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HIV infection and disease is a multistage process that involves a variety of cell types as the virus spreads through the body. Initially, dendritic cells (DCs) present at the mucosal site of infection bind and internalise HIV for degradation and presentation to T cells. As the DCs migrate to lymph nodes and mature, part of the internalised virions remains infective inside endosomal compartments. During formation of the immunological synapse between CD4 T cells and DCs, infective virions from dendritic cells are transferred to CD4 T cells leading to a strong infection of those cells allowing rapid virus dissemination throughout the body and establishment of the typical HIV infection. Various membrane receptors are involved in this process. Initial HIV binding to DCs is mediated by C-type lectin receptors such as the mannose receptor or DC-SIGN (DC specific intracellular adhesion molecule 3 grabbing non integrin) which is followed by virus internalisation and lysis albeit virus induced changes in endocytic routing prevents a proportion from degradation. Productive infection of DCs has also been observed allowing trans infection of CD4 T cells through a different mechanism. HIV infection of CD4 T cells, DCs and other cells is a multistep process initiated by binding of HIV envelope gp120 to the CD4 receptor, a 55 kDa transmembrane glycoprotein. Subsequent conformational changes in gp120 allow binding to a chemokine receptor, either CCR5 or CXCR4, followed by membrane fusion and infection. The aim of this thesis was to investigate protein associations with the HIV receptors DC-SIGN and CD4 in order to elucidate the mechanism of complex formation, virus entry and/or defining target sites for antiretroviral drugs. This thesis used a proteomic approach for studying the receptors with mass spectrometry-based protein identification as its core technology. A range of different approaches were developed and compared for identification of protein interactions and characterisation of the identified protein associations. An affinity purification of the CD4 receptor complex from lymphoid cells was used as the basis for detecting novel CD4-binding proteins. For this approach a strategy based on mass spectrometry identification of CD4 associating proteins using affinity chromatography and affinity-tag mediated purification of tryptic peptides was developed. This method proved successful for the identification of CD4 interacting proteins such as the strongly associated kinase p56lck, however a limited number of non-specifically bound proteins were also identified along the receptor complex. Using one-dimensional SDS-polyacrylamide gel electrophoresis followed by in-gel digests and mass spectrometry analysis, a large number of non-specifically binding proteins were identified along the CD4/lck complex. Evaluation of different lysis buffers in several independent experiments demonstrated that there was a large and inconsistent array of proteins that were obviously non-specifically bound to the receptor. No further specific binding partners were detected. These data suggested that protein interactions of CD4 on this cell type are of weak and/or transient nature. It also demonstrated a need for careful interpretation of proteomic data in the light of the propensity of non-specific binding under these conditions. To overcome dissociation of weak protein interactions, a method was developed using chemical cross-linking to preserve weak protein interactions on lymphoid cells. Affinity purification was used to purify CD4 along with cross-linked associated proteins and mass spectrometry analysis identified an interaction with the transferrin receptor CD71 and the tyrosine phosphatase CD45. The CD45-CD4 interaction is well known. The CD4-CD71 interaction was demonstrated to be a result from colocalization of the two molecules during formation of endocytic vesicles. Flow cytometry-based fluorescence resonance energy transfer (FRET) measurements were applied to confirm colocalization. A similar interaction was suspected for CD4 and DC-SIGN on the plasma membrane of DCs as cis infection of DCs has been demonstrated i.e. initial binding to DC-SIGN then to CD4/CCR5 on the same cell. Therefore, protein associations of DC-SIGN were investigated using the developed techniques. Using cross-linking, DC-SIGN was shown to assemble in large complexes on the surface of immature monocyte-derived DCs. Mass spectrometry analysis of the purified complexes identified them as homo-oligomers of DC-SIGN. The absence of CD4 suggested that the fraction interacting with CD4 at any one time must be small. The complexes of DC-SIGN were further characterised to be tetramers and successfully co-immunoprecipitated with HIV gp120 and mannan. DC-SIGN monomers were not evident demonstrating that the assembly of DC-SIGN into tetramers is required for high affinity binding of its natural and viral ligands. Thus potential antiviral agents aimed at blocking the early stage of HIV binding to DCs must simulate tetramers in order to neutralise the virus efficiently. Overall the thesis provides new information on protein interactions of CD4 and DC-SIGN, a careful investigation of "proteomics" techniques for identifying the proteins in affinity-purified samples and demonstrates the need for multifaceted analytical approaches to probe complex cellular systems.

Erlangen, Nürnberg, University, Diss., 2007. / Enth. 1 Sonderabdr. aus: AIDS ; 19. 2005.

Zugl.: Zürich, Hochsch. für Soziale Arbeit, Diplomarbeit, 2007

<p>Cirka 42 miljoner människor lever med humant immunbristvirus (HIV) i världen. Elva miljoner av dessa är ungdomar som är den grupp där HIV ökar mest. Syftet med denna uppsats var att belysa faktorer som påverkade ungdomars riskbeteende gällande HIV. Uppsatsen gjordes som en litteraturstudie med tolv vetenskapliga artiklar. Analysen resulterade i tre kategorier: ungdomars risktagande, personliga erfarenheter samt kunskap och information. I resultatet framkom att ungdomar fattade flera risktagande beslut som ökade det sexuella riskbeteendet gällande HIV. Det framkom faktorer som bristande kondomanvändning, alkohol och droger.  Personliga erfarenheter visade sig påverka ungdomarna både positiv och negativt, beroende på uppfostran och upplevelser. Bristande kunskap och information var en faktor som kunde påverkade ungdomarna negativt. I diskussionen togs sjuksköterskans roll upp. Genom förmåga att kunna integrera i ungdomskulturen identifierades problem och lösningar frambringades. Sjuksköterskan kunde informera ungdomar om HIV i samband med hälsoundervisning. Det var viktigt att fånga ungdomarnas intresse. Slutsatsen var att kunskap hos ungdomar var viktigt för att förhindra smittspridningen av HIV. Sjuksköterskan hade en viktig roll i arbetet mot HIV med ungdomar.</p>

The Human Immunodeficiency Virus -1 (HIV-1) has been the source of substantial human misery since it was first discovered in the early 1980s. Despite remarkable progress that has been made towards understanding HIV- 1, there is no cure, no vaccine and life-prolonging therapies are beyond the reach of millions in need. Our research sought to both gain insight into potential therapeutic targets as well as the preclinical testing of drug candidates. We demonstrate that a RhoA derived peptide is an effective inhibitor of HIV-1 entry. Although this peptide inhibits HIV-1 due to its poly-anionic nature, it nevertheless demonstrates that endogenous host proteins may be repurposed for novel therapeutic uses. We also characterized the mechanism and effectiveness of Basant, a polyherbal topical microbicide candidate for the prevention of HIV-1 transmission. Our data demonstrate that it inhibits the entry of both CCR5 and CXCR4 tropic HIV-1 at non-toxic concentrations. Finally, data is presented on the characterization of a novel HIV-1 protein expressed from an alternative reading frame of the HIV-1 polymerase gene. We demonstrate that this protein is localized to the nucleolus and is likely expressed from a novel HIV-1 transcript. This work lays the foundation for further studies to target this protein for drug development. / Graduation date: 2011 / Access restricted to the OSU Community at author's request from May 9, 2011 - May 9, 2012

There is a real relationship between the HIV/AIDS pandemic and the world of work. The complexities in the relationship are strengthened by the long asymptomatic period of the disease and its significant presence among a certain age group that forms a major part of the work force. The presence of HIV/AIDS in a workplace, if not effectively managed, leads to unpleasant effects such as an increase in the cost of production, which translates into income loss and capital depletion. Superficial knowledge of these factors often makes the workplace hostile and leads to a culture of discrimination in which the well-being and rights of those affected by the virus are either undermined or out rightly ignored. An effective way of mitigating these negative effects is the provision of workplace reasonable accommodation measures. This paper proposes pragmatic legal ways of meeting the reasonable accommodation needs of people living with HIV/AIDS in Canada.

Women infected with HIV are at an increased risk for infection of Human Papillomavirus (HPV), developing cervical lesions, and developing cervical cancer. Prior research has suggested disruptions in the immune system as well as circulating levels of stress and gonadal hormones as possible explanations for the increase of HPV infection in women with HIV. Additionally, psychosocial factors such as symptoms of depression and distress have also been associated with HPV infection, as well as disruptions in immune and endocrinologial systems, suggesting a psychoneuroimmunological pathway to disease progression. It was hypothesized that HIV+HPV+ women assigned to a Cognitive Behavioral Stress Management (CBSM) intervention will experience improvements in disease status, immune markers, circulating stress hormones, and reductions of depression and distress symptoms. An exploratory investigation of the effects of CBSM in levels circulating reproductive hormones was also tested. Follow-up hypotheses tested whether CBSM effects on immune variables were explained by reductions in symptoms of depression, distress, NE, cortisol, and increases of DHEA-S. Additionally, it was hypothesized that CBSM effects on stress hormones would be mediated by reductions in distress and depression symptoms. Finally, it was hypothesized that improvements in immune parameters would be correlated with decreases in risk of cervical dysplasia at a 9 month follow-up. Participants were 71 women co-infected with HIV and HPV that were mostly of African American, Haitian, Latina, and Caribbean descent. Hierarchical regression analyses were performed and showed a significant CBSM effect in decreases on BDI somatic depression subscale scores and increases in NK cell counts. Additionally, there was a marginally significant effect of CBSM on increases in CD4+ T-cells and decreases in urinary NE output. The bootstrapping method evidenced a mediation model, where the relationship between group assignment and CD4+ cell counts was explained by lower BDI somatic scores. More research is necessary to fully elucidate the psychobiological trajectories of disease as immunological changes in our sample did not explain the reduced odds of dysplasia in the women assigned to the CBSM group.

Cirka 42 miljoner människor lever med humant immunbristvirus (HIV) i världen. Elva miljoner av dessa är ungdomar som är den grupp där HIV ökar mest. Syftet med denna uppsats var att belysa faktorer som påverkade ungdomars riskbeteende gällande HIV. Uppsatsen gjordes som en litteraturstudie med tolv vetenskapliga artiklar. Analysen resulterade i tre kategorier: ungdomars risktagande, personliga erfarenheter samt kunskap och information. I resultatet framkom att ungdomar fattade flera risktagande beslut som ökade det sexuella riskbeteendet gällande HIV. Det framkom faktorer som bristande kondomanvändning, alkohol och droger.  Personliga erfarenheter visade sig påverka ungdomarna både positiv och negativt, beroende på uppfostran och upplevelser. Bristande kunskap och information var en faktor som kunde påverkade ungdomarna negativt. I diskussionen togs sjuksköterskans roll upp. Genom förmåga att kunna integrera i ungdomskulturen identifierades problem och lösningar frambringades. Sjuksköterskan kunde informera ungdomar om HIV i samband med hälsoundervisning. Det var viktigt att fånga ungdomarnas intresse. Slutsatsen var att kunskap hos ungdomar var viktigt för att förhindra smittspridningen av HIV. Sjuksköterskan hade en viktig roll i arbetet mot HIV med ungdomar.

Tuberculosis (TB) is a preventable and a treatable disease yet it is considered to be one of the most common infections seen in HIV. People who are infected with HIV are 20 times more likely to develop TB than those without HIV. Globally, there are nearly 40 million people living with HIV and at least one-third of them are infected with TB. Ontario accounts for the highest number of TB cases in Canada yet HIV-TB co-infection in Ontario is not well described. Despite the close relationship between TB and HIV and increasing efforts to fight both concurrently, TB continues to create economic and social burden in HIV infections. Our study estimates the prevalence of active and latent TB and identifies risk factors associated with TB in a cohort of individuals living with HIV in Ontario. Cases diagnosed with HIV during 2001 to 2009 were extracted from the Ontario HIV Treatment Network Cohort Study (OCS). Reviewing Mantoux test results, diagnoses and medication history, identified active and latent TB cases. Period prevalence was estimated by proportion with TB and multivariate analyses were performed to identify associated factors. One thousand two hundred and ninety-three cases (1293) met our selection criteria. Three hundred and eighty four (384; 29.7%) were 29 years or younger, 805 (62.3%) aged between 30 years and 50 years and 104 (8.0%) aged 50 years or older. One thousand and nine (1009; 78.0%) were males. Four hundred and sixty six (466; 36.0%) had at least one record of a Mantoux skin test. The prevalence of active TB was 76/1293 = 0.0587 or 5.87% (95% CI 4.6% to 7.0%) whereas the prevalence of latent TB varied from 5.26% (68/1293 = 0.0526) 95% (CI 4.0% - 6.5%) to 11.37% (53/466 = 0.1137) 95% CI (8.2% to 13.7%) depending on the methodology. In the multivariate analysis, factors associated with active TB were age and birthplace. Individuals 50 years and older were more likely to have active TB than individuals 30 years and younger (OR 4.3 CI (1.7-12.7), p <0.01). Individuals born in Africa were more likely to have active TB than Canadian born (OR 14; 95% CI (5.9 – 32.8) p < 0.001). Factors associated with latent TB were sex and birthplace. Females were more likely to have latent TB than males (OR 2.4; 95% CI (1.1 – 5.2) p < 0.05). Individuals born in Africa were more likely to have latent TB than Canadian born (OR 12.3; 95% CI (4.7 – 32.1) p < 0.001). TB remains a major problem in persons infected with HIV with rates disproportionally high among the foreign born population. Low rates of Mantoux tests in OCS present a missed opportunity for active TB prevention among individuals with HIV. To identify individuals with higher risk of having TB after HIV diagnosis, better screening tools to identify latent TB are needed. Consideration should be given to data capture systems that would ideally be linked between Public Health and HIV clinics. / UOIT