-infected patients with natural or HAART-induced control of HIV replication. Circulating CXCR5-expressing CD8T-cells exhibited low or null expression of the C-C chemokine receptor type 7 (CCR7) and had a transitional memory phenotype. Particular redistributions of CXCR5-expressing CD8T-cells were found in HIV-infected patients, and they were partially restored by HAART. The frequency of CXCR5CCR7CD8T-cells was higher in spontaneous HIV controllers and negatively correlated with plasma HIV RNA levels. Total and HIV-specific CXCR5CD8T-cells were major producers of interleukin-21, and this function was positively associated with their interferon-γ production. Circulating CXCR5-expressing CD8T-cells are associated with low level HIV replication, could be novel correlates of protection and potentially useful in the eradication of HIV reservoirs.

Potent Inhibition of HIV-1 Replication in Resting CD4 T Cells by Resveratrol and Pterostilbene.

Science.gov (United States)

Chan, Chi N; Trinité, Benjamin; Levy, David N

2017-09-01

HIV-1 infection of resting CD4 T cells plays a crucial and numerically dominant role during virus transmission at mucosal sites and during subsequent acute replication and T cell depletion. Resveratrol and pterostilbene are plant stilbenoids associated with several health-promoting benefits. Resveratrol has been shown to inhibit the replication of several viruses, including herpes simplex viruses 1 and 2, papillomaviruses, severe acute respiratory syndrome virus, and influenza virus. Alone, resveratrol does not inhibit HIV-1 infection of activated T cells, but it does synergize with nucleoside reverse transcriptase inhibitors in these cells to inhibit reverse transcription. Here, we demonstrate that resveratrol and pterostilbene completely block HIV-1 infection at a low micromolar dose in resting CD4 T cells, primarily at the reverse transcription step. The anti-HIV effect was fully reversed by exogenous deoxynucleosides and Vpx, an HIV-1 and simian immunodeficiency virus protein that increases deoxynucleoside triphosphate (dNTP) levels. These findings are consistent with the reported ability of resveratrol to inhibit ribonucleotide reductase and to lower dNTP levels in cells. This study supports the potential use of resveratrol, pterostilbene, or related compounds as adjuvants in anti-HIV preexposure prophylaxis (PrEP) formulations. Copyright © 2017 American Society for Microbiology.

HIV Persistence in Adipose Tissue Reservoirs.

Science.gov (United States)

Couturier, Jacob; Lewis, Dorothy E

2018-02-01

The purpose of this review is to examine the evidence describing adipose tissue as a reservoir for HIV-1 and how this often expansive anatomic compartment contributes to HIV persistence. Memory CD4 T cells and macrophages, the major host cells for HIV, accumulate in adipose tissue during HIV/SIV infection of humans and rhesus macaques. Whereas HIV and SIV proviral DNA is detectable in CD4 T cells of multiple fat depots in virtually all infected humans and monkeys examined, viral RNA is less frequently detected, and infected macrophages may be less prevalent in adipose tissue. However, based on viral outgrowth assays, adipose-resident CD4 T cells are latently infected with virus that is replication-competent and infectious. Additionally, adipocytes interact with CD4 T cells and macrophages to promote immune cell activation and inflammation which may be supportive for HIV persistence. Antiviral effector cells, such as CD8 T cells and NK/NKT cells, are abundant in adipose tissue during HIV/SIV infection and typically exceed CD4 T cells, whereas B cells are largely absent from adipose tissue of humans and monkeys. Additionally, CD8 T cells in adipose tissue of HIV patients are activated and have a late differentiated phenotype, with unique TCR clonotypes of less diversity relative to blood CD8 T cells. With respect to the distribution of antiretroviral drugs in adipose tissue, data is limited, but there may be class-specific penetration of fat depots. The trafficking of infected immune cells within adipose tissues is a common event during HIV/SIV infection of humans and monkeys, but the virus may be mostly transcriptionally dormant. Viral replication may occur less in adipose tissue compared to other major reservoirs, such as lymphoid tissue, but replication competence and infectiousness of adipose latent virus are comparable to other tissues. Due to the ubiquitous nature of adipose tissue, inflammatory interactions among adipocytes and CD4 T cells and macrophages, and

Comparative evaluation of oral and intranasal priming with replication-competent adenovirus 5 host range mutant (Ad5hr)-simian immunodeficiency virus (SIV) recombinant vaccines on immunogenicity and protective efficacy against SIV(mac251).

Science.gov (United States)

Zhou, Qifeng; Hidajat, Rachmat; Peng, Bo; Venzon, David; Aldrich, M Kristine; Richardson, Ersell; Lee, Eun Mi; Kalyanaraman, V S; Grimes, George; Gómez-Román, V Raúl; Summers, L Ebonita; Malkevich, Nina; Robert-Guroff, Marjorie

2007-11-19

Oral, replication-competent Ad-HIV vaccines are advancing to human trials. Previous evaluation of protective efficacy in non-human primates has primarily followed upper respiratory tract administrations. Here we compared sequential oral (O/O) versus intranasal/oral (I/O) priming of rhesus macaques with Ad5 host range mutant-SIV recombinants expressing SIV env/rev, gag, and nef genes followed by boosting with SIV gp120 protein. Cellular immune responses in PBMC were stronger and more frequent after I/O administration. Both groups developed mucosal immunity, including memory cells in bronchial alveolar lavage, and gut-homing receptors on PBMC. Following intrarectal SIV(mac251) challenge, both groups exhibited equivalent, significant protection and robust post-challenge cellular immunity. Our results illustrate the promise of oral replication-competent Ad-recombinant vaccines. Pre-challenge PBMC ELISPOT and proliferative responses did not predict protection in the O/O group, highlighting the need for simple, non-invasive methods to reliably assess mucosal immunity.

Validation of the integration of HIV and AIDS related nursing competencies into the undergraduate nursing curriculum in South Africa

Directory of Open Access Journals (Sweden)

Regis R. Marie Modeste

2015-12-01

Full Text Available Background: Being in its fourth decade, HIV remains an epidemic that requires combined efforts for the global fight. The strategies planned and implemented in the fight against HIV include reversing and halting the spread of HIV, increasing health care access, and strengthening the health care system. South Africa has made the fight one of its top priorities, and has developed plans to increase the role of nurses in the management of HIV, demonstrating its willingness, commitment and progress in the fight against HIV. Objective: This article presents the validation process conducted to confirm the integration and mapping of the HIV and AIDS related nursing competencies into the four-year Bachelor of Nursing programme at a university in South Africa. Methods: This study adopted a constructivist paradigm, using a qualitative approach, applyingthe design step of the process model of curriculum development, to validate the inte gration of the mapped HIV and AIDS related nursing competencies into the undergraduate nursing curriculum. Results: For each competency, outcomes were developed for each year. Participants confirmed completeness of outcomes and appropriateness of the mapping of the HIV and AIDS related outcomes into the nursing curriculum, as well as the feasibility and practicability of the integration. Conclusion: Required resources for integration of HIV and AIDS related nursing competencies, such as human resources and nurse educators’ continued personal development were identified, as well as barriers to integration, and measures to eliminate them were discussed. The importance of integration of HIV and AIDS nursing competencies into the curriculum was reiterated.

Mechanisms of CD8+ T cell-mediated suppression of HIV/SIV replication.

Science.gov (United States)

McBrien, Julia Bergild; Kumar, Nitasha A; Silvestri, Guido

2018-02-10

In this article, we summarize the role of CD8 + T cells during natural and antiretroviral therapy (ART)-treated HIV and SIV infections, discuss the mechanisms responsible for their suppressive activity, and review the rationale for CD8 + T cell-based HIV cure strategies. Evidence suggests that CD8 + T cells are involved in the control of virus replication during HIV and SIV infections. During early HIV infection, the cytolytic activity of CD8 + T cells is responsible for control of viremia. However, it has been proposed that CD8 + T cells also use non-cytolytic mechanisms to control SIV infection. More recently, CD8 + T cells were shown to be required to fully suppress virus production in ART-treated SIV-infected macaques, suggesting that CD8 + T cells are involved in the control of virus transcription in latently infected cells that persist under ART. A better understanding of the complex antiviral activities of CD8 + T cells during HIV/SIV infection will pave the way for immune interventions aimed at harnessing these functions to target the HIV reservoir. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reduced sTWEAK and increased sCD163 levels in HIV-infected patients: modulation by antiretroviral treatment, HIV replication and HCV co-infection.

Directory of Open Access Journals (Sweden)

Luis M Beltrán

Full Text Available Patients infected with the human immunodeficiency virus (HIV have an increased risk of cardiovascular disease due to increased inflammation and persistent immune activation. CD163 is a macrophage scavenger receptor that is involved in monocyte-macrophage activation in HIV-infected patients. CD163 interacts with TWEAK, a member of the TNF superfamily. Circulating levels of sTWEAK and sCD163 have been previously associated with cardiovascular disease, but no previous studies have fully analyzed their association with HIV.The aim of this study was to analyze circulating levels of sTWEAK and sCD163 as well as other known markers of inflammation (hsCRP, IL-6 and sTNFRII and endothelial dysfunction (sVCAM-1 and ADMA in 26 patients with HIV before and after 48 weeks of antiretroviral treatment (ART and 23 healthy subjects.Patients with HIV had reduced sTWEAK levels and increased sCD163, sVCAM-1, ADMA, hsCRP, IL-6 and sTNFRII plasma concentrations, as well as increased sCD163/sTWEAK ratio, compared with healthy subjects. Antiretroviral treatment significantly reduced the concentrations of sCD163, sVCAM-1, hsCRP and sTNFRII, although they remained elevated when compared with healthy subjects. Antiretroviral treatment had no effect on the concentrations of ADMA and sTWEAK, biomarkers associated with endothelial function. The use of protease inhibitors as part of antiretroviral therapy and the presence of HCV-HIV co-infection and/or active HIV replication attenuated the ART-mediated decrease in sCD163 plasma concentrations.HIV-infected patients showed a proatherogenic profile characterized by increased inflammatory, immune-activation and endothelial-dysfunction biomarkers that partially improved after ART. HCV-HIV co-infection and/or active HIV replication enhanced immune activation despite ART.

HIV-1 isolation from infected peripheral blood mononuclear cells

NARCIS (Netherlands)

Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A.; Schuitemaker, Hanneke; Scarlatti, Gabriella

2014-01-01

Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and

Recruitment of a SAP18-HDAC1 complex into HIV-1 virions and its requirement for viral replication.

Directory of Open Access Journals (Sweden)

Masha Sorin

2009-06-01

Full Text Available HIV-1 integrase (IN is a virally encoded protein required for integration of viral cDNA into host chromosomes. INI1/hSNF5 is a component of the SWI/SNF complex that interacts with HIV-1 IN, is selectively incorporated into HIV-1 (but not other retroviral virions, and modulates multiple steps, including particle production and infectivity. To gain further insight into the role of INI1 in HIV-1 replication, we screened for INI1-interacting proteins using the yeast two-hybrid system. We found that SAP18 (Sin3a associated protein 18 kD, a component of the Sin3a-HDAC1 complex, directly binds to INI1 in yeast, in vitro and in vivo. Interestingly, we found that IN also binds to SAP18 in vitro and in vivo. SAP18 and components of a Sin3A-HDAC1 complex were specifically incorporated into HIV-1 (but not SIV and HTLV-1 virions in an HIV-1 IN-dependent manner. Using a fluorescence-based assay, we found that HIV-1 (but not SIV virion preparations harbour significant deacetylase activity, indicating the specific recruitment of catalytically active HDAC into the virions. To determine the requirement of virion-associated HDAC1 to HIV-1 replication, an inactive, transdominant negative mutant of HDAC1 (HDAC1(H141A was utilized. Incorporation of HDAC1(H141A decreased the virion-associated histone deacetylase activity. Furthermore, incorporation of HDAC1(H141A decreased the infectivity of HIV-1 (but not SIV virions. The block in infectivity due to virion-associated HDAC1(H141A occurred specifically at the early reverse transcription stage, while entry of the virions was unaffected. RNA-interference mediated knock-down of HDAC1 in producer cells resulted in decreased virion-associated HDAC1 activity and a reduction in infectivity of these virions. These studies indicate that HIV-1 IN and INI1/hSNF5 bind SAP18 and selectively recruit components of Sin3a-HDAC1 complex into HIV-1 virions. Furthermore, HIV-1 virion-associated HDAC1 is required for efficient early post

Inhibitors of Deubiquitinating Enzymes Block HIV-1 Replication and Augment the Presentation of Gag-Derived MHC-I Epitopes.

Science.gov (United States)

Setz, Christian; Friedrich, Melanie; Rauch, Pia; Fraedrich, Kirsten; Matthaei, Alina; Traxdorf, Maximilian; Schubert, Ulrich

2017-08-12

In recent years it has been well established that two major constituent parts of the ubiquitin proteasome system (UPS)-the proteasome holoenzymes and a number of ubiquitin ligases-play a crucial role, not only in virus replication but also in the regulation of the immunogenicity of human immunodeficiency virus type 1 (HIV-1). However, the role in HIV-1 replication of the third major component, the deubiquitinating enzymes (DUBs), has remained largely unknown. In this study, we show that the DUB-inhibitors (DIs) P22077 and PR-619, specific for the DUBs USP7 and USP47, impair Gag processing and thereby reduce the infectivity of released virions without affecting viral protease activity. Furthermore, the replication capacity of X4- and R5-tropic HIV-1 NL4-3 in human lymphatic tissue is decreased upon treatment with these inhibitors without affecting cell viability. Most strikingly, combinatory treatment with DIs and proteasome inhibitors synergistically blocks virus replication at concentrations where mono-treatment was ineffective, indicating that DIs can boost the therapeutic effect of proteasome inhibitors. In addition, P22077 and PR-619 increase the polyubiquitination of Gag and thus its entry into the UPS and the major histocompatibility complex (MHC)-I pathway. In summary, our data point towards a model in which specific inhibitors of DUBs not only interfere with virus spread but also increase the immune recognition of HIV-1 expressing cells.

HIV-1 isolation from infected peripheral blood mononuclear cells.

Science.gov (United States)

Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A; Schuitemaker, Hanneke; Scarlatti, Gabriella

2014-01-01

Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and not in tumor derived cell lines. The procedure involves culture of PBMCs from an infected patient with phytohemagglutinin (PHA)-stimulated PBMC from seronegative donors, which provide susceptible target cells for HIV replication. HIV can be isolated from the bulk population of PBMCs or after cloning of the cells to obtain viral biological clones. Viral production is determined with p24 antigen (Ag) detection assays or with reverse transcriptase (RT) activity assay. Once isolated, HIV-1 can be propagated by infecting PHA-stimulated PBMCs from healthy donors. Aliquots from culture with a high production of virus are stored for later use.

Rapid quantification of the latent reservoir for HIV-1 using a viral outgrowth assay.

Directory of Open Access Journals (Sweden)

Gregory M Laird

Full Text Available HIV-1 persists in infected individuals in a stable pool of resting CD4(+ T cells as a latent but replication-competent provirus. This latent reservoir is the major barrier to the eradication of HIV-1. Clinical trials are currently underway investigating the effects of latency-disrupting compounds on the persistence of the latent reservoir in infected individuals. To accurately assess the effects of such compounds, accurate assays to measure the frequency of latently infected cells are essential. The development of a simpler assay for the latent reservoir has been identified as a major AIDS research priority. We report here the development and validation of a rapid viral outgrowth assay that quantifies the frequency of cells that can release replication-competent virus following cellular activation. This new assay utilizes bead and column-based purification of resting CD4(+ T cells from the peripheral blood of HIV-1 infected patients rather than cell sorting to obtain comparable resting CD4(+ T cell purity. This new assay also utilizes the MOLT-4/CCR5 cell line for viral expansion, producing statistically comparable measurements of the frequency of latent HIV-1 infection. Finally, this new assay employs a novel quantitative RT-PCR specific for polyadenylated HIV-1 RNA for virus detection, which we demonstrate is a more sensitive and cost-effective method to detect HIV-1 replication than expensive commercial ELISA detection methods. The reductions in both labor and cost make this assay suitable for quantifying the frequency of latently infected cells in clinical trials of HIV-1 eradication strategies.

Mass Cytometric Analysis of HIV Entry, Replication, and Remodeling in Tissue CD4+ T Cells

Directory of Open Access Journals (Sweden)

Marielle Cavrois

2017-07-01

Full Text Available To characterize susceptibility to HIV infection, we phenotyped infected tonsillar T cells by single-cell mass cytometry and created comprehensive maps to identify which subsets of CD4+ T cells support HIV fusion and productive infection. By comparing HIV-fused and HIV-infected cells through dimensionality reduction, clustering, and statistical approaches to account for viral perturbations, we identified a subset of memory CD4+ T cells that support HIV entry but not viral gene expression. These cells express high levels of CD127, the IL-7 receptor, and are believed to be long-lived lymphocytes. In HIV-infected patients, CD127-expressing cells preferentially localize to extrafollicular lymphoid regions with limited viral replication. Thus, CyTOF-based phenotyping, combined with analytical approaches to distinguish between selective infection and receptor modulation by viruses, can be used as a discovery tool.

Replicative phenotyping adds value to genotypic resistance testing in heavily pre-treated HIV-infected individuals - the Swiss HIV Cohort Study

Directory of Open Access Journals (Sweden)

Martinetti Gladys

2011-01-01

Full Text Available Abstract Background Replicative phenotypic HIV resistance testing (rPRT uses recombinant infectious virus to measure viral replication in the presence of antiretroviral drugs. Due to its high sensitivity of detection of viral minorities and its dissecting power for complex viral resistance patterns and mixed virus populations rPRT might help to improve HIV resistance diagnostics, particularly for patients with multiple drug failures. The aim was to investigate whether the addition of rPRT to genotypic resistance testing (GRT compared to GRT alone is beneficial for obtaining a virological response in heavily pre-treated HIV-infected patients. Methods Patients with resistance tests between 2002 and 2006 were followed within the Swiss HIV Cohort Study (SHCS. We assessed patients' virological success after their antiretroviral therapy was switched following resistance testing. Multilevel logistic regression models with SHCS centre as a random effect were used to investigate the association between the type of resistance test and virological response (HIV-1 RNA Results Of 1158 individuals with resistance tests 221 with GRT+rPRT and 937 with GRT were eligible for analysis. Overall virological response rates were 85.1% for GRT+rPRT and 81.4% for GRT. In the subgroup of patients with >2 previous failures, the odds ratio (OR for virological response of GRT+rPRT compared to GRT was 1.45 (95% CI 1.00-2.09. Multivariate analyses indicate a significant improvement with GRT+rPRT compared to GRT alone (OR 1.68, 95% CI 1.31-2.15. Conclusions In heavily pre-treated patients rPRT-based resistance information adds benefit, contributing to a higher rate of treatment success.

Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

Energy Technology Data Exchange (ETDEWEB)

Di Nunzio, Francesca, E-mail: francesca.di-nunzio@pasteur.fr [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Fricke, Thomas [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Miccio, Annarita [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Valle-Casuso, Jose Carlos; Perez, Patricio [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Souque, Philippe [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Rizzi, Ermanno; Severgnini, Marco [Institute of Biomedical Technologies, CNR, Milano (Italy); Mavilio, Fulvio [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Genethon, Evry (France); Charneau, Pierre [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Diaz-Griffero, Felipe, E-mail: felipe.diaz-griffero@einstein.yu.edu [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States)

2013-05-25

The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites.

Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

International Nuclear Information System (INIS)

Di Nunzio, Francesca; Fricke, Thomas; Miccio, Annarita; Valle-Casuso, Jose Carlos; Perez, Patricio; Souque, Philippe; Rizzi, Ermanno; Severgnini, Marco; Mavilio, Fulvio; Charneau, Pierre; Diaz-Griffero, Felipe

2013-01-01

The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites

Natural Plant Alkaloid (Emetine Inhibits HIV-1 Replication by Interfering with Reverse Transcriptase Activity

Directory of Open Access Journals (Sweden)

Ana Luiza Chaves Valadão

2015-06-01

Full Text Available Ipecac alkaloids are secondary metabolites produced in the medicinal plant Psychotria ipecacuanha. Emetine is the main alkaloid of ipecac and one of the active compounds in syrup of Ipecac with emetic property. Here we evaluated emetine’s potential as an antiviral agent against Human Immunodeficiency Virus. We performed in vitro Reverse Transcriptase (RT Assay and Natural Endogenous Reverse Transcriptase Activity Assay (NERT to evaluate HIV RT inhibition. Emetine molecular docking on HIV-1 RT was also analyzed. Phenotypic assays were performed in non-lymphocytic and in Peripheral Blood Mononuclear Cells (PBMC with HIV-1 wild-type and HIV-harboring RT-resistant mutation to Nucleoside Reverse Transcriptase Inhibitors (M184V. Our results showed that HIV-1 RT was blocked in the presence of emetine in both models: in vitro reactions with isolated HIV-1 RT and intravirion, measured by NERT. Emetine revealed a strong potential of inhibiting HIV-1 replication in both cellular models, reaching 80% of reduction in HIV-1 infection, with low cytotoxic effect. Emetine also blocked HIV-1 infection of RT M184V mutant. These results suggest that emetine is able to penetrate in intact HIV particles, and bind and block reverse transcription reaction, suggesting that it can be used as anti-HIV microbicide. Taken together, our findings provide additional pharmacological information on the potential therapeutic effects of emetine.

HMGB1-dependent triggering of HIV-1 replication and persistence in dendritic cells as a consequence of NK-DC cross-talk.

Directory of Open Access Journals (Sweden)

Héla Saïdi

Full Text Available HIV-1 has evolved ways to exploit DCs, thereby facilitating viral dissemination and allowing evasion of antiviral immunity. Recently, the fate of DCs has been found to be extremely dependent on the interaction with autologous NK cells, but the mechanisms by which NK-DC interaction controls viral infections remain unclear. Here, we investigate the impact of NK-DC cross-talk on maturation and functions of HIV-infected immature DCs.Immature DCs were derived from primary monocytes, cultured in the presence of IL-4 and GM-CSF. In some experiments, DCs were infected with R5-HIV-1(BaL or X4-HIV-1(NDK, and viral replication, proviral HIV-DNA and the frequency of infected DCs were measured. Autologous NK cells were sorted and either kept unstimulated in the presence of suboptimal concentration of IL-2, or activated by a combination of PHA and IL-2. The impact of 24 h NK-DC cross-talk on the fate of HIV-1-infected DCs was analyzed. We report that activated NK cells were required for the induction of maturation of DCs, whether uninfected or HIV-1-infected, and this process involved HMGB1. However, the cross-talk between HIV-1-infected DCs and activated NK cells was functionally defective, as demonstrated by the strong impairment of DCs to induce Th1 polarization of naïve CD4 T cells. This was associated with the defective production of IL-12 and IL-18 by infected DCs. Moreover, the crosstalk between activated NK cells and HIV-infected DCs resulted in a dramatic increase in viral replication and proviral DNA expression in DCs. HMGB1, produced both by NK cells and DCs, was found to play a pivotal role in this process, and inhibition of HMGB1 activity by glycyrrhizin, known to bind specifically to HMGB1, or blocking anti-HMGB1 antibodies, abrogated NK-dependent HIV-1 replication in DCs.These observations provide evidence for the crucial role of NK-DC cross-talk in promoting viral dissemination, and challenge the question of the in vivo involvement of HMGB1

An improved protocol for efficient engraftment in NOD/LTSZ-SCIDIL-2Rγnull mice allows HIV replication and development of anti-HIV immune responses.

Directory of Open Access Journals (Sweden)

Maneesh Singh

Full Text Available Cord blood hematopoietic progenitor cells (CB-HPCs transplanted immunodeficient NOD/LtsZ-scidIL2Rγ(null (NSG and NOD/SCID/IL2Rγ(null (NOG mice need efficient human cell engraftment for long-term HIV-1 replication studies. Total body irradiation (TBI is a classical myeloablation regimen used to improve engraftment levels of human cells in these humanized mice. Some recent reports suggest the use of busulfan as a myeloablation regimen to transplant HPCs in neonatal and adult NSG mice. In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3-4 week old NSG mice. In this CB-CD34+transplanted NSG mice engraftment efficiency of human CD45+cell is over 90% in peripheral blood. Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days. These humanized NSG mice have shown long-lasting viremia after HIV-1JRCSF and HIV-1Bal inoculation through intravenous and rectal routes. We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection. Humanized NSG mice reconstituted according to our new protocol produced, moderate cellular and humoral immune responses to HIV-1 postinfection. We believe that NSG mice reconstituted according to our easy to use protocol will provide a better in vivo model for HIV-1 replication and anti-HIV-1 therapy trials.

Epidermal growth factor receptor targeting of replication competent adenovirus enhances cytotoxicity in bladder cancer

NARCIS (Netherlands)

van der Poel, HG; Molenaar, B; van Beusechem, VW; Haisma, HJ; Rodriguez, R; Curiel, DT; Gerritsen, WR

Purpose: We evaluated the delivery and oncolytic potential of targeted replication competent adenoviruses in bladder cancer lines. Materials and Methods: Seven established human bladder cancer tumor lines (5637, SW800, TCCsup, J82, Scaber, T24 and 253J) were studied for the expression of integrins

Exosomes from Human Immunodeficiency Virus Type 1 (HIV-1)-Infected Cells License Quiescent CD4+ T Lymphocytes To Replicate HIV-1 through a Nef- and ADAM17-Dependent Mechanism

OpenAIRE

Arenaccio, Claudia; Chiozzini, Chiara; Columba-Cabezas, Sandra; Manfredi, Francesco; Affabris, Elisabetta; Baur, Andreas; Federico, Maurizio

2014-01-01

Resting CD4+ T lymphocytes resist human immunodeficiency virus (HIV) infection. Here, we provide evidence that exosomes from HIV-1-infected cells render resting human primary CD4+ T lymphocytes permissive to HIV-1 replication. These results were obtained with transwell cocultures of HIV-1-infected cells with quiescent CD4+ T lymphocytes in the presence of inhibitors of exosome release and were confirmed using exosomes purified from supernatants of HIV-1-infected primary CD4+ T lymphocytes. We...

The role of technical assistance in the replication of effective HIV interventions.

Science.gov (United States)

O'Donnell, L; Scattergood, P; Adler, M; Doval, A S; Barker, M; Kelly, J A; Kegeles, S M; Rebchook, G M; Adams, J; Terry, M A; Neumann, M S

2000-01-01

This article examines the role of technical assistance (TA) in supporting the replication of proven HIV interventions. A case study of the replication of the VOICES/VOCES intervention elucidates the level and types of TA provided to support new users through the adoption process. TA included help in garnering administrative support, identifying target audiences, recruiting groups for sessions, maintaining fidelity to the intervention's core elements, tailoring the intervention to meet clients' needs, strengthening staff members' facilitation skills, troubleshooting challenges, and devising strategies to sustain the intervention. Two to four hours per month of TA were provided to each agency adopting the intervention, at an estimated monthly cost of $206 to $412. Findings illustrate how TA supports replication by establishing a conversation between the researcher TA providers experienced with the intervention and new users. This communication helps preserve key program elements and contributes to ongoing refinement of the intervention.

Solving the Blood-Brain Barrier Challenge for the Effective Treatment of HIV Replication in the Central Nervous System.

Science.gov (United States)

Bertrand, Luc; Nair, Madhavan; Toborek, Michal

2016-01-01

Recent decades mark a great progress in the treatment of HIV infection. What was once a deadly disease is now a chronic infection. However, HIV-infected patients are prone to develop comorbidities, which severely affect their daily functions. For example, a large population of patients develop a variety of neurological and cognitive complications, called HIV associated neurological disorders (HAND). Despite efficient repression of viral replication in the periphery, evidence shows that the virus can remain active in the central nervous system (CNS). This low level of replication is believed to result in a progression of neurocognitive dysfunction in infected individuals. Insufficient viral inhibition in the brain results from the inability of several treatment drugs in crossing the blood-brain barrier (BBB) and reaching therapeutic concentrations in the CNS. The current manuscript discusses several strategies that are being developed to enable therapeutics to cross the BBB, including bypassing BBB, inhibition of efflux transporters, the use of active transporters present at the BBB, and nanotechnology. The increased concentration of therapeutics in the CNS is desirable to prevent viral replication; however, potential side effects of anti-retroviral drugs need also to be taken into consideration.

Liver stiffness is not associated with short- and long-term plasma HIV RNA replication in immunocompetent patients with HIV infection and with HIV/HCV coinfection

Science.gov (United States)

Parisi, Saverio Giuseppe; Basso, Monica; Mengoli, Carlo; Scaggiante, Renzo; Andreis, Samantha; Franzetti, Marzia Maria; Cattelan, Anna Maria; Zago, Daniela; Cruciani, Mario; Andreoni, Massimo; Piovesan, Sara; Palù, Giorgio; Alberti, Alfredo

2017-01-01

Background Human immunodeficiency virus (HIV) may be directly responsible for liver damage but there are contrasting data regarding the influence of detectable plasma viremia. We analyzed the influence of plasma HIV RNA (pHIV) detectability and of other clinical and viro-immunological variables on liver stiffness (LS) measurement in adult immunocompetent HIV-monoinfected patients and in patients coinfected with hepatitis C virus (HCV). Methods Logistic regression analysis was performed using the value of LS>7.1 kPa as the dependent variable. A linear regression model was applied using LS measurement after log10 transformation (lkpa) as the dependent variable and we analyzed the predicted values versus the observed lkpa values; pHIV was classified as detectable or undetectable in the 12- and 36-month study periods before LS measurement. Results We studied 251 patients (178 with HIV monoinfection), most of whom were on antiviral treatment; 36-month study time was available for 154 subjects. The mean CD4+ cell count was 634 cells/mm3 in HIV-monoinfected patients and 606 cells/mm3 in coinfected patients. No difference in LS was found between patients with detectable or undetectable pHIV in either the 12- or the 36-month study period before transient elastography. The mean LS was higher in HIV/HCV coinfected patients (P<0.0001) than in the HIV-monoinfected subjects; lkpa was positively correlated with HCV coinfection (P<0.0001) and aspartate aminotransferase levels (P<0.0001). Detectable pHIV failed to reach significance. Eight HIV-monoinfected patients had a predicted LS measurement lower than the observed one, while eight patients had the opposite result. Conclusion LS was not correlated with ongoing HIV replication during the 12- and 36-month study periods in immunocompetent HIV-monoinfected and HIV/HCV-coinfected patients. PMID:28845109

Quantitative comparison of HTLV-1 and HIV-1 cell-to-cell infection with new replication dependent vectors.

Directory of Open Access Journals (Sweden)

Dmitriy Mazurov

2010-02-01

Full Text Available We have developed an efficient method to quantify cell-to-cell infection with single-cycle, replication dependent reporter vectors. This system was used to examine the mechanisms of infection with HTLV-1 and HIV-1 vectors in lymphocyte cell lines. Effector cells transfected with reporter vector, packaging vector, and Env expression plasmid produced virus-like particles that transduced reporter gene activity into cocultured target cells with zero background. Reporter gene expression was detected exclusively in target cells and required an Env-expression plasmid and a viral packaging vector, which provided essential structural and enzymatic proteins for virus replication. Cell-cell fusion did not contribute to infection, as reporter protein was rarely detected in syncytia. Coculture of transfected Jurkat T cells and target Raji/CD4 B cells enhanced HIV-1 infection two fold and HTLV-1 infection ten thousand fold in comparison with cell-free infection of Raji/CD4 cells. Agents that interfere with actin and tubulin polymerization strongly inhibited HTLV-1 and modestly decreased HIV-1 cell-to-cell infection, an indication that cytoskeletal remodeling was more important for HTLV-1 transmission. Time course studies showed that HTLV-1 transmission occurred very rapidly after cell mixing, whereas slower kinetics of HIV-1 coculture infection implies a different mechanism of infectious transmission. HTLV-1 Tax was demonstrated to play an important role in altering cell-cell interactions that enhance virus infection and replication. Interestingly, superantigen-induced synapses between Jurkat cells and Raji/CD4 cells did not enhance infection for either HTLV-1 or HIV-1. In general, the dependence on cell-to-cell infection was determined by the virus, the effector and target cell types, and by the nature of the cell-cell interaction.

SAMHD1 restricts HIV-1 replication and regulates interferon production in mouse myeloid cells.

Directory of Open Access Journals (Sweden)

Ruonan Zhang

Full Text Available SAMHD1 restricts the replication of HIV-1 and other retroviruses in human myeloid and resting CD4(+ T cells and that is counteracted in SIV and HIV-2 by the Vpx accessory protein. The protein is a phosphohydrolase that lowers the concentration of deoxynucleoside triphosphates (dNTP, blocking reverse transcription of the viral RNA genome. Polymorphisms in the gene encoding SAMHD1 are associated with Aicardi-Goutières Syndrome, a neurological disorder characterized by increased type-I interferon production. SAMHD1 is conserved in mammals but its role in restricting virus replication and controlling interferon production in non-primate species is not well understood. We show that SAMHD1 is catalytically active and expressed at high levels in mouse spleen, lymph nodes, thymus and lung. siRNA knock-down of SAMHD1 in bone marrow-derived macrophages increased their susceptibility to HIV-1 infection. shRNA knock-down of SAMHD1 in the murine monocytic cell-line RAW264.7 increased its susceptibility to HIV-1 and murine leukemia virus and increased the levels of the dNTP pool. In addition, SAMHD1 knock-down in RAW264.7 cells induced the production of type-I interferon and several interferon-stimulated genes, modeling the situation in Aicardi-Goutières Syndrome. Our findings suggest that the role of SAMHD1 in restricting viruses is conserved in the mouse. The RAW264.7 cell-line serves as a useful tool to study the antiviral and innate immune response functions of SAMHD1.

Control of HIV replication in astrocytes by a family of highly conserved host proteins with a common Rev-interacting domain (Risp).

Science.gov (United States)

Vincendeau, Michelle; Kramer, Susanne; Hadian, Kamyar; Rothenaigner, Ina; Bell, Jeanne; Hauck, Stefanie M; Bickel, Christian; Nagel, Daniel; Kremmer, Elisabeth; Werner, Thomas; Leib-Mösch, Christine; Brack-Werner, Ruth

2010-10-23

In human astrocytes, restriction of HIV replication involves inhibition of HIV Rev activity. We previously identified a Rev-interacting human protein fragment (16.4.1) that can reduce Rev activity. The 16.4.1 sequence is contained in a group of highly similar host cell proteins, which we call the Risp family. Here we investigate whether the Risp family is connected to HIV replication in astrocytes. Cell/tissue lysates were analyzed for Risp expression by western blot with various anti-Risp antibodies. The interaction of astrocytic Risp members with Rev was investigated by affinity chromatography. Astrocytes were transfected with expression plasmids containing cDNAs encoding full-length Risp or the isolated 16.4.1 region for Risp overexpression or with siRNAs designed for Risp knock-down. Rev activity was investigated with a Rev-reporter assay. RNA levels were quantified by real-time RT-PCR, HIV Gag levels by p24ELISA. Expression of the Risp family was demonstrated in human brain tissues and astrocytes. Astrocytes were shown to produce Risp family members that interact with Rev. Production of HIV Gag proteins and Rev-dependent RNAs in persistently infected astrocytes increased upon Risp knock-down and decreased upon Risp overexpression. Risp knock-down increased Rev activity and raised proportions of Rev proteins in the nucleus of astrocytes. Our results link the Risp family to restriction of HIV production and inhibition of Rev activity in astrocytes. We conclude that the Risp family represents a novel family of host factors that can control HIV replication and may be important for the containment of HIV infection in brain reservoirs.

Early low-titer neutralizing antibodies impede HIV-1 replication and select for virus escape.

Directory of Open Access Journals (Sweden)

Katharine J Bar

Full Text Available Single genome sequencing of early HIV-1 genomes provides a sensitive, dynamic assessment of virus evolution and insight into the earliest anti-viral immune responses in vivo. By using this approach, together with deep sequencing, site-directed mutagenesis, antibody adsorptions and virus-entry assays, we found evidence in three subjects of neutralizing antibody (Nab responses as early as 2 weeks post-seroconversion, with Nab titers as low as 1∶20 to 1∶50 (IC(50 selecting for virus escape. In each of the subjects, Nabs targeted different regions of the HIV-1 envelope (Env in a strain-specific, conformationally sensitive manner. In subject CH40, virus escape was first mediated by mutations in the V1 region of the Env, followed by V3. HIV-1 specific monoclonal antibodies from this subject mapped to an immunodominant region at the base of V3 and exhibited neutralizing patterns indistinguishable from polyclonal antibody responses, indicating V1-V3 interactions within the Env trimer. In subject CH77, escape mutations mapped to the V2 region of Env, several of which selected for alterations of glycosylation. And in subject CH58, escape mutations mapped to the Env outer domain. In all three subjects, initial Nab recognition was followed by sequential rounds of virus escape and Nab elicitation, with Nab escape variants exhibiting variable costs to replication fitness. Although delayed in comparison with autologous CD8 T-cell responses, our findings show that Nabs appear earlier in HIV-1 infection than previously recognized, target diverse sites on HIV-1 Env, and impede virus replication at surprisingly low titers. The unexpected in vivo sensitivity of early transmitted/founder virus to Nabs raises the possibility that similarly low concentrations of vaccine-induced Nabs could impair virus acquisition in natural HIV-1 transmission, where the risk of infection is low and the number of viruses responsible for transmission and productive clinical

High-dose vitamin D3 reduces deficiency caused by low UVB exposure and limits HIV-1 replication in urban Southern Africans

Science.gov (United States)

Coussens, Anna K.; Naude, Celeste E.; Goliath, Rene; Chaplin, George; Wilkinson, Robert J.; Jablonski, Nina G.

2015-06-01

Cape Town, South Africa, has a seasonal pattern of UVB radiation and a predominantly dark-skinned urban population who suffer high HIV-1 prevalence. This coexistent environmental and phenotypic scenario puts residents at risk for vitamin D deficiency, which may potentiate HIV-1 disease progression. We conducted a longitudinal study in two ethnically distinct groups of healthy young adults in Cape Town, supplemented with vitamin D3 in winter, to determine whether vitamin D status modifies the response to HIV-1 infection and to identify the major determinants of vitamin D status (UVB exposure, diet, pigmentation, and genetics). Vitamin D deficiency was observed in the majority of subjects in winter and in a proportion of individuals in summer, was highly correlated with UVB exposure, and was associated with greater HIV-1 replication in peripheral blood cells. High-dosage oral vitamin D3 supplementation attenuated HIV-1 replication, increased circulating leukocytes, and reversed winter-associated anemia. Vitamin D3 therefore presents as a low-cost supplementation to improve HIV-associated immunity.

Rapid transient production in plants by replicating and non-replicating vectors yields high quality functional anti-HIV antibody.

Directory of Open Access Journals (Sweden)

Frank Sainsbury

2010-11-01

Full Text Available The capacity of plants and plant cells to produce large amounts of recombinant protein has been well established. Due to advantages in terms of speed and yield, attention has recently turned towards the use of transient expression systems, including viral vectors, to produce proteins of pharmaceutical interest in plants. However, the effects of such high level expression from viral vectors and concomitant effects on host cells may affect the quality of the recombinant product.To assess the quality of antibodies transiently expressed to high levels in plants, we have expressed and characterised the human anti-HIV monoclonal antibody, 2G12, using both replicating and non-replicating systems based on deleted versions of Cowpea mosaic virus (CPMV RNA-2. The highest yield (approximately 100 mg/kg wet weight leaf tissue of affinity purified 2G12 was obtained when the non-replicating CPMV-HT system was used and the antibody was retained in the endoplasmic reticulum (ER. Glycan analysis by mass-spectrometry showed that the glycosylation pattern was determined exclusively by whether the antibody was retained in the ER and did not depend on whether a replicating or non-replicating system was used. Characterisation of the binding and neutralisation properties of all the purified 2G12 variants from plants showed that these were generally similar to those of the Chinese hamster ovary (CHO cell-produced 2G12.Overall, the results demonstrate that replicating and non-replicating CPMV-based vectors are able to direct the production of a recombinant IgG similar in activity to the CHO-produced control. Thus, a complex recombinant protein was produced with no apparent effect on its biochemical properties using either high-level expression or viral replication. The speed with which a recombinant pharmaceutical with excellent biochemical characteristics can be produced transiently in plants makes CPMV-based expression vectors an attractive option for

A Cell Internalizing Antibody Targeting Capsid Protein (p24 Inhibits the Replication of HIV-1 in T Cells Lines and PBMCs: A Proof of Concept Study.

Directory of Open Access Journals (Sweden)

Syed A Ali

Full Text Available There remains a need for newer therapeutic approaches to combat HIV/AIDS. Viral capsid protein p24 plays important roles in HIV pathogenesis. Peptides and small molecule inhibitors targeting p24 have shown to inhibit virus replication in treated cell. High specificity and biological stability of monoclonal antibodies (mAbs make them an attractive contender for in vivo treatments. However, mAbs do not enter into cells, thus are restricted to target surface molecules. This also makes targeting intracellular HIV-1 p24 a challenge. A mAb specific to p24 that can internalize into the HIV-infected cells is hypothesized to inhibit the virus replication. We selected a mAb that has previously shown to inhibit p24 polymerization in an in vitro assay and chemically conjugated it with cell penetrating peptides (CPP to generate cell internalizing anti-p24 mAbs. Out of 8 CPPs tested, κFGF-MTS -conjugated mAbs internalized T cells most efficiently. At nontoxic concentration, the κFGF-MTS-anti-p24-mAbs reduced the HIV-1 replication up to 73 and 49% in T-lymphocyte and PBMCs respectively. Marked inhibition of HIV-1 replication in relevant cells by κFGF-MTS-anti-p24-mAbs represents a viable strategy to target HIV proteins present inside the cells.

Imperatorin inhibits HIV-1 replication through an Sp1-dependent pathway.

Science.gov (United States)

Sancho, Rocío; Márquez, Nieves; Gómez-Gonzalo, Marta; Calzado, Marco A; Bettoni, Giorgio; Coiras, Maria Teresa; Alcamí, José; López-Cabrera, Manuel; Appendino, Giovanni; Muñoz, Eduardo

2004-09-03

Coumarins and structurally related compounds have been recently shown to present anti-human immunodeficiency virus, type 1 (HIV-1) activity. Among them, the dietary furanocoumarin imperatorin is present in citrus fruits, in culinary herbs, and in some medicinal plants. In this study we report that imperatorin inhibits either vesicular stomatitis virus-pseudotyped or gp160-enveloped recombinant HIV-1 infection in several T cell lines and in HeLa cells. These recombinant viruses express luciferase as a marker of viral replication. Imperatorin did not inhibit the reverse transcription nor the integration steps in the viral cell cycle. Using several 5' long terminal repeat-HIV-1 constructs where critical response elements were either deleted or mutated, we found that the transcription factor Sp1 is critical for the inhibitory activity of imperatorin induced by both phorbol 12-myristate 13-acetate and HIV-1 Tat. Moreover in transient transfections imperatorin specifically inhibited phorbol 12-myristate 13-acetate-induced transcriptional activity of the Gal4-Sp1 fusion protein. Since Sp1 is also implicated in cell cycle progression we further studied the effect of imperatorin on cyclin D1 gene transcription and protein expression and in HeLa cell cycle progression. We found that imperatorin strongly inhibited cyclin D1 expression and arrested the cells at the G(1) phase of the cell cycle. These results highlight the potential of Sp1 transcription factor as a target for natural anti-HIV-1 compounds such as furanocoumarins that might have a potential therapeutic role in the management of AIDS.

Curaxin CBL0100 Blocks HIV-1 Replication and Reactivation through Inhibition of Viral Transcriptional Elongation

Directory of Open Access Journals (Sweden)

Maxime J. Jean

2017-10-01

Full Text Available Despite combination antiretroviral therapy (cART, acquired immunodeficiency syndrome (AIDS, predominantly caused by the human immunodeficiency virus type 1 (HIV-1, remains incurable. The barrier to a cure lies in the virus' ability to establish a latent infection in HIV/AIDS patients. Unsurprisingly, efforts for a sterilizing cure have focused on the “shock and kill” strategy using latency-reversing agents (LRAs to complement cART in order to eliminate these latent reservoirs. However, this method faces numerous challenges. Recently, the “block and lock” strategy has been proposed. It aims to reinforce a deep state of latency and prevent sporadic reactivation (“blip” of HIV-1 using latency-promoting agents (LPAs for a functional cure. Our studies of curaxin 100 (CBL0100, a small-molecule targeting the facilitates chromatin transcription (FACT complex, show that it blocks both HIV-1 replication and reactivation in in vitro and ex vivo models of HIV-1. Mechanistic investigation elucidated that CBL0100 preferentially targets HIV-1 transcriptional elongation and decreases the occupancy of RNA Polymerase II (Pol II and FACT at the HIV-1 promoter region. In conclusion, CBL0100 is a newly identified inhibitor of HIV-1 transcription that can be used as an LPA in the “block and lock” cure strategy.

Role of transmitted Gag CTL polymorphisms in defining replicative capacity and early HIV-1 pathogenesis.

Directory of Open Access Journals (Sweden)

Jessica L Prince

Full Text Available Initial studies of 88 transmission pairs in the Zambia Emory HIV Research Project cohort demonstrated that the number of transmitted HLA-B associated polymorphisms in Gag, but not Nef, was negatively correlated to set point viral load (VL in the newly infected partners. These results suggested that accumulation of CTL escape mutations in Gag might attenuate viral replication and provide a clinical benefit during early stages of infection. Using a novel approach, we have cloned gag sequences isolated from the earliest seroconversion plasma sample from the acutely infected recipient of 149 epidemiologically linked Zambian transmission pairs into a primary isolate, subtype C proviral vector, MJ4. We determined the replicative capacity (RC of these Gag-MJ4 chimeras by infecting the GXR25 cell line and quantifying virion production in supernatants via a radiolabeled reverse transcriptase assay. We observed a statistically significant positive correlation between RC conferred by the transmitted Gag sequence and set point VL in newly infected individuals (p = 0.02. Furthermore, the RC of Gag-MJ4 chimeras also correlated with the VL of chronically infected donors near the estimated date of infection (p = 0.01, demonstrating that virus replication contributes to VL in both acute and chronic infection. These studies also allowed for the elucidation of novel sites in Gag associated with changes in RC, where rare mutations had the greatest effect on fitness. Although we observed both advantageous and deleterious rare mutations, the latter could point to vulnerable targets in the HIV-1 genome. Importantly, RC correlated significantly (p = 0.029 with the rate of CD4+ T cell decline over the first 3 years of infection in a manner that is partially independent of VL, suggesting that the replication capacity of HIV-1 during the earliest stages of infection is a determinant of pathogenesis beyond what might be expected based on set point VL alone.

Supraphysiologic control over HIV-1 replication mediated by CD8 T cells expressing a re-engineered CD4-based chimeric antigen receptor.

Directory of Open Access Journals (Sweden)

Rachel S Leibman

2017-10-01

Full Text Available HIV is adept at avoiding naturally generated T cell responses; therefore, there is a need to develop HIV-specific T cells with greater potency for use in HIV cure strategies. Starting with a CD4-based chimeric antigen receptor (CAR that was previously used without toxicity in clinical trials, we optimized the vector backbone, promoter, HIV targeting moiety, and transmembrane and signaling domains to determine which components augmented the ability of T cells to control HIV replication. This re-engineered CAR was at least 50-fold more potent in vitro at controlling HIV replication than the original CD4 CAR, or a TCR-based approach, and substantially better than broadly neutralizing antibody-based CARs. A humanized mouse model of HIV infection demonstrated that T cells expressing optimized CARs were superior at expanding in response to antigen, protecting CD4 T cells from infection, and reducing viral loads compared to T cells expressing the original, clinical trial CAR. Moreover, in a humanized mouse model of HIV treatment, CD4 CAR T cells containing the 4-1BB costimulatory domain controlled HIV spread after ART removal better than analogous CAR T cells containing the CD28 costimulatory domain. Together, these data indicate that potent HIV-specific T cells can be generated using improved CAR design and that CAR T cells could be important components of an HIV cure strategy.

Towards HIV-1 remission: potential roles for broadly neutralizing antibodies.

Science.gov (United States)

Halper-Stromberg, Ariel; Nussenzweig, Michel C

2016-02-01

Current antiretroviral drug therapies do not cure HIV-1 because they do not eliminate a pool of long-lived cells harboring immunologically silent but replication-competent proviruses - termed the latent reservoir. Eliminating this reservoir and stimulating the immune response to control infection in the absence of therapy remain important but unsolved goals of HIV-1 cure research. Recently discovered broadly neutralizing antibodies (bNAbs) exhibit remarkable breadth and potency in their ability to neutralize HIV-1 in vitro, and recent studies have demonstrated new therapeutic applications for passively administered bNAbs in vivo. This Review discusses the roles bNAbs might play in HIV-1 treatment regimens, including prevention, therapy, and cure.

Redefining the Viral Reservoirs That Prevent HIV-1 Eradication

Science.gov (United States)

Eisele, Evelyn; Siliciano, Robert F.

2014-01-01

Summary This review proposes definitions for key terms in the field of HIV-1 latency and eradication. In the context of eradication, a reservoir is a cell type that allows persistence of replication-competent HIV-1 on a time scale of years in patients on optimal antiretroviral therapy. Reservoirs act as a barrier to eradication in the patient population in whom cure attempts will likely be made. Halting viral replication is essential to eradication, and definitions and criteria for assessing whether this goal has been achieved are proposed. The cell types that may serve as reservoirs for HIV-1 are discussed. Currently, only latently infected resting CD4+ T cells fit the proposed definition of a reservoir, and more evidence is necessary to demonstrate that other cell types including hematopoietic stem cells and macrophages fit this definition. Further research is urgently required on potential reservoirs in the gut-associated lymphoid tissue and the central nervous system. PMID:22999944

Inhibition of HIV Replication by Cyclic and Hairpin PNAs Targeting the HIV-1 TAR RNA Loop

Science.gov (United States)

Upert, Gregory; Di Giorgio, Audrey; Upadhyay, Alok; Manvar, Dinesh; Pandey, Nootan; Pandey, Virendra N.; Patino, Nadia

2012-01-01

Human immunodeficiency virus-1 (HIV-1) replication and gene expression entails specific interaction of the viral protein Tat with its transactivation responsive element (TAR), to form a highly stable stem-bulge-loop structure. Previously, we described triphenylphosphonium (TPP) cation-based vectors that efficiently deliver nucleotide analogs (PNAs) into the cytoplasm of cells. In particular, we showed that the TPP conjugate of a linear 16-mer PNA targeting the apical stem-loop region of TAR impedes Tat-mediated transactivation of the HIV-1 LTR in vitro and also in cell culture systems. In this communication, we conjugated TPP to cyclic and hairpin PNAs targeting the loop region of HIV-1 TAR and evaluated their antiviral efficacy in a cell culture system. We found that TPP-cyclic PNAs containing only 8 residues, showed higher antiviral potency compared to hairpin PNAs of 12 or 16 residues. We further noted that the TPP-conjugates of the 8-mer cyclic PNA as well as the 16-mer linear PNA displayed similar antiviral efficacy. However, cyclic PNAs were shown to be highly specific to their target sequences. This communication emphasizes on the importance of small constrained cyclic PNAs over both linear and hairpin structures for targeting biologically relevant RNA hairpins. PMID:23029603

Inhibition of HIV Replication by Cyclic and Hairpin PNAs Targeting the HIV-1 TAR RNA Loop

Directory of Open Access Journals (Sweden)

Gregory Upert

2012-01-01

Full Text Available Human immunodeficiency virus-1 (HIV-1 replication and gene expression entails specific interaction of the viral protein Tat with its transactivation responsive element (TAR, to form a highly stable stem-bulge-loop structure. Previously, we described triphenylphosphonium (TPP cation-based vectors that efficiently deliver nucleotide analogs (PNAs into the cytoplasm of cells. In particular, we showed that the TPP conjugate of a linear 16-mer PNA targeting the apical stem-loop region of TAR impedes Tat-mediated transactivation of the HIV-1 LTR in vitro and also in cell culture systems. In this communication, we conjugated TPP to cyclic and hairpin PNAs targeting the loop region of HIV-1 TAR and evaluated their antiviral efficacy in a cell culture system. We found that TPP-cyclic PNAs containing only 8 residues, showed higher antiviral potency compared to hairpin PNAs of 12 or 16 residues. We further noted that the TPP-conjugates of the 8-mer cyclic PNA as well as the 16-mer linear PNA displayed similar antiviral efficacy. However, cyclic PNAs were shown to be highly specific to their target sequences. This communication emphasizes on the importance of small constrained cyclic PNAs over both linear and hairpin structures for targeting biologically relevant RNA hairpins.

A combinational CRISPR/Cas9 gene-editing approach can halt HIV replication and prevent viral escape

NARCIS (Netherlands)

Lebbink, Robert Jan; de Jong, Dorien C M; Wolters, Femke; Kruse, Elisabeth M; van Ham, Petra M; Wiertz, Emmanuel J H J; Nijhuis, Monique

2017-01-01

HIV presents one of the highest evolutionary rates ever detected and combination antiretroviral therapy is needed to overcome the plasticity of the virus population and control viral replication. Conventional treatments lack the ability to clear the latent reservoir, which remains the major obstacle

McCoy cell line as a possible model containing CD4+ receptors for the study of HIV-1 replication

Directory of Open Access Journals (Sweden)

Nogueira Yeda L.

2003-01-01

Full Text Available Several studies have recently shown the use of recombinant rabies virus as potential vector-viral vaccine for HIV-1. The sequence homology between gp 120 and rabies virus glycoprotein has been reported. The McCoy cell line has therefore been used to show CD4+ or CD4+ like receptors. Samples of HIV-1 were isolated, when plasma of HIV-1 positive patients was inoculated in the McCoy cell line. The virus infection was then studied during successive virus passages. The proteins released in the extra cellular medium were checked for protein activity, by exposure to SDS Electrophoresis and blotting to nitro-cellulose filter, then reacting with sera of HIV positive and negative patients. Successive passages were performed, and showed viral replication, membrane permeabilization, the syncytium formation, and the cellular lysis (cytopathic effect. Flow cytometry analysis shows clear evidence that CD4+ receptors are present in this cell line, which enhances the likelihood of easy isolation and replication of HIV. The results observed allow the use of this cell line as a possible model for isolating HIV, as well as for carrying out studies of the dynamics of viral infection in several situations, including exposure to drugs in pharmacological studies, and possibly studies and analyses of the immune response in vaccine therapies.

Interleukin-27 is a potent inhibitor of cis HIV-1 replication in monocyte-derived dendritic cells via a type I interferon-independent pathway.

Directory of Open Access Journals (Sweden)

Qian Chen

Full Text Available IL-27, a member of the IL-12 family of cytokines, plays an important and diverse role in the function of the immune system. Whilst generally recognized as an anti-inflammatory cytokine, in addition IL-27 has been found to have broad anti-viral effects. Recently, IL-27 has been shown to be a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages. The main objective of this study was to see whether IL-27 has a similar inhibitory effect on HIV-1 replication in dendritic cells (DCs. Monocytes were differentiated into immature DCs (iDCs and mature DCs (mDCs with standard techniques using a combination of GM-CSF, IL-4 and LPS. Following differentiation, iDCs were infected with HIV-1 and co-cultured in the presence or absence of IL-27. IL-27 treated DCs were shown to be highly potent inhibitors of cis HIV-1, particularly of CCR5 tropic strains. Of note, other IL-12 family members (IL-12, IL-23 and IL-35 had no effect on HIV-1 replication. Microarray studies of IL-27 treated DCs showed no up-regulation of Type I (IFN gene expression. Neutralization of the Type-I IFN receptor had no impact on the HIV inhibition. Lastly, IL-27 mediated inhibition was shown to act post-viral entry and prior to completion of reverse transcription. These results show for the first time that IL-27 is a potent inhibitor of cis HIV-1 infection in DCs by a Type I IFN independent mechanism. IL-27 has previously been reported to inhibit HIV-1 replication in CD4+ T cells and macrophages, thus taken together, this cytokine is a potent anti-HIV agent against all major cell types targeted by the HIV-1 virus and may have a therapeutic role in the future.

Construction and characterisation of a full-length infectious molecular clone from a fast replicating, X4-tropic HIV-1 CRF02.AG primary isolate

International Nuclear Information System (INIS)

Tebit, Denis M.; Zekeng, Leopold; Kaptue, Lazare; Kraeusslich, Hans-Georg; Herchenroeder, Ottmar

2003-01-01

Based on our previous analysis of HIV-1 isolates from Cameroon, we constructed a full-length infectious molecular clone from a primary isolate belonging to the CRF02.AG group of recombinant viruses which dominate the HIV-epidemic in West and Central Africa. The virus derived by transfection of the proviral clone pBD6-15 replicated with similar efficiency compared to its parental isolate and used CXCR4 as coreceptor as well. Furthermore, HIV-1 BD6-15 exhibited similar replication properties and virus yield as the reference B-type HIV-1 strain NL4-3. Sequence analysis revealed open reading frames for all structural and accessory genes apart from vpr. Phylogenetic and bootscanning analyses confirmed that BD6-15 clusters with CRF02.AG recombinant strains from West and Central Africa with similar cross-over points as described for the CRF02.AG prototype strain lbNG. Thus, pBD6-15 represents the first non-subtype B infectious molecular clone of a fast replicating, high producer, X4-tropic primary HIV-1 isolate, which had only been briefly passaged in primary cells

EcoHIV infection of mice establishes latent viral reservoirs in T cells and active viral reservoirs in macrophages that are sufficient for induction of neurocognitive impairment.

Science.gov (United States)

Gu, Chao-Jiang; Borjabad, Alejandra; Hadas, Eran; Kelschenbach, Jennifer; Kim, Boe-Hyun; Chao, Wei; Arancio, Ottavio; Suh, Jin; Polsky, Bruce; McMillan, JoEllyn; Edagwa, Benson; Gendelman, Howard E; Potash, Mary Jane; Volsky, David J

2018-06-01

Suppression of HIV replication by antiretroviral therapy (ART) or host immunity can prevent AIDS but not other HIV-associated conditions including neurocognitive impairment (HIV-NCI). Pathogenesis in HIV-suppressed individuals has been attributed to reservoirs of latent-inducible virus in resting CD4+ T cells. Macrophages are persistently infected with HIV but their role as HIV reservoirs in vivo has not been fully explored. Here we show that infection of conventional mice with chimeric HIV, EcoHIV, reproduces physiological conditions for development of disease in people on ART including immunocompetence, stable suppression of HIV replication, persistence of integrated, replication-competent HIV in T cells and macrophages, and manifestation of learning and memory deficits in behavioral tests, termed here murine HIV-NCI. EcoHIV established latent reservoirs in CD4+ T lymphocytes in chronically-infected mice but could be induced by epigenetic modulators ex vivo and in mice. In contrast, macrophages expressed EcoHIV constitutively in mice for up to 16 months; murine leukemia virus (MLV), the donor of gp80 envelope in EcoHIV, did not infect macrophages. Both EcoHIV and MLV were found in brain tissue of infected mice but only EcoHIV induced NCI. Murine HIV-NCI was prevented by antiretroviral prophylaxis but once established neither persistent EcoHIV infection in mice nor NCI could be reversed by long-acting antiretroviral therapy. EcoHIV-infected, athymic mice were more permissive to virus replication in macrophages than were wild-type mice, suffered cognitive dysfunction, as well as increased numbers of monocytes and macrophages infiltrating the brain. Our results suggest an important role of HIV expressing macrophages in HIV neuropathogenesis in hosts with suppressed HIV replication.

Effect of low-dose gamma radiation on HIV replication in human peripheral blood mononuclear cells

Energy Technology Data Exchange (ETDEWEB)

Xu, Y. [British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada); Conway, B. [British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada)]|[British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada). Dept. of Medicine; Montaner, J.S.G. [British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada)]|[British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada). Dept. of Medicine]|[Canadian HIV Trials Network, Vancouver (Canada); O`Shaughnessy, M.V. [British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada)]|[British Columbia Centre for Excellence in HIV/AIDS, British Columbia (Canada). Faculty of Medicine]|[Canadian HIV Trials Network, Vancouver (Canada); Greenstock, C.L. [AECL Research, Chalk River, Ontario (Canada). Radiation Biology and Health Physics Branch

1996-08-01

Recent studies have demonstrated that UV light and x-irradiation enhance human immunodeficiency virus (HIV) gene expression. There are few published data on related effects of {gamma}-radiation. This may be of clinical relevance, as radiotherapy has been used extensively for the treatment of acquired immunodeficiency syndrome associated conditions. We have studied the effects of {gamma}-radiation on HIV replication in mono-nuclear cells (MC). These cells were obtained from five seronegative healthy donors, exposed to 0-200 cGy {gamma}-radiation, stimulated with phytohemagglutinin-P (PHA-P) for 24 h, infected with a laboratory strain of HIV (HTLV-IIIB, multiplicity of infection = 0.001), then carried in culture for 14 days. Overall, when considering p24 antigen levels on days 7 and 11 in cultures established from cells exposed to 50 cGy, the maximal levels were significantly higher than those measured in the parallel control cultures taken as a whole (P < 0.05), with viral replication enhanced as much as 1000-fold in one case. No significant cytotoxicity was observed following exposure to doses up to 50 cGy. The mechanism of the observed effect remains unknown but may relate to direct gene activation and/or free radical generation, leading to such activation. To date, there is no evidence that viral stimulation occurs following therapeutic radiation in a clinical setting. (author).

Effect of low-dose gamma radiation on HIV replication in human peripheral blood mononuclear cells

International Nuclear Information System (INIS)

Xu, Y.; Conway, B.; O'Shaughnessy, M.V.; Greenstock, C.L.

1996-01-01

Recent studies have demonstrated that UV light and x-irradiation enhance human immunodeficiency virus (HIV) gene expression. There are few published data on related effects of γ-radiation. This may be of clinical relevance, as radiotherapy has been used extensively for the treatment of acquired immunodeficiency syndrome associated conditions. We have studied the effects of γ-radiation on HIV replication in mono-nuclear cells (MC). These cells were obtained from five seronegative healthy donors, exposed to 0-200 cGy γ-radiation, stimulated with phytohemagglutinin-P (PHA-P) for 24 h, infected with a laboratory strain of HIV (HTLV-IIIB, multiplicity of infection = 0.001), then carried in culture for 14 days. Overall, when considering p24 antigen levels on days 7 and 11 in cultures established from cells exposed to 50 cGy, the maximal levels were significantly higher than those measured in the parallel control cultures taken as a whole (P < 0.05), with viral replication enhanced as much as 1000-fold in one case. No significant cytotoxicity was observed following exposure to doses up to 50 cGy. The mechanism of the observed effect remains unknown but may relate to direct gene activation and/or free radical generation, leading to such activation. To date, there is no evidence that viral stimulation occurs following therapeutic radiation in a clinical setting. (author)

Control of HIV-1 in Elite Suppressors despite Ongoing Replication and Evolution in Plasma Virus▿

Science.gov (United States)

O'Connell, Karen A.; Brennan, Timothy P.; Bailey, Justin R.; Ray, Stuart C.; Siliciano, Robert F.; Blankson, Joel N.

2010-01-01

A subset of HIV-1-infected patients known as elite controllers or suppressors (ES) control the virus naturally. We have previously demonstrated sequence discordance between proviral and plasma gag clones in ES, much of which can be attributed to selective pressure from the host (J. R. Bailey, T. M. Williams, R. F. Siliciano, and J. N. Blankson, J. Exp. Med. 203:1357-1369, 2006). However, it is not clear whether ongoing viral replication continues in ES once the control of viremia has been established or whether selective pressure impacts this evolution. The cytotoxic T-lymphocyte (CTL) response in ES often targets Gag and frequently is superior to that of HIV-1 progressors, partially due to the HLA class I alleles B*57/5801 and B*27, which are overrepresented in ES. We therefore examined longitudinal plasma and proviral gag sequences from HLA-B*57/5801 and -B*27 ES. Despite the highly conserved nature of gag, we observed clear evidence of evolution in the plasma virus, largely due to synonymous substitutions. In contrast, evolution was rare in proviral clones, suggesting that ongoing replication in ES does not permit the significant reseeding of the latent reservoir. Interestingly, there was little continual evolution in CTL epitopes, and we detected de novo CTL responses to autologous viral mutants. Thus, some ES control viremia despite ongoing replication and evolution. PMID:20444904

Human cyclin T1 expression ameliorates a T-cell-specific transcriptional limitation for HIV in transgenic rats, but is not sufficient for a spreading infection of prototypic R5 HIV-1 strains ex vivo

Directory of Open Access Journals (Sweden)

Littman Dan R

2009-01-01

Full Text Available Abstract Background Cells derived from native rodents have limits at distinct steps of HIV replication. Rat primary CD4 T-cells, but not macrophages, display a profound transcriptional deficit that is ameliorated by transient trans-complementation with the human Tat-interacting protein Cyclin T1 (hCycT1. Results Here, we generated transgenic rats that selectively express hCycT1 in CD4 T-cells and macrophages. hCycT1 expression in rat T-cells boosted early HIV gene expression to levels approaching those in infected primary human T-cells. hCycT1 expression was necessary, but not sufficient, to enhance HIV transcription in T-cells from individual transgenic animals, indicating that endogenous cellular factors are critical co-regulators of HIV gene expression in rats. T-cells from hCD4/hCCR5/hCycT1-transgenic rats did not support productive infection of prototypic wild-type R5 HIV-1 strains ex vivo, suggesting one or more significant limitation in the late phase of the replication cycle in this primary rodent cell type. Remarkably, we identify a replication-competent HIV-1 GFP reporter strain (R7/3 YU-2 Env that displays characteristics of a spreading, primarily cell-to-cell-mediated infection in primary T-cells from hCD4/hCCR5-transgenic rats. Moreover, the replication of this recombinant HIV-1 strain was significantly enhanced by hCycT1 transgenesis. The viral determinants of this so far unique replicative ability are currently unknown. Conclusion Thus, hCycT1 expression is beneficial to de novo HIV infection in a transgenic rat model, but additional genetic manipulations of the host or virus are required to achieve full permissivity.

Assessing Human Immunodeficiency Virus Type 1 Tropism: Comparison of Assays Using Replication-Competent Virus versus Plasma-Derived Pseudotyped Virions ▿

Science.gov (United States)

Hosoya, Noriaki; Su, Zhaohui; Wilkin, Timothy; Gulick, Roy M.; Flexner, Charles; Hughes, Michael D.; Skolnik, Paul R.; Giguel, Françoise; Greaves, Wayne L.; Coakley, Eoin; Kuritzkes, Daniel R.

2009-01-01

Detection of CXCR4-using human immunodeficiency virus by the Trofile assay was compared to that by assays using virus isolates or replication-competent recombinants. Concordance with the Trofile assay was good, but assays using replicating viruses did not increase substantially the ability to detect the presence of CXCR4-using virus. PMID:19494074

Transgene Expression and Host Cell Responses to Replication-Defective, Single-Cycle, and Replication-Competent Adenovirus Vectors

Directory of Open Access Journals (Sweden)

Catherine M. Crosby

2017-02-01

Full Text Available Most adenovirus (Ad vectors are E1 gene deleted replication defective (RD-Ad vectors that deliver one transgene to the cell and all expression is based on that one gene. In contrast, E1-intact replication-competent Ad (RC-Ad vectors replicate their DNA and their transgenes up to 10,000-fold, amplifying transgene expression markedly higher than RD-Ad vectors. While RC-Ad are more potent, they run the real risk of causing adenovirus infections in vector recipients and those that administer them. To gain the benefits of transgene amplification, but avoid the risk of Ad infections, we developed “single cycle” Ad (SC-Ad vectors. SC-Ads amplify transgene expression and generated markedly stronger and more persistent immune responses than RD-Ad as expected. However, they also unexpectedly generated stronger immune responses than RC-Ad vectors. To explore the basis of this potency here, we compared gene expression and the cellular responses to infection to these vectors in vitro and in vivo. In vitro, in primary human lung epithelial cells, SC- and RC-Ad amplified their genomes more than 400-fold relative to RD-Ad with higher replication by SC-Ad. This replication translated into higher green fluorescent protein (GFP expression for 48 h by SC- and RC-Ad than by RD-Ad. In vitro, in the absence of an immune system, RD-Ad expression became higher by 72 h coincident with cell death mediated by SC- and RC-Ad and release of transgene product from the dying cells. When the vectors were compared in human THP-1 Lucia- interferon-stimulated gene (ISG cells, which are a human monocyte cell line that have been modified to quantify ISG activity, RC-Ad6 provoked significantly stronger ISG responses than RD- or SC-Ad. In mice, intravenous or intranasal injection produced up to 100-fold genome replication. Under these in vivo conditions in the presence of the immune system, luciferase expression by RC and SC-Ad was markedly higher than that by RD-Ad. In

Interleukin-7 induces HIV replication in primary naive T cells through a nuclear factor of activated T cell (NFAT)-dependent pathway

International Nuclear Information System (INIS)

Managlia, Elizabeth Z.; Landay, Alan; Al-Harthi, Lena

2006-01-01

Interleukin (IL)-7 plays several roles critical to T cell maturation, survival, and homeostasis. Because of these functions, IL-7 is under investigation as an immune-modulator for therapeutic use in lymphopenic clinical conditions, including HIV. We reported that naive T cells, typically not permissive to HIV, can be productively infected when pre-treated with IL-7. We evaluated the mechanism by which IL-7-mediates this effect. IL-7 potently up-regulated the transcriptional factor NFAT, but had no effect on NFκB. Blocking NFAT activity using a number of reagents, such as Cyclosporin A, FK-506, or the NFAT-specific inhibitor known as VIVIT peptide, all markedly reduced IL-7-mediated induction of HIV replication in naive T cells. Additional neutralization of cytokines present in IL-7-treated cultures and/or those that have NFAT-binding sequences within their promotors indicated that IL-10, IL-4, and most significantly IFNγ, all contribute to IL-7-induction of HIV productive replication in naive T cells. These data clarify the mechanism by which IL-7 can overcome the block to HIV productive infection in naive T cells, despite their quiescent cell status. These findings are relevant to the treatment of HIV disease and understanding HIV pathogenesis in the naive CD4+ T cell compartment, especially in light of the vigorous pursuit of IL-7 as an in vivo immune modulator

First-in-Human Evaluation of the Safety and Immunogenicity of an Intranasally Administered Replication-Competent Sendai Virus–Vectored HIV Type 1 Gag Vaccine: Induction of Potent T-Cell or Antibody Responses in Prime-Boost Regimens

Science.gov (United States)

Nyombayire, Julien; Anzala, Omu; Gazzard, Brian; Karita, Etienne; Bergin, Philip; Hayes, Peter; Kopycinski, Jakub; Omosa-Manyonyi, Gloria; Jackson, Akil; Bizimana, Jean; Farah, Bashir; Sayeed, Eddy; Parks, Christopher L.; Inoue, Makoto; Hironaka, Takashi; Hara, Hiroto; Shu, Tsugumine; Matano, Tetsuro; Dally, Len; Barin, Burc; Park, Harriet; Gilmour, Jill; Lombardo, Angela; Excler, Jean-Louis; Fast, Patricia; Laufer, Dagna S.; Cox, Josephine H.

2017-01-01

Background. We report the first-in-human safety and immunogenicity assessment of a prototype intranasally administered, replication-competent Sendai virus (SeV)–vectored, human immunodeficiency virus type 1 (HIV-1) vaccine. Methods. Sixty-five HIV-1–uninfected adults in Kenya, Rwanda, and the United Kingdom were assigned to receive 1 of 4 prime-boost regimens (administered at 0 and 4 months, respectively; ratio of vaccine to placebo recipients, 12:4): priming with a lower-dose SeV-Gag given intranasally, followed by boosting with an adenovirus 35–vectored vaccine encoding HIV-1 Gag, reverse transcriptase, integrase, and Nef (Ad35-GRIN) given intramuscularly (SLA); priming with a higher-dose SeV-Gag given intranasally, followed by boosting with Ad35-GRIN given intramuscularly (SHA); priming with Ad35-GRIN given intramuscularly, followed by boosting with a higher-dose SeV-Gag given intranasally (ASH); and priming and boosting with a higher-dose SeV-Gag given intranasally (SHSH). Results. All vaccine regimens were well tolerated. Gag-specific IFN-γ enzyme-linked immunospot–determined response rates and geometric mean responses were higher (96% and 248 spot-forming units, respectively) in groups primed with SeV-Gag and boosted with Ad35-GRIN (SLA and SHA) than those after a single dose of Ad35-GRIN (56% and 54 spot-forming units, respectively) or SeV-Gag (55% and 59 spot-forming units, respectively); responses persisted for ≥8 months after completion of the prime-boost regimen. Functional CD8+ T-cell responses with greater breadth, magnitude, and frequency in a viral inhibition assay were also seen in the SLA and SHA groups after Ad35-GRIN boost, compared with those who received either vaccine alone. SeV-Gag did not boost T-cell counts in the ASH group. In contrast, the highest Gag-specific antibody titers were seen in the ASH group. Mucosal antibody responses were sporadic. Conclusions. SeV-Gag primed functional, durable HIV-specific T

The role of Vif oligomerization and RNA chaperone activity in HIV-1 replication.

Science.gov (United States)

Batisse, Julien; Guerrero, Santiago; Bernacchi, Serena; Sleiman, Dona; Gabus, Caroline; Darlix, Jean-Luc; Marquet, Roland; Tisné, Carine; Paillart, Jean-Christophe

2012-11-01

The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells that involve most natural HIV-1 target cells. Vif counteracts the packaging of two cellular cytidine deaminases named APOBEC3G (A3G) and A3F by diverse mechanisms including the recruitment of an E3 ubiquitin ligase complex and the proteasomal degradation of A3G/A3F, the inhibition of A3G mRNA translation or by a direct competition mechanism. In addition, Vif appears to be an active partner of the late steps of viral replication by participating in virus assembly and Gag processing, thus regulating the final stage of virion formation notably genomic RNA dimerization and by inhibiting the initiation of reverse transcription. Vif is a small pleiotropic protein with multiple domains, and recent studies highlighted the importance of Vif conformation and flexibility in counteracting A3G and in binding RNA. In this review, we will focus on the oligomerization and RNA chaperone properties of Vif and show that the intrinsic disordered nature of some Vif domains could play an important role in virus assembly and replication. Experimental evidence demonstrating the RNA chaperone activity of Vif will be presented. Copyright © 2012 Elsevier B.V. All rights reserved.

Fluorescent reporter signals, EGFP and DsRed, encoded in HIV-1 facilitate the detection of productively infected cells and cell-associated viral replication levels

Directory of Open Access Journals (Sweden)

Kazutaka eTerahara

2012-01-01

Full Text Available Flow cytometric analysis is a reliable and convenient method for investigating molecules at the single cell level. Previously, recombinant human immunodeficiency virus type 1 (HIV-1 strains were constructed that express a fluorescent reporter, either enhanced green fluorescent protein or DsRed, which allow the monitoring of HIV-1-infected cells by flow cytometry. The present study further investigated the potential of these recombinant viruses in terms of whether the HIV-1 fluorescent reporters would be helpful in evaluating viral replication based on fluorescence intensity. When primary CD4+ T cells were infected with recombinant viruses, the fluorescent reporter intensity measured by flow cytometry was associated with the level of CD4 downmodulation and Gag p24 expression in infected cells. Interestingly, some HIV-1-infected cells, in which CD4 was only moderately downmodulated, were reporter-positive but Gag p24-negative. Furthermore, when the activation status of primary CD4+ T cells was modulated by T cell receptor-mediated stimulation, we confirmed the preferential viral production upon strong stimulation and showed that the intensity of the fluorescent reporter within a proportion of HIV-1-infected cells was correlated with the viral replication level. These findings indicate that a fluorescent reporter encoded within HIV-1 is useful for the sensitive detection of productively-infected cells at different stages of infection and for evaluating cell-associated viral replication at the single cell level.

The effect of antiretroviral intensification with dolutegravir on residual virus replication in HIV-infected individuals: a randomised, placebo-controlled, double-blind trial.

Science.gov (United States)

Rasmussen, Thomas A; McMahon, James H; Chang, J Judy; Audsley, Jennifer; Rhodes, Ajantha; Tennakoon, Surekha; Dantanarayana, Ashanti; Spelman, Tim; Schmidt, Tina; Kent, Stephen J; Morcilla, Vincent; Palmer, Sarah; Elliott, Julian H; Lewin, Sharon R

2018-04-06

Whether ongoing virus replication occurs in HIV-infected individuals on antiretroviral therapy (ART) is unclear; therefore, whether residual virus replication is a barrier to achieving a cure for HIV is also unknown. We aimed to establish whether ART intensification with dolutegravir would reveal or affect residual virus replication in HIV-infected individuals on suppressive treatment. In this randomised, placebo-controlled, double-blind trial, we enrolled HIV-infected adults (aged 18 years and older) receiving combination ART (at least three agents) for at least 3 years from the Alfred Hospital and Melbourne Sexual Health Centre, Melbourne, VIC, Australia. Eligible participants had fewer than 50 copies per mL HIV-1 plasma RNA for more than 3 years and fewer than 20 copies per mL at screening and two CD4 counts higher than 350 cells per μL in the previous 24 months including screening. Participants were randomly assigned (1:1) to receive 50 mg oral dolutegravir or placebo once a day for 56 days in addition to background ART. Follow-up was done at days 1, 3, 7, 14, 28, 56, and 84. The primary outcome was the change from baseline in frequency of 2-long terminal repeat (2-LTR) circles in peripheral blood CD4 cells at day 7. This trial is registered with ClinicalTrials.gov, number NCT02500446. Between Sept 21, 2015, and Sept 19, 2016, 46 individuals were screened for inclusion. 40 were eligible for inclusion and were randomly assigned to the dolutegravir (n=21) or placebo group (n=19). All enrolled participants completed the study procedures and no individuals were lost to follow up. All participants were on suppressive ART with 12% receiving protease inhibitors and the others non-nucleoside reverse transcriptase inhibitors. Median 2-LTR circles fold-change from baseline to day 7 was -0·17 (IQR -0·90 to 0·90) in the dolutegravir group and -0·26 (-1·00 to 1·17) in the placebo group (p=0·17). The addition of dolutegravir to pre-existing ART regimens was safe and

Metabolic Abnormalities and Viral Replication is Associated with Biomarkers of Vascular Dysfunction in HIV-Infected Children

Science.gov (United States)

Miller, Tracie L.; Borkowsky, William; DiMeglio, Linda A.; Dooley, Laurie; Geffner, Mitchell E.; Hazra, Rohan; McFarland, Elizabeth J.; Mendez, Armando J.; Patel, Kunjal; Siberry, George K.; Van Dyke, Russell B.; Worrell, Carol J.; Jacobson, Denise L.

2011-01-01

Objectives Human immunodeficiency virus (HIV)-infected children may be at risk for premature cardiovascular disease. We compared levels of biomarkers of vascular dysfunction among HIV-infected children with and without hyperlipidemia to HIV-exposed, uninfected children (HEU) enrolled in the Pediatric HIV/AIDS Cohort Study (PHACS), and determined factors associated with these biomarkers. Design Prospective cohort study Methods Biomarkers of inflammation (C-reactive protein (CRP), interleukin-6 (IL-6), and monocyte chemoattractant protein-1 (MCP1)); coagulant dysfunction (fibrinogen and P-selectin); endothelial dysfunction (soluble intracellular cell adhesion molecule-1 (sICAM), soluble vascular cell adhesion molecule-1 (sVCAM), and E-selectin); and metabolic dysfunction (adiponectin) were measured in 226 HIV-infected and 140 HEU children. Anthropometry, body composition, lipids, glucose, insulin, HIV disease severity, and antiretroviral therapy were recorded. Results The median ages were 12.3 y (HIV-infected) and 10.1 y (HEU). Body mass index (BMI) Z-scores, waist and hip circumference, and percent body fat were lower among HIV-infected. Total and non-HDL cholesterol and triglycerides were higher in HIV-infected children. HIV-infected children had higher MCP-1, fibrinogen, sICAM, and sVCAM levels. In multivariable analyses in the HIV-infected children alone, BMI z-score was associated with higher CRP and fibrinogen, but lower MCP-1 and sVCAM. Unfavorable lipid profiles were positively associated with IL6, MCP1, fibrinogen, and P- and E-selectin, whereas increased HIV viral load was associated with markers of inflammation (MCP1 and CRP) and endothelial dysfunction (sICAM and sVCAM). Conclusions HIV-infected children have higher levels of biomarkers of vascular dysfunction than do HEU children. Risk factors associated with higher biomarkers include unfavorable lipid levels and active HIV replication. PMID:22136114

Integrating Prevention of Mother to Child HIV Transmission competencies into the nursing curriculum: Methodological lessons from a university-based undergraduate programme.

Science.gov (United States)

Mbombo, Nomafrench; Bimerew, Million

2012-11-14

South Africa (SA) has the highest number of women infected with HIV and AIDS during pregnancy, which results in more than 70 000 infected babies being born each year AIDS is the major contributor to maternal and child morbidities and mortalities in the country. To combat this, the SA government has developed a national policy to prevent mother-to-child HIV transmission (PMTCT). However, for effective implementation of this policy, there is a dire need for a competent, skilled health worker to render the service. In response to this, the School of Nursing at the University of the Western Cape has integrated PMTCT competencies into the undergraduate Bachelor of Nursing Science curriculum. In this paper, we described teaching and learning approaches used to integrate PMTCT competencies, including the skills laboratory methodology and case-based learning, as well as a portfolio of evidence assessment tool. A quantitative descriptive design was used to analyse data collected from students in regard to assessment of PMTCT competencies achieved. The study used the conceptual framework of Lenburg's competency outcomes and performance assessment model, which focuses on competency development and assessment in a clinical environment. HIV competencies, including PMTCT, should be integrated both theoretically and at service delivery into other nursing and midwifery competencies, including assessment strategies. Provincial policies in provision of antiretrovirals by nurses and midwives become barriers to successful implementation of PMTCT, resulting in limited learning opportunities for students to practice PMTCT competencies. Further research is required to assess an attribute, affect, which is another prong for competencies.

Clonal expansion of genome-intact HIV-1 in functionally polarized Th1 CD4+ T cells.

Science.gov (United States)

Lee, Guinevere Q; Orlova-Fink, Nina; Einkauf, Kevin; Chowdhury, Fatema Z; Sun, Xiaoming; Harrington, Sean; Kuo, Hsiao-Hsuan; Hua, Stephane; Chen, Hsiao-Rong; Ouyang, Zhengyu; Reddy, Kavidha; Dong, Krista; Ndung'u, Thumbi; Walker, Bruce D; Rosenberg, Eric S; Yu, Xu G; Lichterfeld, Mathias

2017-06-30

HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells.

Enrichment of intersubtype HIV-1 recombinants in a dual infection system using HIV-1 strain-specific siRNAs

Science.gov (United States)

2011-01-01

env regions suggest that other mechanisms are at play. Conclusion These findings show that siRNAs can be used as an efficient in vitro tool for enriching recombinants, to facilitate further study on mechanisms of intersubytpe HIV-1 recombination, and to generate replication-competent intersubtype recombinant proteins with a breadth in HIV-1 diversity for future vaccine studies. PMID:21232148

Integrated and Total HIV-1 DNA Predict Ex Vivo Viral Outgrowth.

Directory of Open Access Journals (Sweden)

Maja Kiselinova

2016-03-01

Full Text Available The persistence of a reservoir of latently infected CD4 T cells remains one of the major obstacles to cure HIV. Numerous strategies are being explored to eliminate this reservoir. To translate these efforts into clinical trials, there is a strong need for validated biomarkers that can monitor the reservoir over time in vivo. A comprehensive study was designed to evaluate and compare potential HIV-1 reservoir biomarkers. A cohort of 25 patients, treated with suppressive antiretroviral therapy was sampled at three time points, with median of 2.5 years (IQR: 2.4-2.6 between time point 1 and 2; and median of 31 days (IQR: 28-36 between time point 2 and 3. Patients were median of 6 years (IQR: 3-12 on ART, and plasma viral load (<50 copies/ml was suppressed for median of 4 years (IQR: 2-8. Total HIV-1 DNA, unspliced (us and multiply spliced HIV-1 RNA, and 2LTR circles were quantified by digital PCR in peripheral blood, at 3 time points. At the second time point, a viral outgrowth assay (VOA was performed, and integrated HIV-1 DNA and relative mRNA expression levels of HIV-1 restriction factors were quantified. No significant change was found for long- and short-term dynamics of all HIV-1 markers tested in peripheral blood. Integrated HIV-1 DNA was associated with total HIV-1 DNA (p<0.001, R² = 0.85, us HIV-1 RNA (p = 0.029, R² = 0.40, and VOA (p = 0.041, R2 = 0.44. Replication-competent virus was detected in 80% of patients by the VOA and it correlated with total HIV-1 DNA (p = 0.039, R² = 0.54. The mean quantification difference between Alu-PCR and VOA was 2.88 log10, and 2.23 log10 between total HIV-1 DNA and VOA. The levels of usHIV-1 RNA were inversely correlated with mRNA levels of several HIV-1 restriction factors (TRIM5α, SAMHD1, MX2, SLFN11, pSIP1. Our study reveals important correlations between the viral outgrowth and total and integrated HIV-1 DNA measures, suggesting that the total pool of HIV-1 DNA may predict the size of the

Experiences with HIV testing, entry, and engagement in care by HIV-infected women of color, and the need for autonomy, competency, and relatedness.

Science.gov (United States)

Quinlivan, E Byrd; Messer, Lynne C; Adimora, Adaora A; Roytburd, Katya; Bowditch, Natasha; Parnell, Heather; Seay, Julia; Bell, Lynda; Pierce, Jonah K

2013-07-01

Self-determination theory examines the needs of people adopting new behaviors but has not been applied to the adoption of HIV healthcare behaviors. The current study applied self-determination theory to descriptions of healthcare behaviors adopted by ethnic minority women after an HIV diagnosis. Women of color were asked to describe their experiences with HIV testing, entry, and engagement-in-care in qualitative interviews and focus groups. Participants were mostly African-American (88%), over 40 years old (70%), had been diagnosed for more than 6 years (87%) and had disclosed their HIV infection to more than 3 people (73%). Women described unmet self-determination needs at different time points along the HIV Continuum of Care. Women experienced a significant loss of autonomy at the time of HIV diagnosis. Meeting competency and relatedness needs assisted women in entry and engagement-in-care. However, re-establishing autonomy was a key element for long-term engagement-in-care. Interventions that satisfy these needs at the optimal time point in care could improve diagnosis, entry-to-care, and retention-in-care for women living with HIV.

The Tat protein of human immunodeficiency virus-1 enhances hepatitis C virus replication through interferon gamma-inducible protein-10

Directory of Open Access Journals (Sweden)

Qu Jing

2012-04-01

Full Text Available Abstract Background Co-infection with human immunodeficiency virus-1 (HIV-1 and hepatitis C virus (HCV is associated with faster progression of liver disease and an increase in HCV persistence. However, the mechanism by which HIV-1 accelerates the progression of HCV liver disease remains unknown. Results HIV-1/HCV co-infection is associated with increased expression of interferon gamma-induced protein-10 (IP-10 mRNA in peripheral blood mononuclear cells (PBMCs. HCV RNA levels were higher in PBMCs of patients with HIV-1/HCV co-infection than in patients with HCV mono-infection. HIV-1 Tat and IP-10 activated HCV replication in a time-dependent manner, and HIV-1 Tat induced IP-10 production. In addition, the effect of HIV-1 Tat on HCV replication was blocked by anti-IP-10 monoclonal antibody, demonstrating that the effect of HIV-1 Tat on HCV replication depends on IP-10. Taken together, these results suggest that HIV-1 Tat protein activates HCV replication by upregulating IP-10 production. Conclusions HIV-1/HCV co-infection is associated with increased expression of IP-10 mRNA and replication of HCV RNA. Furthermore, both HIV-1 Tat and IP-10 activate HCV replication. HIV-1 Tat activates HCV replication by upregulating IP-10 production. These results expand our understanding of HIV-1 in HCV replication and the mechanism involved in the regulation of HCV replication mediated by HIV-1 during co-infection.

Latent HIV reservoirs exhibit inherent resistance to elimination by CD8+ T cells.

Science.gov (United States)

Huang, Szu-Han; Ren, Yanqin; Thomas, Allison S; Chan, Dora; Mueller, Stefanie; Ward, Adam R; Patel, Shabnum; Bollard, Catherine M; Cruz, Conrad Russell; Karandish, Sara; Truong, Ronald; Macedo, Amanda B; Bosque, Alberto; Kovacs, Colin; Benko, Erika; Piechocka-Trocha, Alicja; Wong, Hing; Jeng, Emily; Nixon, Douglas F; Ho, Ya-Chi; Siliciano, Robert F; Walker, Bruce D; Jones, R Brad

2018-02-01

The presence of persistent, latent HIV reservoirs in CD4+ T cells obstructs current efforts to cure infection. The so-called kick-and-kill paradigm proposes to purge these reservoirs by combining latency-reversing agents with immune effectors such as cytotoxic T lymphocytes. Support for this approach is largely based on success in latency models, which do not fully reflect the makeup of latent reservoirs in individuals on long-term antiretroviral therapy (ART). Recent studies have shown that CD8+ T cells have the potential to recognize defective proviruses, which comprise the vast majority of all infected cells, and that the proviral landscape can be shaped over time due to in vivo clonal expansion of infected CD4+ T cells. Here, we have shown that treating CD4+ T cells from ART-treated individuals with combinations of potent latency-reversing agents and autologous CD8+ T cells consistently reduced cell-associated HIV DNA, but failed to deplete replication-competent virus. These CD8+ T cells recognized and potently eliminated CD4+ T cells that were newly infected with autologous reservoir virus, ruling out a role for both immune escape and CD8+ T cell dysfunction. Thus, our results suggest that cells harboring replication-competent HIV possess an inherent resistance to CD8+ T cells that may need to be addressed to cure infection.

A heterologous prime-boosting strategy with replicating Vaccinia virus vectors and plant-produced HIV-1 Gag/dgp41 virus-like particles

Energy Technology Data Exchange (ETDEWEB)

Meador, Lydia R. [Ira A. Fulton School of Engineering, Arizona State University, Tempe, AZ (United States); Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); Kessans, Sarah A. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Kilbourne, Jacquelyn; Kibler, Karen V. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); Pantaleo, Giuseppe [Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois, University of Lausanne, Lausanne (Switzerland); Swiss Vaccine Research Institute, Lausanne (Switzerland); Roderiguez, Mariano Esteban [Department of Molecular and Cellular Biology, Centro Nacional de Biotecnologia – CSIC, Madrid (Spain); Blattman, Joseph N. [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Jacobs, Bertram L., E-mail: bjacobs@asu.edu [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States); Mor, Tsafrir S., E-mail: tsafrir.mor@asu.edu [Center for Infectious Diseases and Vaccinology, The Biodesign Institute, Arizona State University, Tempe, AZ (United States); School of Life Sciences, Arizona State University, Tempe, AZ (United States)

2017-07-15

Showing modest efficacy, the RV144 HIV-1 vaccine clinical trial utilized a non-replicating canarypox viral vector and a soluble gp120 protein boost. Here we built upon the RV144 strategy by developing a novel combination of a replicating, but highly-attenuated Vaccinia virus vector, NYVAC-KC, and plant-produced HIV-1 virus-like particles (VLPs). Both components contained the full-length Gag and a membrane anchored truncated gp41 presenting the membrane proximal external region with its conserved broadly neutralizing epitopes in the pre-fusion conformation. We tested different prime/boost combinations of these components in mice and showed that the group primed with NYVAC-KC and boosted with both the viral vectors and plant-produced VLPs have the most robust Gag-specific CD8 T cell responses, at 12.7% of CD8 T cells expressing IFN-γ in response to stimulation with five Gag epitopes. The same immunization group elicited the best systemic and mucosal antibody responses to Gag and dgp41 with a bias towards IgG1. - Highlights: • We devised a prime/boost anti HIV-1 vaccination strategy modeled after RV144. • We used plant-derived virus-like particles (VLPs) consisting of Gag and dgp41. • We used attenuated, replicating vaccinia virus vectors expressing the same antigens. • The immunogens elicited strong cellular and humoral immune responses.

Differentiating Immune Cell Targets in Gut-Associated Lymphoid Tissue for HIV Cure.

Science.gov (United States)

Khan, Shahzada; Telwatte, Sushama; Trapecar, Martin; Yukl, Steven; Sanjabi, Shomyseh

2017-11-01

The single greatest challenge to an HIV cure is the persistence of latently infected cells containing inducible, replication-competent proviral genomes, which constitute only a small fraction of total or infected cells in the body. Although resting CD4 + T cells in the blood are a well-known source of viral rebound, more than 90% of the body's lymphocytes reside elsewhere. Many are in gut tissue, where HIV DNA levels per million CD4 + T cells are considerably higher than in the blood. Despite the significant contribution of gut tissue to viral replication and persistence, little is known about the cell types that support persistence of HIV in the gut; importantly, T cells in the gut have phenotypic, functional, and survival properties that are distinct from T cells in other tissues. The mechanisms by which latency is established and maintained will likely depend on the location and cytokine milieu surrounding the latently infected cells in each compartment. Therefore, successful HIV cure strategies require identification and characterization of the exact cell types that support viral persistence, particularly in the gut. In this review, we describe the seeding of the latent HIV reservoir in the gut mucosa; highlight the evidence for compartmentalization and depletion of T cells; summarize the immunologic consequences of HIV infection within the gut milieu; propose how the damaged gut environment may promote the latent HIV reservoir; and explore several immune cell targets in the gut and their place on the path toward HIV cure.

First-in-Human Evaluation of the Safety and Immunogenicity of an Intranasally Administered Replication-Competent Sendai Virus-Vectored HIV Type 1 Gag Vaccine: Induction of Potent T-Cell or Antibody Responses in Prime-Boost Regimens.

Science.gov (United States)

Nyombayire, Julien; Anzala, Omu; Gazzard, Brian; Karita, Etienne; Bergin, Philip; Hayes, Peter; Kopycinski, Jakub; Omosa-Manyonyi, Gloria; Jackson, Akil; Bizimana, Jean; Farah, Bashir; Sayeed, Eddy; Parks, Christopher L; Inoue, Makoto; Hironaka, Takashi; Hara, Hiroto; Shu, Tsugumine; Matano, Tetsuro; Dally, Len; Barin, Burc; Park, Harriet; Gilmour, Jill; Lombardo, Angela; Excler, Jean-Louis; Fast, Patricia; Laufer, Dagna S; Cox, Josephine H

2017-01-01

 We report the first-in-human safety and immunogenicity assessment of a prototype intranasally administered, replication-competent Sendai virus (SeV)-vectored, human immunodeficiency virus type 1 (HIV-1) vaccine.  Sixty-five HIV-1-uninfected adults in Kenya, Rwanda, and the United Kingdom were assigned to receive 1 of 4 prime-boost regimens (administered at 0 and 4 months, respectively; ratio of vaccine to placebo recipients, 12:4): priming with a lower-dose SeV-Gag given intranasally, followed by boosting with an adenovirus 35-vectored vaccine encoding HIV-1 Gag, reverse transcriptase, integrase, and Nef (Ad35-GRIN) given intramuscularly (S L A); priming with a higher-dose SeV-Gag given intranasally, followed by boosting with Ad35-GRIN given intramuscularly (S H A); priming with Ad35-GRIN given intramuscularly, followed by boosting with a higher-dose SeV-Gag given intranasally (AS H ); and priming and boosting with a higher-dose SeV-Gag given intranasally (S H S H ).  All vaccine regimens were well tolerated. Gag-specific IFN-γ enzyme-linked immunospot-determined response rates and geometric mean responses were higher (96% and 248 spot-forming units, respectively) in groups primed with SeV-Gag and boosted with Ad35-GRIN (S L A and S H A) than those after a single dose of Ad35-GRIN (56% and 54 spot-forming units, respectively) or SeV-Gag (55% and 59 spot-forming units, respectively); responses persisted for ≥8 months after completion of the prime-boost regimen. Functional CD8 + T-cell responses with greater breadth, magnitude, and frequency in a viral inhibition assay were also seen in the S L A and S H A groups after Ad35-GRIN boost, compared with those who received either vaccine alone. SeV-Gag did not boost T-cell counts in the AS H group. In contrast, the highest Gag-specific antibody titers were seen in the AS H group. Mucosal antibody responses were sporadic.  SeV-Gag primed functional, durable HIV-specific T-cell responses and boosted antibody

Occupational HIV Infection in a Research Laboratory With Unknown Mode of Transmission: A Case Report.

Science.gov (United States)

Soria, Alessandro; Alteri, Claudia; Scarlatti, Gabriella; Bertoli, Ada; Tolazzi, Monica; Balestra, Emanuela; Bellocchi, Maria Concetta; Continenza, Fabio; Carioti, Luca; Biasin, Mara; Trabattoni, Daria; Bandera, Alessandra; Ceccherini-Silberstein, Francesca; Perno, Carlo Federico; Gori, Andrea

2017-03-15

A laboratory worker was infected with human immunodeficiency virus (HIV) type 1 in a biosafety level 2 containment facility, without any apparent breach. Through full-genome sequencing and phylogenetic analyses, we could identify the source of infection in a replication-competent clone that unknowingly contaminated a safe experiment. Mode of transmission remains unclear. Caution is warranted when handling HIV-derived constructs. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Generation of H9 T-cells stably expressing a membrane-bound form of the cytoplasmic tail of the Env-glycoprotein: lack of transcomplementation of defective HIV-1 virions encoding C-terminally truncated Env

Directory of Open Access Journals (Sweden)

Bosch Valerie

2006-05-01

Full Text Available Abstract H9-T-cells do not support the replication of mutant HIV-1 encoding Env protein lacking its long cytoplasmic C-terminal domain (Env-CT. Here we describe the generation of a H9-T-cell population constitutively expressing the HIV-1 Env-CT protein domain anchored in the cellular membrane by it homologous membrane-spanning domain (TMD. We confirmed that the Env-TMD-CT protein was associated with cellular membranes, that its expression did not have any obvious cytotoxic effects on the cells and that it did not affect wild-type HIV-1 replication. However, as measured in both a single-round assay as well as in spreading infections, replication competence of mutant pNL-Tr712, lacking the Env-CT, was not restored in this H9 T-cell population. This means that the Env-CT per se cannot transcomplement the replication block of HIV-1 virions encoding C-terminally truncated Env proteins and suggests that the Env-CT likely exerts its function only in the context of the complete Env protein.

Optical tweezers reveal how proteins alter replication

Science.gov (United States)

Chaurasiya, Kathy

Single molecule force spectroscopy is a powerful method that explores the DNA interaction properties of proteins involved in a wide range of fundamental biological processes such as DNA replication, transcription, and repair. We use optical tweezers to capture and stretch a single DNA molecule in the presence of proteins that bind DNA and alter its mechanical properties. We quantitatively characterize the DNA binding mechanisms of proteins in order to provide a detailed understanding of their function. In this work, we focus on proteins involved in replication of Escherichia coli (E. coli ), endogenous eukaryotic retrotransposons Ty3 and LINE-1, and human immunodeficiency virus (HIV). DNA polymerases replicate the entire genome of the cell, and bind both double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA) during DNA replication. The replicative DNA polymerase in the widely-studied model system E. coli is the DNA polymerase III subunit alpha (DNA pol III alpha). We use optical tweezers to determine that UmuD, a protein that regulates bacterial mutagenesis through its interactions with DNA polymerases, specifically disrupts alpha binding to ssDNA. This suggests that UmuD removes alpha from its ssDNA template to allow DNA repair proteins access to the damaged DNA, and to facilitate exchange of the replicative polymerase for an error-prone translesion synthesis (TLS) polymerase that inserts nucleotides opposite the lesions, so that bacterial DNA replication may proceed. This work demonstrates a biophysical mechanism by which E. coli cells tolerate DNA damage. Retroviruses and retrotransposons reproduce by copying their RNA genome into the nuclear DNA of their eukaryotic hosts. Retroelements encode proteins called nucleic acid chaperones, which rearrange nucleic acid secondary structure and are therefore required for successful replication. The chaperone activity of these proteins requires strong binding affinity for both single- and double-stranded nucleic

HIV Infection: Advances Toward a Cure.

Science.gov (United States)

Douek, Daniel C

2018-04-01

Achieving cure of HIV infection requires eliminating all replication-competent virus from the reservoir of latently infected cells or completely inhibiting infected cells from emerging from latency. Strategies include very early use of antiretroviral therapy; hematopoietic stem cell transplantation; "shock-and-kill" approaches; immune therapy with immune checkpoint inhibitors; gene therapy, including use of CC chemokine receptor 5-modified CD4+ T cells; and broadly neutralizing antibody therapy. Success is likely to require a combination of approaches. This article summarizes a presentation by Daniel C. Douek, MD, PhD, at the IAS-USA continuing education program held in Berkeley, California, in May 2017.

Challenges in detecting HIV persistence during potentially curative interventions: a study of the Berlin patient.

Directory of Open Access Journals (Sweden)

Steven A Yukl

Full Text Available There is intense interest in developing curative interventions for HIV. How such a cure will be quantified and defined is not known. We applied a series of measurements of HIV persistence to the study of an HIV-infected adult who has exhibited evidence of cure after allogeneic hematopoietic stem cell transplant from a homozygous CCR5Δ32 donor. Samples from blood, spinal fluid, lymph node, and gut were analyzed in multiple laboratories using different approaches. No HIV DNA or RNA was detected in peripheral blood mononuclear cells (PBMC, spinal fluid, lymph node, or terminal ileum, and no replication-competent virus could be cultured from PBMCs. However, HIV RNA was detected in plasma (2 laboratories and HIV DNA was detected in the rectum (1 laboratory at levels considerably lower than those expected in ART-suppressed patients. It was not possible to obtain sequence data from plasma or gut, while an X4 sequence from PBMC did not match the pre-transplant sequence. HIV antibody levels were readily detectable but declined over time; T cell responses were largely absent. The occasional, low-level PCR signals raise the possibility that some HIV nucleic acid might persist, although they could also be false positives. Since HIV levels in well-treated individuals are near the limits of detection of current assays, more sensitive assays need to be developed and validated. The absence of recrudescent HIV replication and waning HIV-specific immune responses five years after withdrawal of treatment provide proof of a clinical cure.

Viral persistence, latent reservoir, and blips: a review on HIV-1 dynamics and modeling during HAART and related treatment implications

Energy Technology Data Exchange (ETDEWEB)

Rong, Libin [Los Alamos National Laboratory; Perelson, Alan [Los Alamos National Laboratory

2008-01-01

HIV-1 eradication from infected individuals has not been achieved with the use of highly active antiretroviral therapy (HAART) for a prolonged period of time. The cellular reservoir for HIV-1 in resting memory CD4{sup +} T cells remains a major obstacle to viral elimination. The reservoir does not decay significantly over long periods of time as is able to release replication competent HIV-1 upon cell activation. Residual ongoing viral replication may likely occur in many patients because low levels of virus can be detected in plasma by sensitive assays and transient episodes of viremia, or HIV-1 blips, are often observed in patients even with successful viral suppression for many years. Here we review our current knowledge of the factors contributing to viral persistence, the latent reservoir, and blips, and mathematical models developed to explore them and their relationships. We show how mathematical modeling can help improve our understanding of HIV-1 dynamics in patients on HAART and the quantitative events underlying HIV-1 latency, reservoir stability, low-level viremic persistence, and emergence of intermittent viral blips. We also discuss treatment implications related to these studies.

Comparative analysis of measures of viral reservoirs in HIV-1 eradication studies.

Directory of Open Access Journals (Sweden)

Susanne Eriksson

2013-02-01

Full Text Available HIV-1 reservoirs preclude virus eradication in patients receiving highly active antiretroviral therapy (HAART. The best characterized reservoir is a small, difficult-to-quantify pool of resting memory CD4(+ T cells carrying latent but replication-competent viral genomes. Because strategies targeting this latent reservoir are now being tested in clinical trials, well-validated high-throughput assays that quantify this reservoir are urgently needed. Here we compare eleven different approaches for quantitating persistent HIV-1 in 30 patients on HAART, using the original viral outgrowth assay for resting CD4(+ T cells carrying inducible, replication-competent viral genomes as a standard for comparison. PCR-based assays for cells containing HIV-1 DNA gave infected cell frequencies at least 2 logs higher than the viral outgrowth assay, even in subjects who started HAART during acute/early infection. This difference may reflect defective viral genomes. The ratio of infected cell frequencies determined by viral outgrowth and PCR-based assays varied dramatically between patients. Although strong correlations with the viral outgrowth assay could not be formally excluded for most assays, correlations achieved statistical significance only for integrated HIV-1 DNA in peripheral blood mononuclear cells and HIV-1 RNA/DNA ratio in rectal CD4(+ T cells. Residual viremia was below the limit of detection in many subjects and did not correlate with the viral outgrowth assays. The dramatic differences in infected cell frequencies and the lack of a precise correlation between culture and PCR-based assays raise the possibility that the successful clearance of latently infected cells may be masked by a larger and variable pool of cells with defective proviruses. These defective proviruses are detected by PCR but may not be affected by reactivation strategies and may not require eradication to accomplish an effective cure. A molecular understanding of the discrepancy

An SIV/macaque model targeted to study HIV-associated neurocognitive disorders.

Science.gov (United States)

Beck, Sarah E; Queen, Suzanne E; Metcalf Pate, Kelly A; Mangus, Lisa M; Abreu, Celina M; Gama, Lucio; Witwer, Kenneth W; Adams, Robert J; Zink, M Christine; Clements, Janice E; Mankowski, Joseph L

2018-04-01

Simian immunodeficiency virus (SIV) infection of pigtailed macaques is a highly representative and well-characterized animal model for HIV neuropathogenesis studies that provides an excellent opportunity to study and develop prognostic markers of HIV-associated neurocognitive disorders (HAND) for HIV-infected individuals. SIV studies can be performed in a controlled setting that enhances reproducibility and offers high-translational value. Similar to observations in HIV-infected patients receiving antiretroviral therapy (ART), ongoing neurodegeneration and inflammation are present in SIV-infected pigtailed macaques treated with suppressive ART. By developing quantitative viral outgrowth assays that measure both CD4+ T cells and macrophages harboring replication competent SIV as well as a highly sensitive mouse-based viral outgrowth assay, we have positioned the SIV/pigtailed macaque model to advance our understanding of latent cellular reservoirs, including potential CNS reservoirs, to promote HIV cure. In addition to contributing to our understanding of the pathogenesis of HAND, the SIV/pigtailed macaque model also provides an excellent opportunity to test innovative approaches to eliminate the latent HIV reservoir in the brain.

Genome-Wide Search for Competing Endogenous RNAs Responsible for the Effects Induced by Ebola Virus Replication and Transcription Using a trVLP System

Directory of Open Access Journals (Sweden)

Zhong-Yi Wang

2017-11-01

Full Text Available Understanding how infected cells respond to Ebola virus (EBOV and how this response changes during the process of viral replication and transcription are very important for establishing effective antiviral strategies. In this study, we conducted a genome-wide screen to identify long non-coding RNAs (lncRNAs, circular RNAs (circRNAs, micro RNAs (miRNAs, and mRNAs differentially expressed during replication and transcription using a tetracistronic transcription and replication-competent virus-like particle (trVLP system that models the life cycle of EBOV in 293T cells. To characterize the expression patterns of these differentially expressed RNAs, we performed a series cluster analysis, and up- or down-regulated genes were selected to establish a gene co-expression network. Competing endogenous RNA (ceRNA networks based on the RNAs responsible for the effects induced by EBOV replication and transcription in human cells, including circRNAs, lncRNAs, miRNAs, and mRNAs, were constructed for the first time. Based on these networks, the interaction details of circRNA-chr19 were explored. Our results demonstrated that circRNA-chr19 targeting miR-30b-3p regulated CLDN18 expression by functioning as a ceRNA. These findings may have important implications for further studies of the mechanisms of EBOV replication and transcription. These RNAs potentially have important functions and may be promising targets for EBOV therapy.

Site-directed mutagenesis of HIV-1 vpu gene demonstrates two clusters of replication-defective mutants with distinct ability to down-modulate cell surface CD4 and tetherin

Directory of Open Access Journals (Sweden)

Masako Nomaguchi

2010-11-01

Full Text Available HIV-1 Vpu acts positively on viral infectivity by mediating CD4 degradation in endoplasmic reticulum and enhances virion release by counteracting a virion release restriction factor, tetherin. In order to define the impact of Vpu activity on HIV-1 replication, we have generated a series of site-specific proviral vpu mutants. Of fifteen mutants examined, seven exhibited a replication-defect similar to that of a vpu-deletion mutant in a lymphocyte cell line H9. These mutations clustered in narrow regions within transmembrane domain (TMD and cytoplasmic domain (CTD. Replication-defective mutants displayed the reduced ability to enhance virion release from a monolayer cell line HEp2 without exception. Upon transfection with Vpu expression vectors, neither TMD mutants nor CTD mutants blocked CD4 expression at the cell surface in another monolayer cell line MAGI. While TMD mutants were unable to down-modulate cell surface tetherin in HEp2 cells, CTD mutants did quite efficiently. Confocal microscopy analysis revealed the difference of intracellular localization between TMD and CTD mutants. In total, replication capability of HIV-1 carrying vpu mutations correlates well with the ability of Vpu to enhance virion release and to impede the cell surface expression of CD4 but not with the ability to down-modulate cell surface tetherin. Our results here suggest that efficient viral replication requires not only down-regulation of cell surface tetherin but also its degradation.

Exploring Teacher Competences and Learner Participation in HIV / AIDS Education through the Application of the Investigations Visions Action and Change (IVAC) Approach

DEFF Research Database (Denmark)

Kaki Nyami, Rachael

Within the field of participatory health education, this study made use of the IVAC approach as an intervention in the teaching of HIV/AIDS education. It aimed at exploring teachers competences and learner participation in HIV/AIDS education in two schools i Mutomo District, Kenya....

COMPETÊNCIA TÉCNICA NA PREVENÇÃO DO HIV/AIDS: VALIDAÇÃO DE UM INSTRUMENTO COMPETENCIA TÉCNICA EN LA PREVENCIÓN DE HIV/AIDS: VALIDACIÓN DE UN INSTRUMENTO TECHNICAL COMPETENCE IN THE PREVENTION OF HIV/AIDS: VALIDATION OF AN INSTRUMENT

Directory of Open Access Journals (Sweden)

Gilson de Vasconcelos Torres

2001-11-01

Full Text Available O estudo teve como objetivo construir e validar um instrumento para mensurar a competência técnica das ações educativas desenvolvidas pelo enfermeiro, na prevenção do HIV/AIDS nas unidades básicas de saúde (UBS. As etapas percorridas para a construção e testagem da fidedignidade do instrumento, foram: elaboração de itens relacionados a competência técnica na prevenção do HIV/AIDS; aplicação do instrumento numa amostra de 30 enfermeiros de diferentes instituições hospitalares e de ensino. O instrumento apresentou um nível satisfatório de estabilidade, homogeneidade e reprodutibilidade.El estudio tuvo como objetivo construir y validar un instrumento para medir la competencia de las acciones educativas desarrolladas por el enfermero en la prevención de HIV/AIDS en las unidades básicas de salud (UBS. Las etapas recorridas para la construcción y prueba de fidedignidad del instrumento, fueron: elaboración de ítems relacionados con la competencia técnica en la prevención de HIV/AIDS; aplicación del instrumento en una muestra de 30 enfermeros de diferentes instituciones hospitalarias y de enseñanza. El instrumento presentó un nivel satisfactorio de estabilidad, homogeneidad y reproductibilidad.This study aimed at building and validating an instrument to measure the technical competence of educational actions developed by nurses in the prevention of HIV/AIDS in Basic Health Units (BHU. The phases for constructing and testing the fidelity of the instrument were: elaboration of items related to the technical competence in the prevention of HIV/AIDS; application of the instrument in a sample of 30 male nurses from different hospitals and teaching institutions. The instrument presented a satisfactory level of stability, homogeneity and reproduction.

Development of antibody-modified chitosan nanoparticles for the targeted delivery of siRNA across the blood-brain barrier as a strategy for inhibiting HIV replication in astrocytes.

Science.gov (United States)

Gu, Jijin; Al-Bayati, Karam; Ho, Emmanuel A

2017-08-01

RNA interference (RNAi)-mediated gene silencing offers a novel treatment and prevention strategy for human immunodeficiency virus (HIV) infection. HIV was found to infect and replicate in human brain cells and can cause neuroinfections and neurological deterioration. We designed dual-antibody-modified chitosan/small interfering RNA (siRNA) nanoparticles to deliver siRNA across the blood-brain barrier (BBB) targeting HIV-infected brain astrocytes as a strategy for inhibiting HIV replication. We hypothesized that transferrin antibody and bradykinin B2 antibody could specifically bind to the transferrin receptor (TfR) and bradykinin B2 receptor (B2R), respectively, and deliver siRNA across the BBB into astrocytes as potential targeting ligands. In this study, chitosan nanoparticles (CS-NPs) were prepared by a complex coacervation method in the presence of siRNA, and antibody was chemically conjugated to the nanoparticles. The antibody-modified chitosan nanoparticles (Ab-CS-NPs) were spherical in shape, with an average particle size of 235.7 ± 10.2 nm and a zeta potential of 22.88 ± 1.78 mV. The therapeutic potential of the nanoparticles was evaluated based on their cellular uptake and gene silencing efficiency. Cellular accumulation and gene silencing efficiency of Ab-CS-NPs in astrocytes were significantly improved compared to non-modified CS-NPs and single-antibody-modified CS-NPs. These results suggest that the combination of anti-Tf antibody and anti-B2 antibody significantly increased the knockdown effect of siRNA-loaded nanoparticles. Thus, antibody-mediated dual-targeting nanoparticles are an efficient and promising delivery strategy for inhibiting HIV replication in astrocytes. Graphical abstract Graphic representation of dual-antibody-conjugated chitosan nanoparticles for the targeted delivery of siRNA across the blood-brain barrier (BBB) for inhibiting HIV replication in astrocytes. a Nanoparticle delivery to the BBB and penetration. b Tf

Rational development of radiopharmaceuticals for HIV-1

International Nuclear Information System (INIS)

Lau, Chuen-Yen; Maldarelli, Frank; Eckelman, William C.; Neumann, Ronald D.

2014-01-01

The global battle against HIV-1 would benefit from a sensitive and specific radiopharmaceutical to localize HIV-infected cells. Ideally, this probe would be able to identify latently infected host cells containing replication competent HIV sequences. Clinical and research applications would include assessment of reservoirs, informing clinical management by facilitating assessment of burden of infection in different compartments, monitoring disease progression and monitoring response to therapy. A “rational” development approach could facilitate efficient identification of an appropriate targeted radiopharmaceutical. Rational development starts with understanding characteristics of the disease that can be effectively targeted and then engineering radiopharmaceuticals to hone in on an appropriate target, which in the case of HIV-1 (HIV) might be an HIV-specific product on or in the host cell, a differentially expressed gene product, an integrated DNA sequence specific enzymatic activity, part of the inflammatory response, or a combination of these. This is different from the current approach that starts with a radiopharmaceutical for a target associated with a disease, mostly from autopsy studies, without a strong rationale for the potential to impact patient care. At present, no targeted therapies are available for HIV latency, although a number of approaches are under study. Here we discuss requirements for a radiopharmaceutical useful in strategies targeting persistently infected cells. The radiopharmaceutical for HIV should be developed based on HIV biology, studied in an animal model and then in humans, and ultimately used in clinical and research settings

Structure and possible function of a G-quadruplex in the long terminal repeat of the proviral HIV-1 genome.

Science.gov (United States)

De Nicola, Beatrice; Lech, Christopher J; Heddi, Brahim; Regmi, Sagar; Frasson, Ilaria; Perrone, Rosalba; Richter, Sara N; Phan, Anh Tuân

2016-07-27

The long terminal repeat (LTR) of the proviral human immunodeficiency virus (HIV)-1 genome is integral to virus transcription and host cell infection. The guanine-rich U3 region within the LTR promoter, previously shown to form G-quadruplex structures, represents an attractive target to inhibit HIV transcription and replication. In this work, we report the structure of a biologically relevant G-quadruplex within the LTR promoter region of HIV-1. The guanine-rich sequence designated LTR-IV forms a well-defined structure in physiological cationic solution. The nuclear magnetic resonance (NMR) structure of this sequence reveals a parallel-stranded G-quadruplex containing a single-nucleotide thymine bulge, which participates in a conserved stacking interaction with a neighboring single-nucleotide adenine loop. Transcription analysis in a HIV-1 replication competent cell indicates that the LTR-IV region may act as a modulator of G-quadruplex formation in the LTR promoter. Consequently, the LTR-IV G-quadruplex structure presented within this work could represent a valuable target for the design of HIV therapeutics. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Construction of Nef-positive doxycycline-dependent HIV-1 variants using bicistronic expression elements

Energy Technology Data Exchange (ETDEWEB)

Velden, Yme U. van der; Kleibeuker, Wendy; Harwig, Alex; Klaver, Bep; Siteur-van Rijnstra, Esther; Frankin, Esmay; Berkhout, Ben; Das, Atze T., E-mail: a.t.das@amc.uva.nl

2016-01-15

Conditionally replicating HIV-1 variants that can be switched on and off at will are attractive tools for HIV research. We previously developed a genetically modified HIV-1 variant that replicates exclusively when doxycycline (dox) is administered. The nef gene in this HIV-rtTA variant was replaced with the gene encoding the dox-dependent rtTA transcriptional activator. Because loss of Nef expression compromises virus replication in primary cells and precludes studies on Nef function, we tested different approaches to restore Nef production in HIV-rtTA. Strategies that involved translation via an EMCV or synthetic internal ribosome entry site (IRES) failed because these elements were incompatible with efficient virus replication. Fusion protein approaches with the FMDV 2A peptide and human ubiquitin were successful and resulted in genetically-stable Nef-expressing HIV-rtTA strains that replicate more efficiently in primary T-cells and human immune system (HIS) mice than Nef-deficient variants, thus confirming the positive effect of Nef on in vivo virus replication. - Highlights: • Different approaches to encode additional proteins in the HIV-1 genome were tested. • IRES translation elements are incompatible with efficient HIV-1 replication. • Ubiquitin and 2A fusion protein approaches allow efficient HIV-1 replication. • Doxycycline-controlled HIV-1 variants that encode all viral proteins were developed. • Nef stimulates HIV-rtTA replication in primary cells and human immune system mice.

Old plants newly discovered: Cassia sieberiana D.C. and Cassia abbreviata Oliv. Oliv. root extracts inhibit in vitro HIV-1c replication in peripheral blood mononuclear cells (PBMCs) by different modes of action.

Science.gov (United States)

Leteane, Melvin M; Ngwenya, Barbara N; Muzila, Mbaki; Namushe, Amos; Mwinga, John; Musonda, Rosemary; Moyo, Sikhulile; Mengestu, Yehualashete B; Abegaz, Berhanu M; Andrae-Marobela, Kerstin

2012-05-07

Despite advances in anti-retroviral therapy which has transformed HIV/AIDS from a fatal to a manageable chronic disease, increasing viral drug resistance, side effects and uneven access to anti-retroviral drugs remain considerable therapeutic challenges. Partly as a consequence of these shortcomings and partly based on the fact that HIV/AIDS gives rise to opportunistic infections whose symptoms have been managed in Africa in an HIV/AIDS-independent context by traditional healers for centuries, many HIV/AIDS patients use herbal medicines. The aim of this study was to screen selected medicinal plants from Botswana, used by traditional healers to treat/manage HIV/AIDS, for inhibitory activities on HIV replication. Based on an ethnomedical survey, ethanolic tannin-containing and tannin-free extracts from 10 medicinal plants were tested for inhibitory properties against a clone of HIV-1c (MJ(4)) measuring cytopathic effect protection and levels of viral p24 antigen in infected PBMCs. Cassia sieberiana D.C., Cassia abbreviata Oliv. Oliv. and Plumbago zeylanica L. extracts showed significant inhibition of HIV-1c (MJ(4)) replication. The inhibitory activity of the Plumbago zeylanica extract could be attributed to its tannin content. Anti-HIV activity of Cassia sieberiana root and bark extracts, and Cassia abbreviata root extracts occurred in a concentration-dependent manner with an effective concentration (EC(50)) of 65.1μg/ml, 85.3μg/ml and 102.8μg/ml, respectively. Experiments to elucidate possible mechanism(s) of action revealed that Cassia sieberiana root and bark extracts blocked HIV replication at its binding- (EC(50)=70.2μg/ml and 90.8μg/ml, respectively) and entry stage (EC(50)=88.9μg/ml and 100.5μg/ml, respectively) while Cassia abbreviata extracts did not. We report here for the first time a direct inhibitory effect on HIV-1c replication of extracts from two extremely popular medicinal plants, Cassia sieberiana and Cassia abbreviata. Considering the

The role of community conversations in facilitating local HIV competence: case study from rural Zimbabwe.

Science.gov (United States)

Campbell, Catherine; Nhamo, Mercy; Scott, Kerry; Madanhire, Claudius; Nyamukapa, Constance; Skovdal, Morten; Gregson, Simon

2013-04-17

This paper examines the potential for community conversations to strengthen positive responses to HIV in resource-poor environments. Community conversations are an intervention method through which local people work with a facilitator to collectively identify local strengths and challenges and brainstorm potential strategies for solving local problems. We conducted 18 community conversations (with six groups at three points in time) with a total of 77 participants in rural Zimbabwe (20% HIV positive). Participants were invited to reflect on how they were responding to the challenges of HIV, both as individuals and in community groups, and to think of ways to better support openness about HIV, kindness towards people living with HIV and greater community uptake of HIV prevention and treatment. Community conversations contributed to local HIV competence through (1) enabling participants to brainstorm concrete action plans for responding to HIV, (2) providing a forum to develop a sense of common purpose in relation to implementing these, (3) encouraging and challenging participants to overcome fear, denial and passivity, (4) providing an opportunity for participants to move from seeing themselves as passive recipients of information to active problem solvers, and (5) reducing silence and stigma surrounding HIV. Our discussion cautions that community conversations, while holding great potential to help communities recognize their potential strengths and capacities for responding more effectively to HIV, are not a magic bullet. Poverty, poor harvests and political instability frustrated and limited many participants' efforts to put their plans into action. On the other hand, support from outside the community, in this case the increasing availability of antiretroviral treatment, played a vital role in enabling communities to challenge stigma and envision new, more positive, ways of responding to the epidemic.

Induction and characterization of a replication competent cervid endogenous gammaretrovirus (CrERV) from mule deer cells.

Science.gov (United States)

Fábryová, Helena; Hron, Tomáš; Kabíčková, Hana; Poss, Mary; Elleder, Daniel

2015-11-01

Endogenous retroviruses (ERVs) were acquired during evolution of their host organisms after infection and mendelian inheritance in the germline by their exogenous counterparts. The ERVs can spread in the host genome and in some cases they affect the host phenotype. The cervid endogenous gammaretrovirus (CrERV) is one of only a few well-defined examples of evolutionarily recent invasion of mammalian genome by retroviruses. Thousands of insertionally polymorphic CrERV integration sites have been detected in wild ranging mule deer (Odocoileus hemionus) host populations. Here, we describe for the first time induction of replication competent CrERV by cocultivation of deer and human cells. We characterize the physical properties and tropism of the induced virus. The genomic sequence of the induced virus is phylogenetically related to the evolutionarily young endogenous CrERVs described so far. We also describe the level of replication block of CrERV on deer cells and its capacity to establish superinfection interference. Copyright © 2015 Elsevier Inc. All rights reserved.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

Science.gov (United States)

Perdigão, Pedro; Gaj, Thomas; Santa-Marta, Mariana; Barbas, Carlos F; Goncalves, Joao

2016-01-01

The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE) proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

Directory of Open Access Journals (Sweden)

Pedro Perdigão

Full Text Available The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

Obstacles to local-level AIDS competence in rural Zimbabwe: putting HIV prevention in context.

Science.gov (United States)

Nhamo, Mercy; Campbell, Catherine; Gregson, Simon

2010-01-01

We explore the wider social context of an HIV-prevention programme in rural Zimbabwe. We make no comment on the programme itself, rather seeking to examine the wider community dynamics into which it was inserted, to highlight how pre-existing social dynamics may have influenced community "readiness" to derive optimal benefit from the intervention. Using the concept of "the AIDS competent community", we analysed 44 interviews and 11 focus groups with local people. Despite high levels of HIV/AIDS-related knowledge, there were several ways gender, poverty and low literacy may have undermined its perceived relevance to peoples' lives. Lack of opportunities for dialogue in the social milieu beyond the intervention may have limited opportunities for translating factual AIDS knowledge into action plans, or sharing hidden individual experiences of HIV/AIDS-affected family members or friends, given stigma and denial. The initiative of women and young people to respond effectively to AIDS was limited in a context dominated by adult males. People spoke of HIV/AIDS in a passive and fatalistic way, expecting outsiders to solve the problem. This tendency was exacerbated given the community's previous experiences of HIV/AIDS-related NGOs, which had often regarded local people as unpaid volunteer labour rather than building their capacity to make significant decisions and play leadership roles in health programmes. Despite obstacles, however, there were many potential community strengths and resources. There were high levels of HIV/AIDS-related knowledge. Public denial of HIV/AIDS masked huge reservoirs of private support and kindness to AIDS-affected family and friends. There were many strong community organisations and clubs, potentially forming the springboard for more empowered community responses to HIV/AIDS. HIV/AIDS programmers should pay greater attention to community readiness for interventions, especially around: (1) identifying and anticipating pre-existing obstacles to

Replication capacity in relation to immunologic and virologic outcomes in HIV-1-infected treatment-naive subjects.

Science.gov (United States)

Skowron, Gail; Spritzler, John G; Weidler, Jodi; Robbins, Gregory K; Johnson, Victoria A; Chan, Ellen S; Asmuth, David M; Gandhi, Rajesh T; Lie, Yolanda; Bates, Michael; Pollard, Richard B

2009-03-01

To evaluate the association between baseline (BL) replication capacity (RC) (RCBL) and immunologic/virologic parameters (at BL and after 48 weeks on therapy) in HIV-1-infected subjects initiating antiretroviral therapy. RCBL was determined using a modified Monogram PhenoSense HIV drug susceptibility assay on plasma HIV-1 from 321 treatment-naive subjects from AIDS Clinical Trials Group 384. Univariate and multivariable analyses were performed to determine the association of RCBL with BL and on-therapy virologic and immunologic outcomes. Higher RCBL was associated with lower baseline CD4 (CD4BL) (r = -0.23, P < 0.0001), higher baseline HIV-1 RNA (r = 0.25, P < 0.0001), higher CD4BL activation percent (r = 0.23, P < 0.0001), and lower CD4BL memory count (r = -0.21, P = 0.0002). In a multivariable model, week 48 CD4 increase (DeltaCD448) was associated with lower CD4BL memory count and higher CD4BL-naive percent (P = 0.004, P = 0.015, respectively). The interaction between CD4BL and RCBL was significant (P = 0.018), with a positive association between RCBL and DeltaCD448 in subjects with higher CD4BL and a negative association at lower absCD4BL. At baseline, higher RC was significantly associated with higher HIV-1 RNA, higher CD4 cell activation, lower CD4 cell count, and lower CD4 memory cell count. These factors may interact, directly or indirectly, to modify the extent to which CD4 recovery occurs in patients starting antiretroviral therapy at different CD4BL counts.

Replication-competent infectious hepatitis B virus vectors carrying substantially sized transgenes by redesigned viral polymerase translation.

Directory of Open Access Journals (Sweden)

Zihua Wang

Full Text Available Viral vectors are engineered virus variants able to deliver nonviral genetic information into cells, usually by the same routes as the parental viruses. For several virus families, replication-competent vectors carrying reporter genes have become invaluable tools for easy and quantitative monitoring of replication and infection, and thus also for identifying antivirals and virus susceptible cells. For hepatitis B virus (HBV, a small enveloped DNA virus causing B-type hepatitis, such vectors are not available because insertions into its tiny 3.2 kb genome almost inevitably affect essential replication elements. HBV replicates by reverse transcription of the pregenomic (pg RNA which is also required as bicistronic mRNA for the capsid (core protein and the reverse transcriptase (Pol; their open reading frames (ORFs overlap by some 150 basepairs. Translation of the downstream Pol ORF does not involve a conventional internal ribosome entry site (IRES. We reasoned that duplicating the overlap region and providing artificial IRES control for translation of both Pol and an in-between inserted transgene might yield a functional tricistronic pgRNA, without interfering with envelope protein expression. As IRESs we used a 22 nucleotide element termed Rbm3 IRES to minimize genome size increase. Model plasmids confirmed its activity even in tricistronic arrangements. Analogous plasmids for complete HBV genomes carrying 399 bp and 720 bp transgenes for blasticidin resistance (BsdR and humanized Renilla green fluorescent protein (hrGFP produced core and envelope proteins like wild-type HBV; while the hrGFP vector replicated poorly, the BsdR vector generated around 40% as much replicative DNA as wild-type HBV. Both vectors, however, formed enveloped virions which were infectious for HBV-susceptible HepaRG cells. Because numerous reporter and effector genes with sizes of around 500 bp or less are available, the new HBV vectors should become highly useful tools to

Electrochemistry of deferiprone as an orally active iron chelator and HIV-1 replication inhibitor and its determination

OpenAIRE

Yadegari, H.; Jabbari, A.; Heli, H.; Moosavi-Movahedi, A. A.; Majdi, S.

2008-01-01

The electrochemical behavior of the anti-thalassemia and anti-HIV replication drug, deferiprone, was investigated by cyclic voltammetry (CV) at a platinum electrode. In an acetate buffer solution, pH = 4.0, two irreversible anodic peaks for deferiprone, with E(0)1 = 875 mV and E(0)2 = 1235 mV (vs. Ag/AgCl) appeared at a potential sweep rate of 50 mV s-1. Cyclic voltammetric study indicated that the oxidation process is irreversible and diffusion-controlled. The diffusion and the electron tran...

Molecular cloning of osteoma-inducing replication-competent murine leukemia viruses from the RFB osteoma virus stock

DEFF Research Database (Denmark)

Pedersen, Lene; Behnisch, Werner; Schmidt, Jörg

1992-01-01

We report the molecular cloning of two replication-competent osteoma-inducing murine leukemia viruses from the RFB osteoma virus stock (M. P. Finkel, C. A. Reilly, Jr., B. O. Biskis, and I. L. Greco, p. 353-366, in C. H. G. Price and F. G. M. Ross, ed., Bone--Certain Aspects of Neoplasia, 1973......). Like the original RFB osteoma virus stock, viruses derived from the molecular RFB clones induced multiple osteomas in mice of the CBA/Ca strain. The cloned RFB viruses were indistinguishable by restriction enzyme analysis and by nucleotide sequence analysis of their long-terminal-repeat regions...

Identification of the determinants of efficient Pestivirus replication

DEFF Research Database (Denmark)

Risager, Peter Christian

, and in depth knowledge of the traits that determine the fitness of the virus in this regard are highly valuable. Recent advances in the field of molecular virology with methods to manipulate viral genomes have significantly helped to uncover these core mechanisms responsible for exploitation of the host......, BMC genomics). Manuscript II describes the generation of replicons that express two different types of luciferases (Rluc and Gluc), and their application as a tool for easy monitoring of replication competence (published paper, Journal of General Virology (94), 1739-1748). Manuscript III describes...... the properties of chimeric replicons and infectious clones that include a RNA dependent RNA polymerase (NS5B) from one of three different CSFV strains with distinct virulence properties. The entire NS5B proved to influence replication competence and key residues for replication competence was identified...

Replacement of the murine leukemia virus (MLV) envelope gene with a truncated HIV envelope gene in MLV generates a virus with impaired replication capacity

International Nuclear Information System (INIS)

Nack, Ursula; Schnierle, Barbara S.

2003-01-01

Murine leukemia virus (MLV) capsid particles can be efficiently pseudotyped with a variant of the HIV-1 envelope protein (Env) containing the surface glycoprotein gp120-SU and a carboxyl-terminally truncated transmembrane (TM) protein, with only seven cytoplasmic amino acids. MLV/HIV pseudotyped vector particles acquire the natural host tropism of HIV-1 and their entry is dependent on the presence of CD4 and an appropriate co-receptor on the surface of the target cell. We describe here the construction of chimeric MLV/HIV proviruses containing the truncated HIV envelope gene. The MLV/HIV provirus was generated by direct replacement of the MLV envelope gene with HIV Env coding sequences either with or without the additional inclusion of the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE). Chimeric MLV/HIV particles could be generated from transfected 293T cells and were able to infect CD4/CXCR4-positive target cells. However, the second round of infection of target cells was severely impaired, despite the fact that the WPRE element enhanced the amount of viral mRNA detected. Viral particles released from infected cells showed reduced HIV Env incorporation, indicating that additional factors required for efficient replication of MLV/HIV pseudotyped viruses are missing

A Leu to Ile but not Leu to Val change at HIV-1 reverse transcriptase codon 74 in the background of K65R mutation leads to an increased processivity of K65R+L74I enzyme and a replication competent virus

Directory of Open Access Journals (Sweden)

Crumpacker Clyde S

2011-01-01

Full Text Available Abstract Background The major hurdle in the treatment of Human Immunodeficiency virus type 1 (HIV-1 includes the development of drug resistance-associated mutations in the target regions of the virus. Since reverse transcriptase (RT is essential for HIV-1 replication, several nucleoside analogues have been developed to target RT of the virus. Clinical studies have shown that mutations at RT codon 65 and 74 which are located in β3-β4 linkage group of finger sub-domain of RT are selected during treatment with several RT inhibitors, including didanosine, deoxycytidine, abacavir and tenofovir. Interestingly, the co-selection of K65R and L74V is rare in clinical settings. We have previously shown that K65R and L74V are incompatible and a R→K reversion occurs at codon 65 during replication of the virus. Analysis of the HIV resistance database has revealed that similar to K65R+L74V, the double mutant K65R+L74I is also rare. We sought to compare the impact of L→V versus L→I change at codon 74 in the background of K65R mutation, on the replication of doubly mutant viruses. Methods Proviral clones containing K65R, L74V, L74I, K65R+L74V and K65R+L74I RT mutations were created in pNL4-3 backbone and viruses were produced in 293T cells. Replication efficiencies of all the viruses were compared in peripheral blood mononuclear (PBM cells in the absence of selection pressure. Replication capacity (RC of mutant viruses in relation to wild type was calculated on the basis of antigen p24 production and RT activity, and paired analysis by student t-test was performed among RCs of doubly mutant viruses. Reversion at RT codons 65 and 74 was monitored during replication in PBM cells. In vitro processivity of mutant RTs was measured to analyze the impact of amino acid changes at RT codon 74. Results Replication kinetics plot showed that all of the mutant viruses were attenuated as compared to wild type (WT virus. Although attenuated in comparison to WT virus

Replication-competent human adenovirus 11p vectors can propagate in Vero cells

International Nuclear Information System (INIS)

Gokumakulapalle, Madhuri; Mei, Ya-Fang

2016-01-01

The use of continuous cell lines derived from the African green monkey kidney (AGMK) has led to major advances in virus vaccine development. However, to date, these cells have not been used to facilitate the creation of human adenoviruses because most human adenoviruses undergo abortive infections in them. Here, we report the susceptibility of AGMK-derived cells to adenovirus 11p (Ad11p) infection. First, we showed that CD46 molecules, which act as receptors for Ad11p, are expressed in AGMK cells. We then monitored Ad11p replication by measuring GFP expression as an indicator of viral transcription. We found that AGMK-derived cells were as capable as carcinoma cells at propagating full-length replication-competent Ad11p (RCAd11p) DNA. Of the AGMK cell lines tested, Vero cells had the greatest capacity for adenovirus production. Thus, AGMK cells can be used to evaluate RCAd11p-mediated gene delivery, and Vero cells can be used for the production of RCAd11pGFP vectors at relatively high yields. - Highlights: • Africa green monkey cell lines were monitored for human adenovirus 11p GFP vector infection. • Human CD46 molecules were detectable in these monkey cell lines. • Adenovirus 11p GFP vector can be propagated in Vero cells increases the safety of Ad11p-based vectors for clinical trials. • To use Vero cells for preparation of Ad11p vector avoids the potential inclusion of oncogenes from tumor cells.

Replication-competent human adenovirus 11p vectors can propagate in Vero cells

Energy Technology Data Exchange (ETDEWEB)

Gokumakulapalle, Madhuri; Mei, Ya-Fang, E-mail: ya-fang.mei@umu.se

2016-08-15

The use of continuous cell lines derived from the African green monkey kidney (AGMK) has led to major advances in virus vaccine development. However, to date, these cells have not been used to facilitate the creation of human adenoviruses because most human adenoviruses undergo abortive infections in them. Here, we report the susceptibility of AGMK-derived cells to adenovirus 11p (Ad11p) infection. First, we showed that CD46 molecules, which act as receptors for Ad11p, are expressed in AGMK cells. We then monitored Ad11p replication by measuring GFP expression as an indicator of viral transcription. We found that AGMK-derived cells were as capable as carcinoma cells at propagating full-length replication-competent Ad11p (RCAd11p) DNA. Of the AGMK cell lines tested, Vero cells had the greatest capacity for adenovirus production. Thus, AGMK cells can be used to evaluate RCAd11p-mediated gene delivery, and Vero cells can be used for the production of RCAd11pGFP vectors at relatively high yields. - Highlights: • Africa green monkey cell lines were monitored for human adenovirus 11p GFP vector infection. • Human CD46 molecules were detectable in these monkey cell lines. • Adenovirus 11p GFP vector can be propagated in Vero cells increases the safety of Ad11p-based vectors for clinical trials. • To use Vero cells for preparation of Ad11p vector avoids the potential inclusion of oncogenes from tumor cells.

Hepatitis C virus resistance to broadly neutralizing antibodies measured using replication-competent virus and pseudoparticles.

Science.gov (United States)

Wasilewski, Lisa N; Ray, Stuart C; Bailey, Justin R

2016-11-01

A better understanding of natural variation in neutralization resistance and fitness of diverse hepatitis C virus (HCV) envelope (E1E2) variants will be critical to guide rational development of an HCV vaccine. This work has been hindered by inadequate genetic diversity in viral panels and by a lack of standardization of HCV entry assays. Neutralization assays generally use lentiviral pseudoparticles expressing HCV envelope proteins (HCVpp) or chimeric full-length viruses that are replication competent in cell culture (HCVcc). There have been few systematic comparisons of specific infectivities of E1E2-matched HCVcc and HCVpp, and to our knowledge, neutralization of E1E2-matched HCVpp and HCVcc has never been compared using a diverse panel of human broadly neutralizing monoclonal antibodies (bNAbs) targeting distinct epitopes. Here, we describe an efficient method for introduction of naturally occurring E1E2 genes into a full-length HCV genome, producing replication-competent chimeric HCVcc. We generated diverse panels of E1E2-matched HCVcc and HCVpp and measured the entry-mediating fitness of E1E2 variants using the two systems. We also compared neutralization of E1E2-matched HCVcc and HCVpp by a diverse panel of human bNAbs targeting epitopes across E1E2. We found no correlation between specific infectivities of E1E2-matched HCVcc versus HCVpp, but found a very strong positive correlation between relative neutralization resistance of these same E1E2-matched HCVcc and HCVpp variants. These results suggest that quantitative comparisons of neutralization resistance of E1E2 variants can be made with confidence using either HCVcc or HCVpp, allowing the use of either or both systems to maximize diversity of neutralization panels.

Contribution of the C-terminal region within the catalytic core domain of HIV-1 integrase to yeast lethality, chromatin binding and viral replication

Directory of Open Access Journals (Sweden)

Belhumeur Pierre

2008-11-01

Full Text Available Abstract Background HIV-1 integrase (IN is a key viral enzymatic molecule required for the integration of the viral cDNA into the genome. Additionally, HIV-1 IN has been shown to play important roles in several other steps during the viral life cycle, including reverse transcription, nuclear import and chromatin targeting. Interestingly, previous studies have demonstrated that the expression of HIV-1 IN induces the lethal phenotype in some strains of Saccharomyces cerevisiae. In this study, we performed mutagenic analyses of the C-terminal region of the catalytic core domain of HIV-1 IN in order to delineate the critical amino acid(s and/or motif(s required for the induction of the lethal phenotype in the yeast strain HP16, and to further elucidate the molecular mechanism which causes this phenotype. Results Our study identified three HIV-1 IN mutants, V165A, A179P and KR186,7AA, located in the C-terminal region of the catalytic core domain of IN that do not induce the lethal phenotype in yeast. Chromatin binding assays in yeast and mammalian cells demonstrated that these IN mutants were impaired for the ability to bind chromatin. Additionally, we determined that while these IN mutants failed to interact with LEDGF/p75, they retained the ability to bind Integrase interactor 1. Furthermore, we observed that VSV-G-pseudotyped HIV-1 containing these IN mutants was unable to replicate in the C8166 T cell line and this defect was partially rescued by complementation with the catalytically inactive D64E IN mutant. Conclusion Overall, this study demonstrates that three mutations located in the C-terminal region of the catalytic core domain of HIV-1 IN inhibit the IN-induced lethal phenotype in yeast by inhibiting the binding of IN to the host chromatin. These results demonstrate that the C-terminal region of the catalytic core domain of HIV-1 IN is important for binding to host chromatin and is crucial for both viral replication and the promotion of

Adipose Tissue: Sanctuary for HIV/SIV Persistence and Replication.

Science.gov (United States)

Pallikkuth, Suresh; Mohan, Mahesh

2015-12-01

This commentary highlights new findings from a recent study identifying adipose tissue as a potential HIV reservoir and a major site of inflammation during chronic human/simian immunodeficiency virus (HIV/SIV) infection. A concise discussion about upcoming challenges and new research avenues for reducing chronic adipose inflammation during HIV/SIV infection is presented. Copyright © 2015 Elsevier Ltd. All rights reserved.

HIV: mecanismo de replicação, alvos farmacológicos e inibição por produtos derivados de plantas HIV: replication mechanism, pharmacological targets and inhibition by products derived from plants

Directory of Open Access Journals (Sweden)

Roberta Costa Santos Ferreira

2010-01-01

Full Text Available The AIDS epidemy has spread out and led to the diversification on the research for new antiviral drugs. Natural products, especially those derived from plants, are well-recognized as excellent sources of new drugs. Several of them have inhibitory activity against HIV replication, and some have been already clinically tested, with favorable results. This review presents the biochemical basis of the viral cycle and the research up to date on the identification, determination of the mechanism of biological action together with the therapeutical potential of plants-derived natural products, in the inhibition of HIV.

Replicating rather than nonreplicating adenovirus-human immunodeficiency virus recombinant vaccines are better at eliciting potent cellular immunity and priming high-titer antibodies.

Science.gov (United States)

Peng, Bo; Wang, Liqun Rejean; Gómez-Román, Victor Raúl; Davis-Warren, Alberta; Montefiori, David C; Kalyanaraman, V S; Venzon, David; Zhao, Jun; Kan, Elaine; Rowell, Thomas J; Murthy, Krishna K; Srivastava, Indresh; Barnett, Susan W; Robert-Guroff, Marjorie

2005-08-01

A major challenge in combating the human immunodeficiency virus (HIV) epidemic is the development of vaccines capable of inducing potent, persistent cellular immunity and broadly reactive neutralizing antibody responses to HIV type 1 (HIV-1). We report here the results of a preclinical trial using the chimpanzee model to investigate a combination vaccine strategy involving sequential priming immunizations with different serotypes of adenovirus (Ad)/HIV-1(MN)env/rev recombinants and boosting with an HIV envelope subunit protein, oligomeric HIV(SF162) gp140deltaV2. The immunogenicities of replicating and nonreplicating Ad/HIV-1(MN)env/rev recombinants were compared. Replicating Ad/HIV recombinants were better at eliciting HIV-specific cellular immune responses and better at priming humoral immunity against HIV than nonreplicating Ad-HIV recombinants carrying the same gene insert. Enhanced cellular immunity was manifested by a greater frequency of HIV envelope-specific gamma interferon-secreting peripheral blood lymphocytes and better priming of T-cell proliferative responses. Enhanced humoral immunity was seen in higher anti-envelope binding and neutralizing antibody titers and better induction of antibody-dependent cellular cytotoxicity. More animals primed with replicating Ad recombinants mounted neutralizing antibodies against heterologous R5 viruses after one or two booster immunizations with the mismatched oligomeric HIV-1(SF162) gp140deltaV2 protein. These results support continued development of the replicating Ad-HIV recombinant vaccine approach and suggest that the use of replicating vectors for other vaccines may prove fruitful.

Efficacy and toxicity of replication-competent adenovirus-mediated double suicide gene therapy in combination with radiation therapy in an orthotopic mouse prostate cancer model

International Nuclear Information System (INIS)

Freytag, Svend O.; Paielli, Dell; Wing, Mark; Rogulski, Ken; Brown, Steve; Kolozsvary, Andy; Seely, John; Barton, Ken; Dragovic, Alek; Kim, Jae Ho

2002-01-01

Purpose: The purpose of this study was to evaluate the efficacy and toxicity of replication-competent adenovirus-mediated double suicide gene therapy in an adjuvant setting with external beam radiation therapy (EBRT) in an experimental prostate cancer model in preparation for a Phase I clinical study in humans. Methods: For efficacy studies, i.m. DU145 and intraprostatic LNCaP C4-2 tumors were established in immune-deficient mice. Tumors were injected with the lytic, replication-competent Ad5-CD/TKrep adenovirus containing a cytosine deaminase (CD)/herpes simplex virus thymidine kinase (HSV-1 TK) fusion gene. Two days later, mice were administered 1 week of 5-fluorocytosine + ganciclovir (GCV) prodrug therapy and fractionated doses of EBRT (trimodal therapy). Tumor control rate of trimodal therapy was compared to that of EBRT alone. For toxicology studies, immune-competent male mice received a single intraprostatic injection (10 10 vp) of the replication-competent Ad5-CD/TKrep adenovirus. Two days later, mice were administered 4 weeks of 5-fluorocytosine + GCV prodrug therapy and 56 Gy EBRT to the pelvic region. The toxicity of trimodal therapy was assessed by histopathologic analysis of major organs and clinical chemistries. Results: In both the i.m. DU145 and intraprostatic LNCaP C4-2 tumor models, trimodal therapy significantly improved primary tumor control beyond that of EBRT alone. In the DU145 model, trimodal therapy resulted in a tumor growth delay (70 days) that was more than twice that (32 days) of EBRT alone. Whereas EBRT failed to eradicate DU145 tumors, trimodal therapy resulted in 25% tumor cure. In the LNCaP C4-2 tumor model, EBRT slowed the growth of intraprostatic tumors, but resulted in no tumor cures, and 57% of the mice developed retroperitoneal lymph node metastases at 3 months. By contrast, trimodal therapy resulted in 44% tumor cure and reduced significantly the percentage (13%) of lymph node metastases relative to EBRT alone. Overall

Case report: mechanisms of HIV elite control in two African women.

Science.gov (United States)

Moosa, Yumna; Tanko, Ramla F; Ramsuran, Veron; Singh, Ravesh; Madzivhandila, Mashudu; Yende-Zuma, Nonhlanhla; Abrahams, Melissa-Rose; Selhorst, Philippe; Gounder, Kamini; Moore, Penny L; Williamson, Carolyn; Abdool Karim, Salim S; Garrett, Nigel J; Burgers, Wendy A

2018-01-25

The majority of people living with HIV require antiretroviral therapy (ART) for controlling viral replication, however there are rare HIV controllers who spontaneously and durably control HIV in the absence of treatment. Understanding what mediates viral control in these individuals has provided us with insights into the immune mechanisms that may be important to induce for a vaccine or functional cure for HIV. To date, few African elite controllers from high incidence settings have been described. We identified virological controllers from the CAPRISA 002 cohort of HIV-1 subtype C infected women in KwaZulu Natal, South Africa, two (1%) of whom were elite controllers. We examined the genetic, clinical, immunological and virological characteristics of these two elite HIV controllers in detail, to determine whether they exhibit features of putative viral control similar to those described for elite controllers reported in the literature. In this case report, we present clinical features, CD4 + T cell and viral load trajectories for two African women over 7 years of HIV infection. Viral load became undetectable 10 months after HIV infection in Elite Controller 1 (EC1), and after 6 weeks in Elite Controller 2 (EC2), and remained undetectable for the duration of follow-up, in the absence of ART. Both elite controllers expressed multiple HLA Class I and II haplotypes previously associated with slower disease progression (HLA-A*74:01, HLA-B*44:03, HLA-B*81:01, HLA-B*57:03, HLA-DRB1*13). Fitness assays revealed that both women were infected with replication competent viruses, and both expressed higher mRNA levels of p21, a host restriction factor associated with viral control. HIV-specific T cell responses were examined using flow cytometry. EC1 mounted high frequency HIV-specific CD8+ T cell responses, including a B*81:01-restricted Gag TL9 response. Unusually, EC2 had evidence of pre-infection HIV-specific CD4+ T cell responses. We identified some features typical of

Replication Capacity in Relation to Immunologic and Virologic Outcomes in HIV-1 infected, Treatment-Naïve Subjects

Science.gov (United States)

Skowron, Gail; Spritzler, John G.; Weidler, Jodi; Robbins, Gregory K.; Johnson, Victoria A.; Chan, Ellen S.; Asmuth, David M.; Gandhi, Rajesh T.; Lie, Yolanda; Bates, Michael; Pollard, Richard B.

2012-01-01

Objectives To evaluate the association between baseline (BL) replication capacity (RC) [RCBL] and immunologic/virologic parameters (at BL and after 48 weeks on therapy) in HIV-1 infected subjects initiating antiretroviral therapy. Methods RCBL was determined using a modified Monogram PhenoSense HIV drug susceptibility assay on plasma HIV-1 from 321 treatment-naïve subjects from ACTG384. Univariate and multivariable analyses were performed to determine the association of RCBL with BL and on-therapy virologic and immunologic outcomes. Results Higher RCBL was associated with lower baseline CD4 (CD4BL) (r=−0.23, p<0.0001), higher baseline HIV-1 (RNABL) (r=0.25, p<0.0001), higher CD4BL activation percent (r=0.23, p<0.0001) and lower CD4BL memory count (r=−0.21, p=0.0002). In a multivariable model, week 48 CD4 increase (ΔCD448) was associated with lower CD4BL memory count and higher CD4BL naive percent (p=0.004, p=0.015, respectively). The interaction between CD4BL and RCBL was significant (p=0.018), with a positive association between RCBL and ΔCD448 in subjects with higher CD4BL, and a negative association at lower absCD4BL. Conclusions At baseline, higher RC was significantly associated with higher HIV-1 RNA, higher CD4 cell activation, lower CD4 cell count, and lower CD4 memory cell count. These factors may interact, directly or indirectly, to modify the extent to which CD4 recovery occurs in patients starting antiretroviral therapy at different baseline CD4 counts. PMID:19194319

Selective elimination of HIV-1-infected cells by Env-directed, HIV-1-based virus-like particles

International Nuclear Information System (INIS)

Peretti, Silvia; Schiavoni, Ilaria; Pugliese, Katherina; Federico, Maurizio

2006-01-01

We recently showed that both replicating and resting cells cultivated with ganciclovir (GCV) were killed when challenged with vesicular stomatitis virus G glycoprotein pseudotyped HIV-1-based virus-like particles (VLPs) carrying the Nef7 (i.e., an HIV-1 Nef mutant incorporating in virions at high levels)/herpes simplex virus-1 thymidine kinase (HSV-TK) fusion product. On this basis, a novel anti-HIV therapeutic approach based on Nef7/TK VLPs expressing X4 or R5 HIV cell receptor complexes has been attempted. We here report that (CD4-CXCR4) and (CD4-CCR5) Nef7-based VLPs efficiently enter cells infected by X4- or R5-tropic HIV-1 strains, respectively. Importantly, the delivery of the VLP-associated Nef7/TK led to cell death upon GCV treatment. Of interest, VLPs were effective also against non-replicating, HIV-1-infected primary human monocyte-derived macrophages. HIV-targeted VLPs represent a promising candidate for the treatment of persistently HIV-1-infected cells that are part of virus reservoirs resistant to HAART therapies

Mycobacterium tuberculosis complex enhances susceptibility of CD4 T cells to HIV through a TLR2-mediated pathway.

Directory of Open Access Journals (Sweden)

Seema M Thayil

Full Text Available Among HIV-infected individuals, co-infection with Mycobacterium tuberculosis is associated with faster progression to AIDS. We investigated the hypothesis that M. bovis BCG and M. tuberculosis (Mtb complex could enhance susceptibility of CD4+ cells to HIV infection. Peripheral blood mononuclear cells (PBMCs collected from healthy donors were stimulated with M. bovis BCG, M. tuberculosis CDC1551 and M. smegmatis MC(2155, and stimulated CD4+ cells were infected with R5-and X4-tropic single replication-competent pseudovirus. CD4+ cells stimulated with Mtb complex showed enhanced infection with R5- and X4-tropic HIV, compared to unstimulated cells or cells stimulated with M. smegmatis (p<0.01. Treatment with TLR2 siRNA reversed the increased susceptibility of CD4+ cells with R5- and X4-tropic virus induced by Mtb complex. These findings suggest that TB infection and/or BCG vaccination may be a risk factor for HIV acquisition.

The Depsipeptide Romidepsin Reverses HIV-1 Latency In Vivo.

Directory of Open Access Journals (Sweden)

Ole S Søgaard

2015-09-01

Full Text Available Pharmacologically-induced activation of replication competent proviruses from latency in the presence of antiretroviral treatment (ART has been proposed as a step towards curing HIV-1 infection. However, until now, approaches to reverse HIV-1 latency in humans have yielded mixed results. Here, we report a proof-of-concept phase Ib/IIa trial where 6 aviremic HIV-1 infected adults received intravenous 5 mg/m2 romidepsin (Celgene once weekly for 3 weeks while maintaining ART. Lymphocyte histone H3 acetylation, a cellular measure of the pharmacodynamic response to romidepsin, increased rapidly (maximum fold range: 3.7–7.7 relative to baseline within the first hours following each romidepsin administration. Concurrently, HIV-1 transcription quantified as copies of cell-associated un-spliced HIV-1 RNA increased significantly from baseline during treatment (range of fold-increase: 2.4–5.0; p = 0.03. Plasma HIV-1 RNA increased from <20 copies/mL at baseline to readily quantifiable levels at multiple post-infusion time-points in 5 of 6 patients (range 46–103 copies/mL following the second infusion, p = 0.04. Importantly, romidepsin did not decrease the number of HIV-specific T cells or inhibit T cell cytokine production. Adverse events (all grade 1–2 were consistent with the known side effects of romidepsin. In conclusion, romidepsin safely induced HIV-1 transcription resulting in plasma HIV-1 RNA that was readily detected with standard commercial assays demonstrating that significant reversal of HIV-1 latency in vivo is possible without blunting T cell-mediated immune responses. These finding have major implications for future trials aiming to eradicate the HIV-1 reservoir.clinicaltrials.gov NTC02092116.

Extensive virologic and immunologic characterization in an HIV-infected individual following allogeneic stem cell transplant and analytic cessation of antiretroviral therapy: A case study.

Science.gov (United States)

Cummins, Nathan W; Rizza, Stacey; Litzow, Mark R; Hua, Stephane; Lee, Guinevere Q; Einkauf, Kevin; Chun, Tae-Wook; Rhame, Frank; Baker, Jason V; Busch, Michael P; Chomont, Nicolas; Dean, Patrick G; Fromentin, Rémi; Haase, Ashley T; Hampton, Dylan; Keating, Sheila M; Lada, Steven M; Lee, Tzong-Hae; Natesampillai, Sekar; Richman, Douglas D; Schacker, Timothy W; Wietgrefe, Stephen; Yu, Xu G; Yao, Joseph D; Zeuli, John; Lichterfeld, Mathias; Badley, Andrew D

2017-11-01

Notwithstanding 1 documented case of HIV-1 cure following allogeneic stem cell transplantation (allo-SCT), several subsequent cases of allo-SCT in HIV-1 positive individuals have failed to cure HIV-1 infection. The aim of our study was to describe changes in the HIV reservoir in a single chronically HIV-infected patient on suppressive antiretroviral therapy who underwent allo-SCT for treatment of acute lymphoblastic leukemia. We prospectively collected peripheral blood mononuclear cells (PBMCs) by leukapheresis from a 55-year-old man with chronic HIV infection before and after allo-SCT to measure the size of the HIV-1 reservoir and characterize viral phylogeny and phenotypic changes in immune cells. At day 784 post-transplant, when HIV-1 was undetectable by multiple measures-including PCR measurements of both total and integrated HIV-1 DNA, replication-competent virus measurement by large cell input quantitative viral outgrowth assay, and in situ hybridization of colon tissue-the patient consented to an analytic treatment interruption (ATI) with frequent clinical monitoring. He remained aviremic off antiretroviral therapy until ATI day 288, when a low-level virus rebound of 60 HIV-1 copies/ml occurred, which increased to 1,640 HIV-1 copies/ml 5 days later, prompting reinitiation of ART. Rebounding plasma HIV-1 sequences were phylogenetically distinct from proviral HIV-1 DNA detected in circulating PBMCs before transplantation. The main limitations of this study are the insensitivity of reservoir measurements, and the fact that it describes a single case. allo-SCT led to a significant reduction in the size of the HIV-1 reservoir and a >9-month-long ART-free remission from HIV-1 replication. Phylogenetic analyses suggest that the origin of rebound virus was distinct from the viruses identified pre-transplant in the PBMCs.

Biologic interactions between HSV-2 and HIV-1 and possible implications for HSV vaccine development.

Science.gov (United States)

Schiffer, Joshua T; Gottlieb, Sami L

2017-09-25

Development of a safe and effective vaccine against herpes simplex virus type 2 (HSV-2) has the potential to limit the global burden of HSV-2 infection and disease, including genital ulcer disease and neonatal herpes, and is a global sexual and reproductive health priority. Another important potential benefit of an HSV-2 vaccine would be to decrease HIV infections, as HSV-2 increases the risk of HIV-1 acquisition several-fold. Acute and chronic HSV-2 infection creates ulcerations and draws dendritic cells and activated CD4+ T cells into genital mucosa. These cells are targets for HIV entry and replication. Prophylactic HSV-2 vaccines (to prevent infection) and therapeutic vaccines (to modify or treat existing infections) are currently under development. By preventing or modifying infection, an effective HSV-2 vaccine could limit HSV-associated genital mucosal inflammation and thus HIV risk. However, a vaccine might have competing effects on HIV risk depending on its mechanism of action and cell populations generated in the genital mucosa. In this article, we review biologic interactions between HSV-2 and HIV-1, consider HSV-2 vaccine development in the context of HIV risk, and discuss implications and research needs for future HSV vaccine development. Copyright © 2017 Elsevier Ltd. All rights reserved.

Human endogenous retrovirus K Gag coassembles with HIV-1 Gag and reduces the release efficiency and infectivity of HIV-1.

Science.gov (United States)

Monde, Kazuaki; Contreras-Galindo, Rafael; Kaplan, Mark H; Markovitz, David M; Ono, Akira

2012-10-01

Human endogenous retroviruses (HERVs), which are remnants of ancestral retroviruses integrated into the human genome, are defective in viral replication. Because activation of HERV-K and coexpression of this virus with HIV-1 have been observed during HIV-1 infection, it is conceivable that HERV-K could affect HIV-1 replication, either by competition or by cooperation, in cells expressing both viruses. In this study, we found that the release efficiency of HIV-1 Gag was 3-fold reduced upon overexpression of HERV-K(CON) Gag. In addition, we observed that in cells expressing Gag proteins of both viruses, HERV-K(CON) Gag colocalized with HIV-1 Gag at the plasma membrane. Furthermore, HERV-K(CON) Gag was found to coassemble with HIV-1 Gag, as demonstrated by (i) processing of HERV-K(CON) Gag by HIV-1 protease in virions, (ii) coimmunoprecipitation of virion-associated HERV-K(CON) Gag with HIV-1 Gag, and (iii) rescue of a late-domain-defective HERV-K(CON) Gag by wild-type (WT) HIV-1 Gag. Myristylation-deficient HERV-K(CON) Gag localized to nuclei, suggesting cryptic nuclear trafficking of HERV-K Gag. Notably, unlike WT HERV-K(CON) Gag, HIV-1 Gag failed to rescue myristylation-deficient HERV-K(CON) Gag to the plasma membrane. Efficient colocalization and coassembly of HIV-1 Gag and HERV-K Gag also required nucleocapsid (NC). These results provide evidence that HIV-1 Gag heteromultimerizes with HERV-K Gag at the plasma membrane, presumably through NC-RNA interaction. Intriguingly, HERV-K Gag overexpression reduced not only HIV-1 release efficiency but also HIV-1 infectivity in a myristylation- and NC-dependent manner. Altogether, these results indicate that Gag proteins of endogenous retroviruses can coassemble with HIV-1 Gag and modulate the late phase of HIV-1 replication.

The dual action of poly(ADP-ribose polymerase -1 (PARP-1 inhibition in HIV-1 infection: HIV-1 LTR inhibition and diminution in Rho GTPase activity

Directory of Open Access Journals (Sweden)

Slava eRom

2015-08-01

Full Text Available The transcription of HIV-1 (HIV is regulated by complex mechanisms involving various cellular factors and virus-encoded transactivators. Poly(ADP-ribose polymerase 1 (PARP-1 inhibition has emerged recently as a potent anti-inflammatory tool, since PARP-1 is involved in the regulation of some genes through its interaction with various transcription factors. We propose a novel approach to diminish HIV replication via PARP-1 inhibition using human primary monocyte-derived macrophages (MDM as an in vitro model system. PARP-1 inhibitors were able to reduce HIV replication in MDM by 60-80% after 7 days infection. Long Terminal Repeat (LTR acts as a switch in virus replication and can be triggered by several agents such as: Tat, tumor necrosis factor α (TNFα, and phorbol 12-myristate 13-acetate (PMA. Overexpression of Tat in MDM transfected with an LTR reporter plasmid led to a 4.2-fold increase in LTR activation; PARP inhibition resulted in 70% reduction of LTR activity. LTR activity, which increased 3-fold after PMA or TNFα treatment, was reduced by PARP inhibition (by 85-95%. MDM treated with PARP inhibitors showed 90% reduction in NFκB activity (known to mediate PMA- and TNFα-induced HIV LTR activation. Cytoskeleton rearrangements are important in effective HIV-1 infection. PARP inactivation reduced actin cytoskeleton rearrangements by affecting Rho GTPase machinery. These findings suggest that HIV replication in MDM could be suppressed by PARP inhibition via NFκB suppression, diminution of LTR activation and its effects on the cytoskeleton. PARP appears to be essential for HIV replication and its inhibition may provide a potent approach to treatment of HIV infection.

Efficacy of Tenofovir 1% Vaginal Gel in Reducing the Risk of HIV-1 and HSV-2 Infection

Directory of Open Access Journals (Sweden)

Christopher McConville

2014-01-01

Full Text Available Human Immunodeficiency Virus (HIV is a retrovirus that can result in rare opportunistic infections occurring in humans. The onset of these infections is known as Acquired Immune Deficiency Syndrome (AIDS. Sexual transmission is responsible for the majority of infections 1, resulting in transmission of HIV due to infected semen or vaginal and cervical secretions containing infected lymphocytes. HIV microbicides are formulations of chemical or biological agents that can be applied to the vagina or rectum with the intention of reducing the acquisition of HIV. Tenofovir is an NRTI that is phosphorylated by adenylate kinase to tenofovir diphosphate, which in turn competes with deoxyadeosine 5′-triphosphate for incorporation into newly synthesized HIV DNA. Once incorporated, tenofovir diphosphate results in chain termination, thus inhibiting viral replication. Tenofovir has been formulated into a range of vaginal formulations, such as rings, tablets gels and films. It has been shown to safe and effective in numerous animal models, while demonstrating safety and acceptability in numerous human trials. The most encouraging results came from the CAPRISA 004 clinical trial which demonstrated that a 1% Tenofovir vaginal gel reduced HIV infection by approximately 39%.

CD32 is expressed on cells with transcriptionally active HIV but does not enrich for HIV DNA in resting T cells.

Science.gov (United States)

Abdel-Mohsen, Mohamed; Kuri-Cervantes, Leticia; Grau-Exposito, Judith; Spivak, Adam M; Nell, Racheal A; Tomescu, Costin; Vadrevu, Surya Kumari; Giron, Leila B; Serra-Peinado, Carla; Genescà, Meritxell; Castellví, Josep; Wu, Guoxin; Del Rio Estrada, Perla M; González-Navarro, Mauricio; Lynn, Kenneth; King, Colin T; Vemula, Sai; Cox, Kara; Wan, Yanmin; Li, Qingsheng; Mounzer, Karam; Kostman, Jay; Frank, Ian; Paiardini, Mirko; Hazuda, Daria; Reyes-Terán, Gustavo; Richman, Douglas; Howell, Bonnie; Tebas, Pablo; Martinez-Picado, Javier; Planelles, Vicente; Buzon, Maria J; Betts, Michael R; Montaner, Luis J

2018-04-18

The persistence of HIV reservoirs, including latently infected, resting CD4 + T cells, is the major obstacle to cure HIV infection. CD32a expression was recently reported to mark CD4 + T cells harboring a replication-competent HIV reservoir during antiretroviral therapy (ART) suppression. We aimed to determine whether CD32 expression marks HIV latently or transcriptionally active infected CD4 + T cells. Using peripheral blood and lymphoid tissue of ART-treated HIV + or SIV + subjects, we found that most of the circulating memory CD32 + CD4 + T cells expressed markers of activation, including CD69, HLA-DR, CD25, CD38, and Ki67, and bore a T H 2 phenotype as defined by CXCR3, CCR4, and CCR6. CD32 expression did not selectively enrich for HIV- or SIV-infected CD4 + T cells in peripheral blood or lymphoid tissue; isolated CD32 + resting CD4 + T cells accounted for less than 3% of the total HIV DNA in CD4 + T cells. Cell-associated HIV DNA and RNA loads in CD4 + T cells positively correlated with the frequency of CD32 + CD69 + CD4 + T cells but not with CD32 expression on resting CD4 + T cells. Using RNA fluorescence in situ hybridization, CD32 coexpression with HIV RNA or p24 was detected after in vitro HIV infection (peripheral blood mononuclear cell and tissue) and in vivo within lymph node tissue from HIV-infected individuals. Together, these results indicate that CD32 is not a marker of resting CD4 + T cells or of enriched HIV DNA-positive cells after ART; rather, CD32 is predominately expressed on a subset of activated CD4 + T cells enriched for transcriptionally active HIV after long-term ART. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Nuclear trafficking of the HIV-1 pre-integration complex depends on the ADAM10 intracellular domain

International Nuclear Information System (INIS)

Endsley, Mark A.; Somasunderam, Anoma D.; Li, Guangyu; Oezguen, Numan; Thiviyanathan, Varatharasa; Murray, James L.; Rubin, Donald H.; Hodge, Thomas W.

2014-01-01

Previously, we showed that ADAM10 is necessary for HIV-1 replication in primary human macrophages and immortalized cell lines. Silencing ADAM10 expression interrupted the HIV-1 life cycle prior to nuclear translocation of viral cDNA. Furthermore, our data indicated that HIV-1 replication depends on the expression of ADAM15 and γ-secretase, which proteolytically processes ADAM10. Silencing ADAM15 or γ-secretase expression inhibits HIV-1 replication between reverse transcription and nuclear entry. Here, we show that ADAM10 expression also supports replication in CD4 + T lymphocytes. The intracellular domain (ICD) of ADAM10 associates with the HIV-1 pre-integration complex (PIC) in the cytoplasm and immunoprecipitates and co-localizes with HIV-1 integrase, a key component of PIC. Taken together, our data support a model whereby ADAM15/γ-secretase processing of ADAM10 releases the ICD, which then incorporates into HIV-1 PIC to facilitate nuclear trafficking. Thus, these studies suggest ADAM10 as a novel therapeutic target for inhibiting HIV-1 prior to nuclear entry. - Highlights: • Nuclear trafficking of the HIV-1 pre-integration complex depends on ADAM10. • ADAM10 associates with HIV-1 integrase in the pre-integration complex. • HIV-1 replication depends on the expression of ADAM15 and γ-secretase. • Silencing ADAM15 or γ-secretase expression inhibits nuclear import of viral cDNA. • ADAM10 is important for HIV-1 replication in human macrophages and CD4 + T lymphocytes

Nuclear trafficking of the HIV-1 pre-integration complex depends on the ADAM10 intracellular domain

Energy Technology Data Exchange (ETDEWEB)

Endsley, Mark A., E-mail: maendsle@utmb.edu [Department Internal Medicine, Division of Infectious Diseases, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555 (United States); Somasunderam, Anoma D., E-mail: asomasun@utmb.edu [Department Internal Medicine, Division of Infectious Diseases, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555 (United States); Li, Guangyu, E-mail: LIG001@mail.etsu.edu [Department of Internal Medicine, Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614 (United States); Oezguen, Numan, E-mail: numan.oezguen@bcm.edu [Department of Pathology and Immunology, Microbiome Center, Texas Children' s Hospital, Houston, TX 77030 (United States); Thiviyanathan, Varatharasa, E-mail: Varatharasa.Thiviyanathan@uth.tmc.edu [Institute of Molecular Medicine, University of Texas Health Science Center, Houston, TX 77030 (United States); Murray, James L., E-mail: jmurray100@yahoo.com [GeneTAG Technology, Inc., 3155 Northwoods Place, Norcross, GA 30071 (United States); Rubin, Donald H., E-mail: don.h.rubin@vanderbilt.edu [Research Medicine, VA Tennessee Valley Healthcare System, 1310 24th Ave. South, Nashville, TN 37212 (United States); Departments of Medicine, Pathology, Microbiology and Immunology, Vanderbilt University School of Medicine, 1161 21st Ave South, Nashville, TN 37232 (United States); Hodge, Thomas W., E-mail: twhodge3@gmail.com [Pre-clinical and Antiviral Research, Tamir Biotechnology, Inc., 12625 High Bluff Dr., Suite 113, San Diego, CA 92130 (United States); and others

2014-04-15

Previously, we showed that ADAM10 is necessary for HIV-1 replication in primary human macrophages and immortalized cell lines. Silencing ADAM10 expression interrupted the HIV-1 life cycle prior to nuclear translocation of viral cDNA. Furthermore, our data indicated that HIV-1 replication depends on the expression of ADAM15 and γ-secretase, which proteolytically processes ADAM10. Silencing ADAM15 or γ-secretase expression inhibits HIV-1 replication between reverse transcription and nuclear entry. Here, we show that ADAM10 expression also supports replication in CD4{sup +} T lymphocytes. The intracellular domain (ICD) of ADAM10 associates with the HIV-1 pre-integration complex (PIC) in the cytoplasm and immunoprecipitates and co-localizes with HIV-1 integrase, a key component of PIC. Taken together, our data support a model whereby ADAM15/γ-secretase processing of ADAM10 releases the ICD, which then incorporates into HIV-1 PIC to facilitate nuclear trafficking. Thus, these studies suggest ADAM10 as a novel therapeutic target for inhibiting HIV-1 prior to nuclear entry. - Highlights: • Nuclear trafficking of the HIV-1 pre-integration complex depends on ADAM10. • ADAM10 associates with HIV-1 integrase in the pre-integration complex. • HIV-1 replication depends on the expression of ADAM15 and γ-secretase. • Silencing ADAM15 or γ-secretase expression inhibits nuclear import of viral cDNA. • ADAM10 is important for HIV-1 replication in human macrophages and CD4{sup +} T lymphocytes.

BET bromodomain inhibition as a novel strategy for reactivation of HIV-1.

Science.gov (United States)

Banerjee, Camellia; Archin, Nancie; Michaels, Daniel; Belkina, Anna C; Denis, Gerald V; Bradner, James; Sebastiani, Paola; Margolis, David M; Montano, Monty

2012-12-01

The persistence of latent HIV-1 remains a major challenge in therapeutic efforts to eradicate infection. We report the capacity for HIV reactivation by a selective small molecule inhibitor of BET family bromodomains, JQ1, a promising therapeutic agent with antioncogenic properties. JQ1 reactivated HIV transcription in models of latent T cell infection and latent monocyte infection. We also tested the effect of exposure to JQ1 to allow recovery of replication-competent HIV from pools of resting CD4(+) T cells isolated from HIV-infected, ART-treated patients. In one of three patients, JQ1 allowed recovery of virus at a frequency above unstimulated conditions. JQ1 potently suppressed T cell proliferation with minimal cytotoxic effect. Transcriptional profiling of T cells with JQ1 showed potent down-regulation of T cell activation genes, including CD3, CD28, and CXCR4, similar to HDAC inhibitors, but JQ1 also showed potent up-regulation of chromatin modification genes, including SIRT1, HDAC6, and multiple lysine demethylases (KDMs). Thus, JQ1 reactivates HIV-1 while suppressing T cell activation genes and up-regulating histone modification genes predicted to favor increased Tat activity. Thus, JQ1 may be useful in studies of potentially novel mechanisms for transcriptional control as well as in translational efforts to identify therapeutic molecules to achieve viral eradication.

Murine leukemia virus (MLV replication monitored with fluorescent proteins

Directory of Open Access Journals (Sweden)

Bittner Alexandra

2004-12-01

Full Text Available Abstract Background Cancer gene therapy will benefit from vectors that are able to replicate in tumor tissue and cause a bystander effect. Replication-competent murine leukemia virus (MLV has been described to have potential as cancer therapeutics, however, MLV infection does not cause a cytopathic effect in the infected cell and viral replication can only be studied by immunostaining or measurement of reverse transcriptase activity. Results We inserted the coding sequences for green fluorescent protein (GFP into the proline-rich region (PRR of the ecotropic envelope protein (Env and were able to fluorescently label MLV. This allowed us to directly monitor viral replication and attachment to target cells by flow cytometry. We used this method to study viral replication of recombinant MLVs and split viral genomes, which were generated by replacement of the MLV env gene with the red fluorescent protein (RFP and separately cloning GFP-Env into a retroviral vector. Co-transfection of both plasmids into target cells resulted in the generation of semi-replicative vectors, and the two color labeling allowed to determine the distribution of the individual genomes in the target cells and was indicative for the occurrence of recombination events. Conclusions Fluorescently labeled MLVs are excellent tools for the study of factors that influence viral replication and can be used to optimize MLV-based replication-competent viruses or vectors for gene therapy.

Replication-Competent Influenza A and B Viruses Expressing a Fluorescent Dynamic Timer Protein for In Vitro and In Vivo Studies.

Directory of Open Access Journals (Sweden)

Michael Breen

Full Text Available Influenza A and B viruses (IAV and IBV, respectively cause annual seasonal human respiratory disease epidemics. In addition, IAVs have been implicated in occasional pandemics with inordinate health and economic consequences. Studying influenza viruses in vitro or in vivo requires the use of laborious secondary methodologies to identify infected cells. To circumvent this requirement, replication-competent infectious influenza viruses expressing an easily traceable fluorescent reporter protein can be used. Timer is a fluorescent protein that undergoes a time-dependent color emission conversion from green to red. The rate of spectral change is independent of Timer protein concentration and can be used to chronologically measure the duration of its expression. Here, we describe the generation of replication-competent IAV and IBV where the viral non-structural protein 1 (NS1 was fused to the fluorescent dynamic Timer protein. Timer-expressing IAV and IBV displayed similar plaque phenotypes and growth kinetics to wild-type viruses in tissue culture. Within infected cells, Timer's spectral shift can be used to measure the rate and cell-to-cell spread of infection using fluorescent microscopy, plate readers, or flow cytometry. The progression of Timer-expressing IAV infection was also evaluated in a mouse model, demonstrating the feasibility to characterize IAV cell-to-cell infections in vivo. By providing the ability to chronologically track viral spread, Timer-expressing influenza viruses are an excellent option to evaluate the in vitro and in vivo dynamics of viral infection.

Novel inhibitors of HIV discovered among existing classes of pharmaceutical compounds indicated for unrelated clinical indications.

Science.gov (United States)

Kucherov, I I; Rytik, P G; Podol'skaya, I A; Mistryukova, L O; Korjev, M O

2009-01-01

In vitro screening of 307 drugs with various clinical indications (cardiotropic, neurotropic, antibacterial, etc.) has revealed 6 compounds which displayed remarkable antiretroviral activity. Three of these drugs had a tendency to have undesirable side effects and were thus excluded from further consideration. Remaining three, i.e., Xantinol Nicotinate, Tardiferon, and Trental may become valid candidates for inclusion into antiviral regimens such as HAART. In vitro tests have shown that xantinol and trental display synergistic effect with azidothymidine, inhibit the replication AZT-resistant strains of HIV, and have no competing undesirable activities. These compounds should be evaluated in safety studies to determine optimal doses for patients with HIV. If these studies confirm in vitro results these compounds may become valid candidates as safe and affordable means to be added into the arsenal of antiretroviral drugs.

Pre-clinical evaluation of a replication-competent recombinant adenovirus serotype 4 vaccine expressing influenza H5 hemagglutinin.

Science.gov (United States)

Alexander, Jeff; Ward, Simone; Mendy, Jason; Manayani, Darly J; Farness, Peggy; Avanzini, Jenny B; Guenther, Ben; Garduno, Fermin; Jow, Lily; Snarsky, Victoria; Ishioka, Glenn; Dong, Xin; Vang, Lo; Newman, Mark J; Mayall, Tim

2012-01-01

Influenza virus remains a significant health and social concern in part because of newly emerging strains, such as avian H5N1 virus. We have developed a prototype H5N1 vaccine using a recombinant, replication-competent Adenovirus serotype 4 (Ad4) vector, derived from the U.S. military Ad4 vaccine strain, to express the hemagglutinin (HA) gene from A/Vietnam/1194/2004 influenza virus (Ad4-H5-Vtn). Our hypothesis is that a mucosally-delivered replicating Ad4-H5-Vtn recombinant vector will be safe and induce protective immunity against H5N1 influenza virus infection and disease pathogenesis. The Ad4-H5-Vtn vaccine was designed with a partial deletion of the E3 region of Ad4 to accommodate the influenza HA gene. Replication and growth kinetics of the vaccine virus in multiple human cell lines indicated that the vaccine virus is attenuated relative to the wild type virus. Expression of the HA transgene in infected cells was documented by flow cytometry, western blot analysis and induction of HA-specific antibody and cellular immune responses in mice. Of particular note, mice immunized intranasally with the Ad4-H5-Vtn vaccine were protected against lethal H5N1 reassortant viral challenge even in the presence of pre-existing immunity to the Ad4 wild type virus. Several non-clinical attributes of this vaccine including safety, induction of HA-specific humoral and cellular immunity, and efficacy were demonstrated using an animal model to support Phase 1 clinical trial evaluation of this new vaccine.

HIV-1 gp120 induces NFAT nuclear translocation in resting CD4+ T-cells

International Nuclear Information System (INIS)

Cicala, Claudia; Arthos, James; Censoplano, Nina; Cruz, Catherine; Chung, Eva; Martinelli, Elena; Lempicki, Richard A.; Natarajan, Ven; VanRyk, Donald; Daucher, Marybeth; Fauci, Anthony S.

2006-01-01

The replication of human immunodeficiency virus (HIV) in CD4+ T-cells is strongly dependent upon the state of activation of infected cells. Infection of sub-optimally activated cells is believed to play a critical role in both the transmission of virus and the persistence of CD4+ T-cell reservoirs. There is accumulating evidence that HIV can modulate signal-transduction pathways in a manner that may facilitate replication in such cells. We previously demonstrated that HIV gp120 induces virus replication in resting CD4+ T cells isolated from HIV-infected individuals. Here, we show that in resting CD4+ T-cells, gp120 activates NFATs and induces their translocation into the nucleus. The HIV LTR encodes NFAT recognition sites, and NFATs may play a critical role in promoting viral replication in sub-optimally activated cells. These observations provide insight into a potential mechanism by which HIV is able to establish infection in resting cells, which may have implications for both transmission of HIV and the persistence of viral reservoirs

Clinical value of determination HIV viral load in the cerebrospinal fluid of HIV-infected patients

Directory of Open Access Journals (Sweden)

V. B. Musatov

2015-01-01

Full Text Available Aim. To analyze the concentration of HIV RNA in the cerebrospinal fluid and to evaluate its significance in the pathology of the central nervous system among HIV infected persons.Materials: We examined 36 patients with HIV infection with signs of pathology of the central nervous system. All patients was done completed a standard investigation of cerebrospinal fluid, cytological examination and detection viral load of HIV in the cerebrospinal fluid and serum.Results. A different of opportunistic and HIV-related disease was diagnosed in 29 patients. The most frequent pathology of the nervous system (12 cases is a diffuse HIV-associated brain damage occurring in 7 patients in the form of aseptic non purulent meningitis and in 5 patients in the form of encephalitis. The average value of the absolute and relative count of CD4-lymphocytes in patients amounted 147,0 cells/μl (40,0; 408,75 and 10.0% (4,00; 18,50. Pathological changes in cellular composition and protein concentration of cerebrospinal fluid detected in 19 cases. Replication of HIV in the cerebrospinal fluid are detected in 31 of 32 patients not receiving antiretroviral therapy, including 17 patients with normal values of cerebrospinal fluid. The average HIV viral load in the cerebrospinal fluid was 15 133,0 copies/ml (2501,0; 30624,0 or 4,18 (3,35; 4,48 lg HIV RNA, average HIV viral load in serum – 62 784,0 copies/ml (6027,5; 173869,0 or 4,80 4,80 (3,7; 5,2 lg HIV RNA. The concentration of HIV in the cerebrospinal fluid was significantly lower than in serum (4,18 and 4,80 lg HIV RNA, p=0.027. 4 patients with severe, multietiology damage of the central nervous system viral, microbial and fungal etiology, there was an inverse relationship between the concentration of HIV in the cerebrospinal fluid and in serum, the concentrations of HIV was higher in the cerebrospinal fluid.Conclusion: Among the majority of HIV-infected patients with signs of the central

The Oncolytic Virus MG1 Targets and Eliminates Cells Latently Infected With HIV-1: Implications for an HIV Cure.

Science.gov (United States)

Ranganath, Nischal; Sandstrom, Teslin S; Burke Schinkel, Stephanie C; Côté, Sandra C; Angel, Jonathan B

2018-02-14

Cells latently infected with human immunodeficiency virus (HIV) evade immune- and drug-mediated clearance. These cells harbor intracellular signaling defects, including impairment of the antiviral type I interferon response. Such defects have also been observed in several cancers and have been exploited for the development of therapeutic oncolytic viruses, including the recombinant Maraba virus (MG1). We therefore hypothesized that MG1 would infect and eliminate cells latently infected with HIV-1, while sparing healthy uninfected cells. Preferential infection and elimination by MG1 was first demonstrated in cell lines latently infected with HIV-1. Following this, a reduction in HIV-1 DNA and inducible HIV-1 replication was observed following MG1 infection of latently infected, resting CD4+ T cells generated using an in vitro model of latency. Last, MG1 infection resulted in a reduction in HIV-1 DNA and inducible HIV-1 replication in memory CD4+ T cells isolated from effectively treated, HIV-1-infected individuals. Our results therefore highlight a novel approach to eliminate the latent HIV-1 reservoir. © The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

A Novel Leu92 Mutant of HIV-1 Reverse Transcriptase with a Selective Deficiency in Strand Transfer Causes a Loss of Viral Replication.

Science.gov (United States)

Herzig, Eytan; Voronin, Nickolay; Kucherenko, Nataly; Hizi, Amnon

2015-08-01

The process of reverse transcription (RTN) in retroviruses is essential to the viral life cycle. This key process is catalyzed exclusively by the viral reverse transcriptase (RT) that copies the viral RNA into DNA by its DNA polymerase activity, while concomitantly removing the original RNA template by its RNase H activity. During RTN, the combination between DNA synthesis and RNA hydrolysis leads to strand transfers (or template switches) that are critical for the completion of RTN. The balance between these RT-driven activities was considered to be the sole reason for strand transfers. Nevertheless, we show here that a specific mutation in HIV-1 RT (L92P) that does not affect the DNA polymerase and RNase H activities abolishes strand transfer. There is also a good correlation between this complete loss of the RT's strand transfer to the loss of the DNA clamp activity of the RT, discovered recently by us. This finding indicates a mechanistic linkage between these two functions and that they are both direct and unique functions of the RT (apart from DNA synthesis and RNA degradation). Furthermore, when the RT's L92P mutant was introduced into an infectious HIV-1 clone, it lost viral replication, due to inefficient intracellular strand transfers during RTN, thus supporting the in vitro data. As far as we know, this is the first report on RT mutants that specifically and directly impair RT-associated strand transfers. Therefore, targeting residue Leu92 may be helpful in selectively blocking this RT activity and consequently HIV-1 infectivity and pathogenesis. Reverse transcription in retroviruses is essential for the viral life cycle. This multistep process is catalyzed by viral reverse transcriptase, which copies the viral RNA into DNA by its DNA polymerase activity (while concomitantly removing the RNA template by its RNase H activity). The combination and balance between synthesis and hydrolysis lead to strand transfers that are critical for reverse transcription

ORAL MANIFESTATIONS AMONG ROMANIAN HIV PATIENTS

OpenAIRE

Manuela ARBUNE; Oana-Mirela POTÂRNICHIE; Silvia MARTU

2013-01-01

The objective of the study is to evaluate the oral health problems on HIV youth patients from Galati. Materials and method. A cross-sectional study assessed 102 patients with mean age 22. The most frequent oral manifestations on HIV infected youth under ART are erythema marginatum, periodontitis, candidosis and hypertrophia gingivalis. Results and discussion. Dental decay-missing-filled index on HIV patients is high. Viral HIV replication, long time pediatric exposu...

Electron microscopic analysis of rotavirus assembly-replication intermediates

International Nuclear Information System (INIS)

Boudreaux, Crystal E.; Kelly, Deborah F.; McDonald, Sarah M.

2015-01-01

Rotaviruses (RVs) replicate their segmented, double-stranded RNA genomes in tandem with early virion assembly. In this study, we sought to gain insight into the ultrastructure of RV assembly-replication intermediates (RIs) using transmission electron microscopy (EM). Specifically, we examined a replicase-competent, subcellular fraction that contains all known RV RIs. Three never-before-seen complexes were visualized in this fraction. Using in vitro reconstitution, we showed that ~15-nm doughnut-shaped proteins in strings were nonstructural protein 2 (NSP2) bound to viral RNA transcripts. Moreover, using immunoaffinity-capture EM, we revealed that ~20-nm pebble-shaped complexes contain the viral RNA polymerase (VP1) and RNA capping enzyme (VP3). Finally, using a gel purification method, we demonstrated that ~30–70-nm electron-dense, particle-shaped complexes represent replicase-competent core RIs, containing VP1, VP3, and NSP2 as well as capsid proteins VP2 and VP6. The results of this study raise new questions about the interactions among viral proteins and RNA during the concerted assembly–replicase process. - Highlights: • Rotaviruses replicate their genomes in tandem with early virion assembly. • Little is known about rotavirus assembly-replication intermediates. • Assembly-replication intermediates were imaged using electron microscopy

Electron microscopic analysis of rotavirus assembly-replication intermediates

Energy Technology Data Exchange (ETDEWEB)

Boudreaux, Crystal E.; Kelly, Deborah F. [Virginia Tech Carilion School of Medicine and Research Institute, Roanoke, VA (United States); McDonald, Sarah M., E-mail: mcdonaldsa@vtc.vt.edu [Virginia Tech Carilion School of Medicine and Research Institute, Roanoke, VA (United States); Department of Biomedical Sciences and Pathobiology, Virginia—Maryland Regional College of Veterinary Medicine, Blacksburg, VA (United States)

2015-03-15

Rotaviruses (RVs) replicate their segmented, double-stranded RNA genomes in tandem with early virion assembly. In this study, we sought to gain insight into the ultrastructure of RV assembly-replication intermediates (RIs) using transmission electron microscopy (EM). Specifically, we examined a replicase-competent, subcellular fraction that contains all known RV RIs. Three never-before-seen complexes were visualized in this fraction. Using in vitro reconstitution, we showed that ~15-nm doughnut-shaped proteins in strings were nonstructural protein 2 (NSP2) bound to viral RNA transcripts. Moreover, using immunoaffinity-capture EM, we revealed that ~20-nm pebble-shaped complexes contain the viral RNA polymerase (VP1) and RNA capping enzyme (VP3). Finally, using a gel purification method, we demonstrated that ~30–70-nm electron-dense, particle-shaped complexes represent replicase-competent core RIs, containing VP1, VP3, and NSP2 as well as capsid proteins VP2 and VP6. The results of this study raise new questions about the interactions among viral proteins and RNA during the concerted assembly–replicase process. - Highlights: • Rotaviruses replicate their genomes in tandem with early virion assembly. • Little is known about rotavirus assembly-replication intermediates. • Assembly-replication intermediates were imaged using electron microscopy.

Mentoring the Mentors of Underrepresented Racial/Ethnic Minorities Who are Conducting HIV Research: Beyond Cultural Competency

Science.gov (United States)

Simoni, Jane M.; Evans-Campbell, Teresa (Tessa); Udell, Wadiya; Johnson-Jennings, Michelle; Pearson, Cynthia R.; MacDonald, Meg M.; Duran, Bonnie

2016-01-01

The majority of literature on mentoring focuses on mentee training needs, with significantly less guidance for the mentors. Moreover, many mentoring the mentor models assume generic (i.e. White) mentees with little attention to the concerns of underrepresented racial/ethnic minorities (UREM). This has led to calls for increased attention to diversity in research training programs, especially in the field of HIV where racial/ethnic disparities are striking. Diversity training tends to address the mentees' cultural competency in conducting research with diverse populations, and often neglects the training needs of mentors in working with diverse mentees. In this article, we critique the framing of diversity as the problem (rather than the lack of mentor consciousness and skills), highlight the need to extend mentor training beyond aspirations of cultural competency toward cultural humility and cultural safety, and consider challenges to effective mentoring of UREM, both for White and UREM mentors. PMID:27484060

Inhibition of human immunodeficiency virus type 1 (HIV-1) nuclear import via Vpr-Importin α interactions as a novel HIV-1 therapy

International Nuclear Information System (INIS)

Suzuki, Tatsunori; Yamamoto, Norio; Nonaka, Mizuho; Hashimoto, Yoshie; Matsuda, Go; Takeshima, Shin-nosuke; Matsuyama, Megumi; Igarashi, Tatsuhiko; Miura, Tomoyuki; Tanaka, Rie; Kato, Shingo; Aida, Yoko

2009-01-01

The development of multidrug-resistant viruses compromises the efficacy of anti-human immunodeficiency virus (HIV) therapy and limits treatment options. Therefore, new targets that can be used to develop novel antiviral agents need to be identified. One such target is the interaction between Vpr, one of the accessory gene products of HIV-1 and Importin α, which is crucial, not only for the nuclear import of Vpr, but also for HIV-1 replication in macrophages. We have identified a potential parent compound, hematoxylin, which suppresses Vpr-Importin α interaction, thereby inhibiting HIV-1 replication in a Vpr-dependent manner. Analysis by real-time PCR demonstrated that hematoxylin specifically inhibited nuclear import step of pre-integration complex. Thus, hematoxylin is a new anti-HIV-1 inhibitor that targets the nuclear import of HIV-1 via the Vpr-Importin α interaction, suggesting that a specific inhibitor of the interaction between viral protein and the cellular factor may provide a new strategy for HIV-1 therapy.

Pre-clinical evaluation of a replication-competent recombinant adenovirus serotype 4 vaccine expressing influenza H5 hemagglutinin.

Directory of Open Access Journals (Sweden)

Jeff Alexander

Full Text Available Influenza virus remains a significant health and social concern in part because of newly emerging strains, such as avian H5N1 virus. We have developed a prototype H5N1 vaccine using a recombinant, replication-competent Adenovirus serotype 4 (Ad4 vector, derived from the U.S. military Ad4 vaccine strain, to express the hemagglutinin (HA gene from A/Vietnam/1194/2004 influenza virus (Ad4-H5-Vtn. Our hypothesis is that a mucosally-delivered replicating Ad4-H5-Vtn recombinant vector will be safe and induce protective immunity against H5N1 influenza virus infection and disease pathogenesis.The Ad4-H5-Vtn vaccine was designed with a partial deletion of the E3 region of Ad4 to accommodate the influenza HA gene. Replication and growth kinetics of the vaccine virus in multiple human cell lines indicated that the vaccine virus is attenuated relative to the wild type virus. Expression of the HA transgene in infected cells was documented by flow cytometry, western blot analysis and induction of HA-specific antibody and cellular immune responses in mice. Of particular note, mice immunized intranasally with the Ad4-H5-Vtn vaccine were protected against lethal H5N1 reassortant viral challenge even in the presence of pre-existing immunity to the Ad4 wild type virus.Several non-clinical attributes of this vaccine including safety, induction of HA-specific humoral and cellular immunity, and efficacy were demonstrated using an animal model to support Phase 1 clinical trial evaluation of this new vaccine.

Interferon-alpha mediates restriction of human immunodeficiency virus type-1 replication in primary human macrophages at an early stage of replication.

Directory of Open Access Journals (Sweden)

Kelly M Cheney

2010-10-01

Full Text Available Type I interferons (IFNα and β are induced directly in response to viral infection, resulting in an antiviral state for the cell. In vitro studies have shown that IFNα is a potent inhibitor of viral replication; however, its role in HIV-1 infection is incompletely understood. In this study we describe the ability of IFNα to restrict HIV-1 infection in primary human macrophages in contrast to peripheral blood mononuclear cells and monocyte-derived dendritic cells. Inhibition to HIV-1 replication in cells pretreated with IFNα occurred at an early stage in the virus life cycle. Late viral events such as budding and subsequent rounds of infection were not affected by IFNα treatment. Analysis of early and late HIV-1 reverse transcripts and integrated proviral DNA confirmed an early post entry role for IFNα. First strand cDNA synthesis was slightly reduced but late and integrated products were severely depleted, suggesting that initiation or the nucleic acid intermediates of reverse transcription are targeted. The depletion of integrated provirus is disproportionally greater than that of viral cDNA synthesis suggesting the possibility of a least an additional later target. A role for either cellular protein APOBEC3G or tetherin in this IFNα mediated restriction has been excluded. Vpu, previously shown by others to rescue a viral budding restriction by tetherin, could not overcome this IFNα induced effect. Determining both the viral determinants and cellular proteins involved may lead to novel therapeutic approaches. Our results add to the understanding of HIV-1 restriction by IFNα.

Dynamics of HIV-containing compartments in macrophages reveal sequestration of virions and transient surface connections.

Directory of Open Access Journals (Sweden)

Raphaël Gaudin

Full Text Available During HIV pathogenesis, infected macrophages behave as "viral reservoirs" that accumulate and retain virions within dedicated internal Virus-Containing Compartments (VCCs. The nature of VCCs remains ill characterized and controversial. Using wild-type HIV-1 and a replication-competent HIV-1 carrying GFP internal to the Gag precursor, we analyzed the biogenesis and evolution of VCCs in primary human macrophages. VCCs appear roughly 14 hours after viral protein synthesis is detected, initially contain few motile viral particles, and then mature to fill up with virions that become packed and immobile. The amount of intracellular Gag, the proportion of dense VCCs, and the density of viral particles in their lumen increased with time post-infection. In contrast, the secretion of virions, their infectivity and their transmission to T cells decreased overtime, suggesting that HIV-infected macrophages tend to pack and retain newly formed virions into dense compartments. A minor proportion of VCCs remains connected to the plasma membrane overtime. Surprisingly, live cell imaging combined with correlative light and electron microscopy revealed that such connections can be transient, highlighting their dynamic nature. Together, our results shed light on the late phases of the HIV-1 cycle and reveal some of its macrophage specific features.

Gene Therapy Targeting HIV Entry

Directory of Open Access Journals (Sweden)

Chuka Didigu

2014-03-01

Full Text Available Despite the unquestionable success of antiretroviral therapy (ART in the treatment of HIV infection, the cost, need for daily adherence, and HIV-associated morbidities that persist despite ART all underscore the need to develop a cure for HIV. The cure achieved following an allogeneic hematopoietic stem cell transplant (HSCT using HIV-resistant cells, and more recently, the report of short-term but sustained, ART-free control of HIV replication following allogeneic HSCT, using HIV susceptible cells, have served to both reignite interest in HIV cure research, and suggest potential mechanisms for a cure. In this review, we highlight some of the obstacles facing HIV cure research today, and explore the roles of gene therapy targeting HIV entry, and allogeneic stem cell transplantation in the development of strategies to cure HIV infection.

Quantification of integrated HIV DNA by repetitive-sampling Alu-HIV PCR on the basis of poisson statistics.

Science.gov (United States)

De Spiegelaere, Ward; Malatinkova, Eva; Lynch, Lindsay; Van Nieuwerburgh, Filip; Messiaen, Peter; O'Doherty, Una; Vandekerckhove, Linos

2014-06-01

Quantification of integrated proviral HIV DNA by repetitive-sampling Alu-HIV PCR is a candidate virological tool to monitor the HIV reservoir in patients. However, the experimental procedures and data analysis of the assay are complex and hinder its widespread use. Here, we provide an improved and simplified data analysis method by adopting binomial and Poisson statistics. A modified analysis method on the basis of Poisson statistics was used to analyze the binomial data of positive and negative reactions from a 42-replicate Alu-HIV PCR by use of dilutions of an integration standard and on samples of 57 HIV-infected patients. Results were compared with the quantitative output of the previously described Alu-HIV PCR method. Poisson-based quantification of the Alu-HIV PCR was linearly correlated with the standard dilution series, indicating that absolute quantification with the Poisson method is a valid alternative for data analysis of repetitive-sampling Alu-HIV PCR data. Quantitative outputs of patient samples assessed by the Poisson method correlated with the previously described Alu-HIV PCR analysis, indicating that this method is a valid alternative for quantifying integrated HIV DNA. Poisson-based analysis of the Alu-HIV PCR data enables absolute quantification without the need of a standard dilution curve. Implementation of the CI estimation permits improved qualitative analysis of the data and provides a statistical basis for the required minimal number of technical replicates. © 2014 The American Association for Clinical Chemistry.

Variable fitness impact of HIV-1 escape mutations to cytotoxic T lymphocyte (CTL response.

Directory of Open Access Journals (Sweden)

Ryan M Troyer

2009-04-01

Full Text Available Human lymphocyte antigen (HLA-restricted CD8(+ cytotoxic T lymphocytes (CTL target and kill HIV-infected cells expressing cognate viral epitopes. This response selects for escape mutations within CTL epitopes that can diminish viral replication fitness. Here, we assess the fitness impact of escape mutations emerging in seven CTL epitopes in the gp120 Env and p24 Gag coding regions of an individual followed longitudinally from the time of acute HIV-1 infection, as well as some of these same epitopes recognized in other HIV-1-infected individuals. Nine dominant mutations appeared in five gp120 epitopes within the first year of infection, whereas all four mutations found in two p24 epitopes emerged after nearly two years of infection. These mutations were introduced individually into the autologous gene found in acute infection and then placed into a full-length, infectious viral genome. When competed against virus expressing the parental protein, fitness loss was observed with only one of the nine gp120 mutations, whereas four had no effect and three conferred a slight increase in fitness. In contrast, mutations conferring CTL escape in the p24 epitopes significantly decreased viral fitness. One particular escape mutation within a p24 epitope was associated with reduced peptide recognition and high viral fitness costs but was replaced by a fitness-neutral mutation. This mutation appeared to alter epitope processing concomitant with a reduced CTL response. In conclusion, CTL escape mutations in HIV-1 Gag p24 were associated with significant fitness costs, whereas most escape mutations in the Env gene were fitness neutral, suggesting a balance between immunologic escape and replicative fitness costs.

Mucosal IgA Responses: Damaged in Established HIV Infection—Yet, Effective Weapon against HIV Transmission

Directory of Open Access Journals (Sweden)

Viraj Kulkarni

2017-11-01

Full Text Available HIV infection not only destroys CD4+ T cells but also inflicts serious damage to the B-cell compartment, such as lymphadenopathy, destruction of normal B-cell follicle architecture, polyclonal hypergammaglobulinemia, increased apoptosis of B cells, and irreversible loss of memory B-cell responses with advanced HIV disease. Subepithelial B cells and plasma cells are also affected, which results in loss of mucosal IgG and IgA antibodies. This leaves the mucosal barrier vulnerable to bacterial translocation. The ensuing immune activation in mucosal tissues adds fuel to the fire of local HIV replication. We postulate that compromised mucosal antibody defenses also facilitate superinfection of HIV-positive individuals with new HIV strains. This in turn sets the stage for the generation of circulating recombinant forms of HIV. What can the mucosal B-cell compartment contribute to protect a healthy, uninfected host against mucosal HIV transmission? Here, we discuss proof-of-principle studies we have performed using passive mucosal immunization, i.e., topical administration of preformed anti-HIV monoclonal antibodies (mAbs as IgG1, dimeric IgA1 (dIgA1, and dIgA2 isotypes, alone or in combination. Our data indicate that mucosally applied anti-HIV envelope mAbs can provide potent protection against mucosal transmission of simian-human immunodeficiency virus. Our review also discusses the induction of mucosal antibody defenses by active vaccination and potential strategies to interrupt the vicious cycle of bacterial translocation, immune activation, and stimulation of HIV replication in individuals with damaged mucosal barriers.

The development of the conditionally replication-competent adenovirus: replacement of E4 orf1-4 region by exogenous gene.

Science.gov (United States)

Nam, Jae-Kook; Lee, Mi-Hyang; Seo, Hae-Hyun; Kim, Seok-Ki; Lee, Kang-Huyn; Kim, In-Hoo; Lee, Sang-Jin

2010-05-01

Tumor or tissue specific replicative adenovirus armed with a therapeutic gene has shown a promising anti-cancer therapeutic modality. However, because the genomic packaging capacity is constrained, only a few places inside it are available for transgene insertion. In the present study, we introduce a novel strategy utilizing the early E4 region for the insertion of therapeutic gene(s). We constructed the conditionally replication-competent adenovirus (CRAd), Ad5E4(mRFP) by: (i) replacing the E4/E1a promoter by the prostate-specific enhancer element; (ii) inserting mRFP inside the E4orf1-4 deletion region; and (iii) sub-cloning enhanced green fluorescent protein controlled by cytomegalovirus promoter in the left end of the viral genome. Subsequently, we evaluated its replication abilities and killing activities in vitro, as well as its in vivo anti-tumor efficacy in CWR22rv xenografts. When infected with Ad5E4(mRFP), the number and intensity of the mRFP gene products increased in a prostate cancer cell-specific manner as designed, suggesting that the mRFP gene and E4orfs other than E4orf1-4 were well synthesized from one transcript via alternative splicing as the recombinant adenovirus replicated. As expected from the confirmed virus replication capability, Ad5E4(mRFP) induced cell lysis as potent as the wild-type adenovirus and effectively suppressed tumor growth when tested in the CWR22rv xenografts in nude mice. Furthermore, Ad5E4(endo/angio) harboring an endostatin-angiostatin gene in E4orf1-4 was able to enhance CRAd by replacing mRFP with a therapeutic gene. The approach employed in the present study for the insertion of a therapeutic transgene in CRAd should facilitate the construction of CRAd containing multiple therapeutic genes in the viral genome that may have the potential to serve as highly potent cancer therapeutic reagents. Copyright (c) 2010 John Wiley & Sons, Ltd.

Improving HIV proteome annotation: new features of BioAfrica HIV Proteomics Resource.

Science.gov (United States)

Druce, Megan; Hulo, Chantal; Masson, Patrick; Sommer, Paula; Xenarios, Ioannis; Le Mercier, Philippe; De Oliveira, Tulio

2016-01-01

The Human Immunodeficiency Virus (HIV) is one of the pathogens that cause the greatest global concern, with approximately 35 million people currently infected with HIV. Extensive HIV research has been performed, generating a large amount of HIV and host genomic data. However, no effective vaccine that protects the host from HIV infection is available and HIV is still spreading at an alarming rate, despite effective antiretroviral (ARV) treatment. In order to develop effective therapies, we need to expand our knowledge of the interaction between HIV and host proteins. In contrast to virus proteins, which often rapidly evolve drug resistance mutations, the host proteins are essentially invariant within all humans. Thus, if we can identify the host proteins needed for virus replication, such as those involved in transporting viral proteins to the cell surface, we have a chance of interrupting viral replication. There is no proteome resource that summarizes this interaction, making research on this subject a difficult enterprise. In order to fill this gap in knowledge, we curated a resource presents detailed annotation on the interaction between the HIV proteome and host proteins. Our resource was produced in collaboration with ViralZone and used manual curation techniques developed by UniProtKB/Swiss-Prot. Our new website also used previous annotations of the BioAfrica HIV-1 Proteome Resource, which has been accessed by approximately 10 000 unique users a year since its inception in 2005. The novel features include a dedicated new page for each HIV protein, a graphic display of its function and a section on its interaction with host proteins. Our new webpages also add information on the genomic location of each HIV protein and the position of ARV drug resistance mutations. Our improved BioAfrica HIV-1 Proteome Resource fills a gap in the current knowledge of biocuration.Database URL:http://www.bioafrica.net/proteomics/HIVproteome.html. © The Author(s) 2016. Published

Neurological complication in HIV patients

Science.gov (United States)

Ritarwan, K.

2018-03-01

Human Immunodeficiency Virus (HIV) is neurotropic and immunotropic, making themassive destruction of both systems. Although their amount has been reduced, there is still neurological presentations and complications of HIV remain common in the era of combination antiretroviral therapy (cART). Neurological opportunistic infections (OI) occur in advanced HIV diseases such as primary cerebral lymphoma, cryptococcal meningitis, cerebral toxoplasmosis, and progressive multifocal encephalopathy. Neurological problem directly related to HIV appear at any stage in the progress of HIV disease, from AIDS-associated dementia to the aseptic meningitis of primary HIV infection observed in subjects with an immune deficiency. The replication of peripheral HIV viral is able to be controlled in the era of effective antiretroviral therapy. Non-HIV-related neurological disease such as stroke increased important as the HIV population ages.

Serum selenium status of HIV-infected children on care and ...

African Journals Online (AJOL)

Although the use of HAART has revolutionised the management of. HIV infection ... prevent the replication of HIV and retard the development of AIDS in newly infected ..... Effect of multiple ... Nutrition, HIV, and drug abuse: The molecular basis ...

Differential effect of CLK SR Kinases on HIV-1 gene expression: potential novel targets for therapy

Directory of Open Access Journals (Sweden)

Dobson Wendy

2011-06-01

Full Text Available Abstract Background RNA processing plays a critical role in the replication of HIV-1, regulated in part through the action of host SR proteins. To explore the impact of modulating SR protein activity on virus replication, the effect of increasing or inhibiting the activity of the Cdc2-like kinase (CLK family of SR protein kinases on HIV-1 expression and RNA processing was examined. Results Despite their high homology, increasing individual CLK expression had distinct effects on HIV-1, CLK1 enhancing Gag production while CLK2 inhibited the virus. Parallel studies on the anti-HIV-1 activity of CLK inhibitors revealed a similar discrepant effect on HIV-1 expression. TG003, an inhibitor of CLK1, 2 and 4, had no effect on viral Gag synthesis while chlorhexidine, a CLK2, 3 and 4 inhibitor, blocked virus production. Chlorhexidine treatment altered viral RNA processing, decreasing levels of unspliced and single spliced viral RNAs, and reduced Rev accumulation. Subsequent experiments in the context of HIV-1 replication in PBMCs confirmed the capacity of chlorhexidine to suppress virus replication. Conclusions Together, these findings establish that HIV-1 RNA processing can be targeted to suppress virus replication as demonstrated by manipulating individual CLK function and identified chlorhexidine as a lead compound in the development of novel anti-viral therapies.

Replicative senescence of T cells: does the Hayflick Limit lead to immune exhaustion?

Science.gov (United States)

Effros, R B; Pawelec, G

1997-09-01

Extensive in vitro research on fibroblasts has defined numerous genetic and phenotypic changes associated with replicative senescence. Identification of T-cell replicative senescence as a feature of human immunodeficiency virus (HIV) disease and ageing suggests this phenomenon merits more careful consideration by immunologists, especially with regard to chronic infection, memory and adoptive immunotherapy.

Mechanisms and regulation of DNA replication initiation in eukaryotes.

Science.gov (United States)

Parker, Matthew W; Botchan, Michael R; Berger, James M

2017-04-01

Cellular DNA replication is initiated through the action of multiprotein complexes that recognize replication start sites in the chromosome (termed origins) and facilitate duplex DNA melting within these regions. In a typical cell cycle, initiation occurs only once per origin and each round of replication is tightly coupled to cell division. To avoid aberrant origin firing and re-replication, eukaryotes tightly regulate two events in the initiation process: loading of the replicative helicase, MCM2-7, onto chromatin by the origin recognition complex (ORC), and subsequent activation of the helicase by its incorporation into a complex known as the CMG. Recent work has begun to reveal the details of an orchestrated and sequential exchange of initiation factors on DNA that give rise to a replication-competent complex, the replisome. Here, we review the molecular mechanisms that underpin eukaryotic DNA replication initiation - from selecting replication start sites to replicative helicase loading and activation - and describe how these events are often distinctly regulated across different eukaryotic model organisms.

Modification of a loop sequence between α-helices 6 and 7 of virus capsid (CA protein in a human immunodeficiency virus type 1 (HIV-1 derivative that has simian immunodeficiency virus (SIVmac239 vif and CA α-helices 4 and 5 loop improves replication in cynomolgus monkey cells

Directory of Open Access Journals (Sweden)

Adachi Akio

2009-08-01

Full Text Available Abstract Background Human immunodeficiency virus type 1 (HIV-1 productively infects only humans and chimpanzees but not cynomolgus or rhesus monkeys while simian immunodeficiency virus isolated from macaque (SIVmac readily establishes infection in those monkeys. Several HIV-1 and SIVmac chimeric viruses have been constructed in order to develop an animal model for HIV-1 infection. Construction of an HIV-1 derivative which contains sequences of a SIVmac239 loop between α-helices 4 and 5 (L4/5 of capsid protein (CA and the entire SIVmac239 vif gene was previously reported. Although this chimeric virus could grow in cynomolgus monkey cells, it did so much more slowly than did SIVmac. It was also reported that intrinsic TRIM5α restricts the post-entry step of HIV-1 replication in rhesus and cynomolgus monkey cells, and we previously demonstrated that a single amino acid in a loop between α-helices 6 and 7 (L6/7 of HIV type 2 (HIV-2 CA determines the susceptibility of HIV-2 to cynomolgus monkey TRIM5α. Results In the study presented here, we replaced L6/7 of HIV-1 CA in addition to L4/5 and vif with the corresponding segments of SIVmac. The resultant HIV-1 derivatives showed enhanced replication capability in established T cell lines as well as in CD8+ cell-depleted primary peripheral blood mononuclear cells from cynomolgus monkey. Compared with the wild type HIV-1 particles, the viral particles produced from a chimeric HIV-1 genome with those two SIVmac loops were less able to saturate the intrinsic restriction in rhesus monkey cells. Conclusion We have succeeded in making the replication of simian-tropic HIV-1 in cynomolgus monkey cells more efficient by introducing into HIV-1 the L6/7 CA loop from SIVmac. It would be of interest to determine whether HIV-1 derivatives with SIVmac CA L4/5 and L6/7 can establish infection of cynomolgus monkeys in vivo.

Modeling HIV-associated neurocognitive disorders in mice: new approaches in the changing face of HIV neuropathogenesis

OpenAIRE

Laura B. Jaeger; Avindra Nath

2012-01-01

It is well established that infection with the human immunodeficiency virus (HIV) leads to immune suppression. Less well known is the fact that long-term, progressive HIV disease is associated with the development of cognitive deficits. Since the introduction of combined antiretroviral therapy (cART), the clinical presentation of HIV infection has evolved into a chronic illness with very low levels of viral replication and chronic immune activation, with compliant affected individuals survivi...

HIV restriction by APOBEC3 in humanized mice.

Directory of Open Access Journals (Sweden)

John F Krisko

2013-03-01

Full Text Available Innate immune restriction factors represent important specialized barriers to zoonotic transmission of viruses. Significant consideration has been given to their possible use for therapeutic benefit. The apolipoprotein B mRNA editing enzyme catalytic polypeptide 3 (APOBEC3 family of cytidine deaminases are potent immune defense molecules capable of efficiently restricting endogenous retroelements as well as a broad range of viruses including Human Immunodeficiency virus (HIV, Hepatitis B virus (HBV, Human Papilloma virus (HPV, and Human T Cell Leukemia virus (HTLV. The best characterized members of this family are APOBEC3G (A3G and APOBEC3F (A3F and their restriction of HIV. HIV has evolved to counteract these powerful restriction factors by encoding an accessory gene designated viral infectivity factor (vif. Here we demonstrate that APOBEC3 efficiently restricts CCR5-tropic HIV in the absence of Vif. However, our results also show that CXCR4-tropic HIV can escape from APOBEC3 restriction and replicate in vivo independent of Vif. Molecular analysis identified thymocytes as cells with reduced A3G and A3F expression. Direct injection of vif-defective HIV into the thymus resulted in viral replication and dissemination detected by plasma viral load analysis; however, vif-defective viruses remained sensitive to APOBEC3 restriction as extensive G to A mutation was observed in proviral DNA recovered from other organs. Remarkably, HIV replication persisted despite the inability of HIV to develop resistance to APOBEC3 in the absence of Vif. Our results provide novel insight into a highly specific subset of cells that potentially circumvent the action of APOBEC3; however our results also demonstrate the massive inactivation of CCR5-tropic HIV in the absence of Vif.

Picomolar dichotomous activity of gnidimacrin against HIV-1.

Directory of Open Access Journals (Sweden)

Li Huang

Full Text Available Highly active antiretroviral therapy (HAART has offered a promising approach for controlling HIV-1 replication in infected individuals. However, with HARRT, HIV-1 is suppressed rather than eradicated due to persistence of HIV-1 in latent viral reservoirs. Thus, purging the virus from latent reservoirs is an important strategy toward eradicating HIV-1 infection. In this study, we discovered that the daphnane diterpene gnidimacrin, which was previously reported to have potent anti-cancer cell activity, activated HIV-1 replication and killed persistently-infected cells at picomolar concentrations. In addition to its potential to purge HIV-1 from latently infected cells, gnidimacrin potently inhibited a panel of HIV-1 R5 virus infection of peripheral blood mononuclear cells (PBMCs at an average concentration lower than 10 pM. In contrast, gnidimacrin only partially inhibited HIV-1 ×4 virus infection of PBMCs. The strong anti-HIV-1 R5 virus activity of gnidimacrin was correlated with its effect on down-regulation of the HIV-1 coreceptor CCR5. The anti-R5 virus activity of gnidimacrin was completely abrogated by a selective protein kinase C beta inhibitor enzastaurin, which suggests that protein kinase C beta plays a key role in the potent anti-HIV-1 activity of gnidimacrin in PBMCs. In summary, these results suggest that gnidimacrin could activate latent HIV-1, specifically kill HIV-1 persistently infected cells, and inhibit R5 viruses at picomolar concentrations.

The effect of aging, nutrition, and exercise during HIV infection

Directory of Open Access Journals (Sweden)

Gabriel Somarriba

2010-09-01

Full Text Available Gabriel Somarriba, Daniela Neri, Natasha Schaefer, Tracie L MillerDivision of Pediatric Clinical Research, Department of Pediatrics, Miller School of Medicine, University of Miami, Miami, Florida, USAAbstract: Medical advances continue to change the face of human immunodeficiency virus–acquired immunodeficiency syndrome (HIV/AIDS. As life expectancy increases, the number of people living with HIV rises, presenting new challenges for the management of a chronic condition. Aging, nutrition, and physical activity can influence outcomes in other chronic conditions, and emerging data show that each of these factors can impact viral replication and the immune system in HIV. HIV infection results in a decline of the immune system through the depletion of CD4+ T cells. From initial infection, viral replication is a continuous phenomenon. Immunosenescence, a hallmark of aging, results in an increased susceptibility to infections secondary to a delayed immune response, and this phenomenon may be increased in HIV-infected patients. Optimal nutrition is an important adjunct in the clinical care of patients with HIV. Nutritional interventions may improve the quality and span of life and symptom management, support the effectiveness of medications, and improve the patient’s resistance to infections and other disease complications by altering immunity. Moderate physical activity can improve many immune parameters, reduce the risk of acute infection, and combat metabolic abnormalities. As people with HIV age, alternative therapies such as nutrition and physical activity may complement medical management.Keywords: HIV replication, aging, diet, nutrition, exercise, immunity

Iron chelators ICL670 and 311 inhibit HIV-1 transcription

International Nuclear Information System (INIS)

Debebe, Zufan; Ammosova, Tatyana; Jerebtsova, Marina; Kurantsin-Mills, Joseph; Niu, Xiaomei; Charles, Sharroya; Richardson, Des R.; Ray, Patricio E.; Gordeuk, Victor R.; Nekhai, Sergei

2007-01-01

HIV-1 replication is induced by an excess of iron and iron chelation by desferrioxamine (DFO) inhibits viral replication by reducing proliferation of infected cells. Treatment of cells with DFO and 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (311) inhibit expression of proteins that regulate cell-cycle progression, including cycle-dependent kinase 2 (CDK2). Our recent studies showed that CDK2 participates in HIV-1 transcription and viral replication suggesting that inhibition of CDK2 by iron chelators might also affect HIV-1 transcription. Here we evaluated the effect of a clinically approved orally effective iron chelator, 4-[3,5-bis-(hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid (ICL670) and 311 on HIV-1 transcription. Both ICL670 and 311 inhibited Tat-induced HIV-1 transcription in CEM-T cells, 293T and HeLa cells. Neither ICL670 nor 311 induced cytotoxicity at concentrations that inhibited HIV-1 transcription. The chelators decreased cellular activity of CDK2 and reduced HIV-1 Tat phosphorylation by CDK2. Neither ICL670A or 311 decreased CDK9 protein level but significantly reduced association of CDK9 with cyclin T1 and reduced phosphorylation of Ser-2 residues of RNA polymerase II C-terminal domain. In conclusion, our findings add to the evidence that iron chelators can inhibit HIV-1 transcription by deregulating CDK2 and CDK9. Further consideration should be given to the development of iron chelators for future anti-retroviral therapeutics

Current Peptide and Protein Candidates Challenging HIV Therapy beyond the Vaccine Era

Directory of Open Access Journals (Sweden)

Koollawat Chupradit

2017-09-01

Full Text Available Human immunodeficiency virus (HIV is a causative agent of acquired immune deficiency syndrome (AIDS. Highly active antiretroviral therapy (HAART can slow down the replication of HIV-1, leading to an improvement in the survival of HIV-1-infected patients. However, drug toxicities and poor drug administration has led to the emergence of a drug-resistant strain. HIV-1 immunotherapy has been continuously developed, but antibody therapy and HIV vaccines take time to improve its efficiency and have limitations. HIV-1-specific chimeric antigen receptor (CAR-based immunotherapy founded on neutralizing antibodies is now being developed. In HIV-1 therapy, anti-HIV chimeric antigen receptors showed promising data in the suppression of HIV-1 replication; however, autologous transfusion is still a problem. This has led to the development of effective peptides and proteins for an alternative HIV-1 treatment. In this paper, we provide a comprehensive review of potent anti-HIV-1 peptides and proteins that reveal promising therapeutic activities. The inhibitory mechanisms of each therapeutic molecule in the different stages of the HIV-1 life cycle will be discussed herein.

The anti-HIV-1 effect of scutellarin

International Nuclear Information System (INIS)

Zhang Gaohong; Wang Qian; Chen Jijun; Zhang Xuemei; Tam, S.-C.; Zheng Yongtang

2005-01-01

Scutellarin was purified from the plant Erigeron breviscapus (Vant.) Hand.-Mazz. The activity against 3 strains of human immunodeficiency virus (HIV) was determined in vitro in this study. These were laboratory-derived virus (HIV-1 IIIB ), drug-resistant virus (HIV-1 74V ), and low-passage clinical isolated virus (HIV-1 KM018 ). From syncytia inhibition study, the EC 50 of scutellarin against HIV-1 IIIB direct infection in C8166 cells was 26 μM with a therapeutic index of 36. When the mode of infection changed from acute infection to cell-to-cell infection, this compound became even more potent and the EC 50 reduced to 15 μM. This suggested that cell fusion might be affected by this compound. By comparing the inhibitory effects on p24 antigen, scutellarin was also found to be active against HIV-1 74V (EC 50 253 μM) and HIV-1 KM018 (EC 50 136 μM) infection with significant difference in potency. The mechanism of its action was also explored in this study. At a concentration of 433 μM, scutellarin inhibited 48% of the cell free recombinant HIV-1 RT activity. It also caused 82% inhibition of HIV-1 particle attachment and 45% inhibition of fusion at the concentrations of 54 μM. In summary, scutellarin was found to inhibit several strains of HIV-1 replication with different potencies. It appeared to inhibit HIV-1 RT activity, HIV-1 particle attachment and cell fusion. These are essential activities for viral transmission and replication

Optimization and validation of a neutralizing antibody assay for HIV-1 in A3R5 cells.

Science.gov (United States)

Sarzotti-Kelsoe, Marcella; Daniell, Xiaoju; Todd, Christopher A; Bilska, Miroslawa; Martelli, Amanda; LaBranche, Celia; Perez, Lautaro G; Ochsenbauer, Christina; Kappes, John C; Rountree, Wes; Denny, Thomas N; Montefiori, David C

2014-07-01

A3R5 is a human CD4(+) lymphoblastoid cell line that was engineered to express CCR5 and is useful for the detection of weak neutralizing antibody responses against tier 2 strains of HIV-1. Here we describe the optimization and validation of the HIV-1 neutralizing antibody assay that utilizes A3R5 cells, performed in compliance with Good Clinical Laboratory Practice (GCLP) guidelines. The assay utilizes Renilla luciferase-expressing replication competent infectious molecular clones (IMC) encoding heterologous env genes from different HIV-1 clades. Key assay validation parameters tested included specificity, accuracy, precision, limit of detection and quantitation, specificity, linearity and range, and robustness. Plasma samples demonstrated higher non-specific activity than serum samples in the A3R5 assay. This assay can tolerate a wide range of virus input but is more sensitive to cell concentration. The higher sensitivity of the A3R5 assay in neutralization responses to tier 2 strains of HIV-1 makes it complementary to, but not a substitute for the TZM-bl assay. The validated A3R5 assay is employed as an endpoint immunogenicity test for vaccine-elicited neutralizing antibodies against tier 2 strains of HIV-1, and to identify correlates of protection in HIV-1 vaccine trials conducted globally. Copyright © 2014 Elsevier B.V. All rights reserved.

C–C Chemokines Released by Lipopolysaccharide (LPS)-stimulated Human Macrophages Suppress HIV-1 Infection in Both Macrophages and T Cells

Science.gov (United States)

Verani, Alessia; Scarlatti, Gabriella; Comar, Manola; Tresoldi, Eleonora; Polo, Simona; Giacca, Mauro; Lusso, Paolo; Siccardi, Antonio G.; Vercelli, Donata

1997-01-01

Human immunodeficiency virus-1 (HIV-1) expression in monocyte-derived macrophages (MDM) infected in vitro is known to be inhibited by lipopolysaccharide (LPS). However, the mechanisms are incompletely understood. We show here that HIV-1 suppression is mediated by soluble factors released by MDM stimulated with physiologically significant concentrations of LPS. LPS-conditioned supernatants from MDM inhibited HIV-1 replication in both MDM and T cells. Depletion of C–C chemokines (RANTES, MIP-1α, and MIP-1β) neutralized the ability of LPS-conditioned supernatants to inhibit HIV-1 replication in MDM. A combination of recombinant C–C chemokines blocked HIV-1 infection as effectively as LPS. Here, we report an inhibitory effect of C–C chemokines on HIV replication in primary macrophages. Our results raise the possibility that monocytes may play a dual role in HIV infection: while representing a reservoir for the virus, they may contribute to the containment of the infection by releasing factors that suppress HIV replication not only in monocytes but also in T lymphocytes. PMID:9120386

Discordant Impact of HLA on Viral Replicative Capacity and Disease Progression in Pediatric and Adult HIV Infection.

Directory of Open Access Journals (Sweden)

Emily Adland

2015-06-01

Full Text Available HLA class I polymorphism has a major influence on adult HIV disease progression. An important mechanism mediating this effect is the impact on viral replicative capacity (VRC of the escape mutations selected in response to HLA-restricted CD8+ T-cell responses. Factors that contribute to slow progression in pediatric HIV infection are less well understood. We here investigate the relationship between VRC and disease progression in pediatric infection, and the effect of HLA on VRC and on disease outcome in adult and pediatric infection. Studying a South African cohort of >350 ART-naïve, HIV-infected children and their mothers, we first observed that pediatric disease progression is significantly correlated with VRC. As expected, VRCs in mother-child pairs were strongly correlated (p = 0.004. The impact of the protective HLA alleles, HLA-B*57, HLA-B*58:01 and HLA-B*81:01, resulted in significantly lower VRCs in adults (p<0.0001, but not in children. Similarly, in adults, but not in children, VRCs were significantly higher in subjects expressing the disease-susceptible alleles HLA-B*18:01/45:01/58:02 (p = 0.007. Irrespective of the subject, VRCs were strongly correlated with the number of Gag CD8+ T-cell escape mutants driven by HLA-B*57/58:01/81:01 present in each virus (p = 0.0002. In contrast to the impact of VRC common to progression in adults and children, the HLA effects on disease outcome, that are substantial in adults, are small and statistically insignificant in infected children. These data further highlight the important role that VRC plays both in adult and pediatric progression, and demonstrate that HLA-independent factors, yet to be fully defined, are predominantly responsible for pediatric non-progression.

Alterations in HIV-1 LTR promoter activity during AIDS progression

International Nuclear Information System (INIS)

Hiebenthal-Millow, Kirsten; Greenough, Thomas C.; Bretttler, Doreen B.; Schindler, Michael; Wildum, Steffen; Sullivan, John L.; Kirchhoff, Frank

2003-01-01

HIV-1 variants evolving in AIDS patients frequently show increased replicative capacity compared to those present during early asymptomatic infection. It is known that late stage HIV-1 variants often show an expanded coreceptor tropism and altered Nef function. In the present study we investigated whether enhanced HIV-1 LTR promoter activity might also evolve during disease progression. Our results demonstrate increased LTR promoter activity after AIDS progression in 3 of 12 HIV-1-infected individuals studied. Further analysis revealed that multiple alterations in the U3 core-enhancer and in the transactivation-response (TAR) region seem to be responsible for the enhanced functional activity. Our findings show that in a subset of HIV-1-infected individuals enhanced LTR transcription contributes to the increased replicative potential of late stage virus isolates and might accelerate disease progression

Analyses of prevalence and polymorphisms of six replication-competent and chromosomally assigned porcine endogenous retroviruses in individual pigs and pig subspecies

International Nuclear Information System (INIS)

Niebert, Marcus; Toenjes, Ralf R.

2003-01-01

As porcine endogenous retroviruses (PERV) productively infect human cells in vitro, they pose a serious risk in xenotransplantation and xenogeneic cell therapies. We have analyzed the prevalence of six well-characterized full-length PERV, five of them being replication-competent and four of them being chromosomally assigned (J. Virol. 75 (2001) 5465; J. Virol. 76 (2002) 2714). These analyses revealed a heterogeneous distribution of PERV among individuals and, as no PERV is present in every pig, it seems feasible to generate pigs free of functional PERV by conventional breeding. Conversely, as PERV are polymorphic, single proviruses may have escaped detection and this kind of assay must be performed for every herd used in xenotransplantation or xenogeneic cell therapies. In addition, specific proviruses show internal point mutations which significantly affect their replicational capacities. As there are two different types of PERV LTR structures showing varying levels of transcriptional capacity (J. Virol. 75 (2001) 6933), an analysis of 21 distinct chromosomal locations revealed that PERV which harbor highly active LTRs with repeat elements in U3 are dominant

CD41 T cell recovery during suppression of HIV replication: an international comparison of the immunological efficacy of antiretroviral therapy in North America, Asia and Africa.

Science.gov (United States)

Geng, Elvin H; Neilands, Torsten B; Thièbaut, Rodolphe; Bwana, Mwebesa Bosco; Nash, Denis; Moore, Richard D; Wood, Robin; Zannou, Djimon Marcel; Althoff, Keri N; Lim, Poh Lian; Nachega, Jean B; Easterbrook, Philippa J; Kambugu, Andrew; Little, Francesca; Nakigozi, Gertrude; Nakanjako, Damalie; Kiggundu, Valerian; Ki Li, Patrick Chung; Bangsberg, David R; Fox, Matthew P; Prozesky, HansW; Hunt, Peter W; Davies, Mary-Ann; Reynolds, Steven J; Egger, Matthias; Yiannoutsos, Constantin T; Vittinghoff, Eric V; Deeks, Steven G; Martin, Jeffrey N

2015-02-01

Even among HIV-infected patients who fully suppress plasma HIV RNA replication on antiretroviral therapy, genetic (e.g. CCL3L1 copy number), viral (e.g. tropism) and environmental (e.g. chronic exposure to microbial antigens) factors influence CD4 recovery. These factors differ markedly around the world and therefore the expected CD4 recovery during HIV RNA suppression may differ globally. We evaluated HIV-infected adults from North America, West Africa, East Africa, Southern Africa and Asia starting non-nucleoside reverse transcriptase inhibitorbased regimens containing efavirenz or nevirapine, who achieved at least one HIV RNA level Africa showed diminished CD4 recovery as compared with other regions. Three years after antiretroviral therapy initiation, the mean CD4 count for a prototypical patient with a pre-therapy CD4 count of 150/ml was 529/ml [95% confidence interval (CI): 517–541] in North America, 494/ml (95% CI: 429–559) in West Africa, 515/ml (95% CI: 508–522) in Southern Africa, 503/ml (95% CI: 478–528) in Asia and 437/ml (95% CI: 425–449) in East Africa. CD4 recovery during HIV RNA suppression is diminished in East Africa as compared with other regions of the world, and observed differences are large enough to potentially influence clinical outcomes. Epidemiological analyses on a global scale can identify macroscopic effects unobservable at the clinical, national or individual regional level.

Natural HIV-1 NEF accelerates virus replication in primary human lymphocytes

NARCIS (Netherlands)

de Ronde, A.; Klaver, B.; Keulen, W.; Smit, L.; Goudsmit, J.

1992-01-01

HIV-1 NEF genes were isolated directly from peripheral blood lymphocyte DNA of two HIV-1-infected individuals and cloned into an HXB-2-infectious molecular clone. The effect of NEF on virus production in T-cell lines and primary human lymphocytes was studied. Naturally occurring NEF accelerates

Dynamics of viral replication in blood and lymphoid tissues during SIVmac251 infection of macaques

Directory of Open Access Journals (Sweden)

Mannioui Abdelkrim

2009-01-01

Full Text Available Abstract Background Extensive studies of primary infection are crucial to our understanding of the course of HIV disease. In SIV-infected macaques, a model closely mimicking HIV pathogenesis, we used a combination of three markers -- viral RNA, 2LTR circles and viral DNA -- to evaluate viral replication and dissemination simultaneously in blood, secondary lymphoid tissues, and the gut during primary and chronic infections. Subsequent viral compartmentalization in the main target cells of the virus in peripheral blood during the chronic phase of infection was evaluated by cell sorting and viral quantification with the three markers studied. Results The evolutions of viral RNA, 2LTR circles and DNA levels were correlated in a given tissue during primary and early chronic infection. The decrease in plasma viral load principally reflects a large decrease in viral replication in gut-associated lymphoid tissue (GALT, with viral RNA and DNA levels remaining stable in the spleen and peripheral lymph nodes. Later, during chronic infection, a progressive depletion of central memory CD4+ T cells from the peripheral blood was observed, accompanied by high levels of viral replication in the cells of this subtype. The virus was also found to replicate at this point in the infection in naive CD4+ T cells. Viral RNA was frequently detected in monocytes, but no SIV replication appeared to occur in these cells, as no viral DNA or 2LTR circles were detected. Conclusion We demonstrated the persistence of viral replication and dissemination, mostly in secondary lymphoid tissues, during primary and early chronic infection. During chronic infection, the central memory CD4+ T cells were the major site of viral replication in peripheral blood, but viral replication also occurred in naive CD4+ T cells. The role of monocytes seemed to be limited to carrying the virus as a cargo because there was an observed lack of replication in these cells. These data may have important

The evolutionary ecology of molecular replicators.

Science.gov (United States)

Nee, Sean

2016-08-01

By reasonable criteria, life on the Earth consists mainly of molecular replicators. These include viruses, transposons, transpovirons, coviruses and many more, with continuous new discoveries like Sputnik Virophage. Their study is inherently multidisciplinary, spanning microbiology, genetics, immunology and evolutionary theory, and the current view is that taking a unified approach has great power and promise. We support this with a new, unified, model of their evolutionary ecology, using contemporary evolutionary theory coupling the Price equation with game theory, studying the consequences of the molecular replicators' promiscuous use of each others' gene products for their natural history and evolutionary ecology. Even at this simple expository level, we can make a firm prediction of a new class of replicators exploiting viruses such as lentiviruses like SIVs, a family which includes HIV: these have been explicitly stated in the primary literature to be non-existent. Closely connected to this departure is the view that multicellular organism immunology is more about the management of chronic infections rather than the elimination of acute ones and new understandings emerging are changing our view of the kind of theatre we ourselves provide for the evolutionary play of molecular replicators. This study adds molecular replicators to bacteria in the emerging field of sociomicrobiology.

Chimeric peptide-mediated siRNA transduction to inhibit HIV-1 infection.

Science.gov (United States)

Bivalkar-Mehla, Shalmali; Mehla, Rajeev; Chauhan, Ashok

2017-04-01

Persistent human immunodeficiency virus 1 (HIV-1) infection provokes immune activation and depletes CD4 +  lymphocytes, leading to acquired immunodeficiency syndrome. Uninterrupted administration of combination antiretroviral therapy (cART) in HIV-infected patients suppresses viral replication to below the detectable level and partially restores the immune system. However, cART-unresponsive residual HIV-1 infection and elusive transcriptionally silent but reactivatable viral reservoirs maintain a permanent viral DNA blue print. The virus rebounds within a few weeks after interruption of suppressive therapy. Adjunct gene therapy to control viral replication by ribonucleic acid interference (RNAi) is a post-transcriptional gene silencing strategy that could suppress residual HIV-1 burden and overcome viral resistance. Small interfering ribonucleic acids (siRNAs) are efficient transcriptional inhibitors, but need delivery systems to reach inside target cells. We investigated the potential of chimeric peptide (FP-PTD) to deliver specific siRNAs to HIV-1-susceptible and permissive cells. Chimeric FP-PTD peptide was designed with an RNA binding domain (PTD) to bind siRNA and a cell fusion peptide domain (FP) to enter cells. FP-PTD-siRNA complex entered and inhibited HIV-1 replication in susceptible cells, and could be a candidate for in vivo testing.

Nuclear retention of multiply spliced HIV-1 RNA in resting CD4+ T cells.

Directory of Open Access Journals (Sweden)

Kara G Lassen

2006-07-01

Full Text Available HIV-1 latency in resting CD4+ T cells represents a major barrier to virus eradication in patients on highly active antiretroviral therapy (HAART. We describe here a novel post-transcriptional block in HIV-1 gene expression in resting CD4+ T cells from patients on HAART. This block involves the aberrant localization of multiply spliced (MS HIV-1 RNAs encoding the critical positive regulators Tat and Rev. Although these RNAs had no previously described export defect, we show that they exhibit strict nuclear localization in resting CD4+ T cells from patients on HAART. Overexpression of the transcriptional activator Tat from non-HIV vectors allowed virus production in these cells. Thus, the nuclear retention of MS HIV-1 RNA interrupts a positive feedback loop and contributes to the non-productive nature of infection of resting CD4+ T cells. To define the mechanism of nuclear retention, proteomic analysis was used to identify proteins that bind MS HIV-1 RNA. Polypyrimidine tract binding protein (PTB was identified as an HIV-1 RNA-binding protein differentially expressed in resting and activated CD4+ T cells. Overexpression of PTB in resting CD4+ T cells from patients on HAART allowed cytoplasmic accumulation of HIV-1 RNAs. PTB overexpression also induced virus production by resting CD4+ T cells. Virus culture experiments showed that overexpression of PTB in resting CD4+ T cells from patients on HAART allowed release of replication-competent virus, while preserving a resting cellular phenotype. Whether through effects on RNA export or another mechanism, the ability of PTB to reverse latency without inducing cellular activation is a result with therapeutic implications.

Improved HIV and TB Knowledge and Competence Among Mid-level Providers in a Cluster-Randomized Trial of One-on-One Mentorship for Task Shifting.

Science.gov (United States)

Naikoba, Sarah; Senjovu, Kaggwa D; Mugabe, Pallen; McCarthy, Carey F; Riley, Patricia L; Kadengye, Damazo T; Dalal, Shona

2017-08-15

Health worker shortages pose a challenge to the scale up of HIV care and treatment in Uganda. Training mid-level providers (MLPs) in the provision of HIV and tuberculosis (TB) treatment can expand existing health workforce capacity and access to HIV services. We conducted a cluster-randomized trial of on-site clinical mentorship for HIV and TB care at 10 health facilities in rural Uganda. Twenty MLPs at 5 randomly assigned to an intervention facilities received 8 hours a week of one-on-one mentorship, every 6 weeks over a 9-month period; and another 20 at 5 control facilities received no clinical mentorship. Enrolled MLPs' clinical knowledge and competence in management of HIV and TB was assessed using case scenarios and clinical observation at baseline and immediately after the 9-month intervention. The performance of the study health facilities on 8 TB and HIV care indicators was tracked over the 9-month period using facility patient records. Thirty-nine out 40 enrolled MLPs had case scenario and clinical observation scores for both the baseline and end of intervention assessments. Mentorship was associated with a mean score increase of 16.7% (95% confidence interval: 9.8 to 23.6, P shifting.

HIV communication capacity strengthening: a critical review.

Science.gov (United States)

Lettenmaier, Cheryl; Kraft, Joan Marie; Raisanen, Keris; Serlemitsos, Elizabeth

2014-08-15

HIV communication is most effective and sustainable when it is designed and implemented locally and tailored to the local context. This requires capacity strengthening at national, subnational, and community levels. Through a review of the published and selected "grey" literature, we examine HIV communication capacity strengthening: definitions, measurements, implementation, and effects. We found limited documentation of HIV communication capacity needs or systematic approaches to address them. Most HIV communication capacity strengthening to date has focused on building individual competencies to design and manage social and behavior change communication programs through training courses, often coupled with networking opportunities for participants, post-training mentoring, and technical assistance. A few of these efforts have been evaluated through pre- and post-training tests and qualitative interviews with participants and have shown potential for improvement in individual skills and knowledge. Health communication capacity assessment tools that measure individual and organizational competencies exist, but they have most often been used to identify capacity building needs, not for evaluating capacity strengthening efforts. A new definition of capacity strengthening, grown out of recent efforts to improve effectiveness of international health and development programs, focuses on improving organizational and societal systems that support performance and individual competencies. We propose a holistic model for HIV communication capacity strengthening and call for rigorous documentation and evaluation to determine and scale-up optimal capacity building interventions for strengthening social and behavior change communication for HIV prevention, care, and treatment in developing countries.

Detection of Replication Competent Lentivirus Using a qPCR Assay for VSV-G

Directory of Open Access Journals (Sweden)

Lindsey M. Skrdlant

2018-03-01

Full Text Available Lentiviral vectors are a common tool used to introduce new and corrected genes into cell therapy products for treatment of human diseases. Although lentiviral vectors are ideal for delivery and stable integration of genes of interest into the host cell genome, they potentially pose risks to human health, such as integration-mediated transformation and generation of a replication competent lentivirus (RCL capable of infecting non-target cells. In consideration of the latter risk, all cell-based products modified by lentiviral vectors and intended for patient use must be tested for RCL prior to treatment of the patient. Current Food and Drug Administration (FDA guidelines recommend use of cell-based assays to this end, which can take up to 6 weeks for results. However, qPCR-based assays are a quick alternative for rapid assessment of RCL in products intended for fresh infusion. We describe here the development and qualification of a qPCR assay based on detection of envelope gene sequences (vesicular stomatitis virus G glycoprotein [VSV-G] for RCL in accordance with Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE guidelines. Our results demonstrate the sensitivity, linearity, specificity, and reproducibility of detection of VSV-G sequences, with a low false-positive rate. These procedures are currently being used in our phase 1 clinical investigations.

CD4-specific designed ankyrin repeat proteins are novel potent HIV entry inhibitors with unique characteristics.

Directory of Open Access Journals (Sweden)

Andreas Schweizer

2008-07-01

Full Text Available Here, we describe the generation of a novel type of HIV entry inhibitor using the recently developed Designed Ankyrin Repeat Protein (DARPin technology. DARPin proteins specific for human CD4 were selected from a DARPin DNA library using ribosome display. Selected pool members interacted specifically with CD4 and competed with gp120 for binding to CD4. DARPin proteins derived in the initial selection series inhibited HIV in a dose-dependent manner, but showed a relatively high variability in their capacity to block replication of patient isolates on primary CD4 T cells. In consequence, a second series of CD4-specific DARPins with improved affinity for CD4 was generated. These 2nd series DARPins potently inhibit infection of genetically divergent (subtype B and C HIV isolates in the low nanomolar range, independent of coreceptor usage. Importantly, the actions of the CD4 binding DARPins were highly specific: no effect on cell viability or activation, CD4 memory cell function, or interference with CD4-independent virus entry was observed. These novel CD4 targeting molecules described here combine the unique characteristics of DARPins-high physical stability, specificity and low production costs-with the capacity to potently block HIV entry, rendering them promising candidates for microbicide development.

Fight fire with fire: Gene therapy strategies to cure HIV.

Science.gov (United States)

Huyghe, Jon; Magdalena, Sips; Vandekerckhove, Linos

2017-08-01

Human Immunodeficiency Virus (HIV) to date remains one of the most notorious viruses mankind has ever faced. Despite enormous investments in HIV research for more than 30 years an effective cure for HIV has been elusive. Areas covered: Combination antiretroviral therapy (cART) suppresses active viral replication, but is not able to eliminate the virus completely due to stable integration of HIV inside the host genome of infected cells and the establishment of a latent reservoir, that is insensitive to cART. Nevertheless, this latent HIV reservoir is fully capable to refuel viral replication when treatment is stopped, creating a major obstacle towards a cure for HIV. Several gene therapy approaches ranging from the generation of HIV resistant CD4 + T cells to the eradication of HIV infected cells by immune cell engineering are currently under pre-clinical and clinical investigation and may present a promising road to a cure. In this review, we focus on the status and the prospects of gene therapy strategies to cure/eradicate HIV. Expert commentary: Recent advances in gene therapy for oncology and infectious diseases indicate that gene therapy may be a feasible and very potent cure strategy, and therefore a potential game changer in the search for an effective HIV cure.

Interplay between HIV Entry and Transportin-SR2 Dependency

Directory of Open Access Journals (Sweden)

Gijsbers Rik

2011-01-01

Full Text Available Abstract Background Transportin-SR2 (TRN-SR2, TNPO3, transportin 3 was previously identified as an interaction partner of human immunodeficiency virus type 1 (HIV-1 integrase and functions as a nuclear import factor of HIV-1. A possible role of capsid in transportin-SR2-mediated nuclear import was recently suggested by the findings that a chimeric HIV virus, carrying the murine leukemia virus (MLV capsid and matrix proteins, displayed a transportin-SR2 independent phenotype, and that the HIV-1 N74D capsid mutant proved insensitive to transportin-SR2 knockdown. Results Our present analysis of viral specificity reveals that TRN-SR2 is not used to the same extent by all lentiviruses. The DNA flap does not determine the TRN-SR2 requirement of HIV-1. We corroborate the TRN-SR2 independent phenotype of the chimeric HIV virus carrying the MLV capsid and matrix proteins. We reanalyzed the HIV-1 N74D capsid mutant in cells transiently or stably depleted of transportin-SR2 and confirm that the N74D capsid mutant is independent of TRN-SR2 when pseudotyped with the vesicular stomatitis virus glycoprotein (VSV-G. Remarkably, although somewhat less dependent on TRN-SR2 than wild type virus, the N74D capsid mutant carrying the wild type HIV-1 envelope required TRN-SR2 for efficient replication. By pseudotyping with envelopes that mediate pH-independent viral uptake including HIV-1, measles virus and amphotropic MLV envelopes, we demonstrate that HIV-1 N74D capsid mutant viruses retain partial dependency on TRN-SR2. However, this dependency on TRN-SR2 is lost when the HIV N74D capsid mutant is pseudotyped with envelopes mediating pH-dependent endocytosis, such as the VSV-G and Ebola virus envelopes. Conclusion Here we discover a link between the viral entry of HIV and its interaction with TRN-SR2. Our data confirm the importance of TRN-SR2 in HIV-1 replication and argue for careful interpretation of experiments performed with VSV-G pseudotyped viruses in

Inhibition of HIV-1 entry by extracts derived from traditional Chinese medicinal herbal plants

Directory of Open Access Journals (Sweden)

Song Xinming

2009-08-01

Full Text Available Abstract Background Highly active anti-retroviral therapy (HAART is the current HIV/AIDS treatment modality. Despite the fact that HAART is very effective in suppressing HIV-1 replication and reducing the mortality of HIV/AIDS patients, it has become increasingly clear that HAART does not offer an ultimate cure to HIV/AIDS. The high cost of the HAART regimen has impeded its delivery to over 90% of the HIV/AIDS population in the world. This reality has urgently called for the need to develop inexpensive alternative anti-HIV/AIDS therapy. This need has further manifested by recent clinical trial failures in anti-HIV-1 vaccines and microbicides. In the current study, we characterized a panel of extracts of traditional Chinese medicinal herbal plants for their activities against HIV-1 replication. Methods Crude and fractionated extracts were prepared from various parts of nine traditional Chinese medicinal herbal plants in Hainan Island, China. These extracts were first screened for their anti-HIV activity and cytotoxicity in human CD4+ Jurkat cells. Then, a single-round pseudotyped HIV-luciferase reporter virus system (HIV-Luc was used to identify potential anti-HIV mechanisms of these extracts. Results Two extracts, one from Euphorbiaceae, Trigonostema xyphophylloides (TXE and one from Dipterocarpaceae, Vatica astrotricha (VAD inhibited HIV-1 replication and syncytia formation in CD4+ Jurkat cells, and had little adverse effects on host cell proliferation and survival. TXE and VAD did not show any direct inhibitory effects on the HIV-1 RT enzymatic activity. Treatment of these two extracts during the infection significantly blocked infection of the reporter virus. However, pre-treatment of the reporter virus with the extracts and treatment of the extracts post-infection had little effects on the infectivity or gene expression of the reporter virus. Conclusion These results demonstrate that TXE and VAD inhibit HIV-1 replication likely by blocking

Patients with discordant responses to antiretroviral therapy have impaired killing of HIV-infected T cells.

Directory of Open Access Journals (Sweden)

Sekar Natesampillai

2010-11-01

Full Text Available In medicine, understanding the pathophysiologic basis of exceptional circumstances has led to an enhanced understanding of biology. We have studied the circumstance of HIV-infected patients in whom antiretroviral therapy results in immunologic benefit, despite virologic failure. In such patients, two protease mutations, I54V and V82A, occur more frequently. Expressing HIV protease containing these mutations resulted in less cell death, caspase activation, and nuclear fragmentation than wild type (WT HIV protease or HIV protease containing other mutations. The impaired induction of cell death was also associated with impaired cleavage of procaspase 8, a requisite event for HIV protease mediated cell death. Primary CD4 T cells expressing I54V or V82A protease underwent less cell death than with WT or other mutant proteases. Human T cells infected with HIV containing these mutations underwent less cell death and less Casp8p41 production than WT or HIV containing other protease mutations, despite similar degrees of viral replication. The reductions in cell death occurred both within infected cells, as well as in uninfected bystander cells. These data indicate that single point mutations within HIV protease which are selected in vivo can significantly impact the ability of HIV to kill CD4 T cells, while not impacting viral replication. Therefore, HIV protease regulates both HIV replication as well as HIV induced T cell depletion, the hallmark of HIV pathogenesis.

Mother-to-Child HIV-1 Transmission Events Are Differentially Impacted by Breast Milk and Its Components from HIV-1-Infected Women.

Directory of Open Access Journals (Sweden)

Ruizhong Shen

Full Text Available Breast milk is a vehicle of infection and source of protection in post-natal mother-to-child HIV-1 transmission (MTCT. Understanding the mechanism by which breast milk limits vertical transmission will provide critical insight into the design of preventive and therapeutic approaches to interrupt HIV-1 mucosal transmission. However, characterization of the inhibitory activity of breast milk in human intestinal mucosa, the portal of entry in postnatal MTCT, has been constrained by the limited availability of primary mucosal target cells and tissues to recapitulate mucosal transmission ex vivo. Here, we characterized the impact of skimmed breast milk, breast milk antibodies (Igs and non-Ig components from HIV-1-infected Ugandan women on the major events of HIV-1 mucosal transmission using primary human intestinal cells and tissues. HIV-1-specific IgG antibodies and non-Ig components in breast milk inhibited the uptake of Ugandan HIV-1 isolates by primary human intestinal epithelial cells, viral replication in and transport of HIV-1- bearing dendritic cells through the human intestinal mucosa. Breast milk HIV-1-specific IgG and IgA, as well as innate factors, blocked the uptake and transport of HIV-1 through intestinal mucosa. Thus, breast milk components have distinct and complementary effects in reducing HIV-1 uptake, transport through and replication in the intestinal mucosa and, therefore, likely contribute to preventing postnatal HIV-1 transmission. Our data suggests that a successful preventive or therapeutic approach would require multiple immune factors acting at multiple steps in the HIV-1 mucosal transmission process.

A new strategy to inhibit the excision reaction catalysed by HIV-1 reverse transcriptase: compounds that compete with the template–primer

Science.gov (United States)

Cruchaga, Carlos; Anso, Elena; Font, María; Martino, Virginia S.; Rouzaut, Ana; Martinez-Irujo, Juan J.

2007-01-01

Inhibitors of the excision reaction catalysed by HIV-1 RT (reverse transcriptase) represent a promising approach in the fight against HIV, because these molecules would interfere with the main mechanism of resistance of this enzyme towards chain-terminating nucleotides. Only a limited number of compounds have been demonstrated to inhibit this reaction to date, including NNRTIs (non-nucleoside RT inhibitors) and certain pyrophosphate analogues. We have found previously that 2GP (2-O-galloylpunicalin), an antiviral compound extracted from the leaves of Terminalia triflora, was able to inhibit both the RT and the RNase H activities of HIV-1 RT without affecting cell proliferation or viability. In the present study, we show that 2GP also inhibited the ATP- and PPi-dependent phosphorolysis catalysed by wild-type and AZT (3′-azido-3′-deoxythymidine)-resistant enzymes at sub-micromolar concentrations. Kinetic and direct-binding analysis showed that 2GP was a non-competitive inhibitor against the nucleotide substrate, whereas it competed with the binding of RT to the template–primer (Kd=85 nM). As expected from its mechanism of action, 2GP was active against mutations conferring resistance to NNRTIs and AZT. The combination of AZT with 2GP was highly synergistic when tested in the presence of pyrophosphate, indicating that the inhibition of RT-catalysed phosphorolysis was responsible for the synergy found. Although other RT inhibitors that compete with the template–primer have been described, this is the first demonstration that these compounds can be used to block the excision of chain terminating nucleotides, providing a rationale for their combination with nucleoside analogues. PMID:17355225

The second chance story of HIV-1 DNA: Unintegrated? Not a problem!

Science.gov (United States)

Wu, Yuntao

2008-07-09

Accumulation of high levels of unintegrated viral DNA is a common feature of retroviral infection. It was recently discovered that coinfection of cells with integrated and unintegrated HIV-1 can result in complementation, allowing viral replication in the absence of integration. This new mode of HIV-1 replication has numerous implications for the function of unintegrated viral DNA and its application as a therapeutic vector.

Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus.

Science.gov (United States)

Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W; Gautier, Virginie W

2012-01-01

The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these

First UK case report of kidney transplantation from an HIV-infected deceased donor to two HIV-infected recipients.

Science.gov (United States)

Nolan, Eileen; Karydis, Nikolaos; Drage, Martin; Hilton, Rachel

2018-04-01

Kidney transplantation is now considered the treatment of choice for many human immunodeficiency virus (HIV)-infected patients with end-stage renal disease (ESRD). Graft survival rates using HIV-negative donors and carefully selected HIV-positive ESRD patients are similar to those observed in HIV-uninfected kidney transplant recipients. To address the relative shortfall in donated organs it has been proposed that organs from HIV-infected deceased donors might be allocated to HIV-infected patients on the transplant waiting list. Preliminary experience in South Africa reports promising short-term outcomes in a small number of HIV-infected recipients of kidney transplants from HIV-infected donors. We sought to replicate this experience in the UK by accepting kidney offers from HIV infected deceased donors for patients with HIV-infection on the kidney transplant waiting list. Here we report the UK's first cases of kidney transplantation between HIV-positive donors and recipients.

Sifuvirtide, a potent HIV fusion inhibitor peptide

International Nuclear Information System (INIS)

Wang, Rui-Rui; Yang, Liu-Meng; Wang, Yun-Hua; Pang, Wei; Tam, Siu-Cheung; Tien, Po; Zheng, Yong-Tang

2009-01-01

Enfuvirtide (ENF) is currently the only FDA approved HIV fusion inhibitor in clinical use. Searching for more drugs in this category with higher efficacy and lower toxicity seems to be a logical next step. In line with this objective, a synthetic peptide with 36 amino acid residues, called Sifuvirtide (SFT), was designed based on the crystal structure of gp41. In this study, we show that SFT is a potent anti-HIV agent with relatively low cytotoxicity. SFT was found to inhibit replication of all tested HIV strains. The effective concentrations that inhibited 50% viral replication (EC 50 ), as determined in all tested strains, were either comparable or lower than benchmark values derived from well-known anti-HIV drugs like ENF or AZT, while the cytotoxic concentrations causing 50% cell death (CC 50 ) were relatively high, rendering it an ideal anti-HIV agent. A GST-pull down assay was performed to confirm that SFT is a fusion inhibitor. Furthermore, the activity of SFT on other targets in the HIV life cycle was also investigated, and all assays showed negative results. To further understand the mechanism of action of HIV peptide inhibitors, resistant variants of HIV-1 IIIB were derived by serial virus passage in the presence of increasing doses of SFT or ENF. The results showed that there was cross-resistance between SFT and ENF. In conclusion, SFT is an ideal anti-HIV agent with high potency and low cytotoxicity, but may exhibit a certain extent of cross-resistance with ENF.

The second chance story of HIV-1 DNA: Unintegrated? Not a problem!

Directory of Open Access Journals (Sweden)

Wu Yuntao

2008-07-01

Full Text Available Abstract Accumulation of high levels of unintegrated viral DNA is a common feature of retroviral infection. It was recently discovered that coinfection of cells with integrated and unintegrated HIV-1 can result in complementation, allowing viral replication in the absence of integration. This new mode of HIV-1 replication has numerous implications for the function of unintegrated viral DNA and its application as a therapeutic vector.

Analysis of Select Herpes Simplex Virus 1 (HSV-1) Proteins for Restriction of Human Immunodeficiency Virus Type 1 (HIV-1): HSV-1 gM Protein Potently Restricts HIV-1 by Preventing Intracellular Transport and Processing of Env gp160.

Science.gov (United States)

Polpitiya Arachchige, Sachith; Henke, Wyatt; Pramanik, Ankita; Kalamvoki, Maria; Stephens, Edward B

2018-01-15

Virus-encoded proteins that impair or shut down specific host cell functions during replication can be used as probes to identify potential proteins/pathways used in the replication of viruses from other families. We screened nine proteins from herpes simplex virus 1 (HSV-1) for the ability to enhance or restrict human immunodeficiency virus type 1 (HIV-1) replication. We show that several HSV-1 proteins (glycoprotein M [gM], US3, and UL24) potently restricted the replication of HIV-1. Unlike UL24 and US3, which reduced viral protein synthesis, we observed that gM restriction of HIV-1 occurred through interference with the processing and transport of gp160, resulting in a significantly reduced level of mature gp120/gp41 released from cells. Finally, we show that an HSV-1 gM mutant lacking the majority of the C-terminal domain (HA-gM[Δ345-473]) restricted neither gp160 processing nor the release of infectious virus. These studies identify proteins from heterologous viruses that can restrict viruses through novel pathways. IMPORTANCE HIV-1 infection of humans results in AIDS, characterized by the loss of CD4 + T cells and increased susceptibility to opportunistic infections. Both HIV-1 and HSV-1 can infect astrocytes and microglia of the central nervous system (CNS). Thus, the identification of HSV-1 proteins that directly restrict HIV-1 or interfere with pathways required for HIV-1 replication could lead to novel antiretroviral strategies. The results of this study show that select viral proteins from HSV-1 can potently restrict HIV-1. Further, our results indicate that the gM protein of HSV-1 restricts HIV-1 through a novel pathway by interfering with the processing of gp160 and its incorporation into virus maturing from the cell. Copyright © 2018 American Society for Microbiology.

Stepping toward a Macaque Model of HIV-1 Induced AIDS

Directory of Open Access Journals (Sweden)

Jason T. Kimata

2014-09-01

Full Text Available HIV-1 exhibits a narrow host range, hindering the development of a robust animal model of pathogenesis. Past studies have demonstrated that the restricted host range of HIV-1 may be largely due to the inability of the virus to antagonize and evade effector molecules of the interferon response in other species. They have also guided the engineering of HIV-1 clones that can replicate in CD4 T-cells of Asian macaque species. However, while replication of these viruses in macaque hosts is persistent, it has been limited and without progression to AIDS. In a new study, Hatziioannou et al., demonstrate for the first time that adapted macaque-tropic HIV-1 can persistently replicate at high levels in pigtailed macaques (Macaca nemestrina, but only if CD8 T-cells are depleted at the time of inoculation. The infection causes rapid disease and recapitulates several aspects of AIDS in humans. Additionally, the virus undergoes genetic changes to further escape innate immunity in association with disease progression. Here, the importance of these findings is discussed, as they relate to pathogenesis and model development.

Simian Immunodeficiency Virus (SIV-Specific Chimeric Antigen Receptor-T Cells Engineered to Target B Cell Follicles and Suppress SIV Replication

Directory of Open Access Journals (Sweden)

Kumudhini Preethi Haran

2018-03-01

Full Text Available There is a need to develop improved methods to treat and potentially cure HIV infection. During chronic HIV infection, replication is concentrated within T follicular helper cells (Tfh located within B cell follicles, where low levels of virus-specific CTL permit ongoing viral replication. We previously showed that elevated levels of simian immunodeficiency virus (SIV-specific CTL in B cell follicles are linked to both decreased levels of viral replication in follicles and decreased plasma viral loads. These findings provide the rationale to develop a strategy for targeting follicular viral-producing (Tfh cells using antiviral chimeric antigen receptor (CAR T cells co-expressing the follicular homing chemokine receptor CXCR5. We hypothesize that antiviral CAR/CXCR5-expressing T cells, when infused into an SIV-infected animal or an HIV-infected individual, will home to B cell follicles, suppress viral replication, and lead to long-term durable remission of SIV and HIV. To begin to test this hypothesis, we engineered gammaretroviral transduction vectors for co-expression of a bispecific anti-SIV CAR and rhesus macaque CXCR5. Viral suppression by CAR/CXCR5-transduced T cells was measured in vitro, and CXCR5-mediated migration was evaluated using both an in vitro transwell migration assay, as well as a novel ex vivo tissue migration assay. The functionality of the CAR/CXCR5 T cells was demonstrated through their potent suppression of SIVmac239 and SIVE660 replication in in vitro and migration to the ligand CXCL13 in vitro, and concentration in B cell follicles in tissues ex vivo. These novel antiviral immunotherapy products have the potential to provide long-term durable remission (functional cure of HIV and SIV infections.

Homologous regions of Fen1 and p21Cip1 compete for binding to the same site on PCNA: a potential mechanism to co-ordinate DNA replication and repair.

Science.gov (United States)

Warbrick, E; Lane, D P; Glover, D M; Cox, L S

1997-05-15

Following genomic damage, the cessation of DNA replication is co-ordinated with onset of DNA repair; this co-ordination is essential to avoid mutation and genomic instability. To investigate these phenomena, we have analysed proteins that interact with PCNA, which is required for both DNA replication and repair. One such protein is p21Cip1, which inhibits DNA replication through its interaction with PCNA, while allowing repair to continue. We have identified an interaction between PCNA and the structure specific nuclease, Fen1, which is involved in DNA replication. Deletion analysis suggests that p21Cip1 and Fen1 bind to the same region of PCNA. Within Fen1 and its homologues a small region (10 amino acids) is sufficient for PCNA binding, which contains an 8 amino acid conserved PCNA-binding motif. This motif shares critical residues with the PCNA-binding region of p21Cip1. A PCNA binding peptide from p21Cip1 competes with Fen1 peptides for binding to PCNA, disrupts the Fen1-PCNA complex in replicating cell extracts, and concomitantly inhibits DNA synthesis. Competition between homologous regions of Fen1 and p21Cip1 for binding to the same site on PCNA may provide a mechanism to co-ordinate the functions of PCNA in DNA replication and repair.

Risk factors for late-stage HIV disease presentation at initial HIV diagnosis in Durban, South Africa.

Directory of Open Access Journals (Sweden)

Paul K Drain

Full Text Available After observing persistently low CD4 counts at initial HIV diagnosis in South Africa, we sought to determine risk factors for late-stage HIV disease presentation among adults.We surveyed adults prior to HIV testing at four outpatient clinics in Durban from August 2010 to November 2011. All HIV-infected adults were offered CD4 testing, and late-stage HIV disease was defined as a CD4 count <100 cells/mm(3. We used multivariate regression models to determine the effects of sex, emotional health, social support, distance from clinic, employment, perceived barriers to receiving healthcare, and foregoing healthcare to use money for food, clothing, or housing ("competing needs to healthcare" on presentation with late-stage HIV disease.Among 3,669 adults screened, 830 were enrolled, newly-diagnosed with HIV and obtained a CD4 result. Among those, 279 (33.6% presented with late-stage HIV disease. In multivariate analyses, participants who lived ≥5 kilometers from the test site [adjusted odds ratio (AOR 2.8, 95% CI 1.7-4.7], reported competing needs to healthcare (AOR 1.7, 95% CI 1.2-2.4, were male (AOR 1.7, 95% CI 1.2-2.3, worked outside the home (AOR 1.5, 95% CI 1.1-2.1, perceived health service delivery barriers (AOR 1.5, 95% CI 1.1-2.1, and/or had poor emotional health (AOR 1.4, 95% CI 1.0-1.9 had higher odds of late-stage HIV disease presentation.Independent risk factors for late-stage HIV disease presentation were from diverse domains, including geographic, economic, demographic, social, and psychosocial. These findings can inform various interventions, such as mobile testing or financial assistance, to reduce the risk of presentation with late-stage HIV disease.

Ex vivo activation of CD4+ T-cells from donors on suppressive ART can lead to sustained production of infectious HIV-1 from a subset of infected cells.

Directory of Open Access Journals (Sweden)

John K Bui

2017-02-01

Full Text Available The fate of HIV-infected cells after reversal of proviral latency is not well characterized. Simonetti, et al. recently showed that CD4+ T-cells containing intact proviruses can clonally expand in vivo and produce low-level infectious viremia. We hypothesized that reversal of HIV latency by activation of CD4+ T-cells can lead to the expansion of a subset of virus-producing cells rather than their elimination. We established an ex vivo cell culture system involving stimulation of CD4+ T-cells from donors on suppressive antiretroviral therapy (ART with PMA/ionomycin (day 1-7, followed by rest (day 7-21, and then repeat stimulation (day 21-28, always in the presence of high concentrations of raltegravir and efavirenz to effectively block new cycles of viral replication. HIV DNA and virion RNA in the supernatant were quantified by qPCR. Single genome sequencing (SGS of p6-PR-RT was performed to genetically characterize proviruses and virion-associated genomic RNA. The replication-competence of the virions produced was determined by the viral outgrowth assay (VOA and SGS of co-culture supernatants from multiple time points. Experiments were performed with purified CD4+ T-cells from five consecutively recruited donors who had been on suppressive ART for > 2 years. In all experiments, HIV RNA levels in supernatant increased following initial stimulation, decreased or remained stable during the rest period, and increased again with repeat stimulation. HIV DNA levels did not show a consistent pattern of change. SGS of proviruses revealed diverse outcomes of infected cell populations, ranging from their apparent elimination to persistence and expansion. Importantly, a subset of infected cells expanded and produced infectious virus continuously after stimulation. These findings underscore the complexity of eliminating reservoirs of HIV-infected cells and highlight the need for new strategies to kill HIV-infected cells before they can proliferate.

Prospects for Foamy Viral Vector Anti-HIV Gene Therapy

Directory of Open Access Journals (Sweden)

Arun K. Nalla

2016-03-01

Full Text Available Stem cell gene therapy approaches for Human Immunodeficiency Virus (HIV infection have been explored in clinical trials and several anti-HIV genes delivered by retroviral vectors were shown to block HIV replication. However, gammaretroviral and lentiviral based retroviral vectors have limitations for delivery of anti-HIV genes into hematopoietic stem cells (HSC. Foamy virus vectors have several advantages including efficient delivery of transgenes into HSC in large animal models, and a potentially safer integration profile. This review focuses on novel anti-HIV transgenes and the potential of foamy virus vectors for HSC gene therapy of HIV.

Polyfunctional analysis of Gag and Nef specific CD8+ T-cell responses in HIV-1 infected Indian individuals.

Science.gov (United States)

Mendiratta, Sanjay; Vajpayee, Madhu; Mojumdar, Kamalika; Chauhan, Neeraj K; Sreenivas, Vishnubhatla

2011-02-01

Polyfunctional CD8+ T-cells have been described as most competent in controlling viral replication. We studied the impact of antigen persistence on the polyfunctional immune responses of CD8+ T-lymphocytes to HIV Gag and Nef peptides and polyclonal stimuli in 40 ART naïve HIV infected individuals and analyzed the alterations in T-cell functionality in early and late stages of infection. Significantly elevated level of global response and polyfunctional profile of CD8+ T-cells were observed to polyclonal stimulation, than HIV specific antigens in chronically infected individuals. However no key differences were observed in CD8+ T-cell functional profile in any of the 15 unique subsets for Gag and Nef specific antigens. The subjects in early stage of infection (defined as a gap of 6 months or less between seroconversion and enrolment and with no apparent clinical symptoms) had a higher degree of response functionality (4+ or 3+ different functions simultaneously) than in the late stage infection (defined as time duration since seroconversion greater than 6 months). The data suggest that persistence of antigen during chronic infection leads to functional impairment of HIV specific responses. Copyright © 2010 Elsevier Ltd. All rights reserved.

Genome-wide association studies on HIV susceptibility, pathogenesis and pharmacogenomics

NARCIS (Netherlands)

van Manen, Daniëlle; van 't Wout, Angélique B.; Schuitemaker, Hanneke

2012-01-01

Susceptibility to HIV-1 and the clinical course after infection show a substantial heterogeneity between individuals. Part of this variability can be attributed to host genetic variation. Initial candidate gene studies have revealed interesting host factors that influence HIV infection, replication

Replication of avian influenza viruses in equine tracheal epithelium but not in horses

OpenAIRE

Chambers, Thomas M.; Balasuriya, Udeni B. R.; Reedy, Stephanie E.; Tiwari, Ashish

2013-01-01

We evaluated a hypothesis that horses are susceptible to avian influenza viruses by in vitro testing, using explanted equine tracheal epithelial cultures, and in vivo testing by aerosol inoculation of ponies. Results showed that several subtypes of avian influenza viruses detectably replicated in vitro. Three viruses with high in vitro replication competence were administered to ponies. None of the three demonstrably replicated or caused disease signs in ponies. While these results do not exh...

Is phototherapy safe for HIV-infected individuals?

Energy Technology Data Exchange (ETDEWEB)

Adams, M.L.; Houpt, K.R.; Cruz, P.D. Jr. [Texas Univ., Dallas, TX (United States). Southwestern Medical Center

1996-08-01

Patients infected with human immunodeficiency virus (HIV) have a high prevalence of UV radiation-responsive skin diseases including psoriasis, pruitus, eosinophillic folliculitis and eczemas. On the other hand, UV has been shown to suppress T cell-mediated immune responses and to induce activation and replication of HIV. These developments have prompted clinicians and investigators to question whether phototherapy is safe for HIV-infected individuals. We have reviewed these issues and hereby provide a summary and critique of relevant laboratory and clinical evidence. (Author).

Proviral HIV-genome-wide and pol-gene specific Zinc Finger Nucleases: Usability for targeted HIV gene therapy

Directory of Open Access Journals (Sweden)

Wayengera Misaki

2011-07-01

Full Text Available Abstract Background Infection with HIV, which culminates in the establishment of a latent proviral reservoir, presents formidable challenges for ultimate cure. Building on the hypothesis that ex-vivo or even in-vivo abolition or disruption of HIV-gene/genome-action by target mutagenesis or excision can irreversibly abrogate HIV's innate fitness to replicate and survive, we previously identified the isoschizomeric bacteria restriction enzymes (REases AcsI and ApoI as potent cleavers of the HIV-pol gene (11 and 9 times in HIV-1 and 2, respectively. However, both enzymes, along with others found to cleave across the entire HIV-1 genome, slice (SX at palindromic sequences that are prevalent within the human genome and thereby pose the risk of host genome toxicity. A long-term goal in the field of R-M enzymatic therapeutics has thus been to generate synthetic restriction endonucleases with longer recognition sites limited in specificity to HIV. We aimed (i to assemble and construct zinc finger arrays and nucleases (ZFN with either proviral-HIV-pol gene or proviral-HIV-1 whole-genome specificity respectively, and (ii to advance a model for pre-clinically testing lentiviral vectors (LV that deliver and transduce either ZFN genotype. Methods and Results First, we computationally generated the consensus sequences of (a 114 dsDNA-binding zinc finger (Zif arrays (ZFAs or ZifHIV-pol and (b two zinc-finger nucleases (ZFNs which, unlike the AcsI and ApoI homeodomains, possess specificity to >18 base-pair sequences uniquely present within the HIV-pol gene (ZifHIV-polFN. Another 15 ZFNs targeting >18 bp sequences within the complete HIV-1 proviral genome were constructed (ZifHIV-1FN. Second, a model for constructing lentiviral vectors (LVs that deliver and transduce a diploid copy of either ZifHIV-polFN or ZifHIV-1FN chimeric genes (termed LV- 2xZifHIV-polFN and LV- 2xZifHIV-1FN, respectively is proposed. Third, two preclinical models for controlled testing of

Proviral HIV-genome-wide and pol-gene specific zinc finger nucleases: usability for targeted HIV gene therapy.

Science.gov (United States)

Wayengera, Misaki

2011-07-22

Infection with HIV, which culminates in the establishment of a latent proviral reservoir, presents formidable challenges for ultimate cure. Building on the hypothesis that ex-vivo or even in-vivo abolition or disruption of HIV-gene/genome-action by target mutagenesis or excision can irreversibly abrogate HIV's innate fitness to replicate and survive, we previously identified the isoschizomeric bacteria restriction enzymes (REases) AcsI and ApoI as potent cleavers of the HIV-pol gene (11 and 9 times in HIV-1 and 2, respectively). However, both enzymes, along with others found to cleave across the entire HIV-1 genome, slice (SX) at palindromic sequences that are prevalent within the human genome and thereby pose the risk of host genome toxicity. A long-term goal in the field of R-M enzymatic therapeutics has thus been to generate synthetic restriction endonucleases with longer recognition sites limited in specificity to HIV. We aimed (i) to assemble and construct zinc finger arrays and nucleases (ZFN) with either proviral-HIV-pol gene or proviral-HIV-1 whole-genome specificity respectively, and (ii) to advance a model for pre-clinically testing lentiviral vectors (LV) that deliver and transduce either ZFN genotype. First, we computationally generated the consensus sequences of (a) 114 dsDNA-binding zinc finger (Zif) arrays (ZFAs or ZifHIV-pol) and (b) two zinc-finger nucleases (ZFNs) which, unlike the AcsI and ApoI homeodomains, possess specificity to >18 base-pair sequences uniquely present within the HIV-pol gene (ZifHIV-polFN). Another 15 ZFNs targeting >18 bp sequences within the complete HIV-1 proviral genome were constructed (ZifHIV-1FN). Second, a model for constructing lentiviral vectors (LVs) that deliver and transduce a diploid copy of either ZifHIV-polFN or ZifHIV-1FN chimeric genes (termed LV- 2xZifHIV-polFN and LV- 2xZifHIV-1FN, respectively) is proposed. Third, two preclinical models for controlled testing of the safety and efficacy of either of these

Accessory genes confer a high replication rate to virulent feline immunodeficiency virus.

Science.gov (United States)

Troyer, Ryan M; Thompson, Jesse; Elder, John H; VandeWoude, Sue

2013-07-01

Feline immunodeficiency virus (FIV) is a lentivirus that causes AIDS in domestic cats, similar to human immunodeficiency virus (HIV)/AIDS in humans. The FIV accessory protein Vif abrogates the inhibition of infection by cat APOBEC3 restriction factors. FIV also encodes a multifunctional OrfA accessory protein that has characteristics similar to HIV Tat, Vpu, Vpr, and Nef. To examine the role of vif and orfA accessory genes in FIV replication and pathogenicity, we generated chimeras between two FIV molecular clones with divergent disease potentials: a highly pathogenic isolate that replicates rapidly in vitro and is associated with significant immunopathology in vivo, FIV-C36 (referred to here as high-virulence FIV [HV-FIV]), and a less-pathogenic strain, FIV-PPR (referred to here as low-virulence FIV [LV-FIV]). Using PCR-driven overlap extension, we produced viruses in which vif, orfA, or both genes from virulent HV-FIV replaced equivalent genes in LV-FIV. The generation of these chimeras is more straightforward in FIV than in primate lentiviruses, since FIV accessory gene open reading frames have very little overlap with other genes. All three chimeric viruses exhibited increased replication kinetics in vitro compared to the replication kinetics of LV-FIV. Chimeras containing HV-Vif or Vif/OrfA had replication rates equivalent to those of the virulent HV-FIV parental virus. Furthermore, small interfering RNA knockdown of feline APOBEC3 genes resulted in equalization of replication rates between LV-FIV and LV-FIV encoding HV-FIV Vif. These findings demonstrate that Vif-APOBEC interactions play a key role in controlling the replication and pathogenicity of this immunodeficiency-inducing virus in its native host species and that accessory genes act as mediators of lentiviral strain-specific virulence.

Elimination of cancer stem cells and reactivation of latent HIV-1 via AMPK activation: Common mechanism of action linking inhibition of tumorigenesis and the potential eradication of HIV-1.

Science.gov (United States)

Finley, Jahahreeh

2017-07-01

Although promising treatments are currently in development to slow disease progression and increase patient survival, cancer remains the second leading cause of death in the United States. Cancer treatment modalities commonly include chemoradiation and therapies that target components of aberrantly activated signaling pathways. However, treatment resistance is a common occurrence and recent evidence indicates that the existence of cancer stem cells (CSCs) may underlie the limited efficacy and inability of current treatments to effectuate a cure. CSCs, which are largely resistant to chemoradiation therapy, are a subpopulation of cancer cells that exhibit characteristics similar to embryonic stem cells (ESCs), including self-renewal, multi-lineage differentiation, and the ability to initiate tumorigenesis. Interestingly, intracellular mechanisms that sustain quiescence and promote self-renewal in adult stem cells (ASCs) and CSCs likely also function to maintain latency of HIV-1 in CD4 + memory T cells. Although antiretroviral therapy is highly effective in controlling HIV-1 replication, the persistence of latent but replication-competent proviruses necessitates the development of compounds that are capable of selectively reactivating the latent virus, a method known as the "shock and kill" approach. Homeostatic proliferation in central CD4 + memory T (T CM ) cells, a memory T cell subset that exhibits limited self-renewal and differentiation and is a primary reservoir for latent HIV-1, has been shown to reinforce and stabilize the latent reservoir in the absence of T cell activation and differentiation. HIV-1 has also been found to establish durable and long-lasting latency in a recently discovered subset of CD4 + T cells known as T memory stem (T SCM ) cells. T SCM cells, compared to T CM cells, exhibit stem cell properties that more closely match those of ESCs and ASCs, including self-renewal and differentiation into all memory T cell subsets. It is our hypothesis

Macrophage Resistance to HIV-1 Infection Is Enhanced by the Neuropeptides VIP and PACAP

Science.gov (United States)

Temerozo, Jairo R.; Joaquim, Rafael; Regis, Eduardo G.; Savino, Wilson; Bou-Habib, Dumith Chequer

2013-01-01

It is well established that host factors can modulate HIV-1 replication in macrophages, critical cells in the pathogenesis of HIV-1 infection due to their ability to continuously produce virus. The neuropeptides VIP and PACAP induce well-characterized effects on macrophages through binding to the G protein-coupled receptors VPAC1, VPAC2 and PAC1, but their influence on HIV-1 production by these cells has not been established. Here, we describe that VIP and PACAP reduce macrophage production of HIV-1, acting in a synergistic or additive manner to decrease viral growth. Using receptor antagonists, we detected that the HIV-1 inhibition promoted by VIP is dependent on its ligation to VPAC1/2, whereas PACAP decreases HIV-1 growth via activation of the VPAC1/2 and PAC1 receptors. Specific agonists of VPAC2 or PAC1 decrease macrophage production of HIV-1, whereas sole activation of VPAC1 enhances viral growth. However, the combination of specific agonists mimicking the receptor preference of the natural neuropeptides reproduces the ability of VIP and PACAP to increase macrophage resistance to HIV-1 replication. VIP and PACAP up-regulated macrophage secretion of the β-chemokines CCL3 and CCL5 and the cytokine IL-10, whose neutralization reversed the neuropeptide-induced inhibition of HIV-1 replication. Our results suggest that VIP and PACAP and the receptors VPAC2 and PAC1 could be used as targets for developing alternative therapeutic strategies for HIV-1 infection. PMID:23818986

Competence feedback improves CBT competence in trainee therapists: A randomized controlled pilot study.

Science.gov (United States)

Weck, Florian; Kaufmann, Yvonne M; Höfling, Volkmar

2017-07-01

The development and improvement of therapeutic competencies are central aims in psychotherapy training; however, little is known about which training interventions are suitable for the improvement of competencies. In the current pilot study, the efficacy of feedback regarding therapeutic competencies was investigated in cognitive behavioural therapy (CBT). Totally 19 trainee therapists and 19 patients were allocated randomly to a competence feedback group (CFG) or control group (CG). Two experienced clinicians and feedback providers who were blind to the treatment conditions independently evaluated therapeutic competencies on the Cognitive Therapy Scale at five treatment times (i.e., at Sessions 1, 5, 9, 13, and 17). Whereas CFG and CG included regular supervision, only therapists in the CFG additionally received written qualitative and quantitative feedback regarding their demonstrated competencies in conducting CBT during treatment. We found a significant Time × Group interaction effect (η² = .09), which indicates a larger competence increase in the CFG in comparison to the CG. Competence feedback was demonstrated to be suitable for the improvement of therapeutic competencies in CBT. These findings may have important implications for psychotherapy training, clinical practice, and psychotherapy research. However, further research is necessary to ensure the replicability and generalizability of the findings.

Similarities and differences in the nucleic acid chaperone activity of HIV-2 and HIV-1 nucleocapsid proteins in vitro.

Science.gov (United States)

Pachulska-Wieczorek, Katarzyna; Stefaniak, Agnieszka K; Purzycka, Katarzyna J

2014-07-03

The nucleocapsid domain of Gag and mature nucleocapsid protein (NC) act as nucleic acid chaperones and facilitate folding of nucleic acids at critical steps of retroviral replication cycle. The basic N-terminus of HIV-1 NC protein was shown most important for the chaperone activity. The HIV-2 NC (NCp8) and HIV-1 NC (NCp7) proteins possess two highly conserved zinc fingers, flanked by basic residues. However, the NCp8 N-terminal domain is significantly shorter and contains less positively charged residues. This study characterizes previously unknown, nucleic acid chaperone activity of the HIV-2 NC protein. We have comparatively investigated the in vitro nucleic acid chaperone properties of the HIV-2 and HIV-1 NC proteins. Using substrates derived from the HIV-1 and HIV-2 genomes, we determined the ability of both proteins to chaperone nucleic acid aggregation, annealing and strand exchange in duplex structures. Both NC proteins displayed comparable, high annealing activity of HIV-1 TAR DNA and its complementary nucleic acid. Interesting differences between the two NC proteins were discovered when longer HIV substrates, particularly those derived from the HIV-2 genome, were used in chaperone assays. In contrast to NCp7, NCp8 weakly facilitates annealing of HIV-2 TAR RNA to its complementary TAR (-) DNA. NCp8 is also unable to efficiently stimulate tRNALys3 annealing to its respective HIV-2 PBS motif. Using truncated NCp8 peptide, we demonstrated that despite the fact that the N-terminus of NCp8 differs from that of NCp7, this domain is essential for NCp8 activity. Our data demonstrate that the HIV-2 NC protein displays reduced nucleic acid chaperone activity compared to that of HIV-1 NC. We found that NCp8 activity is limited by substrate length and stability to a greater degree than that of NCp7. This is especially interesting in light of the fact that the HIV-2 5'UTR is more structured than that of HIV-1. The reduced chaperone activity observed with NCp8 may

Colorectal mucus binds DC-SIGN and inhibits HIV-1 trans-infection of CD4+ T-lymphocytes.

Science.gov (United States)

Stax, Martijn J; Mouser, Emily E I M; van Montfort, Thijs; Sanders, Rogier W; de Vries, Henry J C; Dekker, Henk L; Herrera, Carolina; Speijer, Dave; Pollakis, Georgios; Paxton, William A

2015-01-01

Bodily secretions, including breast milk and semen, contain factors that modulate HIV-1 infection. Since anal intercourse caries one of the highest risks for HIV-1 transmission, our aim was to determine whether colorectal mucus (CM) also contains factors interfering with HIV-1 infection and replication. CM from a number of individuals was collected and tested for the capacity to bind DC-SIGN and inhibit HIV-1 cis- or trans-infection of CD4+ T-lymphocytes. To this end, a DC-SIGN binding ELISA, a gp140 trimer competition ELISA and HIV-1 capture/ transfer assays were utilized. Subsequently we aimed to identify the DC-SIGN binding component through biochemical characterization and mass spectrometry analysis. CM was shown to bind DC-SIGN and competes with HIV-1 gp140 trimer for binding. Pre-incubation of Raji-DC-SIGN cells or immature dendritic cells (iDCs) with CM potently inhibits DC-SIGN mediated trans-infection of CD4+ T-lymphocytes with CCR5 and CXCR4 using HIV-1 strains, while no effect on direct infection is observed. Preliminary biochemical characterization demonstrates that the component seems to be large (>100kDa), heat and proteinase K resistant, binds in a α1-3 mannose independent manner and is highly variant between individuals. Immunoprecipitation using DC-SIGN-Fc coated agarose beads followed by mass spectrometry indicated lactoferrin (fragments) and its receptor (intelectin-1) as candidates. Using ELISA we showed that lactoferrin levels within CM correlate with DC-SIGN binding capacity. In conclusion, CM can bind the C-type lectin DC-SIGN and block HIV-1 trans-infection of both CCR5 and CXCR4 using HIV-1 strains. Furthermore, our data indicate that lactoferrin is a DC-SIGN binding component of CM. These results indicate that CM has the potential to interfere with pathogen transmission and modulate immune responses at the colorectal mucosa.

ORAL MANIFESTATIONS AMONG ROMANIAN HIV PATIENTS

Directory of Open Access Journals (Sweden)

Manuela ARBUNE

2013-03-01

Full Text Available The objective of the study is to evaluate the oral health problems on HIV youth patients from Galati. Materials and method. A cross-sectional study assessed 102 patients with mean age 22. The most frequent oral manifestations on HIV infected youth under ART are erythema marginatum, periodontitis, candidosis and hypertrophia gingivalis. Results and discussion. Dental decay-missing-filled index on HIV patients is high. Viral HIV replication, long time pediatric exposure on HIV, male sex, smoking, and oral inflamation are related to dental poor condition. Exodontic therapy is comon among HIV youth. However, persistence of some associated oral infections is related to individual or behavioral risk factors, but also to some newly found mechanisms, such as disfunctional immune reconstruction. Seeing to antiretroviral treatment, the severity and frequency of oral manifestations decreased among HIV patients on antiretroviral treatment. Conclusions. Developing medical, social and educational programs is imperative for improving the oral health of HIV youth of Galati.

Molecular cloning and anti-HIV-1 activities of APOBEC3s from northern pig-tailed macaques (Macaca leonina

Directory of Open Access Journals (Sweden)

Xiao-Liang ZHANG

2016-07-01

Full Text Available Northern pig-tailed macaques (NPMs, Macaca leonina are susceptible to HIV-1 infection largely due to the loss of HIV-1-restricting factor TRIM5α. However, great impediments still exist in the persistent replication of HIV-1 in vivo, suggesting some viral restriction factors are reserved in this host. The APOBEC3 proteins have demonstrated a capacity to restrict HIV-1 replication, but their inhibitory effects in NPMs remain elusive. In this study, we cloned the NPM A3A-A3H genes, and determined by BLAST searching that their coding sequences (CDSs showed 99% identity to the corresponding counterparts from rhesus and southern pig-tailed macaques. We further analyzed the anti-HIV-1 activities of the A3A-A3H genes, and found that A3G and A3F had the greatest anti-HIV-1 activity compared with that of other members. The results of this study indicate that A3G and A3F might play critical roles in limiting HIV-1 replication in NPMs in vivo. Furthermore, this research provides valuable information for the optimization of monkey models of HIV-1 infection.

Demonstration of a novel HIV-1 restriction phenotype from a human T cell line.

Directory of Open Access Journals (Sweden)

Yanxing Han

2008-07-01

Full Text Available Although retroviruses may invade host cells, a productive infection can be established only after the virus counteracts inhibition from different types of host restriction factors. Fv1, APOBEC3G/F, TRIM5alpha, ZAP, and CD317 inhibit the replication of different retroviruses by interfering with viral uncoating, reverse transcription, nuclear import, RNA stability, and release. In humans, although APOBEC3G/3F and CD317 block HIV-1 replication, their antiviral activities are neutralized by viral proteins Vif and Vpu. So far, no human gene has been found to effectively block wild type HIV-1 replication under natural condition. Thus, identification of such a gene product would be of great medical importance for the development of HIV therapies.In this study, we discovered a new type of host restriction against the wild type HIV-1 from a CD4/CXCR4 double-positive human T cell line. We identified a CEM-derived cell line (CEM.NKR that is highly resistant to productive HIV-1 infection. Viral production was reduced by at least 1000-fold when compared to the other permissive human T cell lines such as H9, A3.01, and CEM-T4. Importantly, this resistance was evident at extremely high multiplicity of infection. Further analyses demonstrated that HIV-1 could finish the first round of replication in CEM.NKR cells, but the released virions were poorly infectious. These virions could enter the target cells, but failed to initiate reverse transcription. Notably, this restriction phenotype was also present in CEM.NKR and 293T heterokaryons.These results clearly indicate that CEM.NKR cells express a HIV inhibitory gene(s. Further characterization of this novel gene product(s will reveal a new antiretroviral mechanism that directly inactivates wild type HIV-1.

Surfactant protein D binds to human immunodeficiency virus (HIV) envelope protein gp120 and inhibits HIV replication

DEFF Research Database (Denmark)

Meschi, Joseph; Crouch, Erika C; Skolnik, Paul

2005-01-01

The envelope protein (gp120) of human immunodeficiency virus (HIV) contains highly conserved mannosylated oligosaccharides. These glycoconjugates contribute to resistance to antibody neutralization, and binding to cell surface lectins on macrophages and dendritic cells. Mannose-binding lectin (MBL......) binds to gp120 and plays a role in defence against the virus. In this study it is demonstrated that surfactant protein D (SP-D) binds to gp120 and inhibits HIV infectivity at significantly lower concentrations than MBL. The binding of SP-D was mediated by its calcium-dependent carbohydrate......-binding activity and was dependent on glycosylation of gp120. Native dodecameric SP-D bound to HIV gp120 more strongly than native trimeric SP-D. Since one common polymorphic form of SP-D is predominantly expressed as trimers and associated with lower blood levels, these individuals may have less effective innate...

Antigenic properties of a transport-competent influenza HA/HIV Env chimeric protein

International Nuclear Information System (INIS)

Ye Ling; Sun Yuliang; Lin Jianguo; Bu Zhigao; Wu Qingyang; Jiang, Shibo; Steinhauer, David A.; Compans, Richard W.; Yang Chinglai

2006-01-01

The transmembrane subunit (gp41) of the HIV Env glycoprotein contains conserved neutralizing epitopes which are not well-exposed in wild-type HIV Env proteins. To enhance the exposure of these epitopes, a chimeric protein, HA/gp41, in which the gp41 of HIV-1 89.6 envelope protein was fused to the C-terminus of the HA1 subunit of the influenza HA protein, was constructed. Characterization of protein expression showed that the HA/gp41 chimeric proteins were expressed on cell surfaces and formed trimeric oligomers, as found in the HIV Env as well as influenza HA proteins. In addition, the HA/gp41 chimeric protein expressed on the cell surface can also be cleaved into 2 subunits by trypsin treatment, similar to the influenza HA. Moreover, the HA/gp41 chimeric protein was found to maintain a pre-fusion conformation. Interestingly, the HA/gp41 chimeric proteins on cell surfaces exhibited increased reactivity to monoclonal antibodies against the HIV Env gp41 subunit compared with the HIV-1 envelope protein, including the two broadly neutralizing monoclonal antibodies 2F5 and 4E10. Immunization of mice with a DNA vaccine expressing the HA/gp41 chimeric protein induced antibodies against the HIV gp41 protein and these antibodies exhibit neutralizing activity against infection by an HIV SF162 pseudovirus. These results demonstrate that the construction of such chimeric proteins can provide enhanced exposure of conserved epitopes in the HIV Env gp41 and may represent a novel vaccine design strategy for inducing broadly neutralizing antibodies against HIV

Viral gene products and replication of the human immunodeficiency type 1 virus.

Science.gov (United States)

Morrow, C D; Park, J; Wakefield, J K

1994-05-01

The acquired immunodeficiency syndrome (AIDS) epidemic represents a modern-day plague that has not only resulted in a tragic loss of people from a wide spectrum of society but has reshaped our viewpoints regarding health care, the treatment of infectious diseases, and social issues regarding sexual behavior. There is little doubt now that the cause of the disease AIDS is a virus known as the human immunodeficiency virus (HIV). The HIV virus is a member of a large family of viruses termed retroviruses, which have as a hallmark the capacity to convert their RNA genome into a DNA form that then undergoes a process of integration into the host cell chromosome, followed by the expression of the viral genome and translation of viral proteins in the infected cell. This review describes the organization of the HIV-1 viral genome, the expression of viral proteins, as well as the functions of the accessory viral proteins in HIV replication. The replication of the viral genome is divided into two phases, the early phase and the late phase. The early phase consists of the interaction of the virus with the cell surface receptor (CD4 molecule in most cases), the uncoating and conversion of the viral RNA genome into a DNA form, and the integration into the host cell chromosome. The late phase consists of the expression of the viral proteins from the integrated viral genome, the translation of viral proteins, and the assembly and release of the virus. Points in the HIV-1 life cycle that are targets for therapeutic intervention are also discussed.

Polycyclic aromatic hydrocarbons and cytochrome P450 in HIV pathogenesis

Science.gov (United States)

Rao, P. S. S.; Kumar, Santosh

2015-01-01

High prevalence of cigarette smoking in HIV patients is associated with increased HIV pathogenesis and disease progression. While the effect of smoking on the occurrence of lung cancer has been studied extensively, the association between smoking and HIV pathogenesis is poorly studied. We have recently shown the possible role of cytochrome P450 (CYP) in smoking/nicotine-mediated viral replication. In this review, we focus on the potential role of CYP pathway in polycyclic aromatic hydrocarbons (PAH), important constituents of cigarette smoke, mediated HIV pathogenesis. More specifically, we will discuss the role of CYP1A1 and CYP1B1, which are the major PAH-activating CYP enzymes. Our results have shown that treatment with cigarette smoke condensate (CSC) increases viral replication in HIV-infected macrophages. CSC contains PAH, which are known to be activated by CYP1A1 and CYP1B1 into procarcinogens/toxic metabolites. The expression of these CYPs is regulated by aryl hydrocarbon receptors (AHR), the cellular target of PAH, and an important player in various diseases including cancer. We propose that PAH/AHR-mediated CYP pathway is a novel target to develop new interventions for HIV positive smokers. PMID:26082767

HIV Persistence in Gut-Associated Lymphoid Tissues: Pharmacological Challenges and Opportunities.

Science.gov (United States)

Thompson, Corbin G; Gay, Cynthia L; Kashuba, Angela D M

2017-06-01

An increasing amount of evidence suggests that HIV replication persists in gut-associated lymphoid tissues (GALT), despite treatment with combination antiretroviral therapy (cART). Residual replication in this compartment may propagate infection at other sites in the body and contribute to sustained immune dysregulation and delayed immune recovery. Therefore, it is important to focus efforts on eliminating residual replication at this site. There are several challenges to accomplishing this goal, including low antiretroviral (ARV) exposure at specific tissue locations within GALT, which might be overcome by using the tools of clinical pharmacology. Here, we summarize the evidence for GALT as a site of residual HIV replication, highlight the consequences of persistent infection in tissues, identify current pharmacologic knowledge of drug exposure in GALT, define the challenges that hinder eradication from this site, and propose several avenues for pharmacologic intervention.

Treatment and prevention of HIV infection with long-acting antiretrovirals.

Science.gov (United States)

Benítez-Gutiérrez, Laura; Soriano, Vicente; Requena, Silvia; Arias, Ana; Barreiro, Pablo; de Mendoza, Carmen

2018-05-01

Current antiretroviral therapy allows to achieve and sustain maximal suppression of HIV replication in most treated patients. As result, the life expectancy of HIV-infected persons has improved dramatically and is nowadays similar to that of the HIV-negative population. However, oral antiretrovirals have to be taken daily and indefinitely to avoid resumption of HIV replication and selection of drug resistance. Unfortunately, drug adherence is often suboptimal and tends to decline over time. Areas covered: New drugs, formulations and delivery systems are being developed for extended-release of antiretrovirals. At this time, intramuscular cabotegravir and rilpivirine, dapivirine vaginal rings and tenofovir alafenamide subdermal implants are the products in more advanced stages of clinical development. Their pharmacokinetics/dynamics and safety/efficacy are reviewed. Expert commentary: In the absence of eradicative therapy for individuals with HIV infection and protective vaccines for persons at risk, long-term antiretroviral therapy is the best approach for preventing disease progression in patients and halting transmissions, either as result of 'treatment as prevention' for HIV carriers or 'pre-exposure prophylaxis' for uninfected individuals at risk. In all these scenarios, the advent of long-acting antiretrovirals will expand options for overcoming the challenge of suboptimal drug adherence and reduce the burden of HIV infection.

Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

Science.gov (United States)

Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

2015-01-01

Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-κB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-κB at 276th serine residue. These modifications enhance the interaction of NF-κB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. PMID:25980739

Potential use of rapamycin in HIV infection

DEFF Research Database (Denmark)

Donia, Marco; McCubrey, James A; Bendtzen, Klaus

2010-01-01

The strong need for the development of alternative anti-HIV agents is primarily due to the emergence of strain-resistant viruses, the need for sustained adherence to complex treatment regimens and the toxicity of currently used antiviral drugs. This review analyzes proof of concept studies...... indicating that the immunomodulatory drug rapamycin (RAPA) possesses anti-HIV properties both in vitro and in vivo that qualifies it as a potential new anti-HIV drug. It represents a literature review of published studies that evaluated the in vitro and in vivo activity of RAPA in HIV. RAPA represses HIV-1...... replication in vitro through different mechanisms including, but not limited, to down regulation of CCR5. In addition RAPA synergistically enhances the anti-HIV activity of entry inhibitors such as vicriviroc, aplaviroc and enfuvirtide in vitro. RAPA also inhibits HIV-1 infection in human peripheral blood...

Complete replication-competent adenovirus 11p vectors with E1 or E3 insertions show improved heat stability

Energy Technology Data Exchange (ETDEWEB)

Mei, Ya-Fang, E-mail: ya-fang.mei@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden); Wu, Haidong, E-mail: haidong.wu@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden); Hultenby, Kjell, E-mail: kjell.hultenby@ki.se [Division of Clinical Research Centre, Department of Laboratory Medicine, Karolinska Institute, SE-14186 Stockholm (Sweden); Silver, Jim, E-mail: jim.silver@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden)

2016-10-15

Conventional adenovirus vectors harboring E1 or E3 deletions followed by the insertion of an exogenous gene show considerably reduced virion stability. Here, we report strategies to generate complete replication-competent Ad11p(RCAd11p) vectors that overcome the above disadvantage. A GFP cassette was successfully introduced either upstream of E1A or in the E3A region. The resulting vectors showed high expression levels of the hexon and E1genes and also strongly induced the cytopathic effect in targeted cells. When harboring oversized genomes, the RCAd11pE1 and RCAd11pE3 vectors showed significantly improved heat stability in comparison to Ad11pwt;of the three, RCAd11pE3 was the most tolerant to heat treatment. Electron microscopy showed that RCAd11pE3, RCAd11pE1, Ad11pwt, and Ad11pE1 Delmanifested dominant, moderate, minimum, or no full virus particles after heat treatment at 47 °C for 5 h. Our results demonstrated that both genome size and the insertion site in the viral genome affect virion stability. -- Highlights: •Replicating adenovirus 11p GFP vectors at the E1 or E3 region were generated. •RCAd11pE3 and RCAd11pE1 vectors manifested significantly improved heat stability. •RCAd11pE3 and RCAd11pE1 showed more full viral particles than Ad11pwt after heating. •We demonstrated that both genome size and the insertion site affect virion stability.

Complete replication-competent adenovirus 11p vectors with E1 or E3 insertions show improved heat stability

International Nuclear Information System (INIS)

Mei, Ya-Fang; Wu, Haidong; Hultenby, Kjell; Silver, Jim

2016-01-01

Conventional adenovirus vectors harboring E1 or E3 deletions followed by the insertion of an exogenous gene show considerably reduced virion stability. Here, we report strategies to generate complete replication-competent Ad11p(RCAd11p) vectors that overcome the above disadvantage. A GFP cassette was successfully introduced either upstream of E1A or in the E3A region. The resulting vectors showed high expression levels of the hexon and E1genes and also strongly induced the cytopathic effect in targeted cells. When harboring oversized genomes, the RCAd11pE1 and RCAd11pE3 vectors showed significantly improved heat stability in comparison to Ad11pwt;of the three, RCAd11pE3 was the most tolerant to heat treatment. Electron microscopy showed that RCAd11pE3, RCAd11pE1, Ad11pwt, and Ad11pE1 Delmanifested dominant, moderate, minimum, or no full virus particles after heat treatment at 47 °C for 5 h. Our results demonstrated that both genome size and the insertion site in the viral genome affect virion stability. -- Highlights: •Replicating adenovirus 11p GFP vectors at the E1 or E3 region were generated. •RCAd11pE3 and RCAd11pE1 vectors manifested significantly improved heat stability. •RCAd11pE3 and RCAd11pE1 showed more full viral particles than Ad11pwt after heating. •We demonstrated that both genome size and the insertion site affect virion stability.

Membrane-bound SIV envelope trimers are immunogenic in ferrets after intranasal vaccination with a replication-competent canine distemper virus vector.

Science.gov (United States)

Zhang, Xinsheng; Wallace, Olivia; Wright, Kevin J; Backer, Martin; Coleman, John W; Koehnke, Rebecca; Frenk, Esther; Domi, Arban; Chiuchiolo, Maria J; DeStefano, Joanne; Narpala, Sandeep; Powell, Rebecca; Morrow, Gavin; Boggiano, Cesar; Zamb, Timothy J; Richter King, C; Parks, Christopher L

2013-11-01

We are investigating canine distemper virus (CDV) as a vaccine vector for the delivery of HIV envelope (Env) that closely resembles the native trimeric spike. We selected CDV because it will promote vaccine delivery to lymphoid tissues, and because human exposure is infrequent, reducing potential effects of pre-existing immunity. Using SIV Env as a model, we tested a number of vector and gene insert designs. Vectors containing a gene inserted between the CDV H and L genes, which encoded Env lacking most of its cytoplasmic tail, propagated efficiently in Vero cells, expressed the immunogen on the cell surface, and incorporated the SIV glycoprotein into progeny virus particles. When ferrets were vaccinated intranasally, there were no signs of distress, vector replication was observed in the gut-associated lymphoid tissues, and the animals produced anti-SIV Env antibodies. These data show that live CDV-SIV Env vectors can safely induce anti-Env immune responses following intranasal vaccination. © 2013 Elsevier Inc. All rights reserved.

A novel self-replicating chimeric lentivirus-like particle.

Science.gov (United States)

Jurgens, Christy K; Young, Kelly R; Madden, Victoria J; Johnson, Philip R; Johnston, Robert E

2012-01-01

Successful live attenuated vaccines mimic natural exposure to pathogens without causing disease and have been successful against several viruses. However, safety concerns prevent the development of attenuated human immunodeficiency virus (HIV) as a vaccine candidate. If a safe, replicating virus vaccine could be developed, it might have the potential to offer significant protection against HIV infection and disease. Described here is the development of a novel self-replicating chimeric virus vaccine candidate that is designed to provide natural exposure to a lentivirus-like particle and to incorporate the properties of a live attenuated virus vaccine without the inherent safety issues associated with attenuated lentiviruses. The genome from the alphavirus Venezuelan equine encephalitis virus (VEE) was modified to express SHIV89.6P genes encoding the structural proteins Gag and Env. Expression of Gag and Env from VEE RNA in primate cells led to the assembly of particles that morphologically and functionally resembled lentivirus virions and that incorporated alphavirus RNA. Infection of CD4⁺ cells with chimeric lentivirus-like particles was specific and productive, resulting in RNA replication, expression of Gag and Env, and generation of progeny chimeric particles. Further genome modifications designed to enhance encapsidation of the chimeric virus genome and to express an attenuated simian immunodeficiency virus (SIV) protease for particle maturation improved the ability of chimeric lentivirus-like particles to propagate in cell culture. This study provides proof of concept for the feasibility of creating chimeric virus genomes that express lentivirus structural proteins and assemble into infectious particles for presentation of lentivirus immunogens in their native and functional conformation.

NFAT5 regulates HIV-1 in primary monocytes via a highly conserved long terminal repeat site.

Directory of Open Access Journals (Sweden)

Shahin Ranjbar

2006-12-01

Full Text Available To replicate, HIV-1 capitalizes on endogenous cellular activation pathways resulting in recruitment of key host transcription factors to its viral enhancer. RNA interference has been a powerful tool for blocking key checkpoints in HIV-1 entry into cells. Here we apply RNA interference to HIV-1 transcription in primary macrophages, a major reservoir of the virus, and specifically target the transcription factor NFAT5 (nuclear factor of activated T cells 5, which is the most evolutionarily divergent NFAT protein. By molecularly cloning and sequencing isolates from multiple viral subtypes, and performing DNase I footprinting, electrophoretic mobility shift, and promoter mutagenesis transfection assays, we demonstrate that NFAT5 functionally interacts with a specific enhancer binding site conserved in HIV-1, HIV-2, and multiple simian immunodeficiency viruses. Using small interfering RNA to ablate expression of endogenous NFAT5 protein, we show that the replication of three major HIV-1 viral subtypes (B, C, and E is dependent upon NFAT5 in human primary differentiated macrophages. Our results define a novel host factor-viral enhancer interaction that reveals a new regulatory role for NFAT5 and defines a functional DNA motif conserved across HIV-1 subtypes and representative simian immunodeficiency viruses. Inhibition of the NFAT5-LTR interaction may thus present a novel therapeutic target to suppress HIV-1 replication and progression of AIDS.

Evaluation of a multiple-cycle, recombinant virus, growth competition assay that uses flow cytometry to measure replication efficiency of human immunodeficiency virus type 1 in cell culture.

Science.gov (United States)

Dykes, Carrie; Wang, Jiong; Jin, Xia; Planelles, Vicente; An, Dong Sung; Tallo, Amanda; Huang, Yangxin; Wu, Hulin; Demeter, Lisa M

2006-06-01

Human immunodeficiency virus type 1 (HIV-1) replication efficiency or fitness, as measured in cell culture, has been postulated to correlate with clinical outcome of HIV infection, although this is still controversial. One limitation is the lack of high-throughput assays that can measure replication efficiency over multiple rounds of replication. We have developed a multiple-cycle growth competition assay to measure HIV-1 replication efficiency that uses flow cytometry to determine the relative proportions of test and reference viruses, each of which expresses a different reporter gene in place of nef. The reporter genes are expressed on the surface of infected cells and are detected by commercially available fluorescence-labeled antibodies. This method is less labor-intensive than those that require isolation and amplification of nucleic acids. The two reporter gene products are detected with similar specificity and sensitivity, and the proportion of infected cells in culture correlates with the amount of viral p24 antigen produced in the culture supernatant. HIV replication efficiencies of six different drug-resistant site-directed mutants were reproducibly quantified and were similar to those obtained with a growth competition assay in which the relative proportion of each variant was measured by sequence analysis, indicating that recombination between the pol and reporter genes was negligible. This assay also reproducibly quantified the relative fitness conferred by protease and reverse transcriptase sequences containing multiple drug resistance mutations, amplified from patient plasma. This flow cytometry-based growth competition assay offers advantages over current assays for HIV replication efficiency and should prove useful for the evaluation of patient samples in clinical trials.

Competing or co-existing? Representations of HIV/AIDS by white ...

African Journals Online (AJOL)

This study explores the social representations of HIV and AIDS that circulate among white women teachers in South Africa, a group whose personal risk of the disease is low but who have a major role to play in shaping attitudes to HIV/AIDS among children and young people. The study examines how white women talk ...

Enhanced replication of herpes simplex virus type 1 in human cells

International Nuclear Information System (INIS)

Miller, C.S.; Smith, K.O.

1991-01-01

The effects of DNA-damaging agents on the replication of herpes simplex virus type 1 (HSV-1) were assessed in vitro. Monolayers of human lung fibroblast cell lines were exposed to DNA-damaging agents (methyl methanesulfonate [MMS], methyl methanethiosulfonate [MMTS], ultraviolet light [UV], or gamma radiation [GR]) at specific intervals, before or after inoculation with low levels of HSV-1. The ability of cell monolayers to support HSV-1 replication was measured by direct plaque assay and was compared with that of untreated control samples. In this system, monolayers of different cell lines infected with identical HSV-1 strains demonstrated dissimilar levels of recovery of the infectious virus. Exposure of DNA-repair-competent cell cultures to DNA-damaging agents produced time-dependent enhanced virus replication. Treatment with agent before virus inoculation significantly (p less than 0.025) increased the number of plaques by 10 to 68%, compared with untreated control cultures, while treatment with agent after virus adsorption significantly increased (p less than 0.025) the number of plaques by 7 to 15%. In a parallel series of experiments, cells deficient in DNA repair (xeroderma pigmentosum) failed to support enhanced virus replication. These results suggest that after exposure to DNA-damaging agents, fibroblasts competent in DNA repair amplify the replication of HSV-1, and that DNA-repair mechanisms that act on a variety of chromosomal lesions may be involved in the repair and biological activation of HSV-1 genomes

A chimeric measles virus with a lentiviral envelope replicates exclusively in CD4+/CCR5+ cells

International Nuclear Information System (INIS)

Mourez, Thomas; Mesel-Lemoine, Mariana; Combredet, Chantal; Najburg, Valerie; Cayet, Nadege; Tangy, Frederic

2011-01-01

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 chimeric 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.

Immune defence against HIV-1 infection in HIV-1-exposed seronegative persons.

Science.gov (United States)

Schmechel, S C; Russell, N; Hladik, F; Lang, J; Wilson, A; Ha, R; Desbien, A; McElrath, M J

2001-11-01

Rare individuals who are repeatedly exposed to HIV-1 through unprotected sexual contact fail to acquire HIV-1 infection. These persons represent a unique study population to evaluate mechanisms by which HIV-1 replication is either prevented or controlled. We followed longitudinally a group of healthy HIV-1 seronegative persons each reporting repeated high-risk sexual activities with their HIV-1-infected partner at enrollment. The volunteers were primarily (90%) male homosexuals, maintaining high risk activities with their known infected partner (45%) or multiple other partners (61%). We evaluated the quantity and specificity of HIV-1-specific T cells in 31 exposed seronegatives (ES) using a IFN-gamma ELISPOT assay to enumerate T cells recognizing epitopes within HIV-1 Env, Gag, Pol and Nef. PBMC from only three of the 31 volunteers demonstrated ex vivo HIV-1-specific IFN-gamma secretion, in contrast to nearly 30% exhibiting cytolytic responses in previous studies. These findings suggest that if T cell responses in ES are induced by HIV-1 exposure, the frequency is at low levels in most of them, and below the level of detection using the ELISPOT assay. Alternative approaches to improve the sensitivity of detection may include use of dendritic cells as antigen-presenting cells in the ex vivo assay and more careful definition of the risk behavior and extent of HIV-1 exposure in conjunction with the evaluation of T cell responses.

HIV Genetic Diversity and Drug Resistance

Science.gov (United States)

Santos, André F.; Soares, Marcelo A.

2010-01-01

Most of the current knowledge on antiretroviral (ARV) drug development and resistance is based on the study of subtype B of HIV-1, which only accounts for 10% of the worldwide HIV infections. Cumulative evidence has emerged that different HIV types, groups and subtypes harbor distinct biological properties, including the response and susceptibility to ARV. Recent laboratory and clinical data highlighting such disparities are summarized in this review. Variations in drug susceptibility, in the emergence and selection of specific drug resistance mutations, in viral replicative capacity and in the dynamics of resistance acquisition under ARV selective pressure are discussed. Clinical responses to ARV therapy and associated confounding factors are also analyzed in the context of infections by distinct HIV genetic variants. PMID:21994646

Weak anti-HIV CD8+ T-cell effector activity in HIV primary infection

Science.gov (United States)

Dalod, Marc; Dupuis, Marion; Deschemin, Jean-Christophe; Goujard, Cécile; Deveau, Christiane; Meyer, Laurence; Ngo, Nicole; Rouzioux, Christine; Guillet, Jean-Gérard; Delfraissy, Jean-François; Sinet, Martine; Venet, Alain

1999-01-01

HIV-specific CD8+ T cells play a major role in the control of virus during HIV primary infection (PI) but do not completely prevent viral replication. We used IFN-γ enzyme-linked immunospot assay and intracellular staining to characterize the ex vivo CD8+ T-cell responses to a large variety of HIV epitopic peptides in 24 subjects with early HIV PI. We observed HIV-specific responses in 71% of subjects. Gag and Nef peptides were more frequently recognized than Env and Pol peptides. The number of peptides recognized was low (median 2, range 0–6). In contrast, a much broader response was observed in 30 asymptomatic subjects with chronic infection: all were responders with a median of 5 peptides recognized (range 1–13). The frequency of HIV-specific CD8+ T cells among PBMC for a given peptide was of the same order of magnitude in both groups. The proportion of HIV-specific CD8+CD28– terminally differentiated T cells was much lower in PI than at the chronic stage of infection. The weakness of the immune response during HIV PI could partially account for the failure to control HIV. These findings have potential importance for defining immunotherapeutic strategies and establishing the goals for effective vaccination. J. Clin. Invest. 104:1431–1439 (1999). PMID:10562305

TREX1 Knockdown Induces an Interferon Response to HIV that Delays Viral Infection in Humanized Mice

Directory of Open Access Journals (Sweden)

Lee Adam Wheeler

2016-05-01

Full Text Available Despite their antiviral effect, the in vivo effect of interferons on HIV transmission is difficult to predict, because interferons also activate and recruit HIV-susceptible cells to sites of infection. HIV does not normally induce type I interferons in infected cells, but does if TREX1 is knocked down. Here, we investigated the effect of topical TREX1 knockdown and local interferon production on HIV transmission in human cervicovaginal explants and humanized mice. In explants in which TREX1 was knocked down, HIV induced interferons, which blocked infection. In humanized mice, even though TREX1 knockdown increased infiltrating immune cells, it delayed viral replication for 3–4 weeks. Similarly intravaginal application of type I interferons the day before HIV infection induced interferon responsive genes, reduced inflammation, and decreased viral replication. However, intravenous interferon enhanced inflammation and infection. Thus, in models of human sexual transmission, a localized interferon response inhibits HIV transmission but systemic interferons do not.

Potent nonnucleoside reverse transcriptase inhibitors target HIV-1 Gag-Pol.

Directory of Open Access Journals (Sweden)

Anna Figueiredo

2006-11-01

Full Text Available Nonnucleoside reverse transcriptase inhibitors (NNRTIs target HIV-1 reverse transcriptase (RT by binding to a pocket in RT that is close to, but distinct, from the DNA polymerase active site and prevent the synthesis of viral cDNA. NNRTIs, in particular, those that are potent inhibitors of RT polymerase activity, can also act as chemical enhancers of the enzyme's inter-subunit interactions. However, the consequences of this chemical enhancement effect on HIV-1 replication are not understood. Here, we show that the potent NNRTIs efavirenz, TMC120, and TMC125, but not nevirapine or delavirdine, inhibit the late stages of HIV-1 replication. These potent NNRTIs enhanced the intracellular processing of Gag and Gag-Pol polyproteins, and this was associated with a decrease in viral particle production from HIV-1-transfected cells. The increased polyprotein processing is consistent with premature activation of the HIV-1 protease by NNRTI-enhanced Gag-Pol multimerization through the embedded RT sequence. These findings support the view that Gag-Pol multimerization is an important step in viral assembly and demonstrate that regulation of Gag-Pol/Gag-Pol interactions is a novel target for small molecule inhibitors of HIV-1 production. Furthermore, these drugs can serve as useful probes to further understand processes involved in HIV-1 particle assembly and maturation.

HIV-1 and the macrophage

NARCIS (Netherlands)

Bol, Sebastiaan M.; Cobos-Jimenez, Viviana; Kootstra, Neeltje A.; van 't Wout, Angelique B.

2011-01-01

Macrophages and CD4(+) T cells are natural target cells for HIV-1, and both cell types contribute to the establishment of the viral reservoir that is responsible for continuous residual virus replication during antiretroviral therapy and viral load rebound upon treatment interruption. Scientific

Modeling HIV-associated neurocognitive disorders in mice: new approaches in the changing face of HIV neuropathogenesis.

Science.gov (United States)

Jaeger, Laura B; Nath, Avindra

2012-05-01

It is well established that infection with the human immunodeficiency virus (HIV) leads to immune suppression. Less well known is the fact that long-term, progressive HIV disease is associated with the development of cognitive deficits. Since the introduction of combined antiretroviral therapy (cART), the clinical presentation of HIV infection has evolved into a chronic illness with very low levels of viral replication and chronic immune activation, with compliant affected individuals surviving for decades with a high quality of life. Despite these advances, many HIV-infected individuals develop some degree of neurodegeneration and cognitive impairment. The underlying pathophysiological mechanisms are not well understood, and there are no effective treatments. Thus, there is an unmet need for animal models that enable the study of HIV-associated neurocognitive disorders (HAND) and the testing of new therapeutic approaches to combat them. Here, we review the pros and cons of existing mouse models of HIV infection for addressing these aims and propose a detailed strategy for developing a new mouse model of HIV infection.

Modeling HIV-associated neurocognitive disorders in mice: new approaches in the changing face of HIV neuropathogenesis

Directory of Open Access Journals (Sweden)

Laura B. Jaeger

2012-05-01

Full Text Available It is well established that infection with the human immunodeficiency virus (HIV leads to immune suppression. Less well known is the fact that long-term, progressive HIV disease is associated with the development of cognitive deficits. Since the introduction of combined antiretroviral therapy (cART, the clinical presentation of HIV infection has evolved into a chronic illness with very low levels of viral replication and chronic immune activation, with compliant affected individuals surviving for decades with a high quality of life. Despite these advances, many HIV-infected individuals develop some degree of neurodegeneration and cognitive impairment. The underlying pathophysiological mechanisms are not well understood, and there are no effective treatments. Thus, there is an unmet need for animal models that enable the study of HIV-associated neurocognitive disorders (HAND and the testing of new therapeutic approaches to combat them. Here, we review the pros and cons of existing mouse models of HIV infection for addressing these aims and propose a detailed strategy for developing a new mouse model of HIV infection.

Contrasting roles for TLR ligands in HIV-1 pathogenesis.

Directory of Open Access Journals (Sweden)

Beda Brichacek

2010-09-01

Full Text Available The first line of a host's response to various pathogens is triggered by their engagement of cellular pattern recognition receptors (PRRs. Binding of microbial ligands to these receptors leads to the induction of a variety of cellular factors that alter intracellular and extracellular environment and interfere directly or indirectly with the life cycle of the triggering pathogen. Such changes may also affect any coinfecting microbe. Using ligands to Toll-like receptors (TLRs 5 and 9, we examined their effect on human immunodeficiency virus (HIV-1 replication in lymphoid tissue ex vivo. We found marked differences in the outcomes of such treatment. While flagellin (TLR5 agonist treatment enhanced replication of CC chemokine receptor 5 (CCR 5-tropic and CXC chemokine receptor 4 (CXCR4-tropic HIV-1, treatment with oligodeoxynucleotide (ODN M362 (TLR9 agonist suppressed both viral variants. The differential effects of these TLR ligands on HIV-1 replication correlated with changes in production of CC chemokines CCL3, CCL4, CCL5, and of CXC chemokines CXCL10, and CXCL12 in the ligand-treated HIV-1-infected tissues. The nature and/or magnitude of these changes were dependent on the ligand as well as on the HIV-1 viral strain. Moreover, the tested ligands differed in their ability to induce cellular activation as evaluated by the expression of the cluster of differentiation markers (CD 25, CD38, CD39, CD69, CD154, and human leukocyte antigen D related (HLA-DR as well as of a cell proliferation marker, Ki67, and of CCR5. No significant effect of the ligand treatment was observed on apoptosis and cell death/loss in the treated lymphoid tissue ex vivo. Our results suggest that binding of microbial ligands to TLRs is one of the mechanisms that mediate interactions between coinfected microbes and HIV-1 in human tissues. Thus, the engagement of appropriate TLRs by microbial molecules or their mimetic might become a new strategy for HIV therapy or prevention.

Interleukin-27 is differentially associated with HIV viral load and CD4+ T cell counts in therapy-naive HIV-mono-infected and HIV/HCV-co-infected Chinese.

Directory of Open Access Journals (Sweden)

Lai He

Full Text Available Human Immunodeficiency Virus (HIV infection and the resultant Acquired Immunodeficiency Syndrome (AIDS epidemic are major global health challenges; hepatitis C virus (HCV co-infection has made the HIV/AIDS epidemic even worse. Interleukin-27 (IL-27, a cytokine which inhibits HIV and HCV replication in vitro, associates with HIV infection and HIV/HCV co-infection in clinical settings. However, the impact of HIV and HCV viral loads on plasma IL-27 expression levels has not been well characterized. In this study, 155 antiretroviral therapy-naïve Chinese were recruited. Among them 80 were HIV- and HCV-negative healthy controls, 45 were HIV-mono-infected and 30 were HIV/HCV-co-infected. Plasma level HIV, HCV, IL-27 and CD4+ number were counted and their correlation, regression relationships were explored. We show that: plasma IL-27 level was significantly upregulated in HIV-mono-infected and HIV/HCV-co-infected Chinese; HIV viral load was negatively correlated with IL-27 titer in HIV-mono-infected subjects whereas the relationship was opposite in HIV/HCV-co-infected subjects; and the relationships between HIV viral loads, IL-27 titers and CD4+ T cell counts in the HIV mono-infection and HIV/HCV co-infection groups were dramatically different. Overall, our results suggest that IL-27 differs in treatment-naïve groups with HIV mono-infections and HIV/HCV co-infections, thereby providing critical information to be considered when caring and treating those with HIV mono-infection and HIV/HCV co-infection.

Non-Replicating Adenovirus-Vectored Anthrax Vaccine

International Nuclear Information System (INIS)

Van Kampen, K. R.; Zhang, J.; Jex, E.; Tang, D. C.

2007-01-01

As bioterrorism is emerging as a national threat, it is urgent to develop a new generation of anthrax vaccines that can be rapidly produced and mass administered in an emergency setting. We have demonstrated that protective immunity against anthrax spores could be elicited in mice by intranasal administration of a non-replicating human adenovirus serotype 5 (Ad5)-derived vector encoding Bacillus anthracis protective antigen (PA) in a single-dose regimen. The potency of an Ad5 vector encoding PA was remarkably enhanced by codon optimization of the PA gene to match the tRNA pool found in human cells. This nasal vaccine can be mass-administered by non-medical personnel during a bioterrorist attack. In addition, replication-competent adenovirus (RCA)-free Ad5-vectored anthrax vaccines can be mass produced in PER.C6 cells in serum-free wave bioreactors and purified by column chromatography to meet a surge in demand. The non-replicating nature of this new generation of anthrax vaccine ensures an excellent safety profile for vaccines and the environment.(author)

Microbiological profile of chronic suppurative otits media among HIV ...

African Journals Online (AJOL)

Background: Immune compromised HIV/AIDS infected children have consistently shown a higher prevalence of chronic suppurative otitis media than their immune competent counterparts.This study aimed to compare the microbial isolates from ear discharges in HIV infected and non infected children. Method: This was a ...

The Puzzle of HIV Neutral and Selective Evolution.

Science.gov (United States)

Leitner, Thomas

2018-06-01

HIV is one of the fastest evolving organisms known. It evolves about 1 million times faster than its host, humans. Because HIV establishes chronic infections, with continuous evolution, its divergence within a single infected human surpasses the divergence of the entire humanoid history. Yet, it is still the same virus, infecting the same cell types and using the same replication machinery year after year. Hence, one would think that most mutations that HIV accumulates are neutral. But the picture is more complicated than that. HIV evolution is also a clear example of strong positive selection, that is, mutants have a survival advantage. How do these facts come together?

Epidemiological composition, clinical and treatment characteristics of the patient cohort of the german competence network for HIV/AIDS

Directory of Open Access Journals (Sweden)

Jansen K

2009-09-01

Full Text Available Abstract Objective As its central basis for research, the Competence Network for HIV/AIDS (KompNet established a nationwide cohort study on HIV-positive patients being in medical care in Germany. In this paper, we describe the epidemiological composition, and clinical as well as treatment characteristics of the KompNet cohort over time. Methods The KompNet cohort is an open, retrospective and prospective, multi-center, disease-specific and nationwide cohort study that started gathering data in June 2004. Semiannually, follow up visits of the patients are documented, covering a wide range of clinical and sociodemographic data. At enrolment and three years afterwards, an EDTA-sample is taken; a serum-sample is taken at every follow up. Results As of 20.10.2008, a total of 15,541 patients were enrolled by 44 documenting sites. In September 2007, the cohort size was reduced to ten outpatient clinics and fifteen private practitioners, covering a total of 9,410 patients. The documentation of these patients comprised 24,117 years of follow up-time since enrolment (mean: 2.6 years, 62,862 person years inclusive data documented retrospectively on course of HIV-infection and antiretroviral therapy (ART, mean: 6.7 years. Due to the short period of recruitment till now, rates of death (0.3%-0.8% and losses to follow up (1.1%-5.5% were low. 84.9% of patients were men. Main risk of transmission was sex between men (MSM: 62.9%. Mean age was 45 years. About two third of patients were classified as CDC-stage B or C. Therapy regimens of currently treated patients complied with recent guidelines. Trends of mean CD4 cell count/μl regarding the initial therapy and concerning the population under treatment reflected the developments and the changing standards of antiretroviral therapy over time. Conclusion The KompNet cohort covers about a quarter of all patients estimated as being under treatment in Germany. Its composition can be accounted approximately

Inhibition of HIV-1 replication by chimeric phosphorothioate oligodeoxynucleotides applied in free solution

DEFF Research Database (Denmark)

Lund, O S; Hansen, J E

1998-01-01

Oligodeoxynucleotides (ODNs) containing a variable number of 3' and 5' terminal phosphorothioate linkages were applied in free solution to cells infected by HIV-1. ODNs of 28 nt length were applied at up to 5 microM concentration. The ODNs were found to inhibit HIV-1 infection in a dose dependent...... by these modified chimers....

Stem cell gene therapy for HIV: strategies to inhibit viral entry and replication.

Science.gov (United States)

DiGiusto, David L

2015-03-01

Since the demonstration of a cure of an HIV+ patient with an allogeneic stem cell transplant using naturally HIV-resistant cells, significant interest in creating similar autologous products has fueled the development of a variety of "cell engineering" approaches to stem cell therapy for HIV. Among the more well-studied strategies is the inhibition of viral entry through disruption of expression of viral co-receptors or through competitive inhibitors of viral fusion with the cell membrane. Preclinical evaluation of these approaches often starts in vitro but ultimately is tested in animal models prior to clinical implementation. In this review, we trace the development of several key approaches (meganucleases, short hairpin RNA (shRNA), and fusion inhibitors) to modification of hematopoietic stem cells designed to impart resistance to HIV to their T-cell and monocytic progeny. The basic evolution of technologies through in vitro and in vivo testing is discussed as well as the pros and cons of each approach and how the addition of postentry inhibitors may enhance the overall antiviral efficacy of these approaches.

Crinivirus replication and host interactions

Directory of Open Access Journals (Sweden)

Zsofia A Kiss

2013-05-01

Full Text Available Criniviruses comprise one of the genera within the family Closteroviridae. Members in this family are restricted to the phloem and rely on whitefly vectors of the genera Bemisia and/or Trialeurodes for plant-to-plant transmission. All criniviruses have bipartite, positive-sense ssRNA genomes, although there is an unconfirmed report of one having a tripartite genome. Lettuce infectious yellows virus (LIYV is the type species of the genus, the best studied so far of the criniviruses and the first for which a reverse genetics system was available. LIYV RNA 1 encodes for proteins predicted to be involved in replication, and alone is competent for replication in protoplasts. Replication results in accumulation of cytoplasmic vesiculated membranous structures which are characteristic of most studied members of the Closteroviridae. These membranous structures, often referred to as BYV-type vesicles, are likely sites of RNA replication. LIYV RNA 2 is replicated in trans when co-infecting cells with RNA 1, but is temporally delayed relative to RNA1. Efficient RNA 2 replication also is dependent on the RNA 1-encoded RNA binding protein, P34. No LIYV RNA 2-encoded proteins have been shown to affect RNA replication, but at least four, CP, CPm, Hsp70h, and p59 are virion structural components and CPm is a determinant of whitefly transmissibility. Roles of other LIYV RNA 2-encoded proteins are largely as yet unknown, but P26 is a non-virion protein that accumulates in cells as characteristic plasmalemma deposits which in plants are localized within phloem parenchyma and companion cells over plasmodesmata connections to sieve elements. The two remaining crinivirus-conserved RNA 2-encoded proteins are P5 and P9. P5 is 39 amino acid protein and is encoded at the 5’ end of RNA 2 as ORF1 and is part of the hallmark closterovirus gene array. The orthologous gene in BYV has been shown to play a role in cell-to-cell movement and indicated to be localized to the

Serious Non-AIDS Conditions in HIV

DEFF Research Database (Denmark)

Lundgren, Jens D; Borges, Alvaro H; Neaton, James D

2018-01-01

PURPOSE OF REVIEW: Optimal control of HIV can be achieved by early diagnosis followed by the initiation of antiretroviral therapy (ART). Two large randomised trials (TEMPRANO and START) have recently been published documenting the clinical benefits to HIV-positive adults of early ART initiation....... Main findings are reviewed with a focus on serious non-AIDS (SNA) conditions. RECENT FINDINGS: Data from the two trials demonstrated that initiating ART early in the course of HIV infection resulted in marked reductions in the risk of opportunistic diseases and invasive bacterial infections....... This indicates that HIV causes immune impairment in early infection that is remedied by controlling viral replication. Intriguingly, in START, a marked reduction in risk of cancers, both infection-related and unrelated types of cancers, was observed. Like the findings for opportunistic infections, this anti...

Dendrimers as Potential Therapeutic Tools in HIV Inhibition

Directory of Open Access Journals (Sweden)

Xiangbo Li

2013-07-01

Full Text Available The present treatments for HIV transfection include chemical agents and gene therapies. Although many chemical drugs, peptides and genes have been developed for HIV inhibition, a variety of non-ignorable drawbacks limited the efficiency of these materials. In this review, we discuss the application of dendrimers as both therapeutic agents and non-viral vectors of chemical agents and genes for HIV treatment. On the one hand, dendrimers with functional end groups combine with the gp120 of HIV and CD4 molecule of host cell to suppress the attachment of HIV to the host cell. Some of the dendrimers are capable of intruding into the cell and interfere with the later stages of HIV replication as well. On the other hand, dendrimers are also able to transfer chemical drugs and genes into the host cells, which conspicuously increase the anti-HIV activity of these materials. Dendrimers as therapeutic tools provide a potential treatment for HIV infection.

Selective survival of peripheral blood lymphocytes in children with HIV-1 following delivery of an anti-HIV gene to bone marrow CD34(+) cells.

Science.gov (United States)

Podsakoff, Greg M; Engel, Barbara C; Carbonaro, Denise A; Choi, Chris; Smogorzewska, Elzbieta M; Bauer, Gerhard; Selander, David; Csik, Susan; Wilson, Kathy; Betts, Michael R; Koup, Richard A; Nabel, Gary J; Bishop, Keith; King, Steven; Schmidt, Manfred; von Kalle, Christof; Church, Joseph A; Kohn, Donald B

2005-07-01

Two HIV-1-infected children on antiretroviral therapy were enrolled into a clinical study of retroviral-mediated transfer of a gene that inhibits replication of HIV-1, targeting bone marrow CD34+ hematopoietic stem/progenitor cells. Two retroviral vectors were used, one encoding a "humanized" dominant-negative REV protein (huM10) that is a potent inhibitor of HIV-1 replication and one encoding a nontranslated marker gene (FX) to serve as an internal control for the level of gene marking. Peripheral blood mononuclear cells (PBMC) containing the huM10 gene or FX gene were detected by quantitative PCR at frequencies of approximately 1/10,000 in both subjects for the first 1-3 months following re-infusion of the gene-transduced bone marrow, but then were at or below the limits of detection (<1/1,000,000) at most times over 2 years. In one patient, a reappearance of PBMC containing the huM10 gene, but not the FX gene, occurred concomitant with a rise in the HIV-1 viral load during a period of nonadherence to the antiretroviral regimen. Unique clones of gene-marked PBMC were detected by LAM-PCR during the time of elevated HIV-1 levels. These findings indicate that there was a selective survival advantage for PBMC containing the huM10 gene during the time of increased HIV-1 load.

Superior control of HIV-1 replication by CD8+ T cells targeting conserved epitopes: implications for HIV vaccine design.

Directory of Open Access Journals (Sweden)

Pratima Kunwar

Full Text Available A successful HIV vaccine will likely induce both humoral and cell-mediated immunity, however, the enormous diversity of HIV has hampered the development of a vaccine that effectively elicits both arms of the adaptive immune response. To tackle the problem of viral diversity, T cell-based vaccine approaches have focused on two main strategies (i increasing the breadth of vaccine-induced responses or (ii increasing vaccine-induced responses targeting only conserved regions of the virus. The relative extent to which set-point viremia is impacted by epitope-conservation of CD8(+ T cell responses elicited during early HIV-infection is unknown but has important implications for vaccine design. To address this question, we comprehensively mapped HIV-1 CD8(+ T cell epitope-specificities in 23 ART-naïve individuals during early infection and computed their conservation score (CS by three different methods (prevalence, entropy and conseq on clade-B and group-M sequence alignments. The majority of CD8(+ T cell responses were directed against variable epitopes (p<0.01. Interestingly, increasing breadth of CD8(+ T cell responses specifically recognizing conserved epitopes was associated with lower set-point viremia (r = - 0.65, p = 0.009. Moreover, subjects possessing CD8(+ T cells recognizing at least one conserved epitope had 1.4 log10 lower set-point viremia compared to those recognizing only variable epitopes (p = 0.021. The association between viral control and the breadth of conserved CD8(+ T cell responses may be influenced by the method of CS definition and sequences used to determine conservation levels. Strikingly, targeting variable versus conserved epitopes was independent of HLA type (p = 0.215. The associations with viral control were independent of functional avidity of CD8(+ T cell responses elicited during early infection. Taken together, these data suggest that the next-generation of T-cell based HIV-1 vaccines should focus

Back to the future: revisiting HIV-1 lethal mutagenesis

Science.gov (United States)

Dapp, Michael J.; Patterson, Steven E.; Mansky, Louis M.

2012-01-01

The concept of eliminating HIV-1 infectivity by elevating the viral mutation rate was first proposed over a decade ago, even though the general concept had been conceived earlier for RNA viruses. Lethal mutagenesis was originally viewed as a novel chemotherapeutic approach for treating HIV-1 infection in which use of a viral mutagen would over multiple rounds of replication lead to the lethal accumulation of mutations, rendering the virus population non infectious – known as the slow mutation accumulation model. There have been limitations in obtaining good efficacy data with drug leads, leaving some doubt into clinical translation. More recent studies of the APOBEC3 proteins as well as new progress in the use of nucleoside analogs for inducing lethal mutagenesis have helped to refocus attention on rapid induction of HIV-1 lethal mutagenesis in a single or limited number of replication cycles leading to a rapid mutation accumulation model. PMID:23195922

Is a Pacific Coexistence Between Virus and Host the Unexploited Path That May Lead to an HIV Functional Cure?

Directory of Open Access Journals (Sweden)

Jonathan Fior

2013-02-01

Full Text Available The SupT1 cell line supports optimal HIV-1 replication, and prolonged in vitro replication in SupT1 cells renders the virus significantly less virulent. This raises the question of whether the infusion of SupT1 cells could be used as a cell-based therapy to induce a pacific coexistence between the HIV virus and its human host. In a recent study, I investigated this potential therapeutic strategy in vitro. The results suggested that this approach should be further explored in HIV-susceptible animal models. Such studies may lead to the development of a functional cure for HIV infection.

Colorectal mucus binds DC-SIGN and inhibits HIV-1 trans-infection of CD4+ T-lymphocytes.

Directory of Open Access Journals (Sweden)

Martijn J Stax

Full Text Available Bodily secretions, including breast milk and semen, contain factors that modulate HIV-1 infection. Since anal intercourse caries one of the highest risks for HIV-1 transmission, our aim was to determine whether colorectal mucus (CM also contains factors interfering with HIV-1 infection and replication. CM from a number of individuals was collected and tested for the capacity to bind DC-SIGN and inhibit HIV-1 cis- or trans-infection of CD4+ T-lymphocytes. To this end, a DC-SIGN binding ELISA, a gp140 trimer competition ELISA and HIV-1 capture/ transfer assays were utilized. Subsequently we aimed to identify the DC-SIGN binding component through biochemical characterization and mass spectrometry analysis. CM was shown to bind DC-SIGN and competes with HIV-1 gp140 trimer for binding. Pre-incubation of Raji-DC-SIGN cells or immature dendritic cells (iDCs with CM potently inhibits DC-SIGN mediated trans-infection of CD4+ T-lymphocytes with CCR5 and CXCR4 using HIV-1 strains, while no effect on direct infection is observed. Preliminary biochemical characterization demonstrates that the component seems to be large (>100kDa, heat and proteinase K resistant, binds in a α1-3 mannose independent manner and is highly variant between individuals. Immunoprecipitation using DC-SIGN-Fc coated agarose beads followed by mass spectrometry indicated lactoferrin (fragments and its receptor (intelectin-1 as candidates. Using ELISA we showed that lactoferrin levels within CM correlate with DC-SIGN binding capacity. In conclusion, CM can bind the C-type lectin DC-SIGN and block HIV-1 trans-infection of both CCR5 and CXCR4 using HIV-1 strains. Furthermore, our data indicate that lactoferrin is a DC-SIGN binding component of CM. These results indicate that CM has the potential to interfere with pathogen transmission and modulate immune responses at the colorectal mucosa.

What do we need to do to cure HIV infection.

Science.gov (United States)

Siliciano, Robert F

2010-01-01

Finding a cure for HIV infection requires methods to stop ongoing viral replication, to identify all reservoirs in which nonreplicating HIV persists, and to eliminate each of these reservoirs. Current antiretroviral therapy largely stops ongoing viral replication. This is a reflection of the extremely high antiviral activity of some classes of antiretroviral drugs as revealed in a novel index, the inhibitory potential, which incorporates the slope parameter of the dose-response curve. This index may aid in the rational selection of fully suppressive therapy. At least 2 stable reservoirs of latently infected cells have been identified, and attempts are under way to identify compounds that selectively reactivate latent HIV and allow elimination of these reservoirs. This article summarizes a presentation made by Robert F. Siliciano, MD, PhD, at the International AIDS Society-USA continuing medical education program held in Atlanta in March 2010.

Compartmentalization of the gut viral reservoir in HIV-1 infected patients

Directory of Open Access Journals (Sweden)

Grant Tannika

2007-12-01

Full Text Available Abstract Background Recently there has been an increasing interest and appreciation for the gut as both a viral reservoir as well as an important host-pathogen interface in human immunodefiency virus type 1 (HIV-1 infection. The gut associated lymphoid tissue (GALT is the largest lymphoid organ infected by HIV-1. In this study we examined if different HIV-1 quasispecies are found in different parts of the gut of HIV-1 infected individuals. Results Gut biopsies (esophagus, stomach, duodenum and colorectum were obtained from eight HIV-1 infected preHAART (highly active antiretroviral therapy patients. HIV-1 Nef and Reverse transcriptase (RT encoding sequences were obtained through nested PCR amplification from DNA isolated from the gut biopsy tissues. The PCR fragments were cloned and sequenced. The resulting sequences were subjected to various phylogenetic analyses. Expression of the nef gene and viral RNA in the different gut tissues was determined using real-time RT-PCR. Phylogenetic analysis of the Nef protein-encoding region revealed compartmentalization of viral replication in the gut within patients. Viral diversity in both the Nef and RT encoding region varied in different parts of the gut. Moreover, increased nef gene expression (p Conclusion Our results indicated that different HIV-1 quasispecies populate different parts of the gut, and that viral replication in the gut is compartmentalized. These observations underscore the importance of the gut as a host-pathogen interface in HIV-1 infection.

DNA damage repair machinery and HIV escape from innate immune sensing

Directory of Open Access Journals (Sweden)

Christelle eBregnard

2014-04-01

Full Text Available Viruses have been long known to perturb cell cycle regulators and key players of the DNA damage response to benefit their life cycles. In the case of the human immunodeficiency virus (HIV, the viral auxiliary protein Vpr activates the structure-specific endonuclease SLX4 complex to promote escape from innate immune sensing and, as a side effect, induces replication stress in cycling cells and subsequent cell cycle arrest at the G2/M transition. This novel pathway subverted by HIV to prevent accumulation of viral reverse transcription by-products adds up to facilitating effects of major cellular exonucleases that degrade pathological DNA species. Within this review we discuss the impact of this finding on our understanding of the interplay between HIV replication and nucleic acid metabolism and its implications for cancer-related chronic inflammation.

HIV-Specific B Cell Frequency Correlates with Neutralization Breadth in Patients Naturally Controlling HIV-Infection

Directory of Open Access Journals (Sweden)

Angeline Rouers

2017-07-01

Full Text Available HIV-specific broadly neutralizing antibodies (bnAbs have been isolated from patients with high viremia but also from HIV controllers that repress HIV-1 replication. In these elite controllers (ECs, multiple parameters contribute to viral suppression, including genetic factors and immune responses. Defining the immune correlates associated with the generation of bnAbs may help in designing efficient immunotherapies. In this study, in ECs either positive or negative for the HLA-B*57 protective allele, in treated HIV-infected and HIV-negative individuals, we characterized memory B cell compartments and HIV-specific memory B cells responses using flow cytometry and ELISPOT. ECs preserved their memory B cell compartments and in contrast to treated patients, maintained detectable HIV-specific memory B cell responses. All ECs presented IgG1+ HIV-specific memory B cells but some individuals also preserved IgG2+ or IgG3+ responses. Importantly, we also analyzed the capacity of sera from ECs to neutralize a panel of HIV strains including transmitted/founder virus. 29% and 21% of HLA-B*57+ and HLA-B*57− ECs, respectively, neutralized at least 40% of the viral strains tested. Remarkably, in HLA-B*57+ ECs the frequency of HIV-Env-specific memory B cells correlated positively with the neutralization breadth suggesting that preservation of HIV-specific memory B cells might contribute to the neutralizing responses in these patients.

Barriers, Motivators, and Facilitators to Engagement in HIV Care Among HIV-Infected Ghanaian Men Who have Sex with Men (MSM).

Science.gov (United States)

Ogunbajo, Adedotun; Kershaw, Trace; Kushwaha, Sameer; Boakye, Francis; Wallace-Atiapah, Nii-Dromo; Nelson, LaRon E

2018-03-01

In Ghana, men who have sex with men (MSM) bear a high burden of HIV. Identifying factors that influence engagement in HIV care among HIV-infected Ghanaian MSM is critical to devising novel interventions and strengthening existing programs aimed at improving outcomes across the HIV care continuum. Consequently, we conducted an exploratory qualitative research study with 30 HIV-infected Ghanaian MSM between May 2015 and July 2015. Common barriers were fear of being seen in HIV-related health facility, financial difficulties, and health system challenges. Major motivators for engagement in care included social support, fear of mortality from HIV, and knowledge of effectiveness of HIV treatment. Key facilitators were enrollment in health insurance, prior relationship and familiarity with hospital personnel, and positive experience in healthcare setting. Our findings highlight the need for new and innovative care delivery mediums, affirming and competent healthcare providers, and increased access to health insurance.

CRISPR-Cas9 Can Inhibit HIV-1 Replication but NHEJ Repair Facilitates Virus Escape

NARCIS (Netherlands)

Wang, Gang; Zhao, Na; Berkhout, Ben; Das, Atze T.

2016-01-01

Several recent studies demonstrated that the clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease Cas9 can be used for guide RNA (gRNA)-directed, sequence-specific cleavage of HIV proviral DNA in infected cells. We here demonstrate profound inhibition of HIV-1

immune response can measuring immunity to hiv during ...

African Journals Online (AJOL)

2005-11-01

Nov 1, 2005 ... inhibitors (PIs), have resulted in significant suppression of viral replication. ... thymus, with the potential for immune reconstitution when ..... HIV-exposed but uninfected Gambian women [published erratum appears in. Nat Med ...

Cerebrospinal fluid HIV-1 RNA levels in asymptomatic patients with early stage chronic HIV-1 infection: support for the hypothesis of local virus replication.

Science.gov (United States)

García, F; Niebla, G; Romeu, J; Vidal, C; Plana, M; Ortega, M; Ruiz, L; Gallart, T; Clotet, B; Miró, J M; Pumarola, T; Gatell, J M

1999-08-20

To assess HIV-1 RNA levels in cerebrospinal fluid (CSF) and their potential correlation with plasma viral load and central nervous system (CNS) HIV-1 infection markers in stable asymptomatic patients with a CD4 T cell count >500x10(6) cells/l. Consecutive patients screened for two trials were eligible for lumbar puncture assessment. At day 0, simultaneous samples of CSF and plasma were obtained and levels of total proteins, albumin, IgG, antibodies against HIV-1 p24 antigen, HIV-1 RNA (using the polymerase chain technique) and white cells were measured. The integrity of the blood-brain barrier was preserved (albumin index > or =7) in 59 out of 70 patients (84%). Intrathecal production of antibodies against HIV-1 p24 antigen was demonstrated in 55 out of 70 individuals (78%). Viral load in CSF was significantly lower than plasma values (3.13+/-0.95 versus 4.53+/-0.53, P = 0.0001). HIV-1 RNA was not detected in CSF in only three of the 70 patients (4%). Overall, there was a significant correlation between plasma and CSF HIV-1 RNA levels (r = 0.43, P = 0.0001); however, in 29 patients (41%) there were significant differences (>1.5 log10 copies/ml) between the viral loads in plasma and CSF. In the multivariate analysis, a high level of protein and white cells in CSF, but not the HIV-1 RNA plasma level, were factors independently associated with a higher level of HIV-1 RNA in CSF (P = 0.0001). HIV-1 RNA can be detected almost always in CSF of asymptomatic patients in early stages of HIV-1 infection including those with a preserved integrity of the blood-brain barrier. The important discrepancies between plasma and CSF viral load, and the independent association between CSF abnormalities and CSF viral load, support the hypothesis of local production of HIV-1.

Beneficial effect of TRAIL on HIV burden, without detectable immune consequences.

Directory of Open Access Journals (Sweden)

Brett D Shepard

2008-08-01

Full Text Available During uncontrolled HIV disease, both TNF-related apoptosis inducing ligand (TRAIL and TRAIL receptor expression are increased. Enhanced TRAIL sensitivity is due to TRAIL receptor up-regulation induced by gp120. As a result of successful antiretroviral therapy TRAIL is down-regulated, and there are fewer TRAIL-sensitive cells. In this setting, we hypothesized that all cells that contain virus, including those productively- and latently-infected, have necessarily been "primed" by gp120 and remain TRAIL-sensitive, whereas uninfected cells remain relatively TRAIL-resistant.We evaluated the immunologic and antiviral effects of TRAIL in peripheral blood lymphocytes collected from HIV-infected patients with suppressed viral replication. The peripheral blood lymphocytes were treated with recombinant TRAIL or an equivalent amount of bovine serum albumin as a negative control. Treated cells were then analyzed by quantitative flow cytometry, ELISPOT for CD4+ and CD8+ T-cell function, and limiting dilution microculture for viral burden. Alterations in the cytokine milieu of treated cells were assessed with a multiplex cytokine assay. Treatment with recombinant TRAIL in vitro reduced viral burden in lymphocytes collected from HIV-infected patients with suppressed viral load. TRAIL treatment did not alter the cytokine milieu of treated cells. Moreover, treatment with recombinant TRAIL had no adverse effect on either the quantity or function of immune cells from HIV-infected patients with suppressed viral replication.TRAIL treatment may be an important adjunct to antiretroviral therapy, even in patients with suppressed viral replication, perhaps by inducing apoptosis in cells with latent HIV reservoirs. The absence of adverse effect on the quantity or function of immune cells from HIV-infected patients suggests that there is not a significant level of "bystander death" in uninfected cells.

Cyclophilin B enhances HIV-1 infection.

Science.gov (United States)

DeBoer, Jason; Madson, Christian J; Belshan, Michael

2016-02-01

Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. Copyright © 2015 Elsevier Inc. All rights reserved.

Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

International Nuclear Information System (INIS)

Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

2015-01-01

Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR

Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

Energy Technology Data Exchange (ETDEWEB)

Sahu, Geetaram; Farley, Kalamo [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); El-Hage, Nazira [Virginia Commonwealth University, Richmond, VA (United States); Aiamkitsumrit, Benjamas; Fassnacht, Ryan [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Kashanchi, Fatah [George Mason University, Manassas, VA (United States); Ochem, Alex [ICGEB, Wernher and Beit Building, Anzio Road, Observatory, 7925 Cape Town (South Africa); Simon, Gary L. [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Karn, Jonathan [Case Western Reserve University, Cleveland, OH (United States); Hauser, Kurt F. [Virginia Commonwealth University, Richmond, VA (United States); Tyagi, Mudit, E-mail: tmudit@email.gwu.edu [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington, DC 20037 (United States)

2015-09-15

Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR.

Accumulation of MxB/Mx2-resistant HIV-1 Capsid Variants During Expansion of the HIV-1 Epidemic in Human Populations.

Science.gov (United States)

Wei, Wei; Guo, Haoran; Ma, Min; Markham, Richard; Yu, Xiao-Fang

2016-06-01

Recent studies have identified human myxovirus resistance protein 2 (MxB or Mx2) as an interferon induced inhibitor of HIV-1 replication. However, whether HIV-1 can overcome MxB restriction without compromise of viral fitness has been undefined. Here, we have discovered that naturally occurring capsid (CA) variants can render HIV-1 resistant to the activity of MxB without losing viral infectivity or the ability to escape from interferon induction. Moreover, these MxB resistant HIV-1 variants do not lose MxB recognition. Surprisingly, MxB resistant CA variants are most commonly found in the Clade C HIV-1 that is the most rapidly expanding Clade throughout the world. Accumulation of MxB resistant mutations is also observed during HIV-1 spreading in human populations. These findings support a potential role for MxB as a selective force during HIV-1 transmission and evolution. Copyright © 2016. Published by Elsevier B.V.

Influence of common mucosal co-factors on HIV infection in the female genital tract.

Science.gov (United States)

Ferreira, Victor H; Kafka, Jessica K; Kaushic, Charu

2014-06-01

Women constitute almost half of HIV-infected population globally, and the female genital tract (FGT) accounts for approximately 40% of all new HIV infections worldwide. The FGT is composed of upper and lower parts, distinct in their morphological and functional characteristics. Co-factors in the genital microenvironment, such as presence of hormones, semen, and other sexually transmitted infections, can facilitate or deter HIV infection and play a critical role in determining susceptibility to HIV. In this review, we examine some of these co-factors and their potential influence. Presence of physical and chemical barriers such as epithelial tight junctions, mucus, and anti-microbial peptides can actively block and inhibit viral replication, presenting a significant deterrent to HIV. Upon exposure, HIV and other pathogens first encounter the genital epithelium: cells that express a wide repertoire of pattern recognition receptors that can recognize and directly initiate innate immune responses. These and other interactions in the genital tract can lead to direct and indirect inflammation and enhance the number of local target cells, immune activation, and microbial translocation, all of which promote HIV infection and replication. Better understanding of the dynamics of HIV transmission in the female genital tract would be invaluable for improving the design of prophylactic strategies against HIV. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Lower Self-Reported Quality of Life in HIV-Infected Patients on cART and With Low Comorbidity Compared With Healthy Controls

DEFF Research Database (Denmark)

Pedersen, Karin K; Eiersted, Morten R; Gaardbo, Julie C

2015-01-01

replication and with low comorbidity, compared with healthy controls. We furthermore aimed to identify factors associated with QoL. DESIGN AND METHODS: Cross-sectional study of 52 HIV-infected patients and 23 healthy controls matched on age, gender, education, and comorbidity. HIV-infected patients...... and healthy controls had previously been examined regarding cognitive, physical, metabolic, and immunological parameters. QoL was investigated using the Medical Outcomes Study HIV Health Survey (MOS-HIV). Linear multiple regression models were created to find factors associated with mental health summary......BACKGROUND: Self-reported quality of life (QoL) has previously been found to be impaired in patients living with HIV and associated with viral replication, degree of immunodeficiency, and comorbidity. We aimed at investigating QoL in a group of HIV-infected patients with suppressed viral...

Virus-like-vaccines against HIV

DEFF Research Database (Denmark)

Andersson, Anne Marie C.; Schwerdtfeger, Melanie; Holst, Peter J.

2018-01-01

Protection against chronic infections has necessitated the development of ever-more potent vaccination tools. HIV seems to be the most challenging foe, with a remarkable, poorly immunogenic and fragile surface glycoprotein and the ability to overpower the cell immune system. Virus-like-particle (......Protection against chronic infections has necessitated the development of ever-more potent vaccination tools. HIV seems to be the most challenging foe, with a remarkable, poorly immunogenic and fragile surface glycoprotein and the ability to overpower the cell immune system. Virus...... of HIV. Such vaccines are immunologically perceived as viruses, as they infect cells and produce VLPs in situ, but they only resemble viruses, as the replication defective vectors and VLPs cannot propagate an infection. The inherent safety of such a platform, despite robust particle production...

Factors shaping the HIV-competence of two primary schools in rural Zimbabwe

DEFF Research Database (Denmark)

Campbell, Catherine; Andersen, Louise; Mutsikiwa, Alice

2015-01-01

We present multi-method case studies of two Zimbabwean primary schools - one rural and one small-town. The rural school scored higher than the small-town school on measures of child well-being and school attendance by HIV-affected children. The small-town school had superior facilities, more...... teachers with higher morale, more specialist HIV/AIDS activities, and an explicit religious ethos. The relatively impoverished rural school was located in a more cohesive community with a more critically conscious, dynamic and networking headmaster. The current emphasis on HIV/AIDS-related teacher training...

Family Wellness, Not HIV Prevention

Science.gov (United States)

Swendeman, Dallas; Flannery, Diane

2010-01-01

HIV exceptionalism (and disease-specific programs generally) garner both unbalanced funding and the most talented personnel, distorting local health priorities. In support of HIV exceptionalism, the successful mobilization of significant global health sector resources was not possible prior to HIV. Both sides of the debate have merits; rather than perpetuating polarization, we suggest that sustained improvements in global health require creating a prevention infrastructure to meet multiple health challenges experienced by local communities. We propose four fundamental shifts in HIV and disease prevention: (1) horizontally integrating prevention at one site locally, with priorities tailored to local health challenges and managed by local community leaders; (2) using a family wellness metaphor for services, not disease prevention; (3) implementing evidence-based prevention programs (EBPP) based on common principles, factors, and processes, rather than replication of specific programs; and (4) utilizing the expertise of private enterprise to re-design EBPP into highly attractive, engaging, and accessible experiences. PMID:19148744

Impact of CD4 and CD8 dynamics and viral rebounds on loss of virological control in HIV controllers

DEFF Research Database (Denmark)

Chereau, Fanny; Madec, Yoann; Sabin, Caroline

2017-01-01

OBJECTIVE: HIV controllers (HICs) spontaneously maintain HIV viral replication at low level without antiretroviral therapy (ART), a small number of whom will eventually lose this ability to control HIV viremia. The objective was to identify factors associated with loss of virological control. MET...

Development of an epitope-based HIV-1 vaccine strategy from HIV-1 lipopeptide to dendritic-based vaccines.

Science.gov (United States)

Surenaud, Mathieu; Lacabaratz, Christine; Zurawski, Gérard; Lévy, Yves; Lelièvre, Jean-Daniel

2017-10-01

Development of a safe, effective and globally affordable Human Immunodeficiency Virus strain 1 (HIV-1) vaccine offers the best hope for future control of the HIV-1 pandemic. However, with the exception of the recent RV144 trial, which elicited a modest level of protection against infection, no vaccine candidate has shown efficacy in preventing HIV-1 infection or in controlling virus replication in humans. There is also a great need for a successful immunotherapeutic vaccine since combination antiretroviral therapy (cART) does not eliminate the reservoir of HIV-infected cells. But to date, no vaccine candidate has proven to significantly alter the natural history of an individual with HIV-1 infection. Areas covered: For over 25 years, the ANRS (France Recherche Nord&Sud Sida-HIV hépatites) has been committed to an original program combining basic science and clinical research developing an epitope-based vaccine strategy to induce a multiepitopic cellular response against HIV-1. This review describes the evolution of concepts, based on strategies using HIV-1 lipopeptides towards the use of dendritic cell (DC) manipulation. Expert commentary: Understanding the crucial role of DCs in immune responses allowed moving from the non-specific administration of HIV-1 sequences with lipopeptides to DC-based vaccines. These DC-targeting strategies should improve HIV-1 vaccine efficacy.

A DEAD box protein facilitates HIV-1 replication as a cellular co-factor of Rev

International Nuclear Information System (INIS)

Fang Jianhua; Kubota, Satoshi; Yang Bin; Zhou Naiming; Zhang Hui; Godbout, Roseline; Pomerantz, Roger J.

2004-01-01

HIV-1 Rev escorts unspliced viral mRNAs out of the nucleus of infected cells, which allows formation of infectious HIV-1 virions. We have identified a putative DEAD box (Asp-Glu-Ala-Asp) RNA helicase, DDX1, as a cellular co-factor of Rev, through yeast and mammalian two-hybrid systems using the N-terminal motif of Rev as 'bait'. DDX1 is not a functional homolog of HIV-1 Rev, but down-regulation of DDX1 resulted in an alternative splicing pattern of Rev-responsive element (RRE)-containing mRNA, and attenuation of Gag p24 antigen production from HLfb rev(-) cells rescued by exogenous Rev. Co-transfection of a DDX1 expression vector with HIV-1 significantly increased viral production. DDX1 binding to Rev, as well as to the RRE, strongly suggest that DDX1 affects Rev function through the Rev-RRE axis. Moreover, down-regulation of DDX1 altered the steady state subcellular distribution of Rev, from nuclear/nucleolar to cytoplasmic dominance. These findings indicate that DDX1 is a critical cellular co-factor for Rev function, which maintains the proper subcellular distribution of this lentiviral regulatory protein. Therefore, alterations in DDX1-Rev interactions could induce HIV-1 persistence and targeting DDX1 may lead to rationally designed and novel anti-HIV-1 strategies and therapeutics

Antiviral Activity of HIV gp120 Targeting Bispecific T Cell Engager (BiTE®) Antibody Constructs.

Science.gov (United States)

Brozy, Johannes; Schlaepfer, Erika; Mueller, Christina K S; Rochat, Mary-Aude; Rampini, Silvana K; Myburgh, Renier; Raum, Tobias; Kufer, Peter; Baeuerle, Patrick A; Muenz, Markus; Speck, Roberto F

2018-05-02

Today's gold standard in HIV therapy is the combined antiretroviral therapy (cART). It requires strict adherence by patients and life-long medication, which can lower the viral load below detection limits and prevent HIV-associated immunodeficiency, but cannot cure patients. The bispecific T cell engaging (BiTE®) antibody technology has demonstrated long-term relapse-free outcomes in patients with relapsed and refractory acute lymphocytic leukemia. We here generated BiTE® antibody constructs that target the HIV-1 envelope protein gp120 (HIV gp120) using either the scFv B12 or VRC01, the first two extracellular domains (1+2) of human CD4 alone or joined to the single chain variable fragment (scFv) of the antibody 17b fused to an anti-human CD3ϵ scFv. These engineered human BiTE® antibody constructs showed engagement of T cells for redirected lysis of HIV gp120-transfected CHO cells. Furthermore, they substantially inhibited HIV-1 replication in PBMCs as well as in macrophages co-cultured with autologous CD8+ T-cells, the most potent being the human CD4(1+2) BiTE® antibody construct and the CD4(1+2)L17b BiTE® antibody construct. The CD4(1+2) h BiTE® antibody construct promoted HIV infection of human CD4-/CD8+ T cells. In contrast, the neutralizing B12 and the VRC01 BiTE® antibody constructs as well as the CD4(1+2)L17b BiTE® antibody construct did not. Thus, BiTE® antibody constructs targeting HIV gp120 are very promising for constraining HIV and warrant further development as novel antiviral therapy with curative potential. Importance HIV is a chronic infection well controlled with the current cART. However, we lack cure of HIV, and the HIV pandemic goes on. Here we showed in vitro and ex vivo t hat a bispecific T-cell engaging (BiTE®) antibody construct targeting HIV gp120 resulted in substantially reduced HIV replication. In addition, these BiTE® antibody constructs display efficient killing of gp120 expressing cells and inhibited replication in ex vivo

Cyclophilin B enhances HIV-1 infection

Energy Technology Data Exchange (ETDEWEB)

DeBoer, Jason; Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln, NE (United States)

2016-02-15

Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

Cyclophilin B enhances HIV-1 infection

International Nuclear Information System (INIS)

DeBoer, Jason; Madson, Christian J.; Belshan, Michael

2016-01-01

Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

High Multiplicity Infection by HIV-1 in Men Who Have Sex with Men.

Directory of Open Access Journals (Sweden)

Hui Li

2010-05-01

Full Text Available Elucidating virus-host interactions responsible for HIV-1 transmission is important for advancing HIV-1 prevention strategies. To this end, single genome amplification (SGA and sequencing of HIV-1 within the context of a model of random virus evolution has made possible for the first time an unambiguous identification of transmitted/founder viruses and a precise estimation of their numbers. Here, we applied this approach to HIV-1 env analyses in a cohort of acutely infected men who have sex with men (MSM and found that a high proportion (10 of 28; 36% had been productively infected by more than one virus. In subjects with multivariant transmission, the minimum number of transmitted viruses ranged from 2 to 10 with viral recombination leading to rapid and extensive genetic shuffling among virus lineages. A combined analysis of these results, together with recently published findings based on identical SGA methods in largely heterosexual (HSX cohorts, revealed a significantly higher frequency of multivariant transmission in MSM than in HSX [19 of 50 subjects (38% versus 34 of 175 subjects (19%; Fisher's exact p = 0.008]. To further evaluate the SGA strategy for identifying transmitted/founder viruses, we analyzed 239 overlapping 5' and 3' half genome or env-only sequences from plasma viral RNA (vRNA and blood mononuclear cell DNA in an MSM subject who had a particularly well-documented virus exposure history 3-6 days before symptom onset and 14-17 days before peak plasma viremia (47,600,000 vRNA molecules/ml. All 239 sequences coalesced to a single transmitted/founder virus genome in a time frame consistent with the clinical history, and a molecular clone of this genome encoded replication competent virus in accord with model predictions. Higher multiplicity of HIV-1 infection in MSM compared with HSX is consistent with the demonstrably higher epidemiological risk of virus acquisition in MSM and could indicate a greater challenge for HIV-1

Brain Macrophages in Simian Immunodeficiency Virus-Infected, Antiretroviral-Suppressed Macaques: a Functional Latent Reservoir.

Science.gov (United States)

Avalos, Claudia R; Abreu, Celina M; Queen, Suzanne E; Li, Ming; Price, Sarah; Shirk, Erin N; Engle, Elizabeth L; Forsyth, Ellen; Bullock, Brandon T; Mac Gabhann, Feilim; Wietgrefe, Stephen W; Haase, Ashley T; Zink, M Christine; Mankowski, Joseph L; Clements, Janice E; Gama, Lucio

2017-08-15

A human immunodeficiency virus (HIV) infection cure requires an understanding of the cellular and anatomical sites harboring virus that contribute to viral rebound upon treatment interruption. Despite antiretroviral therapy (ART), HIV-associated neurocognitive disorders (HAND) are reported in HIV-infected individuals on ART. Biomarkers for macrophage activation and neuronal damage in cerebrospinal fluid (CSF) of HIV-infected individuals demonstrate continued effects of HIV in brain and suggest that the central nervous system (CNS) may serve as a viral reservoir. Using a simian immunodeficiency virus (SIV)/macaque model for HIV encephalitis and AIDS, we evaluated whether infected cells persist in brain despite ART. Eight SIV-infected pig-tailed macaques were virally suppressed with ART, and plasma and CSF viremia levels were analyzed longitudinally. To assess whether virus persisted in brain macrophages (BrMΦ) in these macaques, we used a macrophage quantitative viral outgrowth assay (MΦ-QVOA), PCR, and in situ hybridization (ISH) to measure the frequency of infected cells and the levels of viral RNA and DNA in brain. Viral RNA in brain tissue of suppressed macaques was undetectable, although viral DNA was detected in all animals. The MΦ-QVOA demonstrated that the majority of suppressed animals contained latently infected BrMΦ. We also showed that virus produced in the MΦ-QVOAs was replication competent, suggesting that latently infected BrMΦ are capable of reestablishing productive infection upon treatment interruption. This report provides the first confirmation of the presence of replication-competent SIV in BrMΦ of ART-suppressed macaques and suggests that the highly debated issue of viral latency in macrophages, at least in brain, has been addressed in SIV-infected macaques treated with ART. IMPORTANCE Resting CD4 + T cells are currently the only cells that fit the definition of a latent reservoir. However, recent evidence suggests that HIV

Histone deacetylase inhibitor romidepsin induces HIV expression in CD4 T cells from patients on suppressive antiretroviral therapy at concentrations achieved by clinical dosing.

Directory of Open Access Journals (Sweden)

Datsen George Wei

2014-04-01

Full Text Available Persistent latent reservoir of replication-competent proviruses in memory CD4 T cells is a major obstacle to curing HIV infection. Pharmacological activation of HIV expression in latently infected cells is being explored as one of the strategies to deplete the latent HIV reservoir. In this study, we characterized the ability of romidepsin (RMD, a histone deacetylase inhibitor approved for the treatment of T-cell lymphomas, to activate the expression of latent HIV. In an in vitro T-cell model of HIV latency, RMD was the most potent inducer of HIV (EC50 = 4.5 nM compared with vorinostat (VOR; EC50 = 3,950 nM and other histone deacetylase (HDAC inhibitors in clinical development including panobinostat (PNB; EC50 = 10 nM. The HIV induction potencies of RMD, VOR, and PNB paralleled their inhibitory activities against multiple human HDAC isoenzymes. In both resting and memory CD4 T cells isolated from HIV-infected patients on suppressive combination antiretroviral therapy (cART, a 4-hour exposure to 40 nM RMD induced a mean 6-fold increase in intracellular HIV RNA levels, whereas a 24-hour treatment with 1 µM VOR resulted in 2- to 3-fold increases. RMD-induced intracellular HIV RNA expression persisted for 48 hours and correlated with sustained inhibition of cell-associated HDAC activity. By comparison, the induction of HIV RNA by VOR and PNB was transient and diminished after 24 hours. RMD also increased levels of extracellular HIV RNA and virions from both memory and resting CD4 T-cell cultures. The activation of HIV expression was observed at RMD concentrations below the drug plasma levels achieved by doses used in patients treated for T-cell lymphomas. In conclusion, RMD induces HIV expression ex vivo at concentrations that can be achieved clinically, indicating that the drug may reactivate latent HIV in patients on suppressive cART.

Minimal Contribution of APOBEC3-Induced G-to-A Hypermutation to HIV-1 Recombination and Genetic Variation.

Science.gov (United States)

Delviks-Frankenberry, Krista A; Nikolaitchik, Olga A; Burdick, Ryan C; Gorelick, Robert J; Keele, Brandon F; Hu, Wei-Shau; Pathak, Vinay K

2016-05-01

Although the predominant effect of host restriction APOBEC3 proteins on HIV-1 infection is to block viral replication, they might inadvertently increase retroviral genetic variation by inducing G-to-A hypermutation. Numerous studies have disagreed on the contribution of hypermutation to viral genetic diversity and evolution. Confounding factors contributing to the debate include the extent of lethal (stop codon) and sublethal hypermutation induced by different APOBEC3 proteins, the inability to distinguish between G-to-A mutations induced by APOBEC3 proteins and error-prone viral replication, the potential impact of hypermutation on the frequency of retroviral recombination, and the extent to which viral recombination occurs in vivo, which can reassort mutations in hypermutated genomes. Here, we determined the effects of hypermutation on the HIV-1 recombination rate and its contribution to genetic variation through recombination to generate progeny genomes containing portions of hypermutated genomes without lethal mutations. We found that hypermutation did not significantly affect the rate of recombination, and recombination between hypermutated and wild-type genomes only increased the viral mutation rate by 3.9 × 10-5 mutations/bp/replication cycle in heterozygous virions, which is similar to the HIV-1 mutation rate. Since copackaging of hypermutated and wild-type genomes occurs very rarely in vivo, recombination between hypermutated and wild-type genomes does not significantly contribute to the genetic variation of replicating HIV-1. We also analyzed previously reported hypermutated sequences from infected patients and determined that the frequency of sublethal mutagenesis for A3G and A3F is negligible (4 × 10-21 and1 × 10-11, respectively) and its contribution to viral mutations is far below mutations generated during error-prone reverse transcription. Taken together, we conclude that the contribution of APOBEC3-induced hypermutation to HIV-1 genetic

Alterations in the nuclear proteome of HIV-1 infected T-cells

International Nuclear Information System (INIS)

DeBoer, Jason; Jagadish, Teena; Haverland, Nicole A.; Madson, Christian J.; Ciborowski, Pawel; Belshan, Michael

2014-01-01

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines

Alterations in the nuclear proteome of HIV-1 infected T-cells

Energy Technology Data Exchange (ETDEWEB)

DeBoer, Jason [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Jagadish, Teena; Haverland, Nicole A. [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); Ciborowski, Pawel [Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE 68198 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178 (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln 68583 (United States)

2014-11-15

Virus infection of a cell involves the appropriation of host factors and the innate defensive response of the cell. The identification of proteins critical for virus replication may lead to the development of novel, cell-based inhibitors. In this study we mapped the changes in T-cell nuclei during human immunodeficiency virus type 1 (HIV-1) at 20 hpi. Using a stringent data threshold, a total of 13 and 38 unique proteins were identified in infected and uninfected cells, respectively, across all biological replicates. An additional 15 proteins were found to be differentially regulated between infected and control nuclei. STRING analysis identified four clusters of protein–protein interactions in the data set related to nuclear architecture, RNA regulation, cell division, and cell homeostasis. Immunoblot analysis confirmed the differential expression of several proteins in both C8166-45 and Jurkat E6-1 T-cells. These data provide a map of the response in host cell nuclei upon HIV-1 infection. - Highlights: • We identify changes in the expression of nuclear proteins during HIV-1 infection. • 163 nuclear proteins were found differentially regulated during HIV-1 infection. • Bioinformatic analysis identified several nuclear pathways altered by HIV infection. • Candidate factors were validated in two independent cell lines.

Diagnosing HIV “window phase” in routine laboratory: a teaching case

Directory of Open Access Journals (Sweden)

Roberta Valentina Marotta

2011-12-01

Full Text Available Human immunodeficiency virus (HIV is the causative agent of acquired immunodeficiency syndrome (AIDS. It is a retrovirus that inserts itself into the host’s DNA and use the host – cell’s replication mechanisms for its own perpetuation. Two distinct species of HIV (HIV 1 and HIV 2 have been identified and they have a different global distribution. In this paper we describe a case of HIV early infection in an italian man of fourty years old. From this example we want to stress the importance to consider with high attention the validation of a negative index of HIV antibodies.We advise to follow this procedure that allows to avoid the risk to report a false negative.

Synthesis and evaluation of antiviral activities of novel sonochemical silver nanorods against HIV and HSV viruses

Directory of Open Access Journals (Sweden)

Mazyar Etemadzade

2016-11-01

Full Text Available Objective: To evaluate the effect of novel sonochemical silver nanorods on HIV and herpes simplex virus type 1 (HSV-1 viruses in human cervical cancer HeLa cells. Methods: The formation of silver nanorods conjugated with sodium 2-mercaptoethane sulfonate (Ag-MES was characterized by scanning electron microscopy, Fourier transform infrared spectroscopy and thermal gravimetric analysis. The antiviral activity of this Ag-MES was examined against HIV and HSV-1 virus replication. Results: The characterizations of Ag-MES and physiochemical structure were determined by scanning electron microscopy, Fourier transform infrared spectroscopy and thermal gravimetric analysis. Approximately entire viral replication was inhibited by Ag-MES at 10 µmol/mL concentration. About 90% of HSV virions failed to replicate in the present of this concentration of nanorods. However, HIV showed more sensitivity to Ag-MES than HSV-1. Conclusions: According to the obtained data, the synthesized sonochemical silver nanorod in this study is a promising candidate for further drug discovery investigation.

The impact of pregnancy on the HIV-1-specific T cell function in infected pregnant women.

Science.gov (United States)

Hygino, Joana; Vieira, Morgana M; Kasahara, Taissa M; Xavier, Luciana F; Blanco, Bernardo; Guillermo, Landi V C; Filho, Renato G S; Saramago, Carmen S M; Lima-Silva, Agostinho A; Oliveira, Ariane L; Guimarães, Vander; Andrade, Arnaldo F B; Bento, Cleonice A M

2012-12-01

Evidences indicate that pregnancy can alter the Ag-specific T-cell responses. This work aims to evaluate the impact of pregnancy on the in vitro HIV-1-specific immune response. As compared with non-pregnant patients, lower T-cell proliferation and higher IL-10 production were observed in T-cell cultures from pregnant patients following addition of either mitogens or HIV-1 antigens. In our system, the main T lymphocyte subset involved in producing IL-10 was CD4(+)FoxP3(-). Depletion of CD4(+) cells elevated TNF-α and IFN-γ production. Interestingly, the in vitro HIV-1 replication was lower in cell cultures from pregnant patients, and it was inversely related to IL-10 production. In these cultures, the neutralization of IL-10 by anti-IL-10 mAb elevated TNF-α release and HIV-1 replication. In conclusion, our results reveal that pregnancy-related events should favor the expansion of HIV-1-specific IL-10-secreting CD4(+) T-cells in HIV-1-infected women, which should, in the scenario of pregnancy, help to reduce the risk of vertical HIV-1 transmission. Copyright © 2012 Elsevier Inc. All rights reserved.

HIV-Associated Gastrointestinal Cancer.

Science.gov (United States)

Jensen, Björn E-O; Oette, Mark; Haes, Johannes; Häussinger, Dieter

2017-01-01

People living nowadays with HIV and AIDS may be treated effectively regarding virus replication and immunology. However, non-AIDS-defining cancer is of growing relevance due to high incidence and unfavorable outcome. The aim of this review is to summarize current knowledge on gastrointestinal (GI) carcinoma. Although literature on GI cancer is rare, an increased incidence of esophageal, gastric, pancreatic, hepatocellular, and colorectal carcinoma has been demonstrated. However, there are only few reports on therapy strategies and outcome, so that, despite increased occurrence of many GI carcinomas, only little is known about individualized treatment options and outcome in HIV-positive patients. More efforts have to be undertaken to close this gap. © 2017 S. Karger GmbH, Freiburg.

Rational design of micro-RNA-like bifunctional siRNAs targeting HIV and the HIV coreceptor CCR5.

Science.gov (United States)

Ehsani, Ali; Saetrom, Pål; Zhang, Jane; Alluin, Jessica; Li, Haitang; Snøve, Ola; Aagaard, Lars; Rossi, John J

2010-04-01

Small-interfering RNAs (siRNAs) and micro-RNAs (miRNAs) are distinguished by their modes of action. SiRNAs serve as guides for sequence-specific cleavage of complementary mRNAs and the targets can be in coding or noncoding regions of the target transcripts. MiRNAs inhibit translation via partially complementary base-pairing to 3' untranslated regions (UTRs) and are generally ineffective when targeting coding regions of a transcript. In this study, we deliberately designed siRNAs that simultaneously direct cleavage and translational suppression of HIV RNAs, or cleavage of the mRNA encoding the HIV coreceptor CCR5 and suppression of translation of HIV. These bifunctional siRNAs trigger inhibition of HIV infection and replication in cell culture. The design principles have wide applications throughout the genome, as about 90% of genes harbor sites that make the design of bifunctional siRNAs possible.

Olean-18-ene triterpenoids from Celastraceae species inhibit HIV replication targeting NF-kB and Sp1 dependent transcription.

Science.gov (United States)

Osorio, Alex A; Muñóz, Alejandro; Torres-Romero, David; Bedoya, Luis M; Perestelo, Nayra R; Jiménez, Ignacio A; Alcamí, José; Bazzocchi, Isabel L

2012-06-01

In the present study we report the isolation of nine new olean-18-ene triterpenes (1-9), along with three known ones (10-12), from Cassine xylocarpa and Maytenus jelskii. Their stereostructures have been elucidated on the basis of spectroscopic analysis, including 1D and 2D NMR techniques (COSY, ROESY, HSQC and HMBC), and spectrometric methods. The natural compounds and derivatives 13-15 have been tested for their potential as inhibitors of human immunodeficiency virus type 1 replication. Five compounds from this series displayed potent antiviral activity with IC(50)s in the micromolar range (1, 3, 4, 7 and 8) being 1 and 8 the most active compounds. The target of these compounds was different from antiretroviral drugs currently licensed as they act as inhibitors of enhancer-dependent transcription. The structure-activity relationships were established based on the regiosubstitution and oxidation degree of the triterpene scaffold, revealing that these aspects were able to modulate the selectivity and intensity of HIV inhibition. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

HIV vaccine research and discovery in the nonhuman primates model: a unified theory in acquisition prevention and control of SIV infection.

Science.gov (United States)

Lynch, Rebecca M; Yamamoto, Takuya; McDermott, Adrian B

2013-07-01

Here we highlight the latest advances in HIV vaccine concepts that will expand our knowledge on how to elicit effective acquisition-prevention and/or control of simian immunodeficiency virus (SIV) replication in the nonhuman primate (NHP) model. In the context of the promising analyses from the RV144 Thai Trial and the effective control of SIV replication exerted by rhCMV-(SIV) elicited EM CD8 T cells, the HIV field has recently shifted toward vaccine concepts that combine protection from acquisition with effective control of SIV replication. Current studies in the NHP model have demonstrated the efficacy of HIV-neutralizing antibodies via passive transfer, the potential importance of the CD4 Tfh subset, the ability to effectively model the RV144 vaccine trial and the capacity of an Ad26 prime and modified vaccinia Ankara virus boost to elicit Env-specific antibody and cellular responses that both limit acquisition and control heterologous SIVmac251 challenge. The latest work in the NHP model suggests that the next generation HIV-1 vaccines should aim to provoke a comprehensive adaptive immune response for both prevention of SIV acquisition as well as control of replication in breakthrough infection.

Genome-wide association studies on HIV susceptibility, pathogenesis and pharmacogenomics

Directory of Open Access Journals (Sweden)

van Manen Daniëlle

2012-08-01

Full Text Available Abstract Susceptibility to HIV-1 and the clinical course after infection show a substantial heterogeneity between individuals. Part of this variability can be attributed to host genetic variation. Initial candidate gene studies have revealed interesting host factors that influence HIV infection, replication and pathogenesis. Recently, genome-wide association studies (GWAS were utilized for unbiased searches at a genome-wide level to discover novel genetic factors and pathways involved in HIV-1 infection. This review gives an overview of findings from the GWAS performed on HIV infection, within different cohorts, with variable patient and phenotype selection. Furthermore, novel techniques and strategies in research that might contribute to the complete understanding of virus-host interactions and its role on the pathogenesis of HIV infection are discussed.

The cytosolic exonuclease TREX1 inhibits the innate immune response to HIV-1

Science.gov (United States)

Yan, Nan; Regalado-Magdos, Ashton D.; Stiggelbout, Bart; Lee-Kirsch, Min Ae; Lieberman, Judy

2010-01-01

Viral infection triggers innate immune sensors to produce type I interferons (IFN). However, HIV infection of T cells and macrophages does not trip these alarms. How HIV avoids activating nucleic acid sensors is unknown. The cytosolic exonuclease TREX1 suppressed IFN triggered by HIV. In Trex1−/− mouse cells and human CD4+ T cells and macrophages in which TREX1 was inhibited by RNA interference, cytosolic HIV DNA accumulated, and HIV infection induced type I IFN that inhibited HIV replication and spreading. TREX1 bound to cytosolic HIV DNA and digested excess HIV DNA that would otherwise activate IFN expression via a TBK1, STING and IRF3 dependent pathway. HIV-stimulated IFN production in cells deficient in TREX1 did not involve known nucleic acid sensors. PMID:20871604

Raltegravir: first in class HIV integrase inhibitor

Directory of Open Access Journals (Sweden)

Zelalem Temesgen

2008-06-01

Full Text Available Zelalem Temesgen1, Dawd S Siraj21Mayo Clinic, Rochester, MN, USA; 2East Carolina University Greenville, NC, USAAbstract: On October 16, 2007, the US Food and Drug Administration (FDA approved raltegravir for treatment of human immunodeficiency virus (HIV-1 infection in combination with other antiretroviral agents in treatment-experienced adult patients who have evidence of viral replication and HIV-1 strains resistant to multiple antiretroviral agents. Raltegravir is first in a novel class of antiretroviral drugs known as integrase inhibitors. It has demonstrated potent anti HIV activity in both antiretroviral treatment-naïve and experienced patients. The most common adverse events reported with raltegravir during phase 2 and 3 clinical trials were diarrhea, nausea, and headache. Laboratory abnormalities include mild elevations in liver transaminases and creatine phosphokinase.Keywords: raltegravir, HIV, antiretroviral agents, integrase inhibitors

Expression of heterologous genes from an IRES translational cassette in replication-competent murine leukemia virus vectors

DEFF Research Database (Denmark)

Jespersen, T.; Duch, M.; Carrasco, M.L.

1999-01-01

of spliced env mRNA for the SL3-3 derived vector relative to the Akv derived vectors, seemingly contributing to its low replication capacity. The EGFP expressing Akv-MLV was genetically stable for multiple rounds of infection; marker-cassette deletion revertants appeared after several replication rounds...

Antibody-dependent enhancement of HIV-1 infection in human term syncytiotrophoblast cells cultured in vitro.

Science.gov (United States)

Tóth, F D; Mosborg-Petersen, P; Kiss, J; Aboagye-Mathiesen, G; Zdravkovic, M; Hager, H; Aranyosi, J; Lampé, L; Ebbesen, P

1994-06-01

We examined if Fc receptor-mediated antibody-dependent enhancement (FcR-ADE) or complement-mediated antibody-dependent enhancement (C'-ADE) of virus infection can contribute to increasing replication of HIV-1 in human syncytiotrophoblast (ST) cells. Here we report that both FcR-ADE and C'-ADE may result in enhanced virus release from HIV-1-infected ST cells. We show that FcR-ADE of HIV-1 infection in ST cells is mediated by FcRIII and other FcR(s) belonging to undetermined Fc classes and does not require CD4 receptors, whereas C'-ADE uses both CD4 and CR2-like receptors. FcR-ADE seems to be more efficient in enhancing HIV-1 replication than C'-ADE. While FcR-ADE leads to increased internalization of HIV-1, C'-ADE does not result in enhanced endocytosis of the virus. In addition, antibodies mediating FcR-ADE are reactive with the gp120 viral envelope antigen, whereas antibodies involved in C'-ADE react with the viral transmembrane glycoprotein gp41. Data suggest that both FcR-ADE and C'-ADE may contribute to the spread of HIV-1 from mother to the fetus.

Bicyclams, selective antagonists of the human chemokine receptor CXCR4, potently inhibit feline immunodeficiency virus replication

NARCIS (Netherlands)

Horzinek, M.C.; Egberink, H.F.; Clercq, E. de; Vliet, A.L.W. van; Balzarini, J.; Bridger, G.J.; Henson, G.; Schols, D.

1999-01-01

Bicyclams are low-molecular-weight anti-human immunodeficiency virus (HIV) agents that have been shown to act as potent and selective CXC chemokine receptor 4 (CXCR4) antagonists. Here, we demonstrate that bicyclams are potent inhibitors of feline immunodeficiency virus (FIV) replication when

Health-related quality of life of HIV infected adults with and without Visceral Leishmaniasis in Northwest Ethiopia

OpenAIRE

Alemayehu, Mekuriaw; Wubshet, Mamo; Mesfin, Nebiyu; Tamiru, Aschalew; Gebayehu, Abebaw

2017-01-01

Background Health-related quality of life (HRQoL) is an important outcome measure among HIV infected patients receiving antiretroviral therapy (ART). When HIV infected patients coinfected with Visceral Leishmaniasis (VL) the problem become severe because VL accelerates HIV replication and disease progression. The impact of VL on the quality of life of HIV infected patients has not been studied. In this study in Ethiopia, we compared the quality of life of HIV infected patients with and withou...

Association of HIV diversity and survival in HIV-infected Ugandan infants.

Directory of Open Access Journals (Sweden)

Maria M James

2011-04-01

Full Text Available The level of viral diversity in an HIV-infected individual can change during the course of HIV infection, reflecting mutagenesis during viral replication and selection of viral variants by immune and other selective pressures. Differences in the level of viral diversity in HIV-infected infants may reflect differences in viral dynamics, immune responses, or other factors that may also influence HIV disease progression. We used a novel high resolution melting (HRM assay to measure HIV diversity in Ugandan infants and examined the relationship between diversity and survival through 5 years of age.Plasma samples were obtained from 31 HIV-infected infants (HIVNET 012 trial. The HRM assay was used to measure diversity in two regions in the gag gene (Gag1 and Gag2 and one region in the pol gene (Pol.HRM scores in all three regions increased with age from 6-8 weeks to 12-18 months (for Gag1: P = 0.005; for Gag2: P = 0.006; for Pol: P = 0.016. Higher HRM scores at 6-8 weeks of age (scores above the 75(th percentile were associated with an increased risk of death by 5 years of age (for Pol: P = 0.005; for Gag1/Gag2 (mean of two scores: P = 0.003; for Gag1/Gag2/Pol (mean of three scores: P = 0.002. We did not find an association between HRM scores and other clinical and laboratory variables.Genetic diversity in HIV gag and pol measured using the HRM assay was typically low near birth and increased over time. Higher HIV diversity in these regions at 6-8 weeks of age was associated with a significantly increased risk of death by 5 years of age.

PRACTICE OF USING VIRAL PROTEASE INHIBITORS IN CHILDREN WITH HIV INFECTION

Directory of Open Access Journals (Sweden)

V.B. Denisenko

2010-01-01

Full Text Available Selection of the most effective and safest high-active antiretroviral therapies is a critical issue faced by modern HIV medicine. Authors studied 28 children with HIV infection aged from 3 to 7 divided into two groups administered a combination of two HIV reverse transcriptase nucleoside inhibitors with viral protease nelfinavir inhibitors (n = 13 and lopinavir/ritonavir (n = 15. The subjects in both groups demonstrated a decreased frequency of HIV-associated symptoms and opportunistic infections, positive dynamics of immunological indicators, suppression of HIV replication. When lopinavir/ritonavir was administered, there was more even better dynamics in clinical, immunological and virologic parameters, which allows this medication to be recommended as a antiretroviral therapy for children. Key words: HIV infection, lopinavir/ritonavir, nelfinavir, children. (Pediatric Pharmacology. – 2010; 7(1:62-67

Preclinical safety and efficacy of an anti–HIV-1 lentiviral vector containing a short hairpin RNA to CCR5 and the C46 fusion inhibitor

Directory of Open Access Journals (Sweden)

Orit Wolstein

2014-01-01

Full Text Available Gene transfer has therapeutic potential for treating HIV-1 infection by generating cells that are resistant to the virus. We have engineered a novel self-inactivating lentiviral vector, LVsh5/C46, using two viral-entry inhibitors to block early steps of HIV-1 cycle. The LVsh5/C46 vector encodes a short hairpin RNA (shRNA for downregulation of CCR5, in combination with the HIV-1 fusion inhibitor, C46. We demonstrate here the effective delivery of LVsh5/C46 to human T cell lines, peripheral blood mononuclear cells, primary CD4+ T lymphocytes, and CD34+ hematopoietic stem/progenitor cells (HSPC. CCR5-targeted shRNA (sh5 and C46 peptide were stably expressed in the target cells and were able to effectively protect gene-modified cells against infection with CCR5- and CXCR4-tropic strains of HIV-1. LVsh5/C46 treatment was nontoxic as assessed by cell growth and viability, was noninflammatory, and had no adverse effect on HSPC differentiation. LVsh5/C46 could be produced at a scale sufficient for clinical development and resulted in active viral particles with very low mutagenic potential and the absence of replication-competent lentivirus. Based on these in vitro results, plus additional in vivo safety and efficacy data, LVsh5/C46 is now being tested in a phase 1/2 clinical trial for the treatment of HIV-1 disease.

Live attenuated measles vaccine expressing HIV-1 Gag virus like particles covered with gp160ΔV1V2 is strongly immunogenic

International Nuclear Information System (INIS)

Guerbois, Mathilde; Moris, Arnaud; Combredet, Chantal; Najburg, Valerie; Ruffie, Claude; Fevrier, Michele; Cayet, Nadege; Brandler, Samantha; Schwartz, Olivier; Tangy, Frederic

2009-01-01

Although a live attenuated HIV vaccine is not currently considered for safety reasons, a strategy inducing both T cells and neutralizing antibodies to native assembled HIV-1 particles expressed by a replicating virus might mimic the advantageous characteristics of live attenuated vaccine. To this aim, we generated a live attenuated recombinant measles vaccine expressing HIV-1 Gag virus-like particles (VLPs) covered with gp160ΔV1V2 Env protein. The measles-HIV virus replicated efficiently in cell culture and induced the intense budding of HIV particles covered with Env. In mice sensitive to MV infection, this recombinant vaccine stimulated high levels of cellular and humoral immunity to both MV and HIV with neutralizing activity. The measles-HIV virus infected human professional antigen-presenting cells, such as dendritic cells and B cells, and induced efficient presentation of HIV-1 epitopes and subsequent activation of human HIV-1 Gag-specific T cell clones. This candidate vaccine will be next tested in non-human primates. As a pediatric vaccine, it might protect children and adolescents simultaneously from measles and HIV.

Factors influencing cerebrospinal fluid and plasma HIV-1 RNA detection rate in patients with and without opportunistic neurological disease during the HAART era

Directory of Open Access Journals (Sweden)

Aleixo Agdemir W

2007-12-01

Full Text Available Abstract Background In the central nervous system, HIV replication can occur relatively independent of systemic infection, and intrathecal replication of HIV-1 has been observed in patients with HIV-related and opportunistic neurological diseases. The clinical usefulness of HIV-1 RNA detection in the cerebrospinal fluid (CSF of patients with opportunistic neurological diseases, or the effect of opportunistic diseases on CSF HIV levels in patients under HAART has not been well defined. We quantified CSF and plasma viral load in HIV-infected patients with and without different active opportunistic neurological diseases, determined the characteristics that led to a higher detection rate of HIV RNA in CSF, and compared these two compartments. Methods A prospective study was conducted on 90 HIV-infected patients submitted to lumbar puncture as part of a work-up for suspected neurological disease. Seventy-one patients had active neurological diseases while the remaining 19 did not. Results HIV-1 RNA was quantified in 90 CSF and 70 plasma samples. The HIV-1 RNA detection rate in CSF was higher in patients with neurological diseases, in those with a CD4 count lower than 200 cells/mm3, and in those not receiving antiretroviral therapy, as well as in patients with detectable plasma HIV-1 RNA. Median viral load was lower in CSF than in plasma in the total population, in patients without neurological diseases, and in patients with toxoplasmic encephalitis, while no significant difference between the two compartments was observed for patients with cryptococcal meningitis and HIV-associated dementia. CSF viral load was lower in patients with cryptococcal meningitis and neurotoxoplasmosis under HAART than in those not receiving HAART. Conclusion Detection of HIV-1 RNA in CSF was more frequent in patients with neurological disease, a CD4 count lower than 200 cells/mm3 and detectable plasma HIV-1. Median HIV-1 RNA levels were generally lower in CSF than in

Interleukin-7 facilitates HIV-1 transmission to cervico-vaginal tissue ex vivo.

Directory of Open Access Journals (Sweden)

Andrea Introini

2013-02-01

Full Text Available The majority of HIV-1 infections in women occur through vaginal intercourse, in which virus-containing semen is deposited on the cervico-vaginal mucosa. Semen is more than a mere carrier of HIV-1, since it contains many biological factors, in particular cytokines, that may affect HIV-1 transmission. The concentration of interleukin (IL-7, one of the most prominent cytokines in semen of healthy individuals, is further increased in semen of HIV-1-infected men. Here, we investigated the potential role of IL-7 in HIV-1 vaginal transmission in an ex vivo system of human cervico-vaginal tissue. We simulated an in vivo situation by depositing HIV-1 on cervico-vaginal tissue in combination with IL-7 at concentrations comparable with those measured in semen of HIV-1-infected individuals. We found that IL-7 significantly enhanced virus replication in ex vivo infected cervico-vaginal tissue. Similarly, we observed an enhancement of HIV-1 replication in lymphoid tissue explants. Analysis of T cells isolated from infected tissues showed that IL-7 reduced CD4⁺ T cell depletion preventing apoptosis, as shown by the decrease in the number of cells expressing the apoptotic marker APO2.7 and the increase in the expression of the anti-apoptotic protein B-cell lymphoma (Bcl-2. Also, IL-7 increased the fraction of cycling CD4⁺ T cells, as evidenced by staining for the nuclear factor Ki-67. High levels of seminal IL-7 in vivo may be relevant to the survival of the founder pool of HIV-1-infected cells in the cervico-vaginal mucosa at the initial stage of infection, promoting local expansion and dissemination of HIV infection.

Host ESCRT proteins are required for bromovirus RNA replication compartment assembly and function.

Directory of Open Access Journals (Sweden)

Arturo Diaz

2015-03-01

Full Text Available Positive-strand RNA viruses genome replication invariably is associated with vesicles or other rearranged cellular membranes. Brome mosaic virus (BMV RNA replication occurs on perinuclear endoplasmic reticulum (ER membranes in ~70 nm vesicular invaginations (spherules. BMV RNA replication vesicles show multiple parallels with membrane-enveloped, budding retrovirus virions, whose envelopment and release depend on the host ESCRT (endosomal sorting complexes required for transport membrane-remodeling machinery. We now find that deleting components of the ESCRT pathway results in at least two distinct BMV phenotypes. One group of genes regulate RNA replication and the frequency of viral replication complex formation, but had no effect on spherule size, while a second group of genes regulate RNA replication in a way or ways independent of spherule formation. In particular, deleting SNF7 inhibits BMV RNA replication > 25-fold and abolishes detectable BMV spherule formation, even though the BMV RNA replication proteins accumulate and localize normally on perinuclear ER membranes. Moreover, BMV ESCRT recruitment and spherule assembly depend on different sets of protein-protein interactions from those used by multivesicular body vesicles, HIV-1 virion budding, or tomato bushy stunt virus (TBSV spherule formation. These and other data demonstrate that BMV requires cellular ESCRT components for proper formation and function of its vesicular RNA replication compartments. The results highlight growing but diverse interactions of ESCRT factors with many viruses and viral processes, and potential value of the ESCRT pathway as a target for broad-spectrum antiviral resistance.

Synthesis of a Vpr-Binding Derivative for Use as a Novel HIV-1 Inhibitor.

Science.gov (United States)

Hagiwara, Kyoji; Ishii, Hideki; Murakami, Tomoyuki; Takeshima, Shin-nosuke; Chutiwitoonchai, Nopporn; Kodama, Eiichi N; Kawaji, Kumi; Kondoh, Yasumitsu; Honda, Kaori; Osada, Hiroyuki; Tsunetsugu-Yokota, Yasuko; Suzuki, Masaaki; Aida, Yoko

2015-01-01

The emergence of multidrug-resistant viruses compromises the efficacy of anti-human immunodeficiency virus type 1 (HIV-1) therapy and limits treatment options. Therefore, new targets that can be used to develop novel antiviral agents need to be identified. We previously identified a potential parent compound, hematoxylin, which suppresses the nuclear import of HIV-1 via the Vpr-importin α interaction and inhibits HIV-1 replication in a Vpr-dependent manner by blocking nuclear import of the pre-integration complex. However, it was unstable. Here, we synthesized a stable derivative of hematoxylin that bound specifically and stably to Vpr and inhibited HIV-1 replication in macrophages. Furthermore, like hematoxylin, the derivative inhibited nuclear import of Vpr in an in vitro nuclear import assay, but had no effect on Vpr-induced G2/M phase cell cycle arrest or caspase activity. Interestingly, this derivative bound strongly to amino acid residues 54-74 within the C-terminal α-helical domain (αH3) of Vpr. These residues are highly conserved among different HIV strains, indicating that this region is a potential target for drug-resistant HIV-1 infection. Thus, we succeeded in developing a stable hematoxylin derivative that bound directly to Vpr, suggesting that specific inhibitors of the interaction between cells and viral accessory proteins may provide a new strategy for the treatment of HIV-1 infection.

Synthesis of a Vpr-Binding Derivative for Use as a Novel HIV-1 Inhibitor.

Directory of Open Access Journals (Sweden)

Kyoji Hagiwara

Full Text Available The emergence of multidrug-resistant viruses compromises the efficacy of anti-human immunodeficiency virus type 1 (HIV-1 therapy and limits treatment options. Therefore, new targets that can be used to develop novel antiviral agents need to be identified. We previously identified a potential parent compound, hematoxylin, which suppresses the nuclear import of HIV-1 via the Vpr-importin α interaction and inhibits HIV-1 replication in a Vpr-dependent manner by blocking nuclear import of the pre-integration complex. However, it was unstable. Here, we synthesized a stable derivative of hematoxylin that bound specifically and stably to Vpr and inhibited HIV-1 replication in macrophages. Furthermore, like hematoxylin, the derivative inhibited nuclear import of Vpr in an in vitro nuclear import assay, but had no effect on Vpr-induced G2/M phase cell cycle arrest or caspase activity. Interestingly, this derivative bound strongly to amino acid residues 54-74 within the C-terminal α-helical domain (αH3 of Vpr. These residues are highly conserved among different HIV strains, indicating that this region is a potential target for drug-resistant HIV-1 infection. Thus, we succeeded in developing a stable hematoxylin derivative that bound directly to Vpr, suggesting that specific inhibitors of the interaction between cells and viral accessory proteins may provide a new strategy for the treatment of HIV-1 infection.

Replication-dependent 65R→K reversion in human immunodeficiency virus type 1 reverse transcriptase double mutant K65R + L74V

International Nuclear Information System (INIS)

Sharma, Prem L.; Nurpeisov, Viktoria; Lee, Kimberly; Skaggs, Sara; Di San Filippo, Christina Amat; Schinazi, Raymond F.

2004-01-01

Understanding of the mechanisms of interaction among nucleoside reverse transcriptase inhibitor (NRTI)-selected mutations in the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) coding sequence is essential for the design of newer drugs and for enhancing our vision of the structure function relationship among amino acids of the polymerase domain of HIV-1. Although several nucleoside reverse transcriptase inhibitors select RT mutations K65R and L74V, the combination of 65R + 74V is rare in clinics. A novel NRTI (-)-β-D-dioxolane-guanosine (DXG) is known to select in vitro either the 65R or 74V mutant virus (Antimicrob. Agents Chemother. 44 (2000) 1783). These mutations were not selected together during repeated passaging of the HIV-1 in the presence of this drug. To analyze the impact of these RT mutations on viral replication, a double mutant containing K65R + L74V was created by site-directed mutagenesis in a pNL4-3 background. Replication kinetic assays revealed that the mutant K65R + L74V is unstable, and 65R→K reversion occurs during replication of virus in phytohemagglutinin (PHA)-stimulated human peripheral blood mononuclear (PBM) cells in the absence of selection pressure. Replication kinetic assays in MT-2 cells demonstrated that double mutant 65R + 74V is highly attenuated for replication and the initiation of reversion is related to the increase in RT activity. Additionally, the suppression of viral replication in the presence of DXG or under suboptimal human recombinant interleukin-2 leads to minimal or no 65R→K reversion. These observations provide evidence that 65R→K reversion in the double mutant 65R + 74V is dependent on a specific rate of viral replication in a pNL4-3 background. A similar phenomenon may occur in vivo, which may have implications for treatment management strategies

RNA interactions in the 5' region of the HIV-1 genome

DEFF Research Database (Denmark)

Damgaard, Christian Kroun; Andersen, Ebbe Sloth; Knudsen, Bjarne

2004-01-01

The untranslated leader of the dimeric HIV-1 RNA genome is folded into a complex structure that plays multiple and essential roles in the viral replication cycle. Here, we have investigated secondary and tertiary structural elements within the 5' 744 nucleotides of the HIV-1 genome using...... a combination of bioinformatics, enzymatic probing, native gel electrophoresis, and UV-crosslinking experiments. We used a recently developed RNA folding algorithm (Pfold) to predict the common secondary structure of an alignment of 20 divergent HIV-1 sequences. Combining this analysis with biochemical data, we...

Novel HIV-1 knockdown targets identified by an enriched kinases/phosphatases shRNA library using a long-term iterative screen in Jurkat T-cells.

Directory of Open Access Journals (Sweden)

Sylvie Rato

2010-02-01

Full Text Available HIV-1 is a complex retrovirus that uses host machinery to promote its replication. Understanding cellular proteins involved in the multistep process of HIV-1 infection may result in the discovery of more adapted and effective therapeutic targets. Kinases and phosphatases are a druggable class of proteins critically involved in regulation of signal pathways of eukaryotic cells. Here, we focused on the discovery of kinases and phosphatases that are essential for HIV-1 replication but dispensable for cell viability. We performed an iterative screen in Jurkat T-cells with a short-hairpin-RNA (shRNA library highly enriched for human kinases and phosphatases. We identified 14 new proteins essential for HIV-1 replication that do not affect cell viability. These proteins are described to be involved in MAPK, JNK and ERK pathways, vesicular traffic and DNA repair. Moreover, we show that the proteins under study are important in an early step of HIV-1 infection before viral integration, whereas some of them affect viral transcription/translation. This study brings new insights for the complex interplay of HIV-1/host cell and opens new possibilities for antiviral strategies.