RESUMO
The envelope glycoprotein (Env) of the human immunodeficiency virus (HIV), has been the primary target for the development of a protective vaccine against infection. The extensive N-linked glycosylation on Env is an important consideration as it may affect efficacy, stability, and expression yields. The expression host has been shown to influence the extent and type of glycosylation that decorates the protein target. Here, we report the glycosylation profile of the candidate subtype C immunogen CO6980v0c22 gp145, which is currently in Phase I clinical trials, produced in two different host cells: CHO-K1 and Expi293F. The amino acid sequence for both glycoproteins was confirmed to be identical by peptide mass fingerprinting. However, the isoelectric point of the proteins differed; 4.5-5.5 and 6.0-7.0 for gp145 produced in CHO-K1 and Expi293F, respectively. These differences in pI were eliminated by enzymatic treatment with sialidase, indicating a large difference in the incorporation of sialic acid between hosts. This dramatic difference in the number of sialylated glycans between hosts was confirmed by analysis of PNGase F-released glycans using MALDI-ToF MS. These differences in glycosylation, however, did not greatly translate into differences in antibody recognition. Biosensor assays showed that gp145 produced in CHO-K1 had similar affinity toward the broadly neutralizing antibodies, 2G12 and PG16, as the gp145 produced in Expi293F. Additionally, both immunogens showed the same reactivity against plasma of HIV-infected patients. Taken together, these results support the notion that there are sizeable differences in the glycosylation of Env depending on the expression host. How these differences translate to vaccine efficacy remains unknown.
Assuntos
Glicopeptídeos/análise , Anticorpos Anti-HIV/imunologia , HIV-1/metabolismo , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Adulto , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Reações Antígeno-Anticorpo , Células CHO , Cricetinae , Cricetulus , Feminino , Glicosilação , Células HEK293 , Humanos , Pessoa de Meia-Idade , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificação , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Adulto Jovem , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/metabolismoRESUMO
BACKGROUND: Low cardiorespiratory fitness (CRF) is usually observed in people living with HIV. The effect of a low-volume high-intensity interval training (LV-HIIT) on CRF in HIV+ and HIV- Hispanic women was evaluated in this study. SETTING: A nonrandomized clinical trial with pre-test and post-test using a LV-HIIT intervention was conducted in the AIDS Clinical Trials Unit and the Puerto Rico Clinical and Translational Research Consortium at the University of Puerto Rico Medical Sciences Campus. METHODS: Twenty-nine HIV+ and 13 HIV- Hispanic women recruited from community-based programs and clinics, and able to engage in daily physical activities, volunteered to participate. Of these, 20 HIV+ (69%) and 11 HIV- (85%) completed the study and were included in the analyses. LV-HIIT consisted of 6-week, 3 d/wk, 8-10 high-intensity and low-intensity intervals on a cycle ergometer at 80%-90% of heart rate reserve. Main outcome measures were CRF (defined as VO2peak), peak workload, and time to peak exercise. RESULTS: Average peak workload and time to peak exercise increased after training (P < 0.05) in both groups. However, average CRF was significantly higher after training only in the HIV- group. Gains in CRF were observed in 100% of HIV- and 50% of HIV+ women. This was not influenced by exercise testing, habitual physical activity, or anthropometric variables. CONCLUSIONS: Given the lack of change in CRF observed in the HIV+ group after LV-HIIT intervention, it is important to focus on variations that may occur within groups.
Assuntos
Exercício Físico , Infecções por HIV , Treinamento Intervalado de Alta Intensidade/métodos , Adulto , Antropometria , Aptidão Cardiorrespiratória , Feminino , Infecções por HIV/fisiopatologia , Hispânico ou Latino , Humanos , Ensaios Clínicos Controlados não Aleatórios como Assunto , Consumo de Oxigênio , Porto RicoRESUMO
Zika has been one of the most devastating new emerging viruses epidemic due to its effects on the nervous system mostly newborns. It was an unexpected epidemic that jeopardized our population and those with the appropriate environment for the virus to replicate, the tropics. It took us by surprise and challenged our health system. When it arrived in Puerto Rico in December 2015 many questions were raised regarding how the clinical manifestations of the infection will affect our population. Although most of infections go on asymptomatic it was not clear how our population heavily infected with other arboviruses such as dengue and chikungunya will alter the clinical and neurological manifestations of Zika. Other questions regarding our health system preparedness to attend to such an epidemic was questioned. We hope that these experiences will aid in the establishment of procedures to deal with similar epidemics in the future and help to gain a better understanding of the acute and chronic effects of the infection in the clinical manifestations and possible management of the infection in a clinical and public health manner. Puerto Rico possess a special position in the Caribbean and can serve to spearhead the establishment of procedures to detect, treat, and study the effects of these new emerging virus epidemics. Thereby establishing the needed programs to attend to other similar epidemics.
Assuntos
Epidemias , Infecção por Zika virus/epidemiologia , Feminino , Humanos , Recém-Nascido , Saúde Pública , Porto Rico/epidemiologiaRESUMO
OBJECTIVE: HIV-associated cognitive impairment (HACI) continues to persist for HIV-seropositive individuals who are on antiretroviral therapy (ART). HACI develops in part when HIV-infected monocytes (MOs) transmigrate through the blood-brain barrier (BBB) and secrete pro-inflammatory cytokines and chemokines, which leads to neuronal damage. In vitro BBB models are important tools that can elucidate mechanisms of MO transmigration. Previously described in vitro BBB models relied on pathology specimens, resulting in potentially variable and inconsistent results. This project reports on a reliable and consistent alternative in vitro BBB model that has the potential to be used in clinical research intervention studies analyzing the effects of ART on the BBB and on MO transmigration. METHODS: A bilayer BBB model was established with commercially available astrocytes and endothelial cells on a 3µm PET membrane insert to allow the contact of astrocytic foot processes with endothelial cells. Inserts were cultured in growth medium for 7 days before exposure to HIV- or HIV+ peripheral blood mononuclear cells (PBMCs). PBMCs were allowed to transmigrate across the BBB for 24 hours. RESULTS: Confluency and integrity measurements by trans-endothelial electrical resistance (TEER) (136.7 ± 18.3Ω/cm2) and permeability (5.64 ± 2.20%) verified the integrity of the in vitro BBB model. Transmigrated MOs and non-MOs were collected and counted (6.0x104 MOs; 1.1x105 non-MOs). Markers indicative of glial fibrillary acidic protein (GFAP), von Willebrand factor (vWF), and p-glycoprotein (Pgp) were revealed in immunofluorescence staining (IF), indicating BBB phenotype and functionality. CONCLUSION: Potential applications for this model include assessing the HIV DNA copy numbers of transmigrated cells (pre- and post-targeted ART) and understanding the role of oxidative stress related to HIV DNA and HACI.
Assuntos
Barreira Hematoencefálica , Movimento Celular , Leucócitos Mononucleares/fisiologia , Modelos Biológicos , Pesquisa Biomédica , Células Cultivadas , Soropositividade para HIV/sangue , Soropositividade para HIV/tratamento farmacológico , HumanosRESUMO
BACKGROUND: HIV-associated vulnerabilities-especially those linked to psychological issues-and limited mental health-treatment resources have the potential to adversely affect the health statuses of individuals. The concept of resilience has been introduced in the literature to shift the emphasis from vulnerability to protective factors. Resilience, however, is an evolving construct and is measured in various ways, though rarely among underserved, minority populations. Herein, we present the preliminary psychometric properties of a sample of HIV-seropositive Puerto Rican women, measured using a newly developed health-related resilience scale. METHODS AND DESIGN: The Resilience Scales for Children and Adolescents, an instrument with solid test construction properties, acted as a model in the development (in both English and Spanish) of the HRRS, providing the same dimensions and most of the same subscales. The present sample was nested within the Hispanic-Latino longitudinal cohort of women (HLLC), that is part of the NeuroAIDS Research Program at the University of Puerto Rico (UPR), Medical Sciences Campus (MSC). Forty-five consecutively recruited, HIV+ women from the HLLC completed a demographic survey, the HRRS, and the Beck Depression Inventory-I, Spanish version. RESULTS: The results demonstrate excellent overall internal consistency for the total HRRS score (α = 0.95). Each of the dimensional scores also evidenced acceptable internal consistency (α ≥ 0.88). All the dimensional and subscale content validity indices were above the 0.42 cut-off. Analysis revealed a significant negative correlation between the HRRS total score and BDI-I-S (r(45) = -0.453, p < 0.003). CONCLUSION: Albeit preliminary in nature, the present study provides support for the HRRS as a measure to assess resilience among individuals living with chronic medical conditions. Minority populations, especially non-English speaking ones, are understudied across the field of medicine, and when efforts are made to include these patient groups, measurement is rarely tailored to their unique cultural and linguistic experiences. The HRRS is a measure that addresses these notable voids in the medical literature.
Assuntos
Infecções por HIV/psicologia , Soropositividade para HIV/psicologia , Hispânico ou Latino/psicologia , Escalas de Graduação Psiquiátrica , Resiliência Psicológica , Adulto , Feminino , Humanos , Pessoa de Meia-Idade , Psicometria , Porto Rico , Reprodutibilidade dos TestesRESUMO
OBJECTIVE: HIV-infected monocytes can infiltrate the blood brain barrier as differentiated macrophages to the central nervous system, becoming the primary source of viral and cellular neurotoxins. The final outcome is HIV-associated cognitive impairment (HACI), which remain prevalent today, possibly due to the longer life-span of the patients treated with combined anti-retroviral therapy. Our main goal was to characterize the proteome of monocyte-derived macrophages (MDM) from HACI patients, and its association with their cognitive status, to find novel targets for therapy. METHODS: MDM were isolated from the peripheral blood of 14 HIV-seropositive women characterized for neurocognitive function, including: four normal cognition (NC), five asymptomatic (A), and five with cognitive impaired (CI). Proteins from macrophage lysates were isobaric-labeled with the microwave and magnetic (M2) sample preparation method followed by liquid chromatography-tandem mass spectrometry-based protein identification and quantification. Differences in protein abundance across groups classified by HACI status were determined using analysis of variance. RESULTS: A total of 2,519 proteins were identified with 2 or more peptides and 28 proteins were quantified as differentially expressed. Statistical analysis revealed increased abundance of 17 proteins in patients with HACI (p<0.05), including several enzymes associated to the glucose metabolism. Western blot confirmed increased expression of 6-Phosphogluconate dehydrogenase and L-Plastin in A and CI patients over NC and HIV seronegatives. CONCLUSIONS: This is the first quantitative proteomics study exploring the changes in protein abundance of macrophages isolated from patients with HACI. Further studies are warranted to determine if these proteins may be target candidates for therapy development against HACI.
Assuntos
Transtornos Cognitivos/metabolismo , Infecções por HIV/metabolismo , Macrófagos/metabolismo , Proteoma/análise , Proteômica/métodos , Análise de Variância , Western Blotting , Células Cultivadas , Cromatografia Líquida , Transtornos Cognitivos/complicações , Estudos de Coortes , Estudos Transversais , Feminino , Infecções por HIV/complicações , Humanos , Magnetismo , Micro-Ondas , Mapas de Interação de Proteínas , Proteoma/metabolismo , Proteômica/instrumentação , Espectrometria de Massas em TandemRESUMO
BACKGROUND: Zika virus has been associated with increases in Guillain-Barré syndrome (GBS) incidence. A GBS incidence estimation and clinical description was performed to assess baseline GBS epidemiology before the introduction of Zika virus in Puerto Rico. METHODS: Hospitalization administrative data from an island-wide insurance claims database and U.S. Census Bureau population estimates provided a crude GBS incidence for 2013. This estimate was adjusted using the proportion of GBS cases meeting Brighton criteria for confirmed GBS from nine reference hospitals. Characteristics of confirmed GBS cases in the same nine hospitals during 2012-2015 are described. RESULTS: A total of 136 GBS hospitalization claims were filed in 2013 (crude GBS incidence was 3.8 per 100,000 population). The adjusted GBS incidence was 1.7 per 100,000 population. Of 67 confirmed GBS cases during 2012-2015, 66% had an antecedent illness. Median time from antecedent illness to GBS onset was 7days. Most cases (67%) occurred during July-September. CONCLUSIONS: Puerto Rico's GBS incidence for 2013 was estimated using a combination of administrative data and medical records review; this method could be employed in other regions to monitor GBS incidence before and after the introduction of GBS infectious triggers.
Assuntos
Síndrome de Guillain-Barré/epidemiologia , Síndrome de Guillain-Barré/virologia , Infecção por Zika virus/epidemiologia , Bases de Dados Factuais/estatística & dados numéricos , Surtos de Doenças , Feminino , Hospitalização/estatística & dados numéricos , Humanos , Incidência , Seguro Médico Ampliado/estatística & dados numéricos , Masculino , Vigilância da População , Porto Rico/epidemiologiaRESUMO
Antiretroviral therapy partially restores the immune system and markedly increases life expectancy of HIV-infected patients. However, antiretroviral therapy does not restore full health. These patients suffer from poorly understood chronic inflammation that causes a number of AIDS and non-AIDS complications. Here we show that chronic inflammation in HIV+ patients may be due to the disruption of the cholinergic anti-inflammatory pathway by HIV envelope protein gp120IIIB. Our results demonstrate that HIV gp120IIIB induces α7 nicotinic acetylcholine receptor (α7) upregulation and a paradoxical proinflammatory phenotype in macrophages, as activation of the upregulated α7 is no longer capable of inhibiting the release of proinflammatory cytokines. Our results demonstrate that disruption of the cholinergic-mediated anti-inflammatory response can result from an HIV protein. Collectively, these findings suggest that HIV tampering with a natural strategy to control inflammation could contribute to a crucial, unresolved problem of HIV infection: chronic inflammation.
RESUMO
OBJECTIVE: HIV-1 variants with different tropisms are associated with various neuropathologies. This study intends to determine if this correlation is determined by unique viral env sequences. We hypothesize that HIV-1 envelope gene sequence changes are associated with cognition status. METHODS: Viral RNA was extracted from peripheral blood mononuclear cells (PBMCs) co-cultures derived from HIV-1 infected Hispanic women that had been characterized for HIV associated neurocognitive disorders (HAND). RESULTS: Analyses of the C2V4 region of HIV gp120 demonstrated that increased sequence diversity correlates with cognition status as sequences derived from subjects with normal cognition exhibited less diversity than sequences derived from subjects with cognitive impairment. In addition, differences in V3 and V4 loop charges were also noted as well as differences in the N-glycosylation of the V4 region. CONCLUSIONS: Our data suggest that the genetic signature within the C2V4 region may contribute to the pathogenesis of HAND. HIV env sequence characteristics for the isolates grouped in milder forms of HAND can provide insightful information of prognostic value to assess neurocognitive status in HIV+ subjects, particularly during the era of highly prevalent milder forms of HAND.