Health-related quality of life of adult and adolescent patients living with alopecia areata in Australia.

INTRODUCTION: To understand the experiences of adolescent and adult patients living with alopecia areata (AA) in Australia regarding symptom severity and the impact on psychosocial well-being and work/classroom productivity. MATERIALS AND METHODS: A cross-sectional online patient survey among adolescent and adult patients diagnosed with AA was recruited via the Australia Alopecia Areata Foundation. Patient-reported outcomes were also assessed. RESULTS: A total of 337 patients (49 adolescents; 288 adults), with a mean ± standard deviation age of 14.7 ± 1.55 and 38.9 ± 13.31 years for adolescents and adults, respectively, were included. In the group with extensive hair loss (Scalp Hair Assessment Patient-Reported Outcome, categories 3 + 4, n = 172), we observed higher emotional symptom and activity limitation scores (Alopecia Areata Patient Priority Outcomes, emotional symptoms: adults 2.5 ± 1.03, adolescents 2.2 ± 1.15; activity limitations: adults 1.4 ± 1.15, adolescents 1.2 ± 0.99). Additionally, in adults, the Alopecia Areata Symptom Impact Scale global score was 4.0 ± 2.10 (symptoms subscale score 4.1 ± 1.91; interference subscale scores 3.8 ± 2.73). Hospital Anxiety and Depression Scale scores were high across participants, irrespective of hair loss extent (adults: anxiety 9.2 ± 3.85, depression 6.6 ± 3.95; adolescents: anxiety 9.7 ± 4.65, depression 5.2 ± 3.59). Work and classroom productivity were substantially impaired due to AA, with 70.5% of adults and 57.1% of adolescents reporting activity impairment, and overall work/classroom impairment reported at 39.2% and 44.9%, respectively. CONCLUSIONS: AA impacts the physical, emotional and psychosocial well-being of both adult and adolescent patients. More extensive hair loss more profoundly impacts those living with AA. Patients may benefit from patient-centred care approaches addressing the impact of hair loss on mental and emotional well-being, daily activities and work productivity.

Understanding AAV vector immunogenicity: from particle to patient.

Gene therapy holds promise for patients with inherited monogenic disorders, cancer, and rare genetic diseases. Naturally occurring adeno-associated virus (AAV) offers a well-suited vehicle for clinical gene transfer due to its lack of significant clinical pathogenicity and amenability to be engineered to deliver therapeutic transgenes in a variety of cell types for long-term sustained expression. AAV has been bioengineered to produce recombinant AAV (rAAV) vectors for many gene therapies that are approved or in late-stage development. However, ongoing challenges hamper wider use of rAAV vector-mediated therapies. These include immunity against rAAV vectors, limited transgene packaging capacity, sub-optimal tissue transduction, potential risks of insertional mutagenesis and vector shedding. This review focuses on aspects of immunity against rAAV, mediated by anti-AAV neutralizing antibodies (NAbs) arising after natural exposure to AAVs or after rAAV vector administration. We provide an in-depth analysis of factors determining AAV seroprevalence and examine clinical approaches to managing anti-AAV NAbs pre- and post-vector administration. Methodologies used to quantify anti-AAV NAb levels and strategies to overcome pre-existing AAV immunity are also discussed. The broad adoption of rAAV vector-mediated gene therapies will require wider clinical appreciation of their current limitations and further research to mitigate their impact.

Maternal mortality committee and death surveillance in Recife in improving information: ex-ante and ex-post evaluation / Comitê de mortalidade materna e a vigilância do óbito em Recife no aprimoramento das informações: avaliação ex-ante e ex-post

Abstract Objectives: to evaluate the contribution of the Maternal Mortality and Death Surveillance Committee for women of childbearing age (WCA) and maternal mortality in the magnitude of maternal mortality and in the qualification of the causes of death in Recife, Brazil. Methods: ex ante/ex post evaluation, ecological, of the annual indicators of mortality of WCA, maternal and case study of declared maternal deaths according to causes of death before and after surveillance. Deaths of WCA (2010 and 2017) were analyzed. The percentage of investigation of deaths of WCA was calculated; their rates and maternal mortality ratio (MMR) were estimated; the groups of causes of death, classification of death, the moment of death, the proportional variation before and after surveillance, and the relocation of the causes after this process were described. Results: 4.327 (97.0%) of deaths of WCA were investigated (increase of 40.7% of maternal deaths) and MMR of 62.9/100 thousand live births. Improved notifications of immediate/late (75.0%) and remote (300.0%) postpartum; there was a difference in direct obstetric causes, total maternal deaths and late maternal death (p<0.001). Conclusion: the surveillance and the Maternal Mortality Committee showed potential in identifying the magnitude and qualification of causes of maternal death in order to propose the interventions directed to obstetric care.

Resumo Objetivos: avaliar a contribuição do Comitê de Mortalidade Materna e da Vigilância do Óbito de mulheres em idade fértil (MIF) e materno na magnitude da mortalidade materna e na qualificação das causas dos óbitos no Recife, Brasil. Métodos: avaliação ex ante/ex post, ecológico, dos indicadores anuais de mortalidade de MIF, materna e estudo de caso de óbitos maternos declarados segundo causas de morte antes e após a vigilância. Analisaram-se óbitos de MIF (2010-2017) e calculou-se o percentual de investigação; estimaram-se suas taxas e a razão de mortalidade materna (RMM); descreveram-se: grupos de causa, classificação e momento do óbito, variação proporcional antes e após a vigilância/análise do comitê e a realocação das causas após esse processo. Resultados: investigou-se 4.327 (97,0%) dos óbitos de MIF (incremento de 40,7% das mortes maternas), e RMM de 62,9/100 mil nascidos vivos; melhoraram as notificações do puerpério imediato/ tardio (75,0%) e remoto (300,0%); houve diferença nas causas obstétricas diretas, total de óbitos maternos e morte materna tardia (p<0,001). Conclusão: mostrou-se o potencial da vigilância e do Comitê de Mortalidade Materna na identificação da magnitude e qualificação das causas de morte materna para proposição de medidas direcionadas aos cuidados obstétricos.

Sociodemographic and health care profile of maternal death in Recife, PE, Brazil, 2006-2017: a descriptive study. / Perfil sociodemográfico e assistencial da morte materna em Recife, 2006-2017: estudo descritivo.

OBJECTIVE: to describe the sociodemographic and health care characteristics of women dying due to maternal causes in Recife, Pernambuco, Brazil. METHODS: this was a descriptive study using the Mortality Information System, case investigation sheets and summary sheets of early and late maternal deaths occurring between 2006 and 2017, with avoidability assessed by the Municipal Maternal Mortality Committee. RESULTS: we identified 171 deaths, of which 133 were in the puerperium; most deaths occurred among Black women (68.4%), women without partners (60.2%), women who had prenatal care (77.2%), during maternity hospital/general hospital delivery (97.1%), women attended to by obstetricians (82.6%);10.4% of women with puerperal complications had no health care; avoidable/probably avoidable deaths corresponded to 81.9%, for indirect causes (n=80), and direct causes (n=79). CONCLUSION: deaths occurred mainly in the postpartum period, among Black women; care failures were frequent; improved health service surveillance and follow-up is needed in the pregnancy-puerperal period, in Recife.

Real-world evaluation of effectiveness, persistence, and usage patterns of tofacitinib in treatment of rheumatoid arthritis in Australia.

INTRODUCTION: The aim of this study was to describe the real-world evidence for effectiveness, treatment persistence, and treatment patterns among patients in the community with rheumatoid arthritis treated with the JAK inhibitor tofacitinib. METHODS: This was a retrospective, non-interventional cohort study that extracted data for new users of tofacitinib or biologic disease-modifying antirheumatic drugs (bDMARDs) from the Australian Optimizing Patient outcomes in Australian RheumatoLogy (OPAL) dataset between March 2015 and September 2018. Patients were propensity score matched at a 1:2 tofacitinib to bDMARD ratio based on age, sex, and selected baseline treatment combinations. Treatment effectiveness was evaluated using disease status measures. Treatment persistence was calculated and the percentage of patients receiving monotherapy or combination therapy at treatment initiation was evaluated. RESULTS: Data from 2810 patients were extracted and 1950 patients were included in the matched population (1300 bDMARD initiators and 650 tofacitinib initiators). Patients were predominantly aged 55 to 74 years (57.8%) and female (81.2%). After 18 months of treatment, 52.4% and 57.8% of patients had achieved disease activity score (DAS) remission in the bDMARD and tofacitinib groups, respectively. The median treatment persistence for tofacitinib was similar to that for bDMARDs: 34.2 months (95% CI 32.2 to not reached) and 33.8 months (95% CI 28.8 to 40.4), respectively. In the overall population, more patients were prescribed tofacitinib as monotherapy (43.4%) compared with bDMARD monotherapy (33.4%). CONCLUSIONS: Tofacitinib demonstrated treatment effectiveness and persistence similar to bDMARDs. Overall, there was a trend for more use of tofacitinib as monotherapy than bDMARDs. Key Points • This study provides real-world evidence regarding effectiveness, treatment persistence, and treatment patterns, among patients with rheumatoid arthritis (RA) in the community being treated with tofacitinib. • The study suggests that tofacitinib is an effective and enduring intervention in RA with tofacitinib persistence and effectiveness comparable to bDMARDs.

Perfil sociodemográfico e assistencial da morte materna em Recife, 2006-2017: estudo descritivo / Perfil sociodemográfico y asistencial de la muerte materna en Recife, PE, Brasil, 2006-2017: estudio descriptivo / Sociodemographic and health care profile of maternal death in Recife, PE, Brazil, 2006-2017: a descriptive study*

Objetivo: descrever características sociodemográficas e assistenciais de mulheres que morreram por causa materna em Recife, Pernambuco, Brasil. Métodos: estudo descritivo utilizando o Sistema de Informações sobre Mortalidade, fichas de investigação e fichas-síntese de óbitos maternos, precoces e tardios, ocorridos entre 2006 e 2017, com evitabilidade avaliada pelo Comitê Municipal de Mortalidade Materna. Resultados: identificaram-se 171 óbitos, 133 no puerpério; a maior parte dos óbitos ocorreu em negras (68,4%), sem companheiro (60,2%), acompanhadas com atendimento pré-natal (77,2%), de parto em maternidades/hospitais (97,1%), assistidas por obstetras (82,6%); das mulheres com complicações puerperais, 10,4% não tiveram assistência; óbitos evitáveis/provavelmente evitáveis corresponderam a 81,9%, por causas indiretas (n=80) e diretas (n=79). Conclusão: as mortes ocorreram principalmente no puerpério e em negras; falhas assistenciais foram frequentes; é necessária melhor vigilância e acompanhamento dos serviços de saúde no período gravídico-puerperal, em Recife.

Objetivo: describir características sociodemográficas y asistenciales de mujeres que murieron por causa materna en Recife, Pernambuco, Brasil. Métodos: estudio descriptivo utilizando el Sistema de Informaciones sobre Mortalidad, fichas de investigación y síntesis de muertes maternas, tempranas y tardías, entre 2006 y 2017, con evaluación de la evitabilidad por el Comité Municipal de la Mortalidad Materna. Resultados: se identificaron 171 óbitos maternos, 133 en el puerperio; la mayoría de las muertes ocurrió en negras (68,4%), sin compañero (60,2%), acompañadas con atención prenatal (77,2%), de parto en maternidades/hospitales (97,1%), asistidas por obstetras (82,6%); de las mujeres con complicaciones puerperales, el 10,4% no tuvo asistencia; muertes evitables/probablemente evitables correspondieron al 81,9%, por causas indirectas (n=80) y directas (n=79). Conclusión: las muertes ocurrieron principalmente en el período del puerperio y en mujeres negras, con frecuentes fallas en la atención; se requiere una mayor vigilancia y acompañamiento de los servicios de salud en el período de embarazo-puerperio, en Recife.

Objective: to describe the sociodemographic and health care characteristics of women dying due to maternal causes in Recife, Pernambuco, Brazil. Methods: this was a descriptive study using the Mortality Information System, case investigation sheets and summary sheets of early and late maternal deaths occurring between 2006 and 2017, with avoidability assessed by the Municipal Maternal Mortality Committee. Results: we identified 171 deaths, of which 133 were in the puerperium; most deaths occurred among Black women (68.4%), women without partners (60.2%), women who had prenatal care (77.2%), during maternity hospital/general hospital delivery (97.1%), women attended to by obstetricians (82.6%);10.4% of women with puerperal complications had no health care; avoidable/probably avoidable deaths corresponded to 81.9%, for indirect causes (n=80), and direct causes (n=79). Conclusion: deaths occurred mainly in the postpartum period, among Black women; care failures were frequent; improved health service surveillance and follow-up is needed in the pregnancy-puerperal period, in Recife.

Evaluation of the implantation of the Mortality Information System in Pernambuco state, Brazil, in 2012. / Avaliação da implantação do Sistema de Informações sobre Mortalidade no estado de Pernambuco em 2012.

OBJECTIVE: to evaluate the implantation of the Mortality Information System (SIM) in Pernambuco, Brazil. METHODS: this was an evaluation study; primary data (questionnaires) and secondary data (SIM) were used for the municipalities to estimate the degree of implantation (DI), comparing structure and process indicators with outcome indicators; data were consolidated by region and state. RESULTS: SIM was partially implanted in the state (70.6%) and its regions (66.3% to 74.8%); 'management' (75.1%), 'issuing and filling in' (79.1%), and 'processing' (71.7%) were partially implanted; 'collection' (80.7%) was implanted; while 'distribution and control' (49.7%) and 'analysis and dissemination' (58.0%) had incipient implantation; more than 90% coverage was found for deaths with defined underlying causes, as well as for municipalities with monthly data transfer, and death certificates typed and sent on a timely basis; consistency was found between DI and outcome indicators, which improved as DI increased. CONCLUSION: SIM was found to be only partially implanted owing to inadequacies in distribution, control, analysis and dissemination, thus influencing unfavorably the effects observed.

Avaliação da implantação do Sistema de Informações sobre Mortalidade no estado de Pernambuco em 2012 / Evaluación de la implantación del sistema de información sobre mortalidad en el estado de Pernambuco, Brasil, en 2012 / Evaluation of the implantation of the Mortality Information System in Pernambuco state, Brazil, in 2012

Objetivo: avaliar a implantação do Sistema de Informações sobre Mortalidade (SIM) em Pernambuco, Brasil. Métodos: pesquisa avaliativa; utilizaram-se dados primários (questionários) e secundários (SIM) referentes aos municípios para estimar o grau de implantação (GI), confrontando indicadores de estrutura e processo aos de resultado; consolidaram-se os dados por região e estado. Resultados: o SIM estava parcialmente implantado no estado (70,6%) e regiões (66,3 a 74,8%); 'gestão' (75,1%), 'emissão e preenchimento' (79,1%) e 'processamento' (71,7%), parcialmente implantados; 'coleta' (80,7%), implantada; 'distribuição e controle' (49,7%) e 'análise e divulgação' (58,0%), com implantação incipiente; encontraram-se valores superiores a 90% para cobertura, óbitos com causa básica definida, municípios com transferência de dados mensal e declarações de óbito digitadas e enviadas oportunamente; verificou-se coerência entre GI e indicadores de resultado, estes melhores quanto maior o GI. Conclusão: o SIM mostrou-se parcialmente implantado, por inadequações na distribuição, controle, análise e divulgação, influenciando desfavoravelmente os efeitos observados.

Objetivo: evaluar la implantación del Sistema de Informaciones sobre Mortalidad (SIM) en Pernambuco, Brasil. Métodos: investigación evaluativa de implantación; se utilizaron datos primarios (cuestionarios) y secundarios (SIM) de los municipios para estimar el grado de implantación (GI), comparando indicadores de estructura y proceso a los de resultado; los datos fueron consolidados por región y estado. Resultados: el SIM se encontraba parcialmente implantado en el estado (70,6%) y regiones (66,3% a 74,8%); 'gestión' (75,1%), 'emisión y llenado' (79,1%) y 'procesamiento' (71,7%), parcialmente implantados; 'colecta' (80,7%), implantada; 'distribución y control' (49,7%) y 'análisis y divulgación' (58,0%), con implantación incipiente; se observó más del 90% de cobertura para óbitos con causa básica definida, municipios con transferencia de datos mensuales y declaraciones de defunción digitadas y enviadas oportunamente; se verificó coherencia entre GI e indicadores de resultado, siendo estos mejores cuanto mayor es el GI. Conclusión: el SIM se mostró parcialmente implantado, en razón de inadecuaciones en la distribución, control, análisis y divulgación, influenciando desfavorablemente los efectos observados.

Objective: to evaluate the implantation of the Mortality Information System (SIM) in Pernambuco, Brazil. Methods: this was an evaluation study; primary data (questionnaires) and secondary data (SIM) were used for the municipalities to estimate the degree of implantation (DI), comparing structure and process indicators with outcome indicators; data were consolidated by region and state. Results: SIM was partially implanted in the state (70.6%) and its regions (66.3% to 74.8%); 'management' (75.1%), 'issuing and filling in' (79.1%), and 'processing' (71.7%) were partially implanted; 'collection' (80.7%) was implanted; while 'distribution and control' (49.7%) and 'analysis and dissemination' (58.0%) had incipient implantation; more than 90% coverage was found for deaths with defined underlying causes, as well as for municipalities with monthly data transfer, and death certificates typed and sent on a timely basis; consistency was found between DI and outcome indicators, which improved as DI increased. Conclusion: SIM was found to be only partially implanted owing to inadequacies in distribution, control, analysis and dissemination, thus influencing unfavorably the effects observed.

Metabolically active CD4+ T cells expressing Glut1 and OX40 preferentially harbor HIV during in vitro infection.

High glucose transporter 1 (Glut1) surface expression is associated with increased glycolytic activity in activated CD4+ T cells. Phosphatidylinositide 3-kinases (PI3K) activation measured by p-Akt and OX40 is elevated in CD4+Glut1+ T cells from HIV+ subjects. TCR engagement of CD4+Glut1+ T cells from HIV+ subjects demonstrates hyperresponsive PI3K-mammalian target of rapamycin signaling. High basal Glut1 and OX40 on CD4+ T cells from combination antiretroviral therapy (cART)-treated HIV+ patients represent a sufficiently metabolically active state permissive for HIV infection in vitro without external stimuli. The majority of CD4+OX40+ T cells express Glut1, thus OX40 rather than Glut1 itself may facilitate HIV infection. Furthermore, infection of CD4+ T cells is limited by p110γ PI3K inhibition. Modulating glucose metabolism may limit cellular activation and prevent residual HIV replication in 'virologically suppressed' cART-treated HIV+ persons.

Tetraspanin CD37 Regulates ß2 Integrin-Mediated Adhesion and Migration in Neutrophils.

Deciphering the molecular basis of leukocyte recruitment is critical to the understanding of inflammation. In this study, we investigated the contribution of the tetraspanin CD37 to this key process. CD37-deficient mice showed impaired neutrophil recruitment in a peritonitis model. Intravital microscopic analysis indicated that the absence of CD37 impaired the capacity of leukocytes to follow a CXCL1 chemotactic gradient accurately in the interstitium. Moreover, analysis of CXCL1-induced leukocyte-endothelial cell interactions in postcapillary venules revealed that CXCL1-induced neutrophil adhesion and transmigration were reduced in the absence of CD37, consistent with a reduced capacity to undergo ß2 integrin-dependent adhesion. This result was supported by in vitro flow chamber experiments that demonstrated an impairment in adhesion of CD37-deficient neutrophils to the ß2 integrin ligand, ICAM-1, despite the normal display of high-affinity ß2 integrins. Superresolution microscopic assessment of localization of CD37 and CD18 in ICAM-1-adherent neutrophils demonstrated that these molecules do not significantly cocluster in the cell membrane, arguing against the possibility that CD37 regulates ß2 integrin function via a direct molecular interaction. Moreover, CD37 ablation did not affect ß2 integrin clustering. In contrast, the absence of CD37 in neutrophils impaired actin polymerization, cell spreading and polarization, dysregulated Rac-1 activation, and accelerated ß2 integrin internalization. Together, these data indicate that CD37 promotes neutrophil adhesion and recruitment via the promotion of cytoskeletal function downstream of integrin-mediated adhesion.

The role of antigen presenting cells in the induction of HIV-1 latency in resting CD4(+) T-cells.

BACKGROUND: Combination antiretroviral therapy (cART) is able to control HIV-1 viral replication, however long-lived latent infection in resting memory CD4(+) T-cells persist. The mechanisms for establishment and maintenance of latent infection in resting memory CD4(+) T-cells remain unclear. Previously we have shown that HIV-1 infection of resting CD4(+) T-cells co-cultured with CD11c(+) myeloid dendritic cells (mDC) produced a population of non-proliferating T-cells with latent infection. Here we asked whether different antigen presenting cells (APC), including subpopulations of DC and monocytes, were able to induce post-integration latent infection in resting CD4(+) T-cells, and examined potential cell interactions that may be involved using RNA-seq. RESULTS: mDC (CD1c(+)), SLAN(+) DC and CD14(+) monocytes were most efficient in stimulating proliferation of CD4(+) T-cells during syngeneic culture and in generating post-integration latent infection in non-proliferating CD4(+) T-cells following HIV-1 infection of APC-T cell co-cultures. In comparison, plasmacytoid DC (pDC) and B-cells did not induce latent infection in APC-T-cell co-cultures. We compared the RNA expression profiles of APC subpopulations that could and could not induce latency in non-proliferating CD4(+) T-cells. Gene expression analysis, comparing the CD1c(+) mDC, SLAN(+) DC and CD14(+) monocyte subpopulations to pDC identified 53 upregulated genes that encode proteins expressed on the plasma membrane that could signal to CD4(+) T-cells via cell-cell interactions (32 genes), immune checkpoints (IC) (5 genes), T-cell activation (9 genes), regulation of apoptosis (5 genes), antigen presentation (1 gene) and through unknown ligands (1 gene). CONCLUSIONS: APC subpopulations from the myeloid lineage, specifically mDC subpopulations and CD14(+) monocytes, were able to efficiently induce post-integration HIV-1 latency in non-proliferating CD4(+) T-cells in vitro. Inhibition of key pathways involved in mDC-T-cell interactions and HIV-1 latency may provide novel targets to eliminate HIV-1 latency.

Monocytes from HIV-infected individuals show impaired cholesterol efflux and increased foam cell formation after transendothelial migration.

DESIGN: HIV-infected (HIV+) individuals have an increased risk of atherosclerosis and cardiovascular disease which is independent of antiretroviral therapy and traditional risk factors. Monocytes play a central role in the development of atherosclerosis, and HIV-related chronic inflammation and monocyte activation may contribute to increased atherosclerosis, but the mechanisms are unknown. METHODS: Using an in-vitro model of atherosclerotic plaque formation, we measured the transendothelial migration of purified monocytes from age-matched HIV+ and uninfected donors and examined their differentiation into foam cells. Cholesterol efflux and the expression of cholesterol metabolism genes were also assessed. RESULTS: Monocytes from HIV+ individuals showed increased foam cell formation compared with controls (18.9 vs. 0%, respectively, P = 0.004) and serum from virologically suppressed HIV+ individuals potentiated foam cell formation by monocytes from both uninfected and HIV+ donors. Plasma tumour necrosis factor (TNF) levels were increased in HIV+ vs. control donors (5.9 vs. 3.5 pg/ml, P = 0.02) and foam cell formation was inhibited by blocking antibodies to TNF receptors, suggesting a direct effect on monocyte differentiation to foam cells. Monocytes from virologically suppressed HIV+ donors showed impaired cholesterol efflux and decreased expression of key genes regulating cholesterol metabolism, including the cholesterol transporter ABCA1 (P = 0.02). CONCLUSION: Monocytes from HIV+ individuals show impaired cholesterol efflux and are primed for foam cell formation following transendothelial migration. Factors present in HIV+ serum, including elevated TNF levels, further enhance foam cell formation. The proatherogenic phenotype of monocytes persists in virologically suppressed HIV+ individuals and may contribute mechanistically to increased atherosclerosis in this population.

Properties of HIV-1 associated cholesterol in addition to raft formation are important for virus infection.

The overall HIV-1 membrane lipid contents resemble lipid rafts, and we have previously demonstrated that raft-promoting properties of virus-associated cholesterol (with modifications in either the 3ß-OH group or AB rings) are important for HIV-1 infectivity. As cholesterol is present in both rafts and non-rafts domains of HIV-1 membrane, we question whether the interpretation of rafts property of virus-associated cholesterol being an absolute requirement for HIV-1 function is too simplistic. The carbon side chain of cholesterol is the third component of cholesterol that can affect the fluidity of membrane depending on its context within the lipid membrane bilayers. In this work, we have used synthetic cholesterol analogues that have different lengths of carbon side chain for our investigation. In contrast to our previous report, we have found that cholesterol side chain analogues that lack in vitro defined raft promoting-property is able to support HIV-1 replication. More specifically, cholesterol analogues with side chains of intermediate length have greater capacity to support HIV-1 infection, suggesting HIV-1 is able to maintain function using cholesterol variants that promote a range of non-rafts- to rafts-properties. Our data demonstrate cholesterol properties other than raft-promoting function also contribute to the infectivity of HIV-1.

Myeloid dendritic cells induce HIV-1 latency in non-proliferating CD4+ T cells.

Latently infected resting CD4(+) T cells are a major barrier to HIV cure. Understanding how latency is established, maintained and reversed is critical to identifying novel strategies to eliminate latently infected cells. We demonstrate here that co-culture of resting CD4(+) T cells and syngeneic myeloid dendritic cells (mDC) can dramatically increase the frequency of HIV DNA integration and latent HIV infection in non-proliferating memory, but not naïve, CD4(+) T cells. Latency was eliminated when cell-to-cell contact was prevented in the mDC-T cell co-cultures and reduced when clustering was minimised in the mDC-T cell co-cultures. Supernatants from infected mDC-T cell co-cultures did not facilitate the establishment of latency, consistent with cell-cell contact and not a soluble factor being critical for mediating latent infection of resting CD4(+) T cells. Gene expression in non-proliferating CD4(+) T cells, enriched for latent infection, showed significant changes in the expression of genes involved in cellular activation and interferon regulated pathways, including the down-regulation of genes controlling both NF-κB and cell cycle. We conclude that mDC play a key role in the establishment of HIV latency in resting memory CD4(+) T cells, which is predominantly mediated through signalling during DC-T cell contact.

Resiliência em Crianças Institucionalizadas / Resilience in Institutionalized Children

isco, proteção e, sobretudo, resiliência que segundo Anaut (2005) consiste fundamentalmente no desenvolvimento normal de uma criança perante circunstâncias dificeis e adversas, são temas presentes na pesquisa do trabalho que nos propusemos dedicar ao crescimento das crianças confindas numa instituição. O conceito de fator de risco já está delimitado, mas o fator de proteção ainda se misturam. Os fatores de risco referem-se aos fatores ambientais que aumentam a probabilidade de ocorrer algum efeito indesejável no desenvolvimento da criança ou jovem. Por outro lado os fatores de proteção estão associados às características da criança, diminuindo o efeito de risco. A resiliência está relacionada com fatores protetores individuais que trazem consequências positivas na criança exposta a fatores que tornam a criança menos vulnerável. Verifica-se que há crianças que perante uma adversidade, conseguem superar ou atenuar os fatores de risco. Com o objetivo de analisar as estratégias de resiliência em crianças institucionalizadas, realizamos um estudo quantitativo-descritivo, percebendo como a criança perante fatores de risco, isto é, os maus-tratos que sofreram antes da sua institucionalização, ultrapassam esses momentos marcantes. O estudo foi feito num grupo de 92 crianças institucionalizadas, pertencente ao distrito da Guarda com idades compreendidas entre os 10 e 18 anos, resultando que 71 crianças desse grupo estudado revelaram um nível elevado de resiliência, enquanto 21 desse mesmo grupo apresentavam um nível médio. Foram utilizadas como hipóteses de investigação a relação entre as variáveis sociodemográficas o género, idade, e escolaridade e as variáveis quanto aos sentimentos iniciais e atuais face à institucionalização. Concluiu-se que só o género influencia as estratégias de resiliência na criança institucionalizada.

Tissue-type plasminogen activator is an extracellular mediator of Purkinje cell damage and altered gait.

Purkinje neurons are a sensitive and specialised cell type important for fine motor movement and coordination. Purkinje cell damage manifests as motor incoordination and ataxia - a prominent feature of many human disorders including spinocerebellar ataxia and Huntington's disease. A correlation between Purkinje degeneration and excess cerebellar levels of tissue-type plasminogen activator (tPA) has been observed in multiple genetically-distinct models of ataxia. Here we show that Purkinje loss in a mouse model of Huntington's disease also correlates with a 200% increase in cerebellar tPA activity. That elevated tPA levels arise in a variety of ataxia models suggests that tPA is a common mediator of Purkinje damage. To address the specific contribution of tPA to cerebellar dysfunction we studied the T4 mice line that overexpresses murine tPA in postnatal neurons through the Thy1.2 gene promoter, which directs preferential expression to Purkinje cells within the cerebellum. Here we show that T4 mice develop signs of cerebellar damage within 10 weeks of birth including atrophy of Purkinje cell soma and dendrites, astrogliosis, reduced molecular layer volume and altered gait. In contrast, T4 mice displayed no evidence of microgliosis, nor any changes in interneuron density, nor alteration in the cerebellar granular neuron layer. Thus, excess tPA levels may be sufficient to cause targeted Purkinje cell degeneration and ataxia. We propose that elevated cerebellar tPA levels exert a common pathway of Purkinje cell damage. Therapeutically lowering cerebellar tPA levels may represent a novel means of preserving Purkinje cell integrity and motor coordination across a wide range of neurodegenerative diseases.

Visualising single molecules of HIV-1 and miRNA nucleic acids.

BACKGROUND: The scarcity of certain nucleic acid species and the small size of target sequences such as miRNA, impose a significant barrier to subcellular visualization and present a major challenge to cell biologists. Here, we offer a generic and highly sensitive visualization approach (oligo fluorescent in situ hybridization, O-FISH) that can be used to detect such nucleic acids using a single-oligonucleotide probe of 19-26 nucleotides in length. RESULTS: We used O-FISH to visualize miR146a in human and avian cells. Furthermore, we reveal the sensitivity of O-FISH detection by using a HIV-1 model system to show that as little as 1-2 copies of nucleic acids can be detected in a single cell. We were able to discern newly synthesized viral cDNA and, moreover, observed that certain HIV RNA sequences are only transiently available for O-FISH detection. CONCLUSIONS: Taken together, these results suggest that the O-FISH method can potentially be used for in situ probing of, as few as, 1-2 copies of nucleic acid and, additionally, to visualize small RNA such as miRNA. We further propose that the O-FISH method could be extended to understand viral function by probing newly transcribed viral intermediates; and discern the localisation of nucleic acids of interest. Additionally, interrogating the conformation and structure of a particular nucleic acid in situ might also be possible, based on the accessibility of a target sequence.