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Germinal centres are the engines of antibody evolution. Here, using human immunodeficiency virus (HIV) Env protein immunogen priming in rhesus monkeys followed by a long period without further immunization, we demonstrate germinal centre B (B

The structural integrity of vaccine antigens is critical to the generation of protective antibody responses, but the impact of protease activity on vaccination in vivo is poorly understood. We characterized protease activity in lymph nodes and found that antigens were rapidly degraded in the subcapsular sinus, paracortex, and interfollicular regions, whereas low protease activity and antigen degradation rates were detected in the vicinity of follicular dendritic cells (FDCs). Correlated with these findings, immunization regimens designed to target antigen to FDCs led to germinal centers dominantly targeting intact antigen, whereas traditional immunizations led to much weaker responses that equally targeted the intact immunogen and antigen breakdown products. Thus, spatially-compartmentalized antigen proteolysis affects humoral immunity and can be exploited to enhance vaccine-induced antibody production against key pathogen structural epitopes.

To prepare for future coronavirus (CoV) pandemics, it is desirable to generate vaccines capable of eliciting broadly neutralizing antibody responses to CoVs. Here, we show that immunization of macaques with SARS-CoV-2 spike (S) protein with a two-shot protocol generated potent serum receptor binding domain cross-neutralizing antibody responses to both SARS-CoV-2 and SARS-CoV-1. Furthermore, responses were equally effective against most SARS-CoV-2 variants of concern (VOCs) and some were highly effective against Omicron. This result contrasts with human infection or many two-shot vaccination protocols where responses were typically more SARS-CoV-2 specific and where VOCs were less well neutralized. Structural studies showed that cloned macaque neutralizing antibodies, particularly using a given heavy chain germline gene, recognized a relatively conserved region proximal to the angiotensin converting enzyme 2 receptor binding site (RBS), whereas many frequently elicited human neutralizing antibodies targeted more variable epitopes overlapping the RBS. B cell repertoire differences between humans and macaques appeared to influence the vaccine response. The macaque neutralizing antibodies identified a pan-SARS–related virus epitope region less well targeted by human antibodies that could be exploited in rational vaccine design.

To combat the HIV epidemic and emerging threats such as SARS-CoV-2, immunization strategies are needed that elicit protection at mucosal portals of pathogen entry. Immunization directly through airway surfaces is effective in driving mucosal immunity, but poor vaccine uptake across the mucus and epithelial lining is a limitation. The major blood protein albumin is constitutively transcytosed bidirectionally across the airway epithelium through interactions with neonatal Fc receptors (FcRn). Exploiting this biology, here, we demonstrate a strategy of “albumin hitchhiking” to promote mucosal immunity using an intranasal vaccine consisting of protein immunogens modified with an amphiphilic albumin-binding polymer-lipid tail, forming amph-proteins. Amph-proteins persisted in the nasal mucosa of mice and nonhuman primates and exhibited increased uptake into the tissue in an FcRn-dependent manner, leading to enhanced germinal center responses in nasal-associated lymphoid tissue. Intranasal immunization with amph-conjugated HIV Env gp120 or SARS-CoV-2 receptor binding domain (RBD) proteins elicited 100- to 1000-fold higher antigen-specific IgG and IgA titers in the serum, upper and lower respiratory mucosa, and distal genitourinary mucosae of mice compared to unmodified protein. Amph-RBD immunization induced high titers of SARS-CoV-2–neutralizing antibodies in serum, nasal washes, and bronchoalveolar lavage. Furthermore, intranasal amph-protein immunization in rhesus macaques elicited 10-fold higher antigen-specific IgG and IgA responses in the serum and nasal mucosa compared to unmodified protein, supporting the translational potential of this approach. These results suggest that using amph-protein vaccines to deliver antigen across mucosal epithelia is a promising strategy to promote mucosal immunity against HIV, SARS-CoV-2, and other infectious diseases.

Vaccines are one of the most powerful technologies supporting public health. The adaptive immune response induced by immunization arises following appropriate activation and differentiation of T and B cells in lymph nodes. Among many parameters impacting the resulting immune response, the presence of antigen and inflammatory cues for an appropriate temporal duration within the lymph nodes, and further within appropriate subcompartments of the lymph nodes– the right timing and location– play a critical role in shaping cellular and humoral immunity. Here we review recent advances in our understanding of how vaccine kinetics and biodistribution impact adaptive immunity, and the underlying immunological mechanisms that govern these responses. We discuss emerging approaches to engineer these properties for future vaccines, with a focus on subunit vaccines., Graphical abstract Unlabelled Image

Saponins are potent and safe vaccine adjuvants, but their mechanisms of action remain incompletely understood. Here, we explored the properties of several saponin formulations, including immunostimulatory complexes (ISCOMs) formed by the self-assembly of saponin and phospholipids in the absence or presence of the Toll like receptor (TLR) 4 agonist monophosphoryl lipid A (MPLA). We found that MPLA self-assembles with saponins to form particles physically resembling ISCOMs, which we termed saponin/MPLA nanoparticles (SMNP). Saponin-containing adjuvants exhibited distinctive mechanisms of action, altering lymph flow in a mast cell-dependent manner and promoting antigen entry into draining lymph nodes. SMNP was particularly effective, exhibiting even greater potency than the compositionally-related adjuvant AS01(B) in mice, and primed robust germinal center B cell, T(FH), and HIV tier 2 neutralizing antibodies in non-human primates. Altogether, these findings shed new light on mechanisms by which saponin adjuvants act to promote the immune response and suggest SMNP may be a promising adjuvant in the setting of HIV, SARS-CoV-2, and other pathogens.

SummaryGerminal centers (GCs) are the engines of antibody evolution. Using HIV Env protein immunogen priming in rhesus monkeys (RM) followed by a long period without further immunization, we demonstrate GC B cells (BGC) lasted at least 6 months (29 weeks), all the while maintaining rapid proliferation. A 186-fold BGC cell increase was present by week 10 compared to a conventional immunization. Single cell transcriptional profiling revealed that both light zone and dark zone GC states were sustained throughout the 6 months. Antibody somatic hypermutation (SHM) of BGC cells continued to accumulate throughout the 29 week priming period, with evidence of selective pressure. Additionally, Env-binding BGC cells were still 49-fold above baseline 29 weeks after immunization, suggesting that they could be active for significantly longer periods of time. High titers of HIV neutralizing antibodies were generated after a single booster immunization. Fully glycosylated HIV trimer protein is a complex antigen, posing significant immunodominance challenges for B cells, among other difficulties. Memory B cells (BMem) generated under these long priming conditions had higher levels of SHM, and both BMem cells and antibodies were more likely to recognize non-immunodominant epitopes. Numerous BGC cell lineage phylogenies spanning the >6-month GC period were identified, demonstrating continuous GC activity and selection for at least 191 days, with no additional antigen exposure. A long prime, adjuvanted, slow delivery (12-day) immunization approach holds promise for difficult vaccine targets, and suggests that patience can have great value for tuning GCs to maximize antibody responses.

Description, Coanchoring a SARS-CoV-2 immunogen and molecular adjuvants to alum extends vaccine kinetics and amplifies humoral immunity., There is a need for additional rapidly scalable, low-cost vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to achieve global vaccination. Aluminum hydroxide (alum) adjuvant is the most widely available vaccine adjuvant but elicits modest humoral responses. We hypothesized that phosphate-mediated coanchoring of the receptor binding domain (RBD) of SARS-CoV-2 together with molecular adjuvants on alum particles could potentiate humoral immunity by promoting extended vaccine kinetics and codelivery of vaccine components to lymph nodes. Modification of RBD immunogens with phosphoserine (pSer) peptides enabled efficient alum binding and slowed antigen clearance, leading to notable increases in germinal center responses and neutralizing antibody titers in mice. Adding phosphate-containing CpG or saponin adjuvants to pSer-RBD:alum immunizations synergistically enhanced vaccine immunogenicity in mice and rhesus macaques, inducing neutralizing responses against SARS-CoV-2 variants. Thus, phosphate-mediated coanchoring of RBD and molecular adjuvants to alum is an effective strategy to enhance the efficacy of SARS-CoV-2 subunit vaccines.

Broadly neutralizing antibodies (bNAbs) against HIV-1 develop after prolonged virus and antibody co-evolution. Previous studies showed that sequential immunization with a V3-glycan-patch germline-targeting HIV-1 envelope trimer (Env) followed by variant Envs can reproduce this process in mice carrying V3-glycan bNAb precursor B cells. However, eliciting bNAbs in animals with polyclonal antibody repertoires is more difficult. We used a V3-glycan immunogen multimerized on virus-like particles (VLPs), followed by boosting with increasingly native-like Env-VLPs, to elicit weak broadly neutralizing antibodies in non-human primates (NHPs). Structures of antibody/Env complexes following prime and boost vaccinations demonstrated target epitope recognition with apparent maturation to accommodate glycans. However, we also observed increasing off-target antibodies with boosting. Eight vaccinated NHPs were subsequently challenged with simian-human immunodeficiency virus (SHIV), and seven of eight animals became infected. The single NHP that remained uninfected after viral challenge exhibited one of the lowest neutralization titers against the challenge virus. These results demonstrate that weak heterologous neutralization achieved using sequential immunization is insufficient for protection in this animal model. Thus, improved prime-boost regimens to increase bNAb potency and stimulate other immune protection mechanisms are essential for developing anti-HIV-1 vaccines.

SUMMARYThe structural integrity of vaccine antigens is critical, as antigen degradation in vivo could eliminate neutralizing epitopes and create competing B cell responses against irrelevant breakdown products. Using FRET imaging and imaging zymography, we found that protease activity and antigen breakdown are spatially heterogeneous in lymph nodes. Following protein immunization, antigens are rapidly degraded in the subcapsular sinus, paracortex, and interfollicular regions of the tissue. By contrast, the follicles and follicular dendritic cell (FDC) networks exhibit low protease activity and antigen degradation rates. Immunization regimens targeting antigen rapidly to FDCs led to germinal centers (GCs) where responses to intact antigen were highly dominant, while traditional bolus immunizations led to weaker GC responses where more GC B cells bound to breakdown products than intact antigen. Thus, spatially-compartmentalized antigen proteolysis impacts humoral immunity and can be exploited to enhance vaccine-induced production of antibody responses against key pathogen structural epitopes.

[SPA] Las geomembranas de PVC-P se han utilizado para impermeabilizar obras hidráulicas desde los años 40 del siglo XX. En España, las balsas para riego agrícola han sido uno de los campos en los que se ha utilizado este material con mayor profusión. Las barreras geosintéticas han permitido construir balsas impermeables con unos costes muy competitivos y unos niveles de seguridad elevados, pero en ciertos casos con una vida útil limitada, por lo que la durabilidad de los materiales se ha convertido en una cuestión clave a la hora de asegurar la viabilidad de los proyectos de construcción de balsas. A día de hoy, existe un stock importante de láminas instaladas en balsas con 30 y 40 años de antigüedad, lo cual ha permitido (1) conocer los principales procesos de degradación de estos materiales e (2) introducir mejoras en el diseño y fabricación de estos materiales. La tecnología de lacado, consistente en la aplicación de una fina película en la superficie de la lámina, ha permitido aumentar la durabilidad de las geomembranas de PVC de manera radical. Ensayos de envejecimiento acelerado demuestran que la aplicación de estas lacas puede llegar a doblar la vida útil de la lámina, ya que son capaces de desacelerar distintos procesos de degradación. Las balsas de Llanos de Mesa y Valle Molina, en la isla de Tenerife, han adoptado esta tecnología, lo cual ha permitido a sus propietarios disponer de garantías de hasta 20 años de duración, maximizando el retorno de la inversión y resultando en una mayor eficiencia en el uso de recursos. [ENG] PVC-P geomembranes have been used to waterproof hydraulic works since the 1940s. In Spain, agricultural irrigation reservoirs has been one of the fields in which this material has been used with greatest profusion. Geosynthetic barriers have enabled the construction of watertight reservoirs with very competitive costs and high levels of safety, but in certain cases with a limited service life, so the durability of the materials has become a key issue when appraising the viability of reservoir construction projects. To date, there is an important stock of liners installed in rafts 30 and 40 years old, which has allowed (1) to understand the main degradation processes of these materials and (2) to introduce improvements in the design and manufacturing of these materials. Lacquering technology, consisting of the application of a thin film on the surface of the sheet, has made it possible to radically increase the durability of PVC geomembranes. Accelerated aging tests show that the application of these lacquers can double the useful life of the sheet, since they are capable of slowing down different degradation processes. The Llanos de Mesa and Valle Molina rafts, on the island of Tenerife, have adopted this technology, which has allowed their owners to have guarantees of up to 20 years, maximizing the return on investment and resulting in a greater efficiency in the use of resources.

Global containment of COVID-19 still requires accessible and affordable vaccines for low- and middle-income countries (LMICs).1 Recently approved vaccines provide needed interventions, albeit at prices that may limit their global access.2 Subunit vaccines based on recombinant proteins are suited for large-volume microbial manufacturing to yield billions of doses annually, minimizing their manufacturing costs.3 These types of vaccines are well-established, proven interventions with multiple safe and efficacious commercial examples.4–6 Many vaccine candidates of this type for SARS-CoV-2 rely on sequences containing the receptor-binding domain (RBD), which mediates viral entry to cells via ACE2.7,8 Here we report an engineered sequence variant of RBD that exhibits high-yield manufacturability, high-affinity binding to ACE2, and enhanced immunogenicity after a single dose in mice compared to the Wuhan-Hu-1 variant used in current vaccines. Antibodies raised against the engineered protein exhibited heterotypic binding to the RBD from two recently reported SARS-CoV-2 variants of concern (501Y.V1/V2). Presentation of the engineered RBD on a designed virus-like particle (VLP) also reduced weight loss in hamsters upon viral challenge.

Introducción: la trombosis venosa profunda (TVP) es una entidad común que afecta principalmente el sistema venoso profundo de los miembros inferiores, para el cual se han desarrollado múltiples escalas de predicción clínica, las cuales han sido construidas y validadas en pacientes ambulatorios y hospitalizados. Objetivos: validar cinco escalas de predicción clínica para TVP en pacientes atendidos en un centro de tercer nivel en la sabana de Bogotá, Colombia. Métodos: se llevó a cabo un estudio de corte transversal con análisis de prueba diagnóstica en sujetos con sospecha de TVP, incluyendo aquellos que contaran con la realización de ecografía Doppler venosa de miembros inferiores. Se calculó el rendimiento de cinco escalas de predicción clínica para TVP (Wells clásico y modificado, Oudega, CEBI y Constans) para pacientes ambulatorios u hospitalizados, individualizando la población en la que fueron validadas. Resultados: ingresaron al análisis 974 pacientes, de estos 485 (49,7 %) presentaron TVP. La escala de Constans tuvo un mejor rendimiento diagnóstico entre los pacientes hospitalizados y ambulatorios, con un área bajo la curva ROC de 0,73 (95 % 0,70-0,78) al compararla con Wells clásico, Wells modificado, Oudega y CEBI. Al comparar el rendimiento de Constans en ambos grupos de pacientes por separado, también se observó un mejor rendimiento con respecto a las demás escalas. Conclusión: la escala de Constans presenta un mejor rendimiento diagnostico comparado con las demás escalas al ser aplicada en paciente hospitalizados y ambulatorios.

To prepare for future coronavirus (CoV) pandemics, it is desirable to generate vaccines capable of eliciting neutralizing antibody responses against multiple CoVs. Because of the phylogenetic similarity to humans, rhesus macaques are an animal model of choice for many virus-challenge and vaccine-evaluation studies, including SARS-CoV-2. Here, we show that immunization of macaques with SARS-CoV-2 spike (S) protein generates potent receptor binding domain cross- neutralizing antibody (nAb) responses to both SARS-CoV-2 and SARS-CoV-1, in contrast to human infection or vaccination where responses are typically SARS-CoV-2-specific. Furthermore, the macaque nAbs are equally effective against SARS-CoV-2 variants of concern. Structural studies show that different immunodominant sites are targeted by the two primate species. Human antibodies generally target epitopes strongly overlapping the ACE2 receptor binding site (RBS), whereas the macaque antibodies recognize a relatively conserved region proximal to the RBS that represents another potential pan-SARS-related virus site rarely targeted by human antibodies. B cell repertoire differences between the two primates appear to significantly influence the vaccine response and suggest care in the use of rhesus macaques in evaluation of vaccines to SARS-related viruses intended for human use.ONE SENTENCE SUMMARYBroadly neutralizing antibodies to an unappreciated site of conservation in the RBD in SARS- related viruses can be readily induced in rhesus macaques because of distinct properties of the naïve macaque B cell repertoire that suggest prudence in the use of the macaque model in SARS vaccine evaluation and design.

Significance Most of the global population resides in low- and middle-income countries, where current vaccines for COVID-19 remain largely unavailable. For the COVID-19 pandemic, the world will need access to >10 billion doses of vaccines, or more than double the annual volume of vaccines for all other diseases. Many vaccine candidates use the SARS-CoV-2 receptor-binding domain (RBD) antigen. Here, we present an engineered RBD with improved production titers in Pichia pastoris, a yeast commonly used for large-scale, low-cost manufacturing by vaccine manufacturers. The modified RBD also raises an enhanced immune response in mice relative to the Wuhan-Hu-1 sequence used in current candidates. These combined traits make it a promising candidate for next-generation vaccines addressing emerging variants of the virus., Global containment of COVID-19 still requires accessible and affordable vaccines for low- and middle-income countries (LMICs). Recently approved vaccines provide needed interventions, albeit at prices that may limit their global access. Subunit vaccines based on recombinant proteins are suited for large-volume microbial manufacturing to yield billions of doses annually, minimizing their manufacturing cost. These types of vaccines are well-established, proven interventions with multiple safe and efficacious commercial examples. Many vaccine candidates of this type for SARS-CoV-2 rely on sequences containing the receptor-binding domain (RBD), which mediates viral entry to cells via ACE2. Here we report an engineered sequence variant of RBD that exhibits high-yield manufacturability, high-affinity binding to ACE2, and enhanced immunogenicity after a single dose in mice compared to the Wuhan-Hu-1 variant used in current vaccines. Antibodies raised against the engineered protein exhibited heterotypic binding to the RBD from two recently reported SARS-CoV-2 variants of concern (501Y.V1/V2). Presentation of the engineered RBD on a designed virus-like particle (VLP) also reduced weight loss in hamsters upon viral challenge.

HIV Env trimers elicit antibodies that lead to their own destruction., Rationally designed protein subunit vaccines are being developed for a variety of viruses including influenza, RSV, SARS-CoV-2, and HIV. These vaccines are based on stabilized versions of the primary targets of neutralizing antibodies on the viral surface, namely, viral fusion glycoproteins. While these immunogens display the epitopes of potent neutralizing antibodies, they also present epitopes recognized by non-neutralizing or weakly neutralizing (“off-target”) antibodies. Using our recently developed electron microscopy polyclonal epitope mapping approach, we have uncovered a phenomenon wherein off-target antibodies elicited by HIV trimer subunit vaccines cause the otherwise highly stabilized trimeric proteins to degrade into cognate protomers. Further, we show that these protomers expose an expanded suite of off-target epitopes, normally occluded inside the prefusion conformation of trimer, that subsequently elicit further off-target antibody responses. Our study provides critical insights for further improvement of HIV subunit trimer vaccines for future rounds of the iterative vaccine design process.

Los nuevos Planes Hidrológicos plantean una reducción del agua disponible para riego en las próximas décadas. En este escenario, es imprescindible que las infraestructuras hidráulicas maximicen su eficiencia, o dicho de otra manera que minimicen sus pérdidas. Las balsas de riego son estructuras construidas con la máxima economía de materiales. En este sentido, el diseño geométrico de la balsa se suele llevar a cabo bajo el principio de balance neto de tierras: en la medida de lo posible, las tierras excavadas se deben utilizar en los diques de cierre y terraplenes para evitar o minimizar los préstamos. Como resultado, las estructuras de cierre son heterogéneas y contienen partículas de distinto tamaño y naturaleza, por lo que las láminas de impermeabilización se colocan sobre un soporte que puede dar lugar a punzonamientos. La resistencia o susceptibilidad a dicho punzonamiento no depende solamente del sustrato, sino también de las condiciones de operación de la balsa y del tipo de lámina. Mediante ensayos de laboratorio se ha simulado el comportamiento de geomembranas de PVC y PEAD frente a este fenómeno para conocer la magnitud de las perforaciones que podrían sufrir bajo distintas condiciones. En base a estos resultados y a la bibliografía existente se discuten las consecuencias que se derivan de las pérdidas de agua esperables a través de las zonas punzonadas y sus efectos a nivel de seguridad estructural de la balsa y de disponibilidad de recurso.

Following curative immunotherapy of B16F10 tumors, ~60% of mice develop a strong antibody response against cell-surface tumor antigens. Their antisera confer prophylactic protection against intravenous challenge with B16F10 cells, and also cross-react with syngeneic and allogeneic tumor cell lines MC38, EL.4, 4T1, and CT26. We identified the envelope glycoprotein (env) of a murine endogenous retrovirus (ERV) as the antigen accounting for the majority of this humoral response. A systemically administered anti-env monoclonal antibody cloned from such a response protects against tumor challenge, and prophylactic vaccination against the env protein protects a majority of naive mice from tumor establishment following subcutaneous inoculation with B16F10 cells. These results suggest the potential for effective prophylactic vaccination against analogous HERV-K env expressed in numerous human cancers.

An HIV vaccine capable of eliciting durable neutralizing antibody responses continues to be an important unmet need. Multivalent nanoparticles displaying a high density of envelope trimers may be promising immunogen forms to elicit strong and durable humoral responses to HIV, but critical particle design criteria remain to be fully defined. To this end, we developed strategies to covalently anchor a stabilized gp140 trimer, BG505 MD39, on the surfaces of synthetic liposomes to study the effects of trimer density and vesicle stability on vaccine-elicited humoral responses in mice. CryoEM imaging revealed homogeneously distributed and oriented MD39 on the surface of liposomes irrespective of particle size, lipid composition, and conjugation strategy. Immunization with covalent MD39-coupled liposomes led to increased germinal center and antigen-specific T follicular helper cell responses and significantly higher avidity serum MD39-specific IgG responses compared to immunization with soluble MD39 trimers. A priming immunization with liposomal-MD39 was important for elicitation of high avidity antibody responses, regardless of whether booster immunizations were administered with either soluble or particulate trimers. The stability of trimer anchoring to liposomes was critical for these effects, as germinal center and output antibody responses were further increased by liposome compositions incorporating sphingomyelin that exhibited high in vitro stability in the presence of serum. Together these data highlight key liposome design features for optimizing humoral immunity to lipid nanoparticle immunogens.

SUMMARY Rapidly evolving pathogens such as HIV or influenza can quickly mutate their antigenic profiles, reducing the efficacy of conventional vaccines. Despite this challenge, functionally required epitopes are highly conserved among heterologous viral strains and represent a key vulnerability that could be targeted during vaccine development. As the antigenicity of these conserved epitopes is frequently subdominant, there is a critical need for innovative vaccination strategies designed to target these neutralizing epitopes. Here, we immunized mice with antigens containing discrete immunodominant and subdominant moieties and show that treatment with soluble heterologous antigen bearing only the immunodominant epitope selectively suppresses these germinal center (GC) B cells. By exploiting this intrinsic tolerance mechanism, we promote the expansion of subdominant B cells in the GC and the subsequent long-lived components of the humoral response. We propose that this strategy may be applied to elicit preferential expansion of subdominant B cells that recognize weakly immunogenic epitopes on microbial pathogens., In Brief Interclonal competition inhibits full participation of subdominant B cells in the germinal center (GC). Silva et al. demonstrate that selective elimination of immunodominant B cells during an active GC response allows subdominant B cells to expand unimpeded. Without competition, these subdominant cells generate an improved long-lived humoral response.

In radar, sonar and wireless communications, there is a critical need for estimating the direction-of-arrival (DoA) of signals or sources. DoA angles of signals are estimated by processing the measurements from spatially distributed sensor arrays. In all these cases, conventional processing involves using linear processing to combine the measurements from the sensor elements of the spatial array. The DoA angular resolution is determined by the size of the spatial array. In this study, a multiplicative processing technique known as multiplicative array processing (MAP) is proposed to achieve fine angular resolution by using much smaller array sizes. Multiplicative processing of array measurements has the potential to achieve super-resolution capability where the effective resolution is commensurate with significantly larger array sizes. In this study, both simulations and experimental results are used to verify and validate the proposed multiplicative processing technique. Simulations indicate that for moderate to high signal-to-noise ratios, MAP performs as well as linear processing methods with much larger array sizes. MAP is applied to experimental measurements to verify and validate the effectiveness of the proposed technique with field data.

Self-replicating RNAs derived from alphaviruses have high potential for vaccine applications. Utilizing lipid nanoparticle-formulated RNA replicons as a vaccine platform to deliver self-assembling protein immunogens, Melo et al. demonstrate robust anti-HIV Env antibody production and significantly improved antigen-specific B cell activation in vaccinated mice when compared with protein immunization., NIAID (Awards UM1AI100663, AI104715, EB025854 and AI048240), National Cancer Institute (Grant P30-CA14051)

Chimeric antigen receptor-T cell (CAR-T) therapy has been effective in the treatment of hematologic malignancies, but it has shown limited efficacy against solid tumors. Here we demonstrate an approach to enhancing CAR-T function in solid tumors by directly vaccine-boosting donor cells through their chimeric receptor in vivo. We designed amphiphile CAR-T ligands (amph-ligands) that, upon injection, trafficked to lymph nodes and decorated the surfaces of antigen-presenting cells, thereby priming CAR-Ts in the native lymph node microenvironment. Amph-ligand boosting triggered massive CAR-T expansion, increased donor cell polyfunctionality, and enhanced antitumor efficacy in multiple immunocompetent mouse tumor models. We demonstrate two approaches to generalizing this strategy to any chimeric antigen receptor, enabling this simple non-human leukocyte antigen-restricted approach to enhanced CAR-T functionality to be applied to existing CAR-T designs.

Germinal centers (GCs) are anatomic sites where B cells undergo secondary diversification to produce high-affinity, class-switched Abs. We hypothesized that proliferating B cells in GCs create a hypoxic microenvironment that governs their further differentiation. Using molecular markers, we found GCs to be predominantly hypoxic. Compared to normoxia (21% O2), hypoxic culture conditions (1% O2) in vitro accelerated class switching and plasma cell formation and enhanced expression of GL-7 on B and CD4+ T cells. Reversal of GC hypoxia in vivo by breathing 60% O2 during immunization resulted in reduced frequencies of GC B cells, T follicular helper cells, and plasmacytes, as well as lower expression of ICOS on T follicular helper cells. Importantly, this reversal of GC hypoxia decreased Ag-specific serum IgG1 and reduced the frequency of IgG1+ B cells within the Ag-specific GC. Taken together, these observations reveal a critical role for hypoxia in GC B cell differentiation.

O objetivo deste trabalho foi avaliar diferentes híbridos de milho frente ao ataque de S. frugiperda em associação com adubação silicatada e o efeito sobre o predador D. luteipes. O experimento foi realizado durante a safra 2008/09, com três híbridos de milho, Impacto (Syngenta), AS1575 (Agroeste) e o transgênico AG7000YG (Agroceres), em diferentes épocas de aplicação do produto comercial (Sili-K®), contendo silício na concentração 12,2% (SIO2) e 15% de K2O, via foliar na dosagem de 3 L ha-1. O delineamento experimental utilizado foi em blocos casualizados, em parcelas subdivididas com quatro repetições. As parcelas foram constituídas por 3 cultivares de milho e as subparcelas constituídas por 3 épocas diferentes de aplicação foliar de silício (aos 10 e 30 dias após a emergência, aos 10, 20 e 30 dias após a emergência e tratamento controle). Foram avaliados: danos causados pela S. frugiperda, número de lagartas e tesourinhas dentro do cartucho e teor de clorofila. O silício aplicado via foliar não teve efeito na redução ao ataque de S. frugiperda. O híbrido Bt AG7000YG, foi resistente a S. frugiperda e não causa impacto negativo sobre o predador D. luteipes.

Summary How antigen valency affects B cells in vivo during immune responses is not well understood. Here, using HIV immunogens with defined valencies ranging from 1 to 60, we investigated the role of antigen valency during different phases of B cell responses in vivo. Highly multimerized immunogens preferentially rapidly activated cognate B cells, with little affinity discrimination. This led to strong early induction of the transcription factors IRF4 (interferon regulatory factor 4) and Bcl6, driving both early extrafollicular plasma cell and germinal center responses, in a CD4+ T-cell-dependent manner, involving B cells with a broad range of affinities. Low-valency antigens induced smaller effector B cell responses, with preferential recruitment of high-affinity B cells. Thus, antigen valency has multifaceted effects on B cell responses and can dictate affinity thresholds and competitive landscapes for B cells in vivo, with implications for vaccine design.

Este artigo apresenta uma avaliação da viabilidade de se implantar VIMs sob demanda (VIMs como um serviço) no contexto de fatiamento de redes, onde mais de um VIM pode ser instanciado simultaneamente. Para realizar essa validação, utilizamos o sistema de alocação de recursos e experimentação do FIBRE. Os resultados experimentais mostram que é viável instanciar VIMs sob demanda nestes cenários, mesmo quando lidando com VIMs mais complexos, como o OpenStack.

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro Sócio-Econômico, Programa de Pós-Graduação em Relações Internacionais, Florianópolis, 2017. As políticas imigratórias podem ser divididas em políticas de acesso e políticas de acomodação. Nos casos abordados nesta dissertação as políticas de acesso foram responsabilidade exclusiva dos Estados centrais (Espanha e Brasil), ao passo que as políticas de acomodação estiveram a cargo das comunidades autônomas e estados. Catalunha e Paraná, entre outras semelhanças, desenvolveram planos de políticas públicas com múltiplas áreas de intervenção. Os casos, escolhidos inicialmente por suas semelhanças, quando tiveram seus planos comparados, utilizando critérios da Administração Pública, apresentaram divergências estruturais em termos de concretude e especificidade, sendo o Plano Estadual para Promoção e Defesa dos Direitos dos Refugiados, Migrantes e Apátridas do Paraná (2014-2016), em seu conjunto, vago em relação ao Plan de Ciudadanía y de las Migraciones (2013-2016). Assim, buscou-se, através do estudo de cada caso, identificar quais elementos poderiam ter influenciado a tomada de decisão que culminou na adoção dos diferentes planos. Buscou-se no nível nacional delimitar os atores e as instituições em torno do subsistema político-administrativo voltado às políticas de imigração na Espanha e no Brasil. No estudo dos casos, buscou-se delimitar o subsistema político-administrativo em torno das políticas de acomodação na Catalunha e no Paraná. Na Catalunha foi identificada uma maior trajetória no recebimento de fluxos imigratórios (1985-2013) em relação ao caso paranaense (2000-2015), que foi acompanhada por cinco planos sucessivos de acomodação de imigrantes. Tomando aportes do incrementalismo, concluiu-se que a decisão que culminou na adoção do plano catalão de 2013 fez parte de um processo longo de construção contínua em que elementos conferidores de concretude e especificidades foram desenvolvidos ao longo do tempo. No caso paranaense, recorrendo às contribuições teóricas do ?modelo de tomada de decisão da lata de lixo?, chegou-se à possível conclusão de que a adoção de um plano mais vago esteve relacionada ao ambiente incerto e conflituoso em que foi elaborado, também aliado à ausência de tomadores de oficiais de decisão e à carência de dados sobre o público-alvo. Abstract : Immigration policies can be divided into access policies and accommodation policies. In the cases addressed in this dissertation, access policies were the exclusive responsibility of the central states (Spain and Brazil), while accommodation policies were in charge of the autonomous communities and states. Catalonia and Paraná, among other similarities, have developed public policy plans with multiple intervention areas. The cases, initially chosen by their similarities, when they had their plans compared, using Public Administration criterias, presented structural divergences in terms of concreteness and specificity, being the Plano Estadual para Promoção e Defesa dos Direitos dos Refugiados, Migrantes e Apátridas do Paraná (2014-2016), as a whole, vacant in relation to the Plan de Ciudadanía y de las Migraciones (2013-2016). Thus, it was sought, through the study of each case, to identify which elements could have influenced the decision making that culminated in the adoption of the different plans. It was sought at the national level to delimit the actors and institutions around the political-administrative sub-system focused on immigration policies in Spain and Brazil. In the study of the cases, it was sought to delimit the political-administrative sub-system around accommodation policies in Catalonia and Paraná. In Catalonia a greater trajectory was identified in the reception of immigrant flows (1985-2013) in relation to the case of Parana (2000-2015), which was accompanied by five successive plans for the accommodation of immigrants. Taking the contributions of incrementalism, it was concluded that the decision that culminated in the adoption of the Catalan plan of 2013 was part of a long process of continuous construction in which elements that confer concreteness and specificity have been developed over time. In the case of Paraná, using the theoretical contributions of ?garbage can model of decision-making?, it was possible to conclude that the adoption of a more vague plan was related to the uncertain and conflicting environment in which it was elaborated, also associated with the absence decision-makers and the lack of data on the target audience.

T follicular helper (TFH) cells have been shown to be critically required for the germinal center (GC) reaction where B cells undergo class switch recombination and clonal selection to generate high affinity neutralizing antibodies. However, detailed knowledge of the physiological cues within the GC microenvironment that regulate T cell help is limited. The cAMP-elevating, Gs protein-coupled A2a adenosine receptor (A2aR) is an evolutionarily conserved receptor that limits and redirects cellular immunity. However, the role of A2aR in humoral immunity and B cell differentiation is unknown. We hypothesized that the hypoxic microenvironment within the GC facilitates an extracellular adenosine-rich milieu, which serves to limit TFH frequency and function, and also promotes immunosuppressive T follicular regulatory cells (TFR). In support of this hypothesis, we found that following immunization, mice lacking A2aR (A2aRKO) exhibited a significant expansion of T follicular cells, as well as increases in TFH to TFR ratio, GC T cell frequency, GC B cell frequency, and class switching of GC B cells to IgG1. Transfer of CD4 T cells from A2aRKO or wild type donors into T cell-deficient hosts revealed that these increases were largely T cell-intrinsic. Finally, injection of A2aR agonist, CGS21680, following immunization suppressed T follicular differentiation, GC B cell frequency, and class switching of GC B cells to IgG1. Taken together, these observations point to a previously unappreciated role of GS protein-coupled A2aR in regulating humoral immunity, which may be pharmacologically targeted during vaccination or pathological states in which GC-derived autoantibodies contribute to the pathology.