Bovine milk fat enriched in conjugated linoleic and vaccenic acids attenuates allergic airway disease in mice

Background: It has been argued that a reduction in the Western diet of anti-inflammatory unsaturated lipids, such as n-3 polyunsaturated fatty acids, has contributed to the increase in the frequency and severity of allergic diseases.

Objective: We investigated whether feeding milk fat enriched in conjugated linoleic acid and vaccenic acids (VAs) ('enriched' milk fat), produced by supplementing the diet of pasture-fed cows with fish and sunflower oil, will prevent development of allergic airway responses.

Methods: C57BL/6 mice were fed a control diet containing soybean oil and diets supplemented with milk lipids. They were sensitized by intraperitoneal injection of ovalbumin (OVA) on days 14 and 28, and challenged intranasally with OVA on day 42. Bronchoalveolar lavage fluid, lung tissues and serum samples were collected 6 days after the intranasal challenge.

Results: Feeding of enriched milk fat led to marked suppression of airway inflammation as evidenced by reductions in eosinophilia and lymphocytosis in the airways, compared with feeding of normal milk fat and control diet. Enriched milk fat significantly reduced circulating allergen-specific IgE and IgG1 levels, together with reductions in bronchoalveolar lavage fluid of IL-5 and CCL11. Treatment significantly inhibited changes in the airway including airway epithelial cell hypertrophy, goblet cell metaplasia and mucus hypersecretion. The two major components of enriched milk fat, cis-9, trans-11 conjugated linoleic acid and VA, inhibited airway inflammation when fed together to mice, whereas alone they were not effective.

Conclusion: Milk fat enriched in conjugated linoleic and VAs suppresses inflammation and changes to the airways in an animal model of allergic airway disease.

Unique IL-13Rα2-based HIV-1 vaccine strategy to enhance mucosal immunity, CD8(+) T-cell avidity and protective immunity.

We have established that mucosal immunization can generate high-avidity human immunodeficiency virus (HIV)- specific CD8þ T cells compared with systemic immunization, and interleukin (IL)-13 is detrimental to the functional avidity of these T cells. We have now constructed two unique recombinant HIV-1 vaccines that co-express soluble or membrane-bound forms of the IL-13 receptor a2 (IL-13Ra2), which can ‘‘transiently’’ block IL-13 activity at the vaccination site causing wild-type animals to behave similar to an IL-13 KO animal. Following intranasal/intramuscular prime-boost immunization, these IL-13Ra2-adjuvanted vaccines have shown to induce (i) enhanced HIV-specific CD8þ Tcells with higher functional avidity, with broader cytokine/chemokine profiles and greater protective immunity using a surrogate mucosal HIV-1 challenge, and also (ii) excellent multifunctional mucosal CD8þ T-cell responses, in the lung, genito-rectal nodes (GN), and Peyer’s patch (PP). Data revealed that intranasal delivery of these IL-13Ra2-adjuvanted HIV vaccines recruited large numbers of unique antigen-presenting cell subsets to the lung mucosae, ultimately promoting the induction of high-avidity CD8þ Tcells. We believe our novel IL-13R cytokine trap vaccine strategy offers great promise for not only HIV-1, but also as a platform technology against range of chronic infections that require strong sustained high-avidity mucosal/systemic immunity for protection.

Inhibition of A/Human/Hubei/3/2005 (H3N2) influenza virus infection by silver nanoparticles in vitro and in vivo

Abstract
Silver nanoparticles (AgNPs) have attracted much attention as antimicrobial agents and have demonstrated efficient inhibitory activity against various viruses, including human immunodeficiency virus, hepatitis B virus, and Tacaribe virus. In this study, we investigated if AgNPs could have antiviral and preventive effects in A/Human/Hubei/3/2005 (H3N2) influenza virus infection. Madin-Darby canine kidney cells infected with AgNP-treated H3N2 influenza virus showed better viability (P,0.05 versus influenza virus control) and no obvious cytopathic effects compared with an influenza virus control group and a group treated with the solvent used for preparation of the AgNPs. Hemagglutination assay indicated that AgNPs could significantly inhibit growth of the influenza virus in Madin-Darby canine kidney cells (P,0.01 versus the influenza virus control). AgNPs significantly reduced cell apoptosis induced by H3N2 influenza virus at three different treatment pathways (P,0.05 versus influenza virus control). H3N2 influenza viruses treated with AgNPs were analyzed by transmission electron microscopy and found to interact with each other, resulting in destruction of morphologic viral structures in a time-dependent manner in a time range of 30 minutes to 2 hours. In addition, intranasal AgNP administration in mice significantly enhanced survival after infection with the H3N2 influenza virus. Mice treated with AgNPs showed lower lung viral titer levels and minor pathologic lesions in lung tissue, and had a marked survival benefit during secondary intranasal passage in vivo. These results provide evidence that AgNPs have beneficial effects in preventing H3N2 influenza virus infection both in vitro and in vivo, and demonstrate that AgNPs can be used as potential therapeutics for inhibiting outbreaks of influenza.

Recombinant influenza viruses as a vaccine vector for HIV

 Live recombinant influenza viruses were successfully used as HIV vaccine vectors in a mouse model. Following intranasal prime-boost vaccination, HIV-specific CD8+ T cell responses were detected in the spleen, broncho-alveolar lavage, mediastinal and inguinal lymph nodes. HIV+α4β7+ CD8+ T cells contributed to protection in pseudo-challenge experiments using recombinant vaccinia virus expressing HIV antigens. This research highlights the importance of mucosal CD8+ T cells in viral immunity and emphasizes the need for additional studies to provide key insights to underpin future vaccine development.

Epitope-specific CD4+, but not CD8+, T-cell responses induced by recombinant influenza A viruses protect against Mycobacterium tuberculosis infection

Tuberculosis remains a global health problem, in part due to failure of the currently available vaccine, BCG, to protect adults against pulmonary forms of the disease. We explored the impact of pulmonary delivery of recombinant influenza A viruses (rIAVs) on the induction of Mycobacterium tuberculosis (M. tuberculosis)-specific CD4(+) and CD8(+) T-cell responses and the resultant protection against M. tuberculosis infection in C57BL/6 mice. Intranasal infection with rIAVs expressing a CD4(+) T-cell epitope from the Ag85B protein (PR8.p25) or CD8(+) T-cell epitope from the TB10.4 protein (PR8.TB10.4) generated strong T-cell responses to the M. tuberculosis-specific epitopes in the lung that persisted long after the rIAVs were cleared. Infection with PR8.p25 conferred protection against subsequent M. tuberculosis challenge in the lung, and this was associated with increased levels of poly-functional CD4(+) T cells at the time of challenge. By contrast, infection with PR8.TB10.4 did not induce protection despite the presence of IFN-γ-producing M. tuberculosis-specific CD8(+) T cells in the lung at the time of challenge and during infection. Therefore, the induction of pulmonary M. tuberculosis epitope-specific CD4(+), but not CD8(+) T cells, is essential for protection against acute M. tuberculosis infection in the lung.

Oxytocin enhances the appropriate use of human social cues by the domestic dog (Canis familiaris) in an object choice task

It has been postulated that the neuropeptide, oxytocin, is involved in human-dog bonding. This may explain why dogs, compared to wolves, are such good performers on object choice tasks, which test their ability to attend to, and use, human social cues in order to find hidden food treats. The objective of this study was to investigate the effect of intranasal oxytocin administration, which is known to increase social cognition in humans, on domestic dogs' ability to perform such a task. We hypothesised that dogs would perform better on the task after an intranasal treatment of oxytocin. Sixty-two (31 males and 31 females) pet dogs completed the experiment over two different testing sessions, 5-15 days apart. Intranasal oxytocin or a saline control was administered 45 min before each session. All dogs received both treatments in a pseudo-randomised, counterbalanced order. Data were collected as scores out of ten for each of the four blocks of trials in each session. Two blocks of trials were conducted using a momentary distal pointing cue and two using a gazing cue, given by the experimenter. Oxytocin enhanced performance using momentary distal pointing cues, and this enhanced level of performance was maintained over 5-15 days time in the absence of oxytocin. Oxytocin also decreased aversion to gazing cues, in that performance was below chance levels after saline administration but at chance levels after oxytocin administration.

Patogenicidade e vacinologia de amostras brasileiras de Herpesvírus bovino tipo 1.

O herpesvírus bovino tipo 1 (BHV-1) está amplamente disseminado no rebanho bovino brasileiro. Visando aprofundar o conhecimento sobre a patogenicidade para o trato respiratório de amostras dos subtipos deste vírus mais freqüentemente encontrados no Brasil (BHV-1.1 e BHV-1.a), isolados autóctones oriundos de casos de doença respiratória foram inoculados em bovinos suscetíveis. As amostras de ambos os subtipos foram capazes de induzir doença respiratória após inoculação pela via intranasal, em intensidade que variou de moderada a grave, independentemente do subtipo de vírus inoculado. Foi ainda caracterizada a resposta imune humoral induzida por tais amostras quanto ao perfil de classes e subclasses de imunoglobulinas, sendo esta igualmente indistinguível com relação aos vírus inoculados. O perfil de classes de imunoglobulinas apresentadas pelos animais permitiu a determinação do status da infecção nos animais inoculados através da análise da resposta sorológica. Na segunda etapa deste trabalho foram testadas as propriedades vacinais de um vírus recombinante, do qual foi deletada a glicoproteína E (gE; 265gE-), gerado a partir de uma amostra brasileira de BHV-1.2a. Experimentos de inoculação do recombinante 265gE- em animais suscetíveis e o desafio destes animais com a amostra parental virulenta demonstraram a segurança e eficácia desta amostra na prevenção de sinais clínicos da infecção pelo BHV-1. Posteriormente, o recombinante 265gE- foi inoculado em vacas em diferentes estágios da gestação. Não foram observadas quaisquer anormalidades nestas gestações e as 22 vacas vacinadas deram à luz a animais saudáveis. Conclui-se que o recombinante é imunogênico e capaz de conferir significativa proteção frente ao desafio com a amostra parental virulenta do vírus. O recombinante também não causou enfermidade nas vacas inoculadas nem tampouco aos fetos quando inoculado em diferentes estágios da gestação.

Adolescência e infecção pelo HIV : situações de risco e proteção, autoconceito e sintomatologia psiquiátrica

Esta dissertação tem como objetivo identificar, através de um estudo transversal, associações entre as características demográficas, comportamentos de risco sexual e por drogas, sintomatologia psiquiátrica e autoconceito em uma amostra de adolescentes, comparando-os pela soropositividade HIV. Foram entrevistados 388 adolescentes entre 13 e 20 anos que procuraram espontaneamente o Centro de Triagem e Avaliação (CTA) Paulo César Bonfim para realizar o teste anti-HIV. Foram utilizados os questionários CRA (Comportamentos de Risco para AIDS), SCL-90-R (The Symptom Check-List-90-R) e EFA (Escala Fatorial de Autoconceito). A média da idade foi de 17,7 anos (±2,0). A maioria da amostra era do sexo feminino, solteiro ou sem companheiro/a fixo/a, sem trabalho, com baixas escolaridade e renda familiar. Em relação ao teste sorológico, 73% estavam realizandoo pela primeira vez, e a soropositividade geral da amostra foi de 6,2%. A escolaridade foi significativamente mais baixa em soropositivos (OR= 7,8) e não houve diferença estatística entre os grupos nas demais variáveis demográficas. Houve associação direta entre soropositividade e precocidade de início da vida sexual (OR= 3,6), história de gravidez (OR=4,7), prática de aborto prévio (OR= 5,9) e relação sexual com parceiro sem preservativo (OR=3,2). Do total da amostra, 8% dos adolescentes recebeu dinheiro em troca de relação sexual, e isto foi associado à soropositividade (OR=4,5). O mesmo ocorreu nos 13,2% que tiveram relação sexual com paceiro/a possivelmente soropositivo (OR=8,3). Apenas 18,9% da amostra afirmaram usar preservativos nos 6 meses prévios à coleta e 81,4% da amostra não haviam usado preservativo nas relações sexuais ao longo da vida. A prevalência geral de uso de drogas no mês anterior à coleta foi alta, com relato freqüente de uso de múltiplas drogas, sendo as mais freqüentes o álcool (71,6 %), maconha (24,7%), cocaína intranasal (12,4%) e solventes (9,3 %). A soropositividade foi mais alta nos usuários recentes de maconha (OR= 2,8), estimulantes (OR=2,9) e solventes (OR= 7,5). A freqüência a local específico para uso de drogas ocorreu em 22,5% do total da amostra e foi estatisticamente maior nos soropositivos (OR=2,6). Os adolescentes soropositivos apresentaram siginficativamente mais sintomatologia psiquiátrica em todas as dimensões avaliadas. Não houve diferença nas dimensões de autoconceito. O Tamanho de Efeito Padronizado foi maior nas variáveis da sintomatologia psiquiátrica do que nas do autoconceito. Os resultados indicam que os adolescentes estudados ainda não possuem cuidados preventivos suficientes para diminuir o contágio pelo vírus da AIDS. São indivíduos sob risco constante por apresentarem maior freqüência de exposição ao HIV associada à baixa escolaridade, comportamento sexual, uso de drogas e sintomatologia psiquiátrica. A falta de diferenças no autoconceito dos adolescentes soropositivos e soronegativos pode estar associada ao senso de invulnerabilidade próprio da faixa etária. Os achados apontam para a necessidade de programas específicos de prevenção para adolescentes. Medidas de intervenção futuras deverão levar em conta a interação de fatores individuais, ambientais, comportamentais e psicológicos, com o objetivo de implementar mudanças efetivas no comportamento de risco para que se detenha o avanço da epidemia nessa faixa etária.

Surfactant systems for nasal zidovudine delivery: structural, rheological and mucoadhesive properties

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Experimental murine mycobacteriosis: evaluation of the functional activity of alveolar macrophages in thalidomide- treated mice

Thalidomide is a selective inhibitor of tumor necrosis factor-alpha (TNF-alpha), a cytokine involved in mycobacterial death mechanisms. We investigated the role of this drug in the functional activity of alveolar macrophages in the presence of infection induced by intranasal inoculation of Mycobacterium avium in thalidomide-treated and untreated adult Swiss mice. Sixty animals were inoculated with 5 x 10(6) M. avium by the respiratory route. Thirty animals received daily thalidomide (30 mg/kg mouse) and 30 received water by gavage up to sacrifice. Ten non-inoculated mice were used as a control group. Lots of animals from each group were evaluated until 6 weeks after inoculation. Infection resulted in an increased total number of inflammatory cells as well as increased activity of pulmonary macrophages. Histologically, intranasal inoculation of bacilli resulted in small mononuclear infiltrates located at the periphery of the organ. Culture of lung fragments revealed the presence of bacilli only at the beginning and at the end of the experimental period. Thalidomide administration did not affect the microbiological or histological features of the infection. Thalidomide-treated and untreated animals showed the same amount of M. avium colonies 3 weeks after infection. Although it did not affect bacillary clearance, thalidomide administration resulted in a decreased percent of spread cells and release of hydrogen peroxide, suggesting that factors other than TNF-alpha play a role in the killing of mycobacteria by alveolar macrophages. Thalidomide administration also reduced the number of spread cells among resident macrophages, suggesting a direct effect of the drug on this phenomenon.

Effect of annexin-A1 peptide treatment during lung inflammation induced by lipopolysaccharide

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Liposomes and micro/nanoparticles as colloidal carriers for nasal drug delivery

The use of the nasal route for drug delivery has attracted much interest in recent years in the pharmaceutical field. Local and principally systemic drug delivery can be achieved by this route of administration. But the nasal route of delivery is not applicable to all drugs. Polar drugs and some macromolecules are not absorbed in sufficient concentration due to poor membrane permeability, rapid clearance and enzymatic degradation into the nasal cavity. Thus, alternative means that help overcome these nasal barriers are currently in development. Absorption enhancers such as phospholipids and surfactants are constantly used, but care must be taken in relation to their concentration. Drug delivery systems including liposomes, cyclodextrins, micro- and nanoparticles are being investigated to increase the bioavailability of drugs delivered intranasally. This review article discusses recent progress and specific development issues relating to colloidal drug delivery systems in nasal drug delivery. © 2006 Bentham Science Publishers Ltd.

Padronização dos critérios de seleção em estudos sobre medicações nasais

Clinical studies on nasal topical medications require the standardization of nasosinusal normality in order to establish control groups through a specific evaluation of the upper airways. Aim: to standardize the evaluation of candidates for control groups in clinical studies on nasal topical medications. Material and Methods: healthy male volunteers of 18 to 50 years of age, asymptomatic from the nasosinusal standpoint were subjected to a sequential and excluding assessment made up of clinical evaluation, immediate hypersensitivity skin test, saccharin test, flexible nasofibroscopy and nasal cytology. Study design: Crosssectional contemporary cohort. Results: Of the 33 people originally enrolled, 14 (42.4%) were excluded for clinical reasons. Of the 19 remaining, 2 (10.5%) had atopy diagnosed in the skin test and were excluded. 17 were tested with saccharin and presented normal mucociliary clearance. Evaluation by nasal endoscopy showed abnormality in 2 cases (11.8%) and these were excluded. The remaining 15 were submitted to nasal cytology, which proved normal, representing 45.5% of those initially included. Conclusion: The proposed protocol for sequential and excluding evaluation was effective in defining candidates for the establishment of control groups in clinical studies on nasal topical medications. © Revista Brasileira de Otorrinolaringologia. All Rights reserved.

Cell Therapy with Bone Marrow Mononuclear Cells in Elastase-Induced Pulmonary Emphysema

Emphysema is characterized by destruction of alveolar walls with loss of gas exchange surface and consequent progressive dyspnea. This study aimed to evaluate the efficiency of cell therapy with bone marrow mononuclear cells (BMMC) in an animal model of elastase-induced pulmonary emphysema. Emphysema was induced in C57Bl/J6 female mice by intranasal instillation of elastase. After 21 days, the mice received bone marrow mononuclear cells from EGFP male mice with C57Bl/J6 background. The groups were assessed by comparison and statistically significant differences (p & 0. 05) were observed among the groups treated with BMMC and evaluated after 7, 14 and 21 days. Analysis of the mean linear intercept (Lm) values for the different groups allowed to observe that the group treated with BMMC and evaluated after 21 days showed the most significant result. The group that received no treatment showed a statistically significant difference when compared to other groups, except the group treated and evaluated after 21 days, evidencing the efficacy of cell therapy with BMMC in pulmonary emphysema. © 2012 Springer Science+Business Media New York.

Nasal administration of liquid crystal precursor mucoadhesive vehicle as an alternative antiretroviral therapy

The purpose of this study was to develop a mucoadhesive stimuli-sensitive drug delivery system for nasal administration of zidovudine (AZT). The system was prepared by formulating a low viscosity precursor of a liquid crystal phase, taking advantage of its lyotropic phase behavior. Flow rheology measurements showed that the formulation composed of PPG-5-CETETH-20, oleic acid and water (55, 30, 15% w/w), denominated P, has Newtonian flow behavior. Polarized light microscopy (PLM) revealed that formulation P is isotropic, whereas its 1:1 (w/w) dilution with artificial nasal mucus (ANM) changed the system to an anisotropic lamellar phase (PD). Oscillatory frequency sweep analysis showed that PD has a high storage modulus (G′) at nasal temperatures. Measurement of the mucoadhesive force against excised porcine nasal mucosa or a mucin disk proved that the transition to the lamellar phase tripled the work of mucoadhesion. Ex vivo permeation studies across porcine nasal mucosa exhibited an 18-fold rise in the permeability of AZT from the formulation. The Weibull mathematical model suggested that the AZT is released by Fickian diffusion mechanisms. Hence, the physicochemical characterization, combined with ex vivo studies, revealed that the PPG-5-CETETH-20, oleic acid, and water formulation could form a mucoadhesive matrix in contact with nasal mucus that promoted nasal absorption of the AZT. For an in vivo assessment, the plasma concentrations of AZT in rats were determined by HPLC method following intravenous and intranasal administration of AZT-loaded P formulation (PA) and AZT solution, respectively, at a dose of 8 mg/kg. The intranasal administration of PA resulted in a fast absorption process (Tmax = 6.7 min). Therefore, a liquid crystal precursor formulation administered by the nasal route might represent a promising novel tool for the systemic delivery of AZT and other antiretroviral drugs. In the present study, the uptake of AZT absorption in the nasal mucosa was demonstrated, providing new foundations for clinical trials in patients with AIDS. © 2012 Elsevier B.V. All rights reserved.