Search

In SARS-CoV-2-infected humans, disease progression is often associated with acute respiratory distress syndrome involving severe lung injury, coagulopathy, and thrombosis of the alveolar capillaries. The pathogenesis of these pulmonary complications in COVID-19 patients has not been elucidated. Autopsy study of these patients showed SARS-CoV-2 virions in pulmonary vessels and sequestrated leukocytes infiltrates associated with endotheliopathy and microvascular thrombosis. Since SARS-CoV-2 enters and infects target cells by binding its spike (S) protein to cellular angiotensin-converting enzyme 2 (ACE2), and there is evidence that vascular endothelial cells and neutrophils express ACE2, we investigated the effect of S-proteins and cell–cell communication on primary human lung microvascular endothelial cells (HLMEC) and neutrophils expression of thrombogenic factors and the potential mechanisms. Using S-proteins of two different SARS-CoV-2 variants (Wuhan and Delta), we demonstrate that exposure of HLMEC or neutrophils to S-proteins, co-culture of HLMEC exposed to S-proteins with non-exposed neutrophils, or co-culture of neutrophils exposed to S-proteins with non-exposed HLMEC induced transcriptional upregulation of tissue factor (TF), significantly increased the expression and secretion of factor (F)-V, thrombin, and fibrinogen and inhibited tissue factor pathway inhibitor (TFPI), the primary regulator of the extrinsic pathway of blood coagulation, in both cell types. Recombinant (r)TFPI and a thiol blocker (5,5′-dithio-bis-(2-nitrobenzoic acid)) prevented S-protein-induced expression and secretion of Factor-V, thrombin, and fibrinogen. Thrombomodulin blocked S-protein-induced expression and secretion of fibrinogen but had no effect on S-protein-induced expression of Factor-V or thrombin. These results suggests that following SARS-CoV-2 contact with the pulmonary endothelium or neutrophils and endothelial–neutrophil interactions, viral S-proteins induce coagulopathy via the TF pathway and mechanisms involving functional thiol groups. These findings suggest that using rTFPI and/or thiol-based drugs could be a viable therapeutic strategy against SARS-CoV-2-induced coagulopathy and thrombosis.

Background Neurocognitive impairment is present in 50% of HIV-infected individuals and is often associated with Alzheimer’s Disease (AD)-like brain pathologies, including increased amyloid-beta (Aβ) and Tau hyperphosphorylation. Here, we aimed to determine whether HIV-1 infection causes AD-like pathologies in an HIV/AIDS humanized mouse model, and whether the CCR5 antagonist maraviroc alters HIV-induced pathologies. Methods NOD/scid–IL-2Rγcnull mice engrafted with human blood leukocytes were infected with HIV-1, left untreated or treated with maraviroc (120 mg/kg twice/day). Human cells in animal’s blood were quantified weekly by flow cytometry. Animals were sacrificed at week-3 post-infection; blood and tissues viral loads were quantified using p24 antigen ELISA, RNAscope, and qPCR. Human (HLA-DR+) cells, Aβ-42, phospho-Tau, neuronal markers (MAP 2, NeuN, neurofilament-L), gamma-secretase activating protein (GSAP), and blood-brain barrier (BBB) tight junction (TJ) proteins expression and transcription were quantified in brain tissues by immunohistochemistry, immunofluorescence, immunoblotting, and qPCR. Plasma Aβ-42, Aβ-42 cellular uptake, release and transendothelial transport were quantified by ELISA. Results HIV-1 significantly decreased human (h)CD4+ T-cells and hCD4/hCD8 ratios; decreased the expression of BBB TJ proteins claudin-5, ZO-1, ZO-2; and increased HLA-DR+ cells in brain tissues. Significantly, HIV-infected animals showed increased plasma and brain Aβ-42 and phospho-Tau (threonine181, threonine231, serine396, serine199), associated with transcriptional upregulation of GSAP, an enzyme that catalyzes Aβ formation, and loss of MAP 2, NeuN, and neurofilament-L. Maraviroc treatment significantly reduced blood and brain viral loads, prevented HIV-induced loss of neuronal markers and TJ proteins; decreased HLA-DR+ cells infiltration in brain tissues, significantly reduced HIV-induced increase in Aβ-42, GSAP, and phospho-Tau. Maraviroc also reduced Aβ retention and increased Aβ release in human macrophages; decreased the receptor for advanced glycation end products (RAGE) and increased low-density lipoprotein receptor–related protein-1 (LRP1) expression in human brain endothelial cells. Maraviroc induced Aβ transendothelial transport, which was blocked by LRP1 antagonist but not RAGE antagonist. Conclusions Maraviroc significantly reduced HIV-induced amyloidogenesis, GSAP, phospho-Tau, neurodegeneration, BBB alterations, and leukocytes infiltration into the CNS. Maraviroc increased cellular Aβ efflux and transendothelial Aβ transport via LRP1 pathways. Thus, therapeutically targeting CCR5 could reduce viremia, preserve the BBB and neurons, increased brain Aβ efflux, and reduce AD-like neuropathologies.

Variations in the HIV genome influence HIV/AIDS epidemiology. We report here a novel HIV-1 unique recombinant form (URF) isolated from an HIV-infected female (NACMR092) in Cameroon, based on the analyses of near-full-length viral genome (partial gag, full-length pol, env, tat, rev, vif, vpr, vpu, and nef genes, and partial 3'-long terminal repeat). Phylogeny, recombination breakpoints, and recombination map analyses showed that NACMR092 was infected with a mosaic URF that had eight breakpoints (two in gag, one in pol, one in vpr, two in env, and two in the nef regions), nine subgenomic regions, and included fragments that had important similarities with HIV-1 subtypes A1, CRF02_AG, and CRF01_AE. This novel mosaic URF underscores complex recombination events occurring between HIV-1 subtypes circulating in Cameroon. Continued monitoring and detection of such recombinants and accurate classification of HIV genotype is important for tracking viral molecular epidemiology and antigenic diversity.

HIV subtypes distribution varies by geographic regions; this is likely associated with differences in viral fitness but the predictors and underlying mechanisms are unknown. Using in-vitro, in-vivo, and ex-vivo approaches, we found significantly higher transactivation and replication of HIV-1-CRF02_AG (prevalent throughout West-Central Africa), compared to subtype-B. While CRF02_AG-infected animals showed higher viremia, subtype-B-infected animals showed significantly more weight loss, lower CD4+ T-cells and lower CD4/CD8 ratios, suggesting that factors other than viremia contribute to immunosuppression and wasting syndrome in HIV/AIDS. Compared to HIV-1-subtype-B and its Tat proteins(Tat.B), HIV-1-CRF02_AG and Tat.AG significantly increased histone acetyl-transferase activity and promoter histones H3 and H4 acetylation. Silencing N-myrystoyltransferase(NMT)-1 and casein-kinase-(CK)-II-alpha prevented Tat.AG- and HIV-1-CRF02_AG-mediated viral transactivation and replication, but not Tat.B- or HIV-1-subtype-B-mediated effects. Tat.AG and HIV-1-CRF02_AG induced the expression of NMT-1 and CKII-alpha in human monocytes and macrophages, but Tat.B and HIV-1-subtype-B had no effect. These data demonstrate that NMT1, CKII-alpha, histone acetylation and histone acetyl-transferase modulate the increased replication of HIV-1-CRF02_AG. These novel findings demonstrate that HIV genotype influence viral replication and provide insights into the molecular mechanisms of differential HIV-1 replication. These studies underline the importance of considering the influence of viral genotypes in HIV/AIDS epidemiology, replication, and eradication strategies.

HIV-associated neurocognitive deficits include impaired speed-of-information processing (SIP) and motor functions. There is lack of Cameroonian adult norms for assessing SIP or motor functions. This study of 683 Cameroonians (320 HIV+, 363 HIV−) establishes demographically-adjusted norms for six SIP [Wechsler-Adult-Intelligence-Scale (WAIS)-III Digit Symbol (WAIS-IIIDS) and Symbol Search (WAIS-IIISS), Stroop Color-Naming, Stroop Word-Reading, Trail-Making Test-A (TMT-A), Color Trails-1 (CTT1)], and two motor function [Grooved Pegboard-dominant (GP-DH) and non-dominant (GP-NDH) hands] tests. We assessed viral effects on SIP and motor functions. HIV-infected persons had significantly lower (worse) T scores on GP-DH, WAIS-IIIDS, Stroop Word-Reading, TMT-A; lower motor and SIP summary T scores. Significantly higher proportion of cases (20.7%) than controls (10.3%) had impaired SIP. Male cases had better T scores than female cases on GP-NDH, WAIS-IIIDS, WAIS-IIISS, TMT-A, CTT1; better SIP summary T scores. Antiretroviral therapy (ART) was associated with significantly better T scores on GP-NDH, WAIS-IIIDS, Stroop Color-Naming; better motor and SIP summary T scores. Cases with higher CD4 had better T scores on WAIS-IIIDS, TMT-A, CTT1; better SIP summary T scores. Overall, we demonstrate that HIV infection in Cameroon is associated with deficits in SIP and motor functions; ART and higher CD4 are associated with better cognitive performance. We provide SIP and psychomotor functions normative standards, which will be useful for neurobehavioral studies in Cameroon of diseases affecting the brain.

HIV-associated neurocognitive disorders (HAND) are frequently associated with impaired executive function and verbal fluency. Given limited knowledge concerning HAND in Sub-Saharan-Africa and lack of Cameroonian adult neuropsychological (NP) test norms, we administered four executive function [Halstead Category Test (HCT), Wisconsin Card Sorting Test (WCST), Color Trails-II (CTT2), and Stroop Color-Word-Interference (SCWT)] and three verbal fluency (Category, Action, and Letter Fluency) tests to 742 adult Cameroonians (395 HIV−, 347 HIV+). We developed demographically-corrected NP test norms and examined the effects of HIV and related variables on subjects’ executive function and verbal fluency. HIV+ subjects had significantly lower T-scores on CTT2 (P = 0.005), HCT (P = 0.032), WCST (P

An urgent need to deliver therapeutics across the blood-brain barrier (BBB) underlies a paucity of effective therapies currently available for treatment of degenerative, infectious, traumatic, chemical, and metabolic disorders of the nervous system. With an eye toward achieving this goal, an in vitro BBB model was employed to simulate biodegradable polyanhydride nanoparticle-based drug delivery to the brain. Using a combination of confocal microscopy, flow cytometry, and high performance liquid chromatography, we examined the potential of polyanhydride nanoparticles containing the anti-oxidant, mito-apocynin, to be internalized and then transferred from monocytes to human brain microvascular endothelial cells. The efficacy of this nanoparticle-based delivery platform was demonstrated by neuronal protection against oxidative stress. Taken together, this polyanhydride nanoparticle-based delivery system holds promise for enhancing neuroprotection by facilitating drug transport across the BBB. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2881-2890, 2018.

HIV-1-associated neurocognitive disorders (HAND) is associated with blood-brain-barrier (BBB) inflammation, and inflammation involves toll-like receptors (TLRs) signaling. It is not known whether primary human brain microvascular endothelial cells (HBMEC), the major BBB component, express TLRs or whether TLRs are involved in BBB dysfunction and HAND. We demonstrate that HBMEC express TLR3, 4, 5, 7, 9, and 10, and TLR3 was the most abundant. HIV-1 and TLR3 activation increased endothelial TLR3 transcription and expression. HIV-1-positive human subjects showed significantly higher TLR3 expression in brain tissues and blood vessels, with higher TLR3 levels in subjects with HAND. HIV-1 and TLR3 activation increased endothelial IL6 expression by 6-to-127-fold (P

Integrase strand-transfer inhibitors (INSTIs) are now included in preferred first-line antiretroviral therapy (ART) for HIV-infected adults. Studies of Western clade-B HIV-1 show increased resistance to INSTIs following mutations in integrase and nef 3&rsquo, polypurine tract (3&rsquo, PPT). With anticipated shifts in Africa (where 25.6-million HIV-infected people resides) to INSTIs-based ART, it is critical to monitor patients in African countries for resistance-associated mutations (RAMs) affecting INSTIs efficacy. We analyzed HIV-1 integrase and 3&rsquo, PPT sequences in 345 clinical samples from INSTIs-na&iuml, ve HIV-infected Cameroonians for polymorphisms and RAMs that affect INSTIs. Phylogeny showed high genetic diversity, with the predominance of HIV-1 CRF02_AG. Major INSTIs RAMs T66A and N155K were found in two (0.6%) samples. Integrase polymorphic and accessory RAMs found included T97A, E157Q, A128T, M50I, S119R, L74M, L74I, S230N, and E138D (0.3%-23.5% of samples). Ten (3.2%) samples had both I72V+L74M, L74M+T97A, or I72V+T97A mutations, thirty-one (9.8%) had 3&rsquo, PPT mutations. The low frequency of major INSTIs RAMs shows that INSTIs-based ART can be successfully used in Cameroon. Several samples had 1 INSTIs accessory RAMs known to reduce INSTIs efficacy, thus, INSTIs-based ART would require genetic surveillance. The 3&rsquo, PPT mutations could also affect INSTIs. For patients failing INSTIs-based ART with no INSTIs RAMs, monitoring 3&rsquo, PPT sequences could reveal treatment failure etiology.

In HIV-1 subtype-B, specific mutations in Gag cleavage sites (CS) are associated with treatment failure, with limited knowledge among non-B subtypes. We analyzed non-B HIV-1 gag and pol (protease/reverse-transcriptase) sequences from Cameroonians for drug resistance mutations (DRMs) in the gag P2/NC CS, and pol major DRMs. Phylogeny of the 141 sequences revealed a high genetic diversity (12 subtypes): 67.37% CRF02_AG versus 32.6% non-CRF02_AG. Overall, 7.3% transmitted and 34.3% acquired DRMs were found, including M184V, thymidine analogue mutations (T215F, D67N, K70R, K219Q), NNRTIs (L100I, Y181C, K103N, V108I, Y188L), and PIs (V82L). Twelve subjects [10 with HIV-1 CRF02_AG, 8 treatment-naïve and 4 on 3TC-AZT-NVP] showed 3 to 4 mutations in the Gag P2/NC CS: S373Q/T/A, A374T/S/G/N, T375S/A/N/G, I376V, G381S, and R380K. Subjects with or without Gag P2/NC CS mutations showed no significant difference in viral loads. Treatment-naïve subjects harboring NRTI-DRMs had significantly lower CD4 cells than those with NRTI-DRMs on ART (p = 0.042). Interestingly, two subjects had major DRMs to NRTIs, NNRTIs, and 4 mutations in the Gag P2/NC CS. In this prevailing CRF02_AG population with little exposure to PIs (~3%), mutations in the Gag P2/NC CS could increase the risk of treatment failure if there is increased use of PIs-based therapy.

Potent antiretroviral activities and a barrier to viral resistance characterize the human immunodeficiency virus type one (HIV-1) integrase strand transfer inhibitor dolutegravir (DTG). Herein, a long-acting parenteral DTG was created through chemical modification to improve treatment outcomes. A hydrophobic and lipophilic modified DTG prodrug is encapsulated into poloxamer nanoformulations (NMDTG) and characterized by size, shape, polydispersity, and stability. Retained intracytoplasmic NMDTG particles release drug from macrophages and attenuate viral replication and spread of virus to CD4+ T cells. Pharmacokinetic tests in Balb/cJ mice show blood DTG levels at, or above, its inhibitory concentration90 of 64 ng/mL for 56 days, and tissue DTG levels for 28 days. NMDTG protects humanized mice from parenteral challenge of the HIV-1ADA strain for two weeks. These results are a first step towards producing a long-acting DTG for human use by affecting drug apparent half-life, cell and tissue drug penetration, and antiretroviral potency., Current ART for treatment of HIV-1 infection requires a strict daily regimen adherence. Herein, the authors report the manufacture and characterization of a nanoformulated dolutegravir prodrug with improved cell and tissue penetration, a remarkable apparent half-life and the potential for bimonthly drug administration.

HIV-1 genetic differences influence viral replication and progression to AIDS. HIV-1 circulating recombinant form (CRF)02_AG is the predominant viral subtype infecting humans in West and Central Africa, but its effects on HIV neuropathogenesis are not known. In the present study, we investigated the effects of Tat proteins from HIV-1 subtype B (Tat.B) and HIV-1 CRF02_AG (Tat.AG) on primary human brain microvascular endothelial cells (HBMEC), the major component of the blood–brain barrier (BBB). Using Affymetrix GeneChip Human Gene 1.0.ST arrays, we showed that Tat.AG had minimal effects while Tat.B induced transcriptional upregulation of 90 genes in HBMEC, including proinflammatory chemokines, complement components C3, C7, and complement factor B, matrix metalloproteinases (MMP)-3, MMP-10, and MMP-12. These results were confirmed by real-time PCR. Compared with Tat.AG, Tat.B significantly increased MMP-3, MMP-10, and MMP-12 activities in HBMEC, and the MMPs tissue inhibitor of metalloproteinase-2 blocked Tat-induced increase in MMPs activity. Western blot analyses also showed that Tat increased the expression of C3 and its cleaved fragment C3b in HBMEC. These data suggest that genetic differences between HIV-1 subtypes B and CRF02_AG influence the effects of Tat proteins from these two clades on HBMEC, including molecular and cellular functions, and canonical pathways, which would affect BBB dysfunction and viral neuropathogenesis.

Following global efforts to increase antiretroviral therapy (ART) access in Sub-Saharan Africa, ART coverage among HIV-infected Cameroonians increased from 0% in 2003 to 22% in 2014. However, the success of current HIV treatment programs depends not only on access to ART, but also on retention in care and good treatment adherence. This is necessary to achieve viral suppression, prevent virologic failure, and reduce viral transmission and HIV/AIDS-related deaths. Previous studies in Cameroon showed poor adherence, treatment interruption, and loss to follow-up among HIV+ subjects on ART, but the factors that influence ART adherence are not well known. In the current cross-sectional study, patient/self-reported questionnaires and pharmacy medication refill data were used to quantify ART adherence and determine the factors associated with increased risk of non-adherence among HIV-infected Cameroonians. We demonstrated that drug side-effects, low CD4 cell counts and higher viral loads are associated with increased risk of non-adherence, and compared to females, males were more likely to forego ART because of side effects (p40 years) were less likely to be non-adherent (p

Depression is a leading cause of HIV/AIDS disease burden; it worsens health outcomes and quality of life. Addressing this problem requires accurate quantification of the extra burden of depression to HIV/AIDS in a given population, and knowledge of the baseline depression prevalence in the general population. There has been no previous study of depression in the general Cameroonian population. The current study attempts to address that important need. We used the Beck Depression Inventory-II to assess the prevalence and severity of depressive symptoms in 270 HIV-infected and seronegative Cameroonians. Univariate analyses showed a trend toward higher depressive symptoms among cases, compared to controls (p = 0.055), and among older subjects (>40 years), compared to younger subjects (≤40 years) (p = 0.059). Analysis of depression severity showed that 33.73% of cases had moderate-to-severe depressive symptoms, compared to 19.8% of controls (p

In May 2012, the Division of AIDS Research at the National Institute of Mental Health (NIMH) organized the "Global NeuroAIDS Roundtable" in conjunction with the 11th International Symposium on Neurovirology and the 2012 Conference on HIV in the Nervous System. The meeting was held in New York, NY, USA and brought together NIMH-funded investigators who are currently working on projects related to the neurological complications of AIDS (NeuroAIDS) in Africa, Asia, Eastern Europe, and Latin America in order to provide an opportunity to share their recent findings and discuss the challenges encountered within each country. The major goals of the roundtable were to evaluate HIV-associated neurocognitive impairment and determine if it may be directly attributable to distinct HIV subtypes or clades and to discuss the future priorities for global NeuroAIDS research. At the "Global NeuroAIDS Roundtable", presentations of preliminary research indicated that HIV-associated neurocognitive impairment is prevalent in all countries examined regardless of which HIV clade is present in the region. The only clear-cut difference between HIV-1 clades was in relation to subtypes A and D in Uganda. However, a key point that emerged from the discussions was that there is an urgent need to standardize neurocognitive assessment methodologies across the globe before definitive conclusions can be drawn regarding the relationship between HIV clade diversity and neuropathogenesis. Future research directions were also discussed at the roundtable with particular emphasis on the potential of viral and host factor molecular interactions to impact the pathophysiology of HIV-associated neurocognitive disorders (HAND) from a global perspective. © 2013 Journal of NeuroVirology, Inc. (outside the USA).

The recombinant HIV-1 CRF02_AG is prevalent in West-Central Africa but its effects on the blood-brain barrier (BBB) and HIV-associated neurocognitive disorders (HAND) are not known. We analyzed the effects of Tat from HIV-1 subtype-B (Tat.B) and CRF02_AG (Tat.AG) on primary human brain microvascular endothelial cells (HBMEC), the major BBB component. Exposure of HBMEC to Tat.B increased IL-6 expression and transcription by 9- (P

Despite major advances in HIV-1 treatment, the prevalence of HIV-associated neurocognitive disorders (HAND) remains a problem, particularly as individuals on suppressive treatment continue to live longer. To facilitate discussion on emerging and future directions in HAND research, a meeting was held in Durban, South Africa in March 2015 as part of the Society of Neuroscientists of Africa (SONA) conference. The objective of the meeting was to assess the impact of HIV subtype diversity on HAND and immunological dysfunction. The meeting brought together international leaders in the area of neurological complications of HIV-1 infection with special focus on the African population. Research presentations indicated that HAND was highly prevalent and that inflammatory cytokines and immune-activation played important roles in progression of neurocognitive impairment. Furthermore, children on antiretroviral therapy were also at risk for developing neurocognitive impairment. With respect to the effect of HIV-1 subtype diversity, analyses of HIV-1 clade C infection among South Africans revealed that clade C infection induced cognitive impairment that was independent of the substitution in HIV-1 Trans-Activator of Transcription (Tat; C31S). At the cellular level, a Zambian study showed that clade C infection resulted in reduced brain cell death compared with clade B infection suggesting clade specific variations in mediating brain cell injury. Furthermore, ex vivo Tat protein from clade CRF02_AG, prevalent in West/ Central Africa, exhibited reduced disruption of brain endothelium compared with clade B Tat protein. Discussions shed light on future research directions aimed at understanding biomarkers and disease mechanisms critical for HAND.

HIV-1 Tat plays a critical role in viral transactivation. Subtype-B Tat has potential use as a therapeutic vaccine. However, viral genetic diversity and population genetics would significantly impact the efficacy of such a vaccine. Over 70% of the 37-million HIV-infected individuals are in sub-Saharan Africa (SSA) and harbor non-subtype-B HIV-1. Using specimens from 100 HIV-infected Cameroonians, we analyzed the sequences of HIV-1 Tat exon-1, its functional domains, post-translational modifications (PTMs), and human leukocyte antigens (HLA)-binding epitopes. Molecular phylogeny revealed a high genetic diversity with nine subtypes, CRF22_01A1/CRF01_AE, and negative selection in all subtypes. Amino acid mutations in Tat functional domains included N24K (44%), N29K (58%), and N40K (30%) in CRF02_AG, and N24K in all G subtypes. Motifs and phosphorylation analyses showed conserved amidation, N-myristoylation, casein kinase-2 (CK2), serine and threonine phosphorylation sites. Analysis of HLA allelic frequencies showed that epitopes for HLAs A*0205, B*5301, Cw*0401, Cw*0602, and Cw*0702 were conserved in 58%–100% of samples, with B*5301 epitopes having binding affinity scores > 100 in all subtypes. This is the first report of N-myristoylation, amidation, and CK2 sites in Tat; these PTMs and mutations could affect Tat function. HLA epitopes identified could be useful for designing Tat-based vaccines for highly diverse HIV-1 populations, as in SSA.

Shantanu Balkundi1, Ari S Nowacek1, Ram S Veerubhotla1, Han Chen2, Andrea Martinez-Skinner1, Upal Roy1, R Lee Mosley1,3, Georgette Kanmogne1, Xinming Liu1,3,4, Alexander V Kabanov3,4, Tatiana Bronich3,4, JoEllyn McMillan1, Howard E Gendelman1,31Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA; 2Center for Biotechnology, University of Nebraska-Lincoln, Lincoln, NE, USA; 3Center for Drug Delivery and Nanomedicine, University of Nebraska Medical Center, Omaha, NE, USA; 4Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE, USAAbstract: Nanoformulations of crystalline indinavir, ritonavir, atazanavir, and efavirenz were manufactured by wet milling, homogenization or sonication with a variety of excipients. The chemical, biological, immune, virological, and toxicological properties of these formulations were compared using an established monocyte-derived macrophage scoring indicator system. Measurements of drug uptake, retention, release, and antiretroviral activity demonstrated differences amongst preparation methods. Interestingly, for drug cell targeting and antiretroviral responses the most significant difference among the particles was the drug itself. We posit that the choice of drug and formulation composition may ultimately affect clinical utility.Keywords: human immunodeficiency virus type one, nanotoxicology, monocyte-derived macrophage, nanoformulated antiretroviral therapy, manufacturing techniques

Limitations inherent to antiretroviral therapy (ART) in its pharmacokinetic properties remain despite over 15 years of broad use. Our laboratory has pioneered a means to improve ART delivery through monocyte-macrophage carriage of nanoformulated drug-encapsulated particles (nanoART). To this end, our prior works sought to optimize nanoART size, structure, and physical properties for cell uptake and antiretroviral activities. To test the functional consequences of indinavir, ritonavir, and efavirenz formulations we investigated relationships between human monocyte and macrophage cytotoxicities and nanoART dose, size, surfactant, and preparation. Wet-milled particles were significantly more cytotoxic to monocytes-macrophages than those prepared by homogenization; with concurrent induction of tumor necrosis factor-alpha. Interestingly, pure suspensions of indinavir and ritonavir at 0.5mM, and efavirenz at 0.1mM and 0.5mM also proved cytotoxic. Individual surfactants and formulated fluconazole neither affected cell function or viability. Although nanoART did not alter brain tight junction proteins ZO-2 and occludin, 0.5mM ritonavir formulations did alter brain transendothelial electric resistance. These results underscore the importance of evaluating the physicochemical and functional properties of nanoART before human evaluations.

How neuroinflammation affects signaling pathways leading to human blood–brain barrier (BBB) dysfunction during HIV-1 infection is incompletely understood. We previously demonstrated that signal transducers and activators of transcription-1 (STAT1) signaling is involved in HIV-1 induced BBB damage and is relevant to viral neuropathogenesis. The objective of this study was to delineate the signaling pathways upstream and downstream of STAT1 involved in HIV-1-induced endothelial dysfunction. We show that HIV-1 activation of STAT1 and STAT3 in human brain microvascular endothelial cells (HBMEC) is associated with induction of promoter activity of the interferon-stimulated response element (ISRE)/interferon-γ-activated sequence (GAS). The STAT1 inhibitor fludarabine diminished HIV-1-induced ISRE/GAS promoter activity. CCR5 neutralizing antibodies and the phosphoinositide-3-kinase (PI3K) inhibitor LY-294002 diminished HIV-1-induced phosphorylation of STAT1 and STAT3, significantly diminished HIV-1-induced ISRE/GAS promoter activity, and diminished virus-induced monocyte adhesion and transendothelial migration. HIV-1 infection did not phosphorylate janus kinases but induced activation of the phosphoinositide-dependent kinase-1 (PDK1) and the serine-threonine protein kinase AKT, both downstream effectors of PI3K. CCR5 antibodies also diminished virus-induced phosphorylation of PDK1 and AKT. These results suggest that the chemokine receptor CCR5 is partially involved in HIV-1 binding to HBMEC and show cross-talk between STAT1 and PI3K pathways in HIV-1-induced BBB dysfunction.

The prevalence of HIV-associated neurocognitive disorders (HAND), especially HIV-associated dementia (HAD) is influenced by several risk factors. The prevalence as well as risk factors for HAD are not well known in sub-Saharan Africa (SSA). We have shown that the International HIV Dementia Scale (IHDS) is a useful screening tool for HAND in Yaoundé [Njamnshi AK, Djientcheu VdP, Fonsah JY, Yepnjio FN, Njamnshi DM, Muna WFT. The IHDS is a useful screening tool for HAD/Cognitive Impairment in HIV-infected adults in Yaoundé-Cameroon. Journal of Acquired Immune Deficiency Syndromes 2008;49(4):393-397], but no study in Cameroon has yet investigated the risk factors for HAND or HAD.A cross-sectional study was conducted in Yaoundé, the capital of Cameroon from September to December 2006. One hundred and eighty-five HIV-positive subjects were included. Diagnosis of HAND was done using the IHDS with a scoreor = 10 considered as abnormal. Age, sex, level of education, IV drug use, body mass index (BMI), CDC clinical stage, CD4 counts, hemoglobin levels, administration of highly active antiretroviral therapy (HAART) and type of regimen used, were considered in univariate analysis, with level of significance set at Por = 0.05. A binary logistic regression was used to determine independent risk factors.The following factors were independent predictors of HAND: advanced clinical stage (OR=7.43, P=0.001), low CD4 count especially CD4or = 200 cells/microL (OR=4.88, P=0.045) and low hemoglobin concentration (OR=1.16, P=0.048).This first study of the risk factors for HAND in Yaoundé-Cameroon shows findings similar to those described in other studies. These results call for rapid action by policy makers to include HAND prevention strategies such as providing early universal access to HAART based on these risk factors, in the management of HIV patients at risk of HAND in resource-limited settings of SSA like ours.

How neuroinflammatory activities affect signaling pathways leading to blood–brain barrier (BBB) injury during HIV/AIDS are currently unknown. Our previous work demonstrated that HIV-1 exposure activates pro-inflammatory genes in human brain microvascular endothelial cells (HBMEC) and showed that these genes are linked to the janus kinase (JAK)/signal transducers and activators of transcription (STAT) pathway. Here, we report that HIV-1 gp120 protein activated STAT1 and induced interleukin (IL)-6 and IL-8 secretion in HBMEC. IL-6, IL-8, and gp120 increased monocyte adhesion and migration across in vitro BBB models. The STAT1 inhibitor, fludarabine, prevented gp120-induced IL-6 and IL-8 secretion. Inhibitors of STAT1, mitogen activated protein kinase kinase (MEK) (PD98059), and phosphatidyl inositol 3 kinase (PI3K) (LY294002), blocked gp120-induced STAT1 activation and significantly diminished IL-8-, IL-6-, and gp120-induced monocyte adhesion and migration across in vitro BBB models. These data support the notion that STAT1 plays an important role in gp120-induced inflammation and BBB dysfunction associated with viral infection. Results also suggest crosstalk between STAT1, MEK, and PI3K pathways in gp120-induced BBB dysfunction. Inhibition of STAT1 activation could provide a unique therapeutic strategy to decrease neuroinflammation and BBB dysfunction in HIV/AIDS.

The relationship among neuroinflammation, blood-brain barrier (BBB) dysfunction, and progressive HIV-1 infection as they affect the onset and development of neuroAIDS is incompletely understood. One possible link is signal transducers and activators of transcription (STATs) pathways. These respond to proinflammatory and regulatory factors and could affect neuroinflammatory responses induced from infected cells and disease-affected brain tissue. Our previous works demonstrated that HIV-1 activates pro-inflammatory and interferon-alpha–inducible genes in human brain microvascular endothelial cells (HBMECs) and that these genes are linked to the Janus kinase (JAK)/STAT pathway. We now demonstrate that HIV-1 activates STAT1, induces IL-6 expression, and diminishes expression of claudin-5, ZO-1, and ZO-2 in HBMECs. The STAT1 inhibitor, fludarabine, blocked HIV-1–induced IL-6, diminished HIV-1–induced claudin-5 and ZO-1 down-regulation, and blocked HIV-1– and IL-6–induced monocyte migration across a BBB model. Enhanced expression and activation of STAT1 and decreased claudin-5 were observed in microvessels from autopsied brains of patients with HIV-1–associated dementia. These data support the notion that STAT1 plays an integral role in HIV-1–induced BBB damage and is relevant to viral neuropathogenesis. Inhibition of STAT1 activation could provide a unique therapeutic strategy to attenuate HIV-1–induced BBB compromise and as such improve clinical outcomes.

The mechanisms underlying blood—brain barrier (BBB) dysfunction seen in human immunodeficiency virus 1 (HIV-1) infection are poorly understood; however, they are believed to be caused by interactions of human brain microvascular endothelial cells (HBMEC) with virus-infected macrophages. Using a transwell system and Affymetrix arrays, we investigated HIV-1-induced genomic changes in HBMEC after coculture with HIV-1-infected or -uninfected monocyte-derived macrophages (MDM). Differentially expressed genes were determined by linear modeling and then were grouped by hierarchical clustering. Compared to HBMEC cocultured with noninfected MDM, 184 probe sets corresponding to 84 genes were differentially expressed in HBMEC cocultured with HIV-infected MDM. Genes activated in HIV-1 MDM-exposed HBMEC included proinflammatory cytokines and chemokines, tumor necrosis factor-α-induced proteins, interferon (IFN)-inducible genes, intercellular adhesion molecule-1, transcription factors of the nuclear factor-κB family, and signal transducer and activator of transcription 1. Analysis of molecular networks and canonical pathways associated with differentially expressed genes suggest that HIV-1 causes BBB impairment by mechanisms involving inflammation, cytokine, and IFN signaling in HBMEC.

TLR3 has been implicated in the pathogenesis of several viral infections, including SIV- and HIV-1-induced inflammation and AIDS. However the molecular mechanisms of these TLR3-mediated effects are not known, and it is not known whether HIV interacts with cellular TLR3 to affect disease process. Here we investigate the effects of TLR3 ligands on HIV-1 transactivation using both primary human macrophages and cells containing integrated copies of the HIV-1 promoter. We demonstrate that TLR3 activation induced upregulation of transcription factors such as c-Jun, CCAAT/enhancer-binding protein alpha (CEBPA), signal transducer and activator of transcription (STAT)-1, STAT-2, RELB, and nuclear factor kappa-B1 (NFκB1), most of which are known to regulate the HIV promoter activity. We also demonstrate that TLR3 activation increased HIV-1 transactivation via the c-Jun N-terminal kinase (JNK) and NFκB pathways. This was associated with epigenetic modifications, including decreased histone deacetylase activity, increased histone acetyl transferase (HAT) activity, and increased acetylation of histones H3 and H4 at lysine residues in the nucleosome-0 and nucleosome-1 of the HIV-1 promoter. However, prolonged TLR3 activation decreased HIV-1 transactivation, decreased HAT activity and Tat transcription, and suppressed viral replication. Overall, data suggests that TLR3 can act as viral sensor to mediate viral transactivation, cellular signaling, innate immune response, and inflammation in HIV-infected humans. Our study provides novel insights into the molecular basis for these TLR3-mediated effects.

Breakdown of the blood-brain barrier (BBB) is commonly seen in patients with HIV-associated dementia (HAD) despite the lack of productive infection of the brain endothelium. It is likely that secreted viral products play a major role in BBB damage and the development of HAD. The objective of this study is to determine the effects of gp120 proteins on brain endothelial cell permeability and junctional protein expression. Our results showed that treatment of cultured human brain endothelial cells with gp120 for 24 hours results in increased permeability of the endothelial monolayer. Also, gp120 proteins caused disruption and downregulation of the tight junction proteins ZO-1, ZO-2, and occludin in these cells. Other junctional proteins such as claudin-1 and claudin-5 were unaffected by gp120 treatment. These data demonstrate that HIV gp120 proteins alter both the functional and molecular properties of the BBB, which could increase trafficking of HIV, infected cells, and toxic humoral factors into the central nervous system and contribute to the pathogenesis of HAD.

Purpose of review This article reviews recent findings on noninfectious pulmonary complications of HIV/AIDS, with a focus on HIV/AIDS-related lung malignancies and pulmonary hypertension, and discusses their incidence in the highly active antiretroviral therapy (HAART) era. Recent findings Noninfectious pulmonary complications of HIV/AIDS are now recognized as important contributors to morbidity and mortality in HIV-infected patients. This is especially the case for HIV-related lung cancer and other non-AIDS-defining malignancies, which are now being diagnosed with increased frequency in HIV-infected patients. The incidence of Kaposi sarcoma and AIDS-related lymphoma has decreased in the HAART era, but compared with the general population, the risk of these malignancies and pulmonary hypertension is still very high in HIV-infected patients. Concurrent use of HAART and chemotherapy improves prognosis and survival of patients with AIDS-related lymphoma. For patients with HIV-related pulmonary hypertension, some studies show no beneficial effect of HAART whereas other reports show that HAART improves patient survival and response to antihypertensive treatment. Summary The beneficial effect of HAART and improved immune response on the treatment of Kaposi sarcoma and AIDS-related lymphoma suggests that HIV or viral-induced immunosuppression plays an important role in the development of these malignancies. Evidence from current studies suggests that HAART does not protect against HIV-related lung cancer. The full impact of HAART on HIV pulmonary hypertension remains to be determined.

Breakdown of the blood-brain barrier is commonly seen in patients with human immunodeficiency virus (HIV)-associated dementia, despite the lack of productive HIV-infection of the brain endothelium. Through this damaged blood-brain barrier, HIV and HIV-infected monocytes/macrophages infiltrate the brain and further infect microglia and brain macrophages. Neuronal cell death and dysfunction are the underlying cause of HIV-associated dementia, but no productive HIV-infection of neurons has been documented. It is likely that secreted viral products play a major role in blood-brain barrier damage and neuronal cell death. The aim of the present study was to examine the effect of HIV-1 gp160 peptides and gp120 proteins on brain microvascular endothelial cells and neurons from both human and rats. Four of the 7 gp160 peptides tested evoked significant neurotoxicity. Two different full-length recombinant HIV gp120 proteins (HIV-1CM235 gp120 and HIV-1MN gp120) also induced neuronal and brain endothelial cell death, and concentrations as little as 1 ng/ml evoked pronounced morphological changes in these cells and marked cytotoxicity. This study suggests that HIV proteins and peptides that are shed in vivo may be directly involved in blood-brain barrier damage and neuronal cell death in HIV-associated dementia.

Microglia constitute the primary cell type infected with HIV in the brain and play a major role in viral persistence in the CNS and in the development of AIDS dementia. Lack of a suitable animal model and limitations in the availability of human tissues hinder most HIV/AIDS studies investigating the neuropathogenesis of AIDS dementia. The aims of this study were to determine whether baboon microglia can be productively infected with SIV-HIV (SHIV) recombinant viruses in vitro and whether they express HIV-1 receptors and coreceptors. Our results show the presence of mRNA for CD4, CCR5, and CXCR4 chemokine receptors on baboon microglial cells. Microglia lacked mRNA for the CCR3 chemokine receptor. We also show productive infection of baboon microglial cells by two SHIV isolates, SHIV-KU and SHIV-89.6P, and blockade of the infection with soluble CD4 protein, CCR5, and CXCR4 monoclonal antibodies. This study demonstrating the feasibility of infecting baboon microglia with SHIV isolates is an important first step in using the baboon as an alternative nonhuman primate model to study HIV neuropathogenesis.

The PCR technique, developed over 3 decades ago, has revolutionized molecular biology. PCR applications encompass several fields, including genetics, molecular cloning, diagnosis of infectious and hereditary diseases, and forensic sciences. Improvements of the PCR technique have resulted in its expansion to include variants such as reverse-transcription PCR, real-time PCR, and real-time reverse-transcription PCR. This chapter provides an overview of current PCR and PCR variants methods.

BackgroundHIV infection has commonly been found to affect lipid profile and antioxidant defense.ObjectivesTo determine the effects of Human Immunodeficiency Virus (HIV) infection and viral subtype on patient's cholesterol and oxidative stress markers, and determine whether in the absence of Highly Active Antiretroviral Therapy (HAART), these biochemical parameters could be useful in patient's management and monitoring disease progression in Cameroon. For this purpose, we measured total cholesterol (TC), LDL cholesterol (LDLC), HDL cholesterol (HDLC), total antioxidant ability (TAA), lipid peroxidation indices (LPI), and malondialdehyde (MDA) in HIV negative persons and HIV positive HAART-naïve patients infected with HIV-1 group M subtypes.MethodsWe measured serum TC, LDLC, HDLC, plasma MDA, and TAA concentrations, and calculated LPI indices in 151 HIV-positive HAART-naïve patients and 134 seronegative controls. We also performed gene sequence analysis on samples from 30 patients to determine the effect of viral genotypes on these biochemical parameters. We also determined the correlation between CD4 cell count and the above biochemical parameters.ResultsWe obtained the following controls/patients values for TC (1.96±0.54/1. 12±0. 48 g/l), LDLC (0. 67±0. 46/0. 43±0. 36 g/l), HDLC (105. 51±28. 10/46. 54±23. 36 mg/dl) TAA (0. 63±0. 17/0. 16±0. 16 mM), MDA (0. 20±0. 07/0. 41±0. 10 µM) and LPI (0. 34±0. 14/26. 02±74. 40). In each case, the difference between the controls and patients was statistically significant (pConclusionThese results show that HIV infection in Cameroon is associated with significant decrease in TAA, LDLC, HDLC and TC, and increased MDA concentration and LPI indices which seem to be linked to the severity of HIV infection as assessed by CD4 cell count. The data suggests increased oxidative stress and lipid peroxidation in HIV-infected patients in Cameroon, and an influence of CRFs on TC and MDA levels.

Lack of adherence, inaccessibility to viral reservoirs, long-term drug toxicities, and treatment failures are limitations of current antiretroviral therapy (ART). These limitations lead to increased viral loads, medicine resistance, immunocompromise, and comorbid conditions. To this end, we developed long-acting nanoformulated ART (nanoART) through modifications of existing atazanavir, ritonavir, and efavirenz suspensions in order to establish cell and tissue drug depots to achieve sustained antiretroviral responses. NanoART's abilities to affect immune and antiviral responses, before or following human immunodeficiency virus type 1 infection were tested in nonobese severe combined immune-deficient mice reconstituted with human peripheral blood lymphocytes. Weekly subcutaneous injections of drug nanoformulations at doses from 80 mg/kg to 250 mg/kg, 1 day before and/or 1 and 7 days after viral exposure, elicited drug levels that paralleled the human median effective concentration, and with limited toxicities. NanoART treatment attenuated viral replication and preserved CD4(+) Tcell numbers beyond that seen with orally administered native drugs. These investigations bring us one step closer toward using long-acting antiretrovirals in humans.

Georgette D Kanmogne1, Sangya Singh1, Upal Roy1, Xinming Liu1, JoEllyn McMillan1, Santhi Gorantla1, Shantanu Balkundi1, Nathan Smith1, Yazen Alnouti2, Nagsen Gautam2, You Zhou3, Larisa Poluektova1, Alexander Kabanov2, Tatiana Bronich2, Howard E Gendelman11Departments of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, 2Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE; 3Center for Biotechnology, University of Nebraska-Lincoln, Lincoln, NE, USAAbstract: Despite the successes of antiretroviral therapy (ART), HIV-associated neurocognitive disorders remain prevalent in infected people. This is due, in part, to incomplete ART penetration across the blood–brain barrier (BBB) and lymph nodes and to the establishment of viral sanctuaries within the central nervous system. In efforts to improve ART delivery, our laboratories developed a macrophage-carriage system for nanoformulated crystalline ART (nanoART) (atazanavir, ritonavir, indinavir, and efavirenz). We demonstrate that nanoART transfer from mononuclear phagocytes (MP) to human brain microvascular endothelial cells (HBMEC) can be realized through cell-to-cell contacts, which can facilitate drug passage across the BBB. Coculturing of donor MP containing nanoART with recipient HBMEC facilitates intercellular particle transfer. NanoART uptake was observed in up to 52% of HBMEC with limited cytotoxicity. Folate coating of nanoART increased MP to HBMEC particle transfer by up to 77%. To translate the cell assays into relevant animal models of disease, ritonavir and atazanavir nanoformulations were injected into HIV-1-infected NOD/scid-γcnull mice reconstituted with human peripheral blood lymphocytes. Atazanavir and ritonavir levels in brains of mice treated with folate-coated nanoART were three- to four-fold higher than in mice treated with noncoated particles. This was associated with decreased viral load in the spleen and brain, and diminished brain CD11b-associated glial activation. We postulate that monocyte-macrophage transfer of nanoART to brain endothelial cells could facilitate drug entry into the brain.Keywords: nanoART, folate, monocyte-endothelial cell interactions, blood–brain barrier, antiretroviral therapy, nanomedicine

Rationale: HIV-1–induced interstitial pneumonitis (IP) is a serious complication of HIV-1 infection, characterized by inflammation and cellular infiltration in lungs, often leading to respiratory failure and death. The barrier function of the pulmonary endothelium is caused in part by tight junction (TJ) proteins, such as claudin-5. Peroxisome proliferator–activated receptor (PPAR)-γ is expressed in lung tissues and regulates inflammation. We hypothesize that HIV-1 induces vascular lung injury, and HIV-1–mediated damage of the pulmonary endothelium and IP is associated with dysregulation of PPAR-γ. Objectives: Investigate the effects of HIV-1 infection on the pulmonary microvasculature and the modulatory effects of the PPAR-γ ligands. Methods: Using human lung tissues, we demonstrated down-regulation of claudin-5 (marker of pulmonary barrier integrity), down-regulation of PPAR-γ transcription, and expression in lung tissues of HIV-1–infected humans with IP. Measurements and Main Results: Human lung microvascular endothelial cells expressed the TJ proteins claudin-5, ZO-1, and ZO-2; HIV-1 decreased TJ proteins expression and induced nuclear factor-κB promoter activity, which was reversed by PPAR-γ agonist. Using two murine HIV/AIDS models, we demonstrated decreased claudin-5 expression and increased macrophage infiltration in the lungs of HIV-1–infected animals. Activation of PPAR-γ prevented HIV-1–induced claudin-5 down-regulation and significantly reduced viremia and pulmonary macrophage infiltration. Conclusions: HIV-induced IP is associated with injury to the lung vascular endothelium, with decreased TJ and PPAR-γ expression, and increased pulmonary macrophage infiltration. PPAR-γ ligands abrogated these effects. Thus, regulation of PPAR-γ can be a therapeutic approach against HIV-1–induced vascular damage and IP in infected humans. Removal of Expression of Concern: Issues leading to the previous expression of concern for this article have been resolved after further revisions and editorial review. No further concerns exist.

Background: Macrophage-carried nanoformulated catalase (‘nanozyme’) attenuates neuroinflammation and protects nigrostriatal neurons from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine intoxication. This is facilitated by effective enzyme transfer from blood-borne macrophages to adjacent endothelial cells and neurons leading to the decomposition of reactive oxygen species. Materials & methods: We examined the intra- and inter-cellular trafficking mechanisms of nanozymes by confocal microscopy. Improved neuronal survival mediated by nanozyme-loaded macrophages was demonstrated by fluorescence-activated cell sorting. Results: In macrophages, nanozymes were internalized mainly by clathrin-mediated endocytosis then trafficked to recycling endosomes. The enzyme is subsequently released in exosomes facilitated by bridging conduits. Nanozyme transfer from macrophages to adjacent cells by endocytosis-independent mechanisms diffusing broadly throughout the recipient cells. In contrast, macrophage-free nanozymes were localized in lysosomes following endocytic entry. Conclusion: Facilitated transfer of nanozyme from cell to cell can improve neuroprotection against oxidative stress commonly seen during neurodegenerative disease processes. Original submitted 29 June 2011; Revised submitted 14 September 2011; Published online 11 January 2012

Long-term antiretroviral therapy (ART) for human immunodeficiency virus type one (HIV-1) infection shows limitations in pharmacokinetics and biodistribution while inducing metabolic and cytotoxic aberrations. In turn, ART commonly requires complex dosing schedules and leads to the emergence of viral resistance and treatment failures. We posit that the development of nanoformulated ART could preclude such limitations and affect improved clinical outcomes. To this end, we wet-milled 20 nanoparticle formulations of crystalline indinavir, ritonavir, atazanavir, and efavirenz, collectively referred to as "nanoART," then assessed their performance using a range of physicochemical and biological tests. These tests were based on cell-nanoparticle interactions using monocyte-derived macrophages and their abilities to uptake and release nanoformulated drugs and affect viral replication. We demonstrate that physical characteristics such as particle size, surfactant coating, surface charge, and most importantly shape are predictors of cell uptake and antiretroviral efficacy. These studies bring this line of research a step closer to developing nanoART that can be used in the clinic to affect the course of HIV-1 infection.

The human immunodeficiency virus-1 (HIV-1) enters target cells by binding its envelope glycoprotein gp120 to the CD4 receptor and/or coreceptors such as C-C chemokine receptor type 5 (CCR5; R5) and C-X-C chemokine receptor type 4 (CXCR4; X4), and R5-tropic viruses predominate during the early stages of infection. CCR5 antagonists bind to CCR5 to prevent viral entry. Maraviroc (MVC) is the only CCR5 antagonist currently approved by the United States Food and Drug Administration, the European Commission, Health Canada, and several other countries for the treatment of patients infected with R5-tropic HIV-1. MVC has been shown to be effective at inhibiting HIV-1 entry into cells and is well tolerated. With expanding MVC use by HIV-1-infected humans, different clinical outcomes post-approval have been observed with MVC monotherapy or combination therapy with other antiretroviral drugs, with MVC use in humans infected with dual-R5- and X4-tropic HIV-1, infected with different HIV-1 genotype or infected with HIV-2. This review discuss the role of CCR5 in HIV-1 infection, the development of the CCR5 antagonist MVC, its pharmacokinetics, pharmacodynamics, drug–drug interactions, and the implications of these interactions on treatment outcomes, including viral mutations and drug resistance, and the mechanisms associated with the development of resistance to MVC. This review also discusses available studies investigating the use of MVC in the treatment of other diseases such as cancer, graft-versus-host disease, and inflammatory diseases.

Blood-brain barrier (BBB) compromise and transendothelial migration of HIV-infected leukocytes into the central nervous system (CNS) underlies the neuropathogenesis of HIV-1 infection. How this occurs is incompletely understood. We used a proteomic platform integrating difference gel electrophoresis and tandem mass spectrometry peptide sequencing to determine the effects that HIV-1-infected macrophages have on human brain microvascular endothelial cell (HBMEC) protein profiles. HIV-1 infected monocyte-derived macrophages (MDM) induced the upregulation of over 200 HBMEC proteins. These included metabolic, voltage-gated ion channels, heat shock, transport, cytoskeletal, regulatory, and calcium binding proteins. Results were validated by Western blot analysis. We conclude that HIV-1-infected MDM affect the HBMEC proteome and, in this way, affect BBB dysfunction and the development of HIV-1 CNS disease.

Human immunodeficiency virus-1 (HIV-1) encephalitis is characterized by brain infiltration of virus-infected monocytes and macrophages. Cellular products and viral proteins secreted by infected cells likely play an important role in blood–brain barrier (BBB) impairment and the development of HIV-1-associated dementia (HAD). We previously demonstrated that HIV-1 envelope glycoprotein gp120 induces toxicity and alters expression of tight junction proteins in human brain microvascular endothelial cells (HBMECs). Here, we delineate the mechanisms of gp120-induced BBB dysfunction. Human brain microvascular endothelial cells expressed HIV-1 co-receptors (CCR5 and CXCR4). Exposure of HBMECs to gp120 derived from macrophage (CCR5) or lymphocyte (CXCR4)-tropic viruses decreased BBB tightness, increased permeability, and enhanced monocyte migration across in vitro BBB models. Blood–brain barrier integrity was restored after gp120 removal. CCR5 antibodies and inhibitors of myosin light chain kinase or protein kinase C (PKC) blocked gp120-enhanced monocyte migration and permeability of BBB in vitro. Exposure of HBMECs to gp120 induced release of intracellular calcium ([Ca2+]i) that was prevented by CCR5 antibody and partially blocked by CXCR4 antagonist. Human immunodeficiency virus-1 gp120 activated three PKC isoforms in HBMECs [PKC-α/βII, PKC(pan)-βII and PKC-ζλ]. Furthermore, specific PKC inhibitors (acting at the ATP-binding and calcium release site) blocked gp120-induced PKC activation and prevented increase in BBB permeability, supporting the biologic significance of these results. Thus, gp120 can cause dysfunction of BBB via PKC pathways and receptor mediated [Ca2+]irelease leading to cytoskeletal alterations and increased monocyte migration.

The blood–brain barrier (BBB) is the specialized system of brain microvascular endothelial cells (BMVEC) that shields the brain from toxic substances in the blood, supplies brain tissues with nutrients, and filters harmful compounds from the brain back to the bloodstream. The close interaction between BMVEC and other components of the neurovascular unit (astrocytes, pericytes, neurons, and basement membrane) ensures proper function of the central nervous system (CNS). Transport across the BBB is strictly limited through both physical (tight junctions) and metabolic barriers (enzymes, diverse transport systems). A functional polarity exists between the luminal and abluminal membrane surfaces of the BMVEC. As a result of restricted permeability, the BBB is a limiting factor for the delivery of therapeutic agents into the CNS. BBB breakdown or alterations in transport systems play an important role in the pathogenesis of many CNS diseases (HIV-1 encephalitis, Alzheimer's disease, ischemia, tumors, multiple sclerosis, and Parkinson's disease). Proinflammatory substances and specific disease-associated proteins often mediate such BBB dysfunction. Despite seemingly diverse underlying causes of BBB dysfunction, common intracellular pathways emerge for the regulation of the BBB structural and functional integrity. Better understanding of tight junction regulation and factors affecting transport systems will allow the development of therapeutics to improve the BBB function in health and disease.

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.