Showing papers in "Current HIV Research in 2003"

Astrocyte infection by HIV-1: Mechanisms of restricted virus replication, and role in the pathogenesis of HIV-1-associated dementia

Abstract: Astrocytes are the most numerous cell type in the brain, and their physiological roles are essential for normal brain function. Studies of post-mortem brain tissue samples from individuals with AIDS have revealed that a small proportion of astrocytes are infected by HIV-1 which is linked to the development of HIVassociated dementia (HIVD), a frequent clinical manifestation of HIV-1 disease affecting up to 20% of infected adults. However, astrocyte infection by HIV-1 in vivo is generally non-productive, and can only be readily detected by sensitive techniques that detect HIV-1 RNA or proviral DNA. Similarly, primary astrocyte cultures and astrocytic cell lines can be permissive to infection by HIV-1 strains, but are refractory to efficient HIV-1 expression. In efforts to delineate the molecular mechanisms underlying the restricted infection, several studies have demonstrated that efficient HIV-1 replication is blocked in astrocytes at different steps of the virus life cycle, including virus entry, reverse transcription, nucleocytoplasmic HIV-1 RNA transport, translation of viral RNA, and maturation of progeny virions. However, the relative importance of each of these possible replication blocks in restricting HIV-1 replication in astrocytes is unclear. Moreover, how restricted astrocyte infection contributes to the development of HIVD is unknown. This review surveys the current in vitro models of restricted HIV-1 replication in astrocytes, and provides an analysis of the available evidence supporting a role for astrocyte infection in the pathogenesis of HIVD. A greater understanding of the fate of HIV-1 in astrocytes may assist in the identification of viral reservoirs in the central nervous system, novel therapies for the treatment of HIVD, and also novel strategies to suppress HIV-1 replication in CD4+ cells of the immune system.

Transmission and Immunopathogenesis of FIV in Cats as a Model for HIV

Abstract: The feline immunodeficiency virus (FIV) model provides a system to study lentivirus transmission, virus kinetics, pathogenesis, host responses, and immune dysfunction in a natural, out-bred host, under controlled conditions with specific-pathogen-free animals. The diversity of primary FIV strains can be exploited to mirror the range of disease manifestations associated with HIV infection. FIV is infectious via intravenous, intraperitoneal, intradermal, or subcutaneous injection as well as by atraumatic instillation onto the oral, vaginal, or rectal mucosa. Together, these features allow investigators to model specific aspects of HIV infection in a highly relevant and relatively inexpensive animal model. Well-developed areas of the FIV model include: (1) transmission of cell-associated as well as cell-free virus; (2) mucosal infectivity and immunopathogenesis; (3) vertical transmission; (4) acquired immunodeficiency including defects of the innate immune system; (5) thymic dysfunction; (6) neurotropism and neuropathogenesis; (7) host-virus interactions and the role of specific gene products; (8) efficacy of antiviral therapy; and (9) efficacy and immune correlates of experimental vaccines. This review will encompass areas specific to transmission and immunopathogenesis.

HIV-1 coreceptor usage, transmission, and disease progression.

Abstract: HIV-1 coreceptor usage is believed to play a critical role in pathogenesis. To initiate infection, HIV-1 interacts with two cell surface receptors; CD4 is the primary receptor and the beta-chemokine receptors CCR5 and CXCR4 usually serve as secondary receptors. HIV-1 strains transmitted in vivo generally use CCR5. Viruses that use CCR5 (R5 viruses) appear to be associated with relatively stable infection. Years after chronic infection is established, CXCR4 utilizing strains emerge in approximately 50% of infected individuals. Viruses that use the coreceptor CXCR4 (X4 viruses) are associated with rapid CD4+ cell decline and disease progression. However, the mechanism by which X4 viruses are associated with accelerated disease progression has never been properly elucidated. For example, the association between X4 virus and acceleration of HIV-1 disease progression has been ascribed to the expanded spectrum of CXCR4+ precursor cells susceptible to infection by X4 strains. It has also been postulated that the decline of the host immune system associated with clinical AIDS may allow X4 viruses to evolve and replicate freely in late-stage infection. Discriminating between these and other alternatives is central to increasing our understanding of the fundamental pathogenic processes involved in HIV-1 infection. In this article, we critically review those studies published over the last few years that purport to examine the relationship between HIV-1 coreceptor usage, transmission, CD4+ T-cell depletion, and disease progression.

The physiological relevance of CD4 receptor down-modulation during HIV infection.

Abstract: Upon binding to the CD4 receptor the HIV envelope protein undergoes conformational changes that culminate in the fusion of the viral and cellular membranes. A few hours later, a sophisticated set of processes is initiated to ensure the down-modulation of the viral receptor. Three viral proteins participate in this process: Nef, Env, and Vpu, suggesting that this function is critical for virus replication. The mechanisms of action of these proteins have been extensively characterized. However, the physiological relevance of the virus-induced CD4 down-modulation remains a focus of controversy, and the impact of this function on the viral life cycle has been underestimated. This review summarizes current hypotheses explaining why HIV needs to reduce expression of its own receptor, and discusses the experimental evidence supporting them. Recent findings indicate that efficient CD4 down-modulation is essential for the production of infectious particles, and highlight the importance of this function in HIV pathogenesis in vivo. Progression to disease correlates with enhanced viral induced CD4 down-modulation, and a subset of long-term nonprogressors carry viruses defective in this function. To date, the HIV-induced CD4 down-modulation has not been targeted for therapeutic intervention. Addressing the reasons why this function is so critical and understanding the interplay between viral and host factors governing surface expression of CD4 may provide clues for the development of new antiviral strategies.

Measuring the infectiousness of persons with HIV-1: opportunities for preventing sexual HIV-1 transmission.

Abstract: Methods to reduce sexual transmission of HIV-1 are urgently needed to slow the global HIV-1 epidemic. These methods should include interventions that minimize susceptibility in uninfected populations at risk, as well as interventions that decrease the infectiousness of HIV-1 infected individuals. Surprisingly few interventions to prevent HIV-1 transmission have been targeted at persons who are already infected, although such interventions could have a significant impact on population-wide HIV-1 spread. This review summarizes current knowledge of factors that influence HIV-1 infectiousness, with special attention on the role of genital tract HIV-1 as a surrogate marker of infectiousness. Epidemiologic studies have demonstrated good agreement between factors associated with increased HIV-1 transmission and increased HIV-1 genital tract shedding. Treatment of sexually transmitted diseases and initiation of antiretroviral therapy have been shown to reduce HIV-1 shedding and thus are promising interventions that may reduce HIV-1 transmission. Unresolved issues related to optimal measurement techniques for genital HIV-1, promises and limitations of antiretroviral therapy and vaccines to reduce infectiousness, and other intervention strategies still in development are reviewed.

Chromatin remodeling and modification during HIV-1 Tat-activated transcription.

Abstract: Human immunodeficiency virus type 1 (HIV-1) is the etiologic agent of AIDS. Following entry into the host cell, the viral RNA is reverse transcribed into DNA and subsequently integrated into the host genome as a chromatin template. Chromatin structure may be responsible for silencing retroviral gene expression. Transcriptional activation occurs after ATP-dependent chromatin remodeling complexes alter chromatin structure and positioning of nucleosomes. Histone acetyltransferases (HATs), histone deacetylases (HDACs), kinases, and methyltransferases (HMTs), covalently modify nucleosomes by adding or removing chemical moieties in the N-terminal tails of histones. Recent advances have indicated that HIV-1 encoded proteins interact with chromatin remodeling complexes and histone modifying enzymes, implying that chromatin remodeling plays an important role in the HIV-1 life cycle. Nucleosomes are positioned on the HIV-1 LTR and are barriers to transcription. Following cellular activation, these nucleosomes are modified and repositioned allowing for activation of viral gene expression. Tat recruits various HATs to the HIV-1 promoter region and can also be acetylated by some of these enzymes. Unmodified Tat is involved in binding to the CBP/p300 and cdk9/cyclin T complexes and facilitates transcription initiation. Acetylated Tat dissociates from the TAR RNA structure and recruits bromodomain-containing chromatin modifying complexes such as p/CAF and SWI/SNF to facilitate transcription elongation. This review summarizes our current knowledge and understanding of chromatin remodeling complexes and their regulation of HIV-1 replication, and highlights the important contributions HIV-1 research has made to further our understanding of the transcription process.

Molecular mechanisms and biological significance of CTL avidity.

Abstract: CD8 CTLs are a major effector for protection against cancer as well as many infectious diseases, including HIV/AIDS. CD8 CTL recognize antigenic peptides in the context of class I MHC. CTL functional avidity has been shown to be an important determinant of in vivo efficacy. CTL that can recognize peptide/MHC only at high antigen density are termed low avidity CTL, while those that can recognize their cognate antigen at low densities are termed high avidity CTL. Recent studies have demonstrated that high avidity CTLs are essential for the effective clearance of viral infections and for the elimination of tumor cells. At this time, approaches that can target high avidity cells for expansion in vivo are not well defined; however, new insights are beginning to emerge. A recent study has shown that prime-boost immunization may be an effective method to generate high avidity CTLs that recognize HIV antigens. In addition, we recently found that high levels of costimulation (signal 2) can skew the CTL response toward higher avidity cells. Thus, vectors expressing a triad of costimulatory molecules (TRICOM) or dendritic cells expressing higher levels of costimulatory molecules, can be used to induce high avidity CTL. Finally a critical role for CD4+ T cell help in the generation of high avidity cells has recently been identified (Palmer, manuscript submitted). While high avidity CTLs are superior for viral and tumor clearance, they also have a greater sensitivity to antigen induced cell death. In some types of chronic infections, such as HIV and HCV, as well as in cancer, the host may lose, by clonal exhaustion or other apoptotic mechanisms, the effector cells that are most critical to viral or tumor clearance. In this review, we examine the current knowledge concerning CTL avidity. We discuss the factors that may distinguish high avidity CTLs from low avidity CTLs and describe some of the mechanisms these cells use to clear viral infections. In addition, we study possible immunization strategies that may be used to elicit higher avidity CTLs and describe what is known about the factors that render these cells more susceptible to apoptosis than low avidity CTLs. Finally, we will incorporate these various elements into a general discussion of possible approaches for induction and maintenance of an effective immune response that can result in clearance of tumors or chronic viral infections and the relevance to vaccine development.

Regulation of TAK/P-TEFb in CD4+ T lymphocytes and macrophages.

Abstract: HIV replication occurs principally in activated CD4+ T cells and macrophages. The HIV-1 Tat protein is essential for HIV replication and requires a cellular protein kinase activity termed TAK/P-TEFb, composed of CDK9 and cyclin T1, for its transactivation function. This article reviews recent work indicating that under some circumstances TAK/P-TEFb is likely to be limiting for HIV replication in CD4+ T cells and macrophages, and discusses mechanisms of regulation of the TAK/P-TEFb subunits in these cell types. In resting CD4+ T lymphocytes, TAK/P-TEFb function is low. Following lymphocyte activation, even under conditions of minimal activation in which activation markers and cellular proliferation are not induced, both CDK9 and cyclin T1 mRNA and protein levels are increased, leading to an induction of TAK/P-TEFb kinase activity that correlates with increased viral replication. In macrophages, regulation of TAK/P-TEFb involves mechanisms distinct from those in lymphocytes. In freshly isolated monocytes, CDK9 protein levels are high, while cyclin T1 protein levels are low to undetectable. Cyclin T1 protein expression is up-regulated during early macrophage differentiation by a mechanism that involves post-transcriptional regulation. Later during differentiation, cyclin T1 expression becomes shut off by a post-transcriptional mechanism, and this correlates with a decrease in Tat transactivation. Interestingly, cyclin T1 can be re-induced with lipopolysaccharide (LPS). These findings suggest that changes in cyclin T1 expression can influence HIV-1 replication levels in monocytes and macrophages. Important areas for future research on Tat and TAK/P-TEFb function are discussed.

Advances in vaccine adjuvants for infectious diseases

Abstract: A HIV Vaccine, particularly that based on recombinant proteins and plasmid DNA, is likely to be less reactogenic than traditional vaccines, but also less immunogenic. Therefore, there is an urgent need for the development of new and improved adjuvants and delivery system for combination with HIV vaccine antigens. Adjuvants can be broadly separated into two classes, based on their principal mechanisms of action; "vaccine delivery systems" and "immunostimulatory adjuvants". Vaccine delivery systems are generally particulate formulations e.g. emulsions, microparticles, iscoms and liposomes, and mainly function to target associated antigens into antigen presenting cells (APC). In contrast, immunostimulatory adjuvants are predominantly derived from pathogens and often represent pathogen associated molecular patterns (PAMP) e.g. LPS, MPL, CpG DNA, which activate cells of the innate immune system. The discovery of more potent adjuvants may allow the development of prophylactic and therapeutic vaccines against HIV. In addition, new adjuvants may also allow vaccines to be delivered mucosally.

Mother-to-Child Transmission of HIV Infection and its Prevention

Abstract: An estimated 800000 children acquired HIV-infection in 2002 most as a result of mother-to-child transmission (MTCT) and vertically-acquired HIV infection continues to be of major public health importance. Prevention of MTCT is possible with a combination of interventions including antiretroviral therapy (ART) (usually in highly active combinations) elective caesarean section and avoidance of breastfeeding and where infected women are identified before or in pregnancy and have access to these interventions risk of MTCT is now below 1- 2%. However prompt identification of pregnant women with HIV infection remains pressing in many developed countries; additionally concerns have arisen regarding adherence to complex treatment regimens in pregnancy and the potential impact of HIV drug resistance. More disturbingly most HIV-infected women live in developing countries where many pregnant women even when tested do not return for their HIV results for a variety of reasons including stigma and where most if not all strategies for prevention of MTCT have been of limited accessibility and/or feasibility. However the Global Fund to Fight AIDS Tuberculosis and Malaria and other initiatives including pharmaceutical companies donation programmes and generic antiretroviral prophylaxis including combination therapy to prevent MTCT. Research is additionally focussing on reducing the risk of postnatal transmission through breastfeeding with exclusive breastfeeding early cessation and antiretroviral prophylaxis to breastfeeding women or breastfed infants under investigation. However the key to prevention of paediatric HIV infections is adequate prevention of infection in women of reproductive age. (authors)

HIV vaccine development: lessons from the past and promise for the future.

Abstract: The global HIV epidemic continues to expand, exceeding previous predictions and causing tremendous suffering. An effective vaccine represents the best hope to curtail the HIV epidemic. The past fifteen years of HIV vaccine clinical trials have not identified an ideal HIV vaccine, but have provided many valuable lessons that contribute to the current generation of promising HIV vaccine regimens. An enhanced understanding of HIV and SIV immunopathogenesis has facilitated the design of vaccination regimens that elicit specific immune responses and effector mechanisms. Intensive investigation of recombinant gp120 subunit vaccines has revealed a previously unexpected complexity in eliciting neutralizing antibodies that are active against primary isolate viruses. The importance of CD8+ CTL responses in controlling HIV and SIV viremia has led to a series of vaccine candidates that effectively induce these responses. Proof that vaccination can prevent SIV/HIV disease has now been obtained in simian models of AIDS. A number of promising HIV vaccine regimens are currently being evaluated in human trials, and the pipeline of new vaccine vectors and combination regimens appears robust. Although challenges to the development of a safe and effective global HIV vaccine remain, the outlook for HIV vaccines in the future is bright.

Rodent models for HIV-1 infection and disease.

Abstract: The development of a predictive, small animal model for human immunodeficiency virus type 1 (HIV-1) disease would greatly facilitate the analysis of many aspects of viral infection, pathogenesis and treatment. While numerous small animal models exist which emulate various aspects of HIV-1 infection and/or disease in humans, none of these models support robust HIV-1 replication within the context of an intact immune system. Despite this major limitation, these models have helped to elucidate different aspects of HIV-1 pathogenesis in humans. Moreover, recent advances regarding the underlying nature of the blocks to viral replication in non-human cells have raised the possibility that rodents may be engineered to support HIV-1 infection. This review will focus on recent attempts to develop a rodent model for HIV-1 disease, and will also describe currently available systems for studying HIV-1.

Recombinant rhabdoviruses as potential vaccines for HIV-1 and other diseases.

Abstract: The failure to develop vaccines to protect against important infectious diseases such as human immunodeficiency virus type I (HIV-1) or Hepatitis C virus (HCV) has increased the interest in new vaccine strategies. One of these methods is immunization with an attenuated recombinant viral vector expressing a foreign antigen, which could protect individuals from later exposure to the respective pathogen. A new method to recover a non-segmented negative-stranded RNA virus (NNSV) from cDNA was described for the first time for rabies virus (RV), a member of the rhabdovirus family. The same approach was successfully used for another rhabdovirus, vesicular stomatitis virus (VSV), and opened the possibility to use rhabdoviruses as vaccine vehicles and biomedical tools. Further research showed that the genomes of rhabdoviruses are highly flexible, easy to manipulate, and able to express large and even multiple foreign genes, and therefore are excellent vaccine candidates. In addition, it has been shown for both RV and VSV that their single surface glycoprotein G, which is responsible for attachment and fusion to the host cell, can functionally be replaced by other viral or cellular glycoproteins. This review gives an overview of the use of RV and VSV as promising new candidates in the fight against HIV-1 and other human diseases.

Pharmacological cyclin-dependent kinase inhibitors as HIV-1 antiviral therapeutics.

Abstract: Human immunodeficiency virus type 1 (HIV-1) can infect quiescent cells; however, viral production is restricted to actively proliferating cells. Recent evidence has indicated that HIV-1 viral proteins, Vpr and Tat, perturb the cell cycle to optimize HIV-1 replication. Vpr arrests the cell cycle at G2 by inactivating the cyclin B/cdk1 complex. Tat regulates the cell cycle by altering factors involved in proliferation and differentiation (i.e. the cdk inhibitor p21/waf1) and associating with cyclin/cdk complexes (i.e. cyclin E/cdk2, cyclin H/cdk7, and cyclin T/cdk9). These studies indicate the importance of host cellular factors, such as cyclin/cdk complexes, in regulating HIV-1 replication and therefore represent novel targets for antiviral therapeutics. Recently, the efficacy of pharmalogical cdk inhibitors (PCIs) in abrogating viral replication has been under development. To date there are 25-30 PCIs that have been synthesized against known cdks, several of which have been shown to inhibit HIV-1 and other AIDS-associated viruses in vitro and in vivo. Targeting these critical cyclin/cdk complexes needed for viral propagation may solve the problems inherent in current HAART therapy, including the emergence of drug-resistant viruses. Thus, PCIs have the potential to become novel therapeutic antiviral drugs that can inhibit HIV-1 transcription and opens the possibility of new avenues of treatment.

Replicative senescence: the final stage of memory T cell differentiation?

Abstract: One of the major obstacles to effective prolonged CD8 T cell control over HIV and other latent infections may be the intrinsic, genetically programmed barrier to unlimited proliferation that is characteristic of all normal human somatic cells. Replicative senescence, characterized extensively in cell culture for a variety of cell types, comprises both irreversible cell cycle arrest and striking changes in function. CD8 T cells with features similar to senescent CD8 T cell cultures (i.e., absence of CD28, inability to proliferate, telomeres in the 5-7 kb range, resistance to apoptosis) increase progressively during aging and in chronic HIV infection, suggesting that replicative senescence may be occurring in vivo, and, in fact, may constitute the final stage in the normal differentiation of human T cells. CD8 T cells with characteristics suggestive of senescence have also been implicated in modulating immune function and altering bone homeostasis. Further characterization of the underlying mechanism leading to the generation of senescent memory CD8 T cells and analysis of their functional attributes will help elucidate their role in HIV disease pathogenesis.

In vivo analysis of Nef function.

Abstract: The nef gene is present in all primate lentiviruses (HIV-1, HIV-2, and SIVs). In vivo, Nef has been shown to be a major determinant of virus pathogenicity. In vitro, many different Nef activities have been reported, including CD4 and MHC I downregulation, enhanced virion infectivity, and T-cell activation. These four different activities have been extensively investigated and appear to increase the pathogenicity of the virus. However, the contribution that these activities (individually or together) make to the in vivo phenotype has not been elucidated. The mechanism(s) by which Nef modulates distinct host cell properties has provided great insights into the intricate interaction between virus and host. In this manuscript, we review the different model systems that have been used to study Nef function in vivo and the information that they have provided regarding the best characterized in vitro Nef activities. The knowledge that has been accumulated has provided clues to our understanding of Nef function but they have also left us with many unanswered questions that should be the focus of future in vivo analysis of Nef function.

Update on D-ala-peptide T-amide (DAPTA): a viral entry inhibitor that blocks CCR5 chemokine receptors.

Abstract: Peptide T, named for its high threonine content (ASTTTNYT), was derived by a database search which assumed that a relevant receptor binding epitope within env (gp120) would have sequence homology to a known signaling peptide. Binding of radiolabeled gp120 to brain membranes was displaced by peptide T and three octapeptide analogs (including “DAPTA”, Dala1-peptide T-amide, the protease- resistant analog now in Phase II clinical trials) with the same potency that these four octapeptides blocked infectivity of an early passage patient isolate. This 1986 report was controversial due to a number of laboratories failure to find peptide T antiviral effects; we now know that peptide T is a potent HIV entry inhibitor selectively targeting CCR5 receptors with minimal effects on the X4 tropic lab adapted virus exclusively in use at that time. Early clinical trials, which demonstrated lack of toxicity and focused on neurological and neurocognitive benefits, are reviewed and data from a small ongoing Phase II trial---the first to assess peptide Ts antiviral effects---are presented. Studies using infectivity, receptor binding, chemotaxis, and blockade of gp120-induced neurotoxicity in vitro and in vivo are reviewed, discussed and presented here. Peptide T and analogs of its core pentapeptide, present near the V2 stem of numerous gp120 isolates, are potent ligands for CCR5. Clinical data showing peptide Ts immunomodulation of plasma cytokine levels and increases in the percentage of IFN? secreting CD8+ T cells in patients with HIV disease are presented and suggests additional therapeutic mechanisms via regulation of specific immunity.

Neuroprotective therapy for HIV dementia.

Abstract: Despite the development and use of effective antiretroviral therapy, HIV dementia persists and has important socio-economic consequences. Significant progress has been made in our current understanding of the neuropathogenesis of HIV infection, and it is clear that adjunctive neuroprotective therapy in addition to antiretroviral therapy are necessary for prevention and treatment of this entity. In this manuscript, we discuss the rationale and the pathophysiological mechanisms that support the development of neuroprotective therapy for HIV dementia. We review all the placebo controlled clinical trials conducted to date with neuromodulatory / neuroprotective therapy in patients with HIV dementia and discuss their outcomes. We also provide a thorough review of potential new treatments for HIV dementia based on the experimental literature. We hope that this manuscript will serve as an important guide for future approaches for clinical trials and drug development for patients with HIV dementia.

Role of FAS in HIV Infection

Abstract: Direct cytopathic effects cannot explain the massive CD4+ T cell depletion in acquired immunodeficiency syndrome (AIDS) patients and several indirect mechanisms may be involved. A role has been proposed for apoptosis of uninfected lymphocytes, since lymphocytes from human immunodeficiency virus-1+ (HIV-1) individuals display increased levels of spontaneous apoptosis. This process may be ascribed in part to cell exhaustion by the chronic uncontrolled infection, but can also be directly induced by viral components, such as gp120, tat or nef. A key role is played by the death receptor Fas, but a role can also be played by other death receptors, such as the TNF and TRAIL receptors. By contrast, death of HIV-infected cells seems to be Fas-independent and driven by other viral components such as vpr and HIV proteases. A further role may be played by depletion of CD4+ T cell itself and hence the withdrawal of survival factors such as cytokines. Different ability of HIV strains to induce death of infected and uninfected cells might play a role in the clinical and biological differences displayed by HIV strains. A further variability may be ascribed to the intrinsic resistance of cells to apoptosis, which may depend on the individual genetic background or the use of drugs inhibiting apoptosis. The observation that when progression of HIV infection is slow due to "apoptosis-resistant" genetic backgrounds of the patients, or defective HIV-1 strains, or successful highly active antiretroviral therapy (HAART), generally also T cell apoptosis is low, suggests that HIV-infected subjects may benefit from therapies aimed to inhibit Fas function and/or spontaneous apoptosis.

The Impact of Human Allelic Variation on HIV-1 Disease

Abstract: Human allelic variants influence the susceptibility to HIV-1 infection and/or the subsequent rates of disease progression towards AIDS that average ten years, although they vary greatly among infected subjects. In this respect, studies involving multiply exposed persons who remain uninfected, long-term nonprogressors (who remain asymptomatic for fifteen years or more) or, in contrast, rapid progressors (who develop AIDS within two to three years post-infection) as well as seroincident cohorts of patients with defined seroconversion dates have contributed to our comprehension of the effects of different natural human polymorphisms on HIV-1 disease. The current article aims at providing an up-to-date review on these polymorphisms that may be broadly classified into three general categories: (1) those that control viral entry into susceptible cells (namely, chemokine and chemokine receptor polymorphisms), (2) mutational variants of genes involved in immune regulation, such as interleukin-10 (IL-10), interleukin-4 (IL-4), tumor necrosis factor-alpha (TNF-alpha), and mannose-binding lectin (MBL), and (3) polymorphisms in genes involved in the adaptive immune recognition by T cells, [human leukocyte antigen (HLA) type]. Particular emphasis has been placed on the state-of-the-art biotechnological methodologies, such as "spectral genotyping" that utilizes molecular beacons in conjunction with polymerase chain reaction in real-time (real-time-PCR), which were developed to assist with the characterization of some of these determinants. Elucidating the functional role of these factors via the application of such biotechnological assays is expected to further enhance our understanding of the pathogenesis of HIV-1 infection, and, eventually, to enrich our therapeutic arsenal with novel antiviral agents or strategic approaches.

HIV capsid assembly.

Abstract: HIV Gag assembly is the first and most essential step in the formation of virus particles. Following protein synthesis, Gag relocates from ribosomes and forms a virus particle at the plasma membrane, using host factors and machinery. Early studies focused on mapping the regions within Gag required for assembly and identified three distinct domains (M, I, and L), although their precise locations within the three-dimensional structure of Gag awaited later study. In this review, I summarize the mapping results in the light of recent progress on Gag structures made by nuclear magnetic resonance and X-ray crystallography as well as further functional analysis. These data are largely consistent and provide sufficient information for an understanding of the interactions and functions of the assembly domains at a macromolecular level. Current studies have moved on to the identification of the host factors and machinery used in the process of Gag assembly. Cumulative data suggest that the dynamics of Gag assembly and transport are achieved not by simply using, but rather by taking control of, cellular machinery. Key area in the process include interactions with TSG101, L domain receptor which normally functions in the endosomal sorting pathway and with lipid rafts, a type of M domain receptor, which has been suggested to be the sites for effective concentration of Gag. The review provides a summary of these data and discusses the likely direction of future studies.

Use of fusion proteins and procaryotic display systems for delivery of HIV-1 antigens: development of novel vaccines for HIV-1 infection.

Abstract: Two non-pathogenic scaffolds (represented by the filamentous bacteriophage fd and the dihydrolipoyl acetyltransferase E2 protein of the Bacillus stearothermophilus pyruvate dehydrogenase (PDH) complex) able to deliver human immunodeficiency virus (HIV)-1 antigenic determinants, were designed in our laboratories and investigated in controlled assay conditions. Based on a modification of the phage display technology, we developed an innovative concept for a safe and inexpensive vaccine in which conserved antigenic determinants of HIV-1 reverse transcriptase (RTase) were inserted into the N-terminal region of the major pVIII coat protein of bacteriophagefd virions. Analogously, we developed another antigen delivery system based on the E2 component from the PDH complex and capable of displaying large intact proteins on the surface of an icosahedral lattice. Our data show that both of these systems can deliver B and T epitopes to their respective presentation compartments in target cells and trigger a humoral response as well as a potent helper and cytolytic response in vitro and in vivo.

Sugar and Spice: Viral Envelope-DC-SIGN Interactions in HIV Pathogenesis

Abstract: DC-SIGN is a calcium dependent lectin that binds to HIV envelope, gp120, with high affinity. Its expression on dendritic cells, coupled with its ability to facilitate the binding and subsequent transfer of virions to permissive T-cells, has led to the hypothesis that DC-SIGN may serve as a conduit the transfer of HIV from the peripheral mucosa to secondary lymphoid organs. Studies have shown that DC-SIGN bound virions can maintain their infectivity for prolonged periods of time despite evidence that DC-SIGN itself may serve as an antigen receptor. How HIV subverts the normal function of DC-SIGN to establish a primary infection in the host is unclear. Therefore, understanding the structural and immunological basis for DCSIGNs function will help us realize the role that DC-SIGN may play in viral transmission and pathogenesis. Importantly, DC-SIGN / envelope interactions may represent a new target for microbicide and vaccine development efforts. Here, we review recent studies on DC-SIGNs structure and function in an effort to present testable models of DC-SIGNs role in HIV pathogenesis.

Resistance to Apoptosis: Mechanism for the Development of HIV Reservoirs

Abstract: New insights into the physiological cell death program in mammalian cells have further confounded the central issue of T lymphocyte destruction during HIV infection. Although infected and uninfected cells die following infection, recent evidence indicates that infected and uninfected cells may have unique pathways controlling death. Two widely accepted models for apoptosis have been described, namely the intrinsic and extrinsic apoptotic pathway. In the extrinsic pathway, ligation of TNF death ligands to their receptors causes oligomerization of the death receptors and recruitment of adaptor proteins typically involving caspase 8 activation. In the intrinsic pathway, apoptotic signals converge on mitochondria to cause activation and subsequent release of cytochrome c, which leads to formation of a multiprotein complex containing Apaf-1, cytochrome c, dATP and procaspase 9. Expression of HIV proteins alters these pathways resulting in enhanced levels of apoptosis. Although HIV infection results in T cell apoptosis, under some circumstances a small fraction of CD4+ T cells and macrophages do not die following infection, indicating that this may be a critical step in the development of viral reservoirs. In addition, monocyte differentiation into macrophages leads to an apoptosis resistant phenotype characterized by upregulation of antiapoptotic molecules and lower levels of proapoptotic molecules. The development of these stable antiretroviral resistant cells represents the major obstacle in achieving a complete sterile cure. However, this provides a unique opportunity to further understand the regulation of apoptosis and may facilitate novel immune based therapies aimed at modifying apoptosis in HIV disease.

The evolutionary adaptation of HIV-1 to specific immunity.

Abstract: Recent evidence of the evolutionary adaptation of HIV-1 to the specific immune system is reviewed. Longitudinal studies of patients show that a neutralizing antibody (NAb) response specific to autologous virus is detectable within 1-2 months of infection and that viral variants resistant to neutralization arise and spread in the viral population within the subsequent three months, and that this general pattern is repeated. There is strong evidence that amino acid replacements in gp120 glycan-binding motifs affect viral sensitivity to neutralization and are selected by NAbs. Longitudinal studies of humans have also provided good evidence of amino acid replacements in cytotoxic T lymphocyte (CTL) epitopes that allow the virus to escape detection by CTLs. But, the clearest evidence of adaptation to CTL surveillance at the molecular level comes from experiments with SIV-infected rhesus macaques. These show unequivocally that amino acid replacements in CTL epitopes are the result of positive selection and that these escape mutants have a lower class I major histocompatibility complex (MHC) binding affinity or are less likely to be recognized by CTLs than non-escape variants. To improve our ability to predict HIV's evolutionary responses to selection by the specific immune system it is suggested that future work focus on the details of the adaptive response to antibody surveillance, the temporal dynamics of specific immune responses, the relative importance of antibody and CTL selection, and the effects of superinfection, viral recombination, and viral protein functional constraints on immune escape.

HIV-1 Replication and Pathogenesis in the Human Thymus

Abstract: How HIV replicates and causes destruction of the thymus, and how to restore thymic function, are among the most important questions of HIV-1 pathogenesis and therapy in adult as well as pediatric patients. The thymus appears to function, albeit at reduced levels, throughout the life of adults, to respond to T cell depletion induced by HIV and to be suppressed by HIV. In this review, we summarize recent findings concerning HIV replication and pathogenesis in the human thymus, focusing on mechanistic insights gleaned from studies in the SCID-hu Thy / Liv mouse and human fetal-thymus organ culture (HF-TOC) models. First, we discuss HIV viral determinants and host factors involved in the replication of HIV in the thymus. Second, we consider evidence that both viral factors and host factors contribute to HIV-induced thymocyte depletion. We thus propose that multiple mechanisms, including depletion and suppression of progenitor cells, paracrine and direct lytic depletion of thymocytes, and altered thymocyte selection are involved in HIV-induced pathology in the thymus. With the SCID-hu Thy / Liv mouse and HF-TOC models, it will be important in the coming years to further clarify the virological, cell biological, and immunological mechanisms of HIV replication and pathogenesis in human thymus, and to correlate their significance in HIV disease progression.

T cell immunity to HIV: defining parameters of protection.

Abstract: In recent years, CD4 and CD8 T cell responses to HIV and SIV infection have been increasingly measured with the use of single-cell assays such as ELISPOT, MHC-peptide oligomers, and cytokine flow cytometry. The results of these assays have been compared to those obtained with traditional bulk assays such as lymphoproliferation (by 3H-thymidine incorporation) and cytotoxicity (by 51Cr release). Such comparisons have led to some general understanding of the T cell responses that characterize progressive disease, long-term non-progressors, and individuals with viral suppression achieved by anti-retroviral therapy. In addition, prophylactic and therapeutic vaccine trials have also begun to use these assays of T cell immunity to gauge the immunogenicity of the vaccines. Whether such analyses will allow us to pick the best vaccine constructs, and whether they will provide us with an improved understanding of what constitutes protective cellular immunity to HIV, are major questions for the field. These questions will be examined in this review from the standpoint of current data and comparisons to other viral diseases. It is hypothesized that sophisticated multiparametric assays will be required to sort out the factors relevant for protective immunity in this complex disease. These parameters may include functional avidity, epitope breadth and specificity, proliferative capacity, cytokine repertoire, degree of anergy, and differentiation phenotype, as well as magnitude, of HIV-specific CD4 and CD8 T cells.

HIV-1 vectors: fulfillment of expectations, further advancements, and still a way to go.

Abstract: The ability of lentiviral vectors to transduce and stably integrate their genomes into non-dividing cells was the major reason for the development of the HIV-1 based vector gene delivery system. The first VSV-G pseudotyped lentiviral vectors fulfilled these expectations by ferrying large genetic payloads to non-dividing cells in vitro and in vivo. Here we discuss advances in HIV-1 vector systems which lead to improvement in biosafety, transduction efficiency, longevity and regulation of transgene expression, and vector production. The successful use of the advanced HIV-1 based vector system opened new avenues in establishing transgenic animal models for basic research. Additionally, we describe accomplishments using HIV-1 based vectors to correct pathological courses of incurable diseases in preclinical animal models including Parkinson's disease and beta-thalassemia.

Is HIV Involved in the Pathogenesis of Non-Infectious Pulmonary Complications in Infected Patients?

Abstract: Pulmonary complications of AIDS are one of the major causes of morbidity and mortality in HIV-infected individuals, and are often direct indicators of enhanced progression to an AIDS-defining illness. HIV can be detected in the lung during asymptomatic disease, as well as during the advanced stages of AIDS. With prolonged survival of HIV-infected patients, non-infectious pulmonary complications are becoming more common and can lead to respiratory compromise and failure. The pathogenesis of non-infectious pulmonary complications, following HIV infection is not fully understood, and the role that HIV-induced immunosuppression plays in the development of these complications is not known. In this review, we will discuss non-infectious pulmonary complications of HIV infection, focusing on HIV-associated malignancies of the lung, inflammatory disorders, pulmonary hypertension and the effect of antiretroviral therapy on the incidence of these diseases. The possible role that HIV plays in the pathogenesis of these pulmonary disorders is also addressed.

HIV's evasion of host's NK cell response and novel ways of its countering and boosting anti-HIV immunity.

Abstract: NK cells were characterized by their ability to spontaneously kill certain tumor and virus-infected cells. They constitute first line of defense against invading pathogens and usually become activated in viral infections particularly in early phases. Activated NK cells play a crucial role in the induction and amplification of virus-specific immunity by providing IFN-gamma and "danger signal". The functional activities of NK cells are regulated by a balance between the engagement of their inhibitory and activating receptors. In recent years, the discovery of several MHC and non-MHC binding NK receptors has provided important insights regarding NK cell biology and its role in viral infections. These receptors are increasingly being viewed as important regulators of immune response. Like many other viruses, HIV also seems to activate NK cells. However, several studies have reported compromised NK cell functions in HIV-infected individuals. The virus employs several strategies to counter the host's NK cell response, e.g., a differential downregulation of MHC class I molecules on the surface of infected cells, a dysregulated production of NK cell function-enhancing cytokines, direct inhibitory effects of certain viral proteins on NK cell functions, and changes in the expression of NK cell receptors, etc. The individuals expressing activating NK cell receptors and their cognate MHC ligands have activated NK cells. The development of AIDS is significantly delayed in these individuals after HIV infection. The discovery of NK receptors and their ligands has opened new avenues of developing AIDS vaccine and boosting innate and adaptive antiviral immunity in HIV-infected individuals.