HIV-1 increases extracellular amyloid-beta levels through neprilysin regulation in primary cultures of human astrocytes.

Since the success of combined antiretroviral therapy, HIV-1-infected individuals are now living much longer. This increased life expectancy is accompanied by a higher prevalence of HIV-1 associated neurocognitive disorders. Rising too is the incidence in these patients of pathological hallmarks of Alzheimer's disease such as increased deposition of amyloid beta protein (Aß). Although neurons are major sources of Aß in the brain, astrocytes are the most numerous glial cells, therefore, even a small level of astrocytic Aß metabolism could make a significant contribution to brain pathology. Neprilysin (NEP) is a decisive/crucial regulator of Aß levels. We evaluated the effects of HIV-1 on Aß deposition and the expression and activity of NEP in primary human astrocytes. Specifically, no differences in intracellular amyloid deposits were found between infected and control cells. However, primary cultures of infected astrocytes showed more extracellular Aß levels compared to controls. This was accompanied by reduced expression of NEP and to a significant decrease in its activity. These results indicate that the presence of HIV-1 in the brain could contribute to the increase in the total burden of cerebral Aß.

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

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

Cyclosporine A in addition to standard ART during primary HIV-1 infection: pilot randomized clinical trial.

Background: Initiating ART during acute/recent HIV-1 infection reduces viral reservoir formation. It has been proposed that, during this phase, the size of the viral reservoir could be further reduced by the association of immunomodulatory therapy with ART. Contradictory results have emerged, however, from two trials evaluating the impact on immune recovery and the viral reservoir of adding cyclosporine A to ART during primary HIV-1 infection. Patients and methods: Twenty patients with acute/recent HIV-1 infection were randomized to receive ART alone (tenofovir, emtricitabine and lopinavir/ritonavir) or associated with 8 weeks of cyclosporine A (0.3-0.6 mg/kg twice daily). The impact on viral load, immune response and integrated and non-integrated DNA viral reservoir at 0, 8 and 36 weeks of treatment was evaluated. Results: The estimated median time from HIV-1 infection to ART onset was 63 days (IQR 53; 79.5) with 90% of patients at Fiebig V stage. No significant differences were observed in viral load decay, CD4 T cell recovery, immune response markers or the evolution of integrated DNA at week 8 (end of cyclosporine A) and week 36 between groups. However, non-integrated DNA significantly increased in the cyclosporine A arm between weeks 0 and 36. Cyclosporine A was well tolerated. Conclusions: Adding cyclosporine A to ART during acute/recent infection did not improve immune recovery. However, unintegrated DNA increased in the cyclosporine A group, suggesting an anti-integration effect, a point warranting further research (ClinicalTrials.gov Identifier: NCT00979706).

CCR5/CD4/CXCR4 oligomerization prevents HIV-1 gp120IIIB binding to the cell surface.

CCR5 and CXCR4, the respective cell surface coreceptors of R5 and X4 HIV-1 strains, both form heterodimers with CD4, the principal HIV-1 receptor. Using several resonance energy transfer techniques, we determined that CD4, CXCR4, and CCR5 formed heterotrimers, and that CCR5 coexpression altered the conformation of both CXCR4/CXCR4 homodimers and CD4/CXCR4 heterodimers. As a result, binding of the HIV-1 envelope protein gp120IIIB to the CD4/CXCR4/CCR5 heterooligomer was negligible, and the gp120-induced cytoskeletal rearrangements necessary for HIV-1 entry were prevented. CCR5 reduced HIV-1 envelope-induced CD4/CXCR4-mediated cell-cell fusion. In nucleofected Jurkat CD4 cells and primary human CD4(+) T cells, CCR5 expression led to a reduction in X4 HIV-1 infectivity. These findings can help to understand why X4 HIV-1 strains infection affect T-cell types differently during AIDS development and indicate that receptor oligomerization might be a target for previously unidentified therapeutic approaches for AIDS intervention.

Dendronized Anionic Gold Nanoparticles: Synthesis, Characterization, and Antiviral Activity.

Anionic carbosilane dendrons decorated with sulfonate functions and one thiol moiety at the focal point have been used to synthesize water-soluble gold nanoparticles (AuNPs) through the direct reaction of dendrons, gold precursor, and reducing agent in water, and also through a place-exchange reaction. These nanoparticles have been characterized by NMR spectroscopy, TEM, thermogravimetric analysis, X-ray photoelectron spectroscopy (XPS), UV/Vis spectroscopy, elemental analysis, and zeta-potential measurements. The interacting ability of the anionic sulfonate functions was investigated by EPR spectroscopy with copper(II) as a probe. Different structures and conformations of the AuNPs modulate the availability of sulfonate and thiol groups for complexation by copper(II). Toxicity assays of AuNPs showed that those produced through direct reaction were less toxic than those obtained by ligand exchange. Inhibition of HIV-1 infection was higher in the case of dendronized AuNPs than in dendrons.

Improved Efficiency of Ibuprofen by Cationic Carbosilane Dendritic Conjugates.

In order to improve the efficiency of the anti-inflammatory drug ibuprofen, cationic carbosilane dendrimers and dendrons with ibuprofen at their periphery or at their focal point, respectively, have been synthesized, and the release of the drug was studied using HPLC. Macrophages were used to evaluate the anti-inflammatory effect of the ibuprofen-conjugated dendritic systems and compared with mixtures of non-ibuprofen dendritic systems in the presence of the drug. The cationic ibuprofen-conjugated dendron was the compound that showed higher anti-inflammatory properties. It reduces the LPS-induced COX-2 expression and decreases the release of several inflammatory cytokines such as TNFα, IL-1ß, IL-6, and CCL3. These results open new perspectives in the use of these compounds as drug carriers.

Polyanionic carbosilane dendrimer-conjugated antiviral drugs as efficient microbicides: Recent trends and developments in HIV treatment/therapy.

Polyanionic carbosilane dendrimers (PCDs) are potential candidates for the development of new microbicides for the prevention of HIV transmission. Tenofovir (TFV), which has dual antiviral activity (anti-HIV/HSV-2), and maraviroc (MRV) are the most studied antiretrovirals as microbicides. Here, we introduce developments in the design of innovative dendrimer-based microbicides. We also review and discuss the combination of various PCDs with TFV and/or MRV for their anti-HIV-1 activity and synergistic combinatory potential. Well-defined combinations blocking HIV-1 infection in early steps of HIV-1 replication provide greater efficacy than monotherapy, as reflected by the decrease in concentration and increase in HIV-1 inhibition. These combinations are characterized by lower doses, which minimize toxic side-effects and the emergence of multi-drug resistant mutants. The above facts suggest that the combination of first- and second-generation PCDs with TFV and/or MRV represents a promising candidate microbicide for preventing HIV-1 sexual transmission and simultaneously suppressing HSV-2. FROM THE CLINICAL EDITOR: HIV infection remains a significant and unresolved problem for humankind, despite the development of combination antiretroviral therapy. It has been found that polyanionic carbosilane dendrimers have efficacy in preventing HIV transmission. In this comprehensive review article, the authors discuss the current status and latest development of the use of dendrimers in combination with other antiretroviral drugs as microbicides, which should stimulate others into further research in the fight against HIV.

Prevention vaginally of HIV-1 transmission in humanized BLT mice and mode of antiviral action of polyanionic carbosilane dendrimer G2-S16.

The development of a safe, effective, and low-priced topical microbicide to prevent HIV-1 sexual transmission is urgently needed. The emerging field of nanotechnology plays an important role in addressing this challenge. We demonstrate that topical vaginal administration of 3% G2-S16 prevents HIV-1JR-CSF transmission in humanized (h)-BLT mice in 84% with no presence of HIV-1 RNA and vaginal lesions. Second-generation polyanionic carbosilane dendrimer G2-S16 with silica core and 16 sulfonate end-groups exerts anti-HIV-1 activity at an early stage of viral replication, blocking the gp120/CD4 interaction, acting on the virus, and inhibiting the cell-to-cell HIV-1 transmission, confirming its multifactorial and non-specific ability. This study represents the first demonstration that transmission of HIV-1 can be efficiently blocked by vaginally applied G2-S16 in h-BLT mice. These findings provide a step forward in the development of G2-S16-based vaginal microbicides to prevent vaginal HIV-1 transmission in humans. FROM THE CLINICAL EDITOR: HIV infections remain a significant problem worldwide and the major route of transmission is through sexual activity. In this article, the authors developed an antiviral agent containing polyanionic carbosilane dendrimer with silica core and 16 sulfonate end-groups. When applied vaginally, this was shown to exert anti-HIV protection. These positive findings may offer hope in the fight against the spread of HIV epidemic.

Synergistic activity profile of carbosilane dendrimer G2-STE16 in combination with other dendrimers and antiretrovirals as topical anti-HIV-1 microbicide.

Polyanionic carbosilane dendrimers represent opportunities to develop new anti-HIV microbicides. Dendrimers and antiretrovirals (ARVs) acting at different stages of HIV replication have been proposed as compounds to decrease new HIV infections. Thus, we determined the potential use of our G2-STE16 carbosilane dendrimer in combination with other carbosilane dendrimers and ARVs for the use as topical microbicide against HIV-1. We showed that these combinations obtained 100% inhibition and displayed a synergistic profile against different HIV-1 isolates in our model of TZM.bl cells. Our results also showed their potent activity in the presence of an acidic vaginal or seminal fluid environment and did not activate an inflammatory response. This study is the first step toward exploring the use of different anionic carbosilane dendrimers in combination and toward making a safe microbicide. Therefore, our results support further studies on dendrimer/dendrimer or dendrimer/ARV combinations as topical anti-HIV-1 microbicide. FROM THE CLINICAL EDITOR: This paper describes the first steps toward the use of anionic carbosilane dendrimers in combination with antivirals to address HIV-1, paving the way to further studies on dendrimer/dendrimer or dendrimer/ARV combinations as topical anti-HIV-1 microbicides.

[Psychosocial aspects in a cohort of vertically transmitted human immunodeficiency virus-infected adolescents]. / Aspectos psicosociales en una cohorte de adolescentes con infección por el virus de la inmunodeficiencia humana por transmisión vertical. NeuroCoRISpeS.

INTRODUCTION: Thanks to advances in antiretroviral treatment, children with HIV infections through vertical transmission have improved their life expectancy. However, new challenges have emerged. We propose this study in order to determine the psychosocial aspects and knowledge of infections in a cohort of adolescents with vertically transmitted HIV infections. METHODS: Patients with vertically-acquired HIV infection between 12 and 19 years old were included. Data were obtained through semi-structured interviews and a Strengths and Difficulties Questionnaire for emotional and behavioral disorders screening. RESULTS: We evaluated 96 patients (58% females) with a median age of 15 years (11-19.1) and a median age at diagnosis of 1.70 years (0-12.2). The median CD4 count was 626cells/mm(3) (132-998), and the viral load was<50cp/ml in 72% of patients. Among them, 90% attended school and 60% repeated at least one course. Although 81% of them knew of their diagnosis, only 30% understood their disease, with 18.2% having discussed it with friends. Six unwanted pregnancies occurred during the study period. Strengths and Difficulties Questionnaire showed hyperactivity risk in 33%. CONCLUSION: A high percentage of adolescents show difficulties in several areas (disease knowledge, peer relationship, school failure...) that can have an impact on their adult lives. Further studies are needed to evaluate their origin and development in depth, as well as interventions to modify this situation.

HIV-1 infection and neurocognitive impairment in the current era.

Brain HIV-1-infection may result in a syndrome of profound cognitive, behavioral and motor impairment known as AIDS dementia complex (ADC) in adults and HIV-related encephalopathy in children. Although the introduction of highly active antiretroviral therapy (HAART) has prolonged and improved the lives of infected individuals, it is clear that HAART does not provide complete protection against neurological damage in HIV/AIDS. HIV-1 associated dementia is a complex phenomenon, which could be the result of several mechanisms caused by those players using different intracellular signaling pathways. Understanding the causes of neurodegeneration during HIV-1 infection and the factors which certain individuals develop disease can provide researches on new therapeutic targets to positively affect disease outcomes. Controlling CNS viral replication with HAART is an essential primary approach, but it should be complemented with adjunctive CNS-directed therapeutics. Understanding the nature of HIV-1 infection within the CNS as well as inflammatory responses will ultimately lead to the elimination of HIV-associated neurocognitive disorders.

Carbosilane dendrimer 2G-NN16 represses Tc17 differentiation in primary T CD8+ lymphocytes.

We studied changes in gene expression induced by the carbosilane dendrimer 2G-NN16 to evaluate their potential as a vehicle for gene therapy and as medication. Global gene expression profiles on CD8+ T lymphocytes reveal that ribosomal proteins are induced in the presence of 2G-NN16. IL17A and IL17F, the principal interleukins secreted by Tc17 cells, a subset of CD8+ T lymphocytes, were down-regulated when cultured in the presence of this dendrimer. Microarray results were confirmed by real time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). 2G-NN16 also showed a high potential for in vitro inhibition of Tc17 differentiation of CD8+ T lymphocytes in the presence of the Tc17 differentiation molecules IL6 and TGF-B1. These findings suggest that 2G-NN16 could facilitate drug delivery and may be used to treat inflammatory processes driven by Tc17 cells.

Preterm neonates show marked leukopenia and lymphopenia that are associated with increased regulatory T-cell values and diminished IL-7.

INTRODUCTION: Current advances in neonatology have improved survival among preterm and low-birth-weight infants. However, the risk of neonatal death in preterm infants is much greater than in full-term neonates and is frequently associated with infections. METHODS: Little is known about the immune status of preterm neonates; therefore, we analyzed the frequency and absolute counts of different immune populations in 211 cord blood samples taken from very-preterm to full-term neonates. RESULTS: We found that absolute counts of all the immune subsets analyzed (i.e., monocytes, granulocytes, B cells, natural killer (NK) cells, CD4(+), and CD8(+) T cells) were markedly lower in preterm infants than in full-term infants. Surprisingly, we observed that regulatory T cells (Tregs) were the only cell subset that did not decrease in preterm infants, and their frequency was even higher than in full-term infants. DISCUSSION: Tregs are crucial to maternal-fetal tolerance, but their suppressive role could be also implicated in the leukopenia observed in preterm infants. We did not observe differences in thymic function, but we found that plasma levels of interleukin (IL)-7 and the frequency of its receptor were significantly decreased in preterm infants. Our results could help to identify leukopenia and to implement immune therapies that significantly diminish mortality in preterm neonates.

Sustained virological response to interferon-α plus ribavirin decreases inflammation and endothelial dysfunction markers in HIV/HCV co-infected patients.

OBJECTIVES: Hepatitis C virus (HCV) antiviral therapy might lead to decreased chronic immune activation and endothelial dysfunction associated with cardiovascular risk. The aim was to evaluate the effect of HCV eradication on serum markers of inflammation and endothelial dysfunction markers in HIV/HCV co-infected patients. METHODS: We carried out a retrospective study of 69 HIV/HCV co-infected patients on interferon (IFN)-α plus ribavirin. In addition, 47 HIV-infected subjects were selected as a control group. A sustained virological response (SVR) was defined as an undetectable HCV viral load up to 24 weeks after the end of treatment. Tumour necrosis factor (TNF) receptor-1 (TNF-R1), soluble E-selectin (sE-selectin), soluble P-selectin (sP-selectin), soluble intercellular adhesion molecule 1 (sICAM-1) and soluble vascular cell adhesion molecule 1 (sVCAM-1) were measured using a multiplex immunoassay kit. RESULTS: HIV/HCV co-infected patients had higher values of soluble TNF-R1 (sTNF-R1), sE-selectin and sICAM-1 than HIV mono-infected patients (P < 0.05). SVR patients had a decrease in sTNF-R1, sP-selectin, sE-selectin and sICAM-1 during anti-HCV treatment (P < 0.05) and, at the end of treatment, SVR patients had lower values of sTNF-R1, sE-selectin and sVCAM-1 than non-responder patients (P < 0.05), although the values of sTNF-R1, sP-selectin, sE-selectin and sICAM-1 remained higher than in HIV mono-infected patients (P < 0.05). Moreover, we found a significant positive relationship between an increase in sTNF-R1 and increases in sP-selectin, sE-selectin and sICAM-1 during anti-HCV therapy. CONCLUSIONS: Chronic hepatitis C infection induces alterations of markers of inflammation and endothelial dysfunction. Eradication of HCV, following IFN-α and ribavirin therapy, reduces immune activation as well as markers of inflammation and endothelial dysfunction.

High plasma fractalkine (CX3CL1) levels are associated with severe liver disease in HIV/HCV co-infected patients with HCV genotype 1.

BACKGROUND: Inappropriate persistence of chemokines expression in hepatitis C virus (HCV) infection can drive tissue damage, intrahepatic inflammation, and liver cell injury. The aim of study was to study the association of plasma fractalkine (CX3CL1) levels with fibrosis stage and necroinflammatory activity grade of liver biopsies in human immunodeficiency virus (HIV)/HCV co-infected patients with HCV genotype 1. METHODS: We carried out a cross-sectional study on 125 patients. Grading and staging of liver biopsies were carried out by METAVIR score. Plasma CX3CL1 was measured using an immunoassay kit. RESULTS: Patients with advanced fibrosis had higher CX3CL1 levels than those with mild or no fibrosis (p=0.010); and patients with severe activity grade had higher CX3CL1 levels than those with low activity grade (p=0.040). Plasma CX3CL1 levels were significantly associated with increased odds of significant fibrosis (odds ratio (OR): 3.47 (95% of confidence interval (95%CI): 1.04; 11.58)), advanced fibrosis (OR: 6.78 (95%CI: 1.70; 26.93)), and moderate necroinflammatory activity grade (OR: 4.09 (95%CI: 1.21; 13.87)). When we analyzed fibrosis stages and activity grades of METAVIR score together, we found a positive significant association of CX3CL1 levels with moderate activity grade/significant fibrosis (OR: 5.49 (95%CI: 1.46; 20.58)) and moderate activity grade/advanced fibrosis (OR: 8.99 (95%CI: 2.06; 39.23)). CONCLUSION: Plasma CX3CL1 levels were independently associated with several characteristics of severe liver disease in HIV/HCV coinfected patients with HCV-genotype 1, suggesting a role of CX3CL1 in the pathogenesis of HCV infection.

BMS Derivatives C7-Linked to ß-Cyclodextrin and Hyperbranched Polyglycerol Retain Activity against R5-HIV-1NLAD8 Isolates and Can Be Deemed Potential Microbicides.

Amides from indole-3-glyoxylic acid and 4-benzoyl-2-methylpiperazine, which are related to entry inhibitors developed by Bristol-Myers Squibb (BMS), have been synthesized with aliphatic chains located at the C7 position of the indole ring. These spacers contain an azido group suitable for the well-known Cu(I)-catalyzed (3+2)-cycloaddition or an activated triple bond for the nucleophilic addition of thiols under physiological conditions. Reaction with polyols (ß-cyclodextrin and hyperbranched polyglycerol) decorated with complementary click partners has afforded polyol-BMS-like conjugates that are not cytotoxic (TZM.bl cells) and retain the activity against R5-HIV-1NLAD8 isolates. Thus, potential vaginal microbicides based on entry inhibitors, which can be called of 4th generation, are reported here for the first time.

Nuclear factor-kappaB activation regulates cyclooxygenase-2 induction in human astrocytes in response to CXCL12: role in neuronal toxicity.

Neurodegenerative and neuroinflammatory disorders are commonly associated with local chemokine release. In other way, emerging data indicate that the prostaglandin E2 (PGE(2)), one of the major prostaglandins produced in the brain, play a central role in several pathological diseases. In this study, we investigated the relationship between CXCL12, cyclooxygenase (COX)-2 and PGE(2) in human brain cells. CXCL12 induced COX-2 and secretion of PGE(2) in a dose-dependent manner in human astrocytes. This induction was abolished by treatment with pertussis toxin and AMD3100, confirming the role of CXCR4 signaling. The nuclear factor-kappaB involvement was confirmed by using pyrrolidine dithiocarbamate, and with transient transfection assays. Over-expression of inhibitory proteins of nuclear factor-kappaB abrogated COX-2 induction, and CXCL12 induced p65/relA translocation. Culture supernatants from CXCL12-treated astrocytes reduced viability of neuroblastoma cells, and COX inhibitors abrogated this toxicity. Therefore, the relationship between chemokines and PGs could differentially influence the pathogenic network responsible for neurodegeneration.

New synthetic procedure for the antiviral sulfonate carbosilane dendrimer G2-S16 and its fluorescein-labelled derivative for biological studies.

The anionic carbosilane (CBS) dendrimer with sulfonate groups G2-S16 is a promising compound for the preparation of a microbicide gel to prevent HIV infection. However, until now its synthesis required aggressive conditions. Hence, a reliable synthetic procedure is very important to face GMP conditions and clinical trials. In this study, G2-S16 has been prepared by a new approach that involves the addition of an amine-terminated dendrimer to ethenesulfonyl fluoride (C2H3SO3F, ESF) and then transformation to the sulfonate dendrimer by treatment with a base. This strategy also makes feasible the synthesis of a labelled sulfonate dendrimer (G2-S16-FITC) to be used as a molecular probe for in vivo experiments. Interestingly, G2-S16-FITC enters into human peripheral blood mononuclear cells (PBMCs).

Combination of G2-S16 dendrimer/dapivirine antiretroviral as a new HIV-1 microbicide.

Aim: To research the synergistic activity of G2-S16 dendrimer and dapivirine (DPV) antiretroviral as microbicide candidate to prevent HIV-1 infection. Materials & methods: We assess the toxicity of DPV on cell lines by MTT assay, the anti-HIV-1 activity of G2-S16 and DPV alone or combined at several fixed ratios. Finally, their ability to inhibit the bacterial growth in vitro was assayed. The analysis of combinatorial effects and the effective concentrations were performed with CalcuSyn software. Conclusion: Our results represent the first proof-of-concept study of G2-S16/DPV combination to develop a safe microbicide.