Long-term Impact of Planovalgus Foot on Activities of Daily Living in Patients With Down Syndrome.

INTRODUCTION: Planovalgus foot (PVF) is the most common orthopaedic abnormality in children with Down syndrome (DS), and as a result these patients rarely develop an adequate plantar arch in adulthood. The present study aims to evaluate the impact of PVF on activities of daily living and participation in sports among young adults with DS and determine whether this impact is related to the degree of foot deformity based on clinical and imaging studies. METHODS: Observational analytical study examining a database of 649 patients with DS from a pediatric referral center, identifying those individuals over age 20 years at the time of the study with a childhood diagnosis of PVF. Finally, 51 patients (102 feet) were evaluated based on clinical and imaging studies, and function was assessed using the The Foot and Ankle Outcome Score (FAOS) and the Visual Analogue Scale (VAS) pain scale. A correlation analysis was performed to determine the clinical and radiographic variables associated with functional outcomes. Linear regression models were obtained to quantify the impact of these variables on function. RESULTS: Patients had a mean age of 26.14±3.88 years and body mass index of 24.51±4.57. Clinically, 63.65% presented grade 3 or 4 PVF, and most were flexible. Radiographically, midfoot flattening was mild-moderate in 92.16%, 58.82% had medial talo-navicular uncoverage, and 30.39% had an increased hallux valgus (HV) angle. Mean scores for all FAOS subscales were between 65 and 71% and the mean VAS score was 1.45±1.96. An association analysis revealed a tendency toward lower scores on all FAOS subscales and greater pain according to the VAS scale in more severe PVF and in cases of moderate HV with asymmetry between feet. Linear regression models showed that major contributors to functional scores were radiographic evidence of hindfoot valgus, midfoot abduction, and flattening, and HV. CONCLUSIONS: Young adults with DS who are diagnosed with PVF in childhood have acceptable functional scores and low pain. Alteration of radiographic parameters toward flatter, more valgus and abducted feet and greater and asymmetric HV tend to be associated with worse long-term functional scores in activities of daily living and sports participation and increased pain. Therefore, non-operative management of these patients is justified, although individualized treatment is recommended. LEVEL OF EVIDENCE: Level IV, Case series.

Synovial Sarcoma of the Foot as an Atypical Cause of Persistent Metatarsalgia: A Report of 3 Cases.

CASE: Three cases of atypical metatarsalgia are presented, all diagnosed with foot synovial sarcomas (SSs) at different stages of evolution, after a year of medical consultations. One case was treated with marginal excision without requiring bone excision; the second patient required amputation of the first ray; and the third patient, with advanced disease, required amputation through Chopart's joint. CONCLUSION: Metatarsalgia is a recurrent reason for consultation in orthopaedics. Even so, patients with persistent symptoms should be studied further in depth. Computed tomography or magnetic resonance imaging can detect tumor pathology, such as SS, of insidious development.

Simultaneous bilateral extracapsular proximal femur fractures in the elderly. A case report.

Simultaneous and bilateral proximal femoral fractures (PFF) are rare and have scarcely been reported in the literature. A case of a bilateral extracapsular PFF is herein presented. Besides, an exhaustive review of the literature was performed, analyzing the information of all previously reported cases. An 81-year-old woman, who suffered a casual fall, was diagnosed with bilateral PFF consisting of both a subtrochanteric and an intertrochanteric fracture. She underwent concurrent intramedullary fixation for both fractures without any relevant complication and started early ambulation. Simultaneous bilateral extracapsular PFF are exceptional, with only 23 cases described in the current literature. In the elderly, they deserve special attention with treatment strategies in between the ones for unilateral hip fractures and those provided to old multiply injured patients. PFF management is not well established. Minimal reaming and careful nailing can be safely performed bilaterally under close monitoring, in order to start an early functional recovery.

TLR activation, immune response and viral protection elicited in cattle by a commercial vaccine against Bovine Herpesvirus-1.

The innate and acquired immune response induced by a commercial inactivated vaccine against Bovine Herpesvirus-1 (BoHV-1) and protection conferred against the virus were analyzed in cattle. Vaccination induced high levels of BoHV-1 antibodies at 30, 60, and 90 days post-vaccination (dpv). IgG1 and IgG2 isotypes were detected at 90 dpv, as well as virus-neutralizing antibodies. An increase of anti-BoHV-1 IgG1 in nasal swabs was detected 6 days post-challenge in vaccinated animals. After viral challenge, lower virus excretion and lower clinical score were observed in vaccinated as compared to unvaccinated animals, as well as BoHV-1-specific proliferation of lymphocytes and production of IFNγ, TNFα, and IL-4. Downregulation of the expression of endosome Toll-like receptors 8-9 was detected after booster vaccination. This is the first thorough study of the immunity generated by a commercial vaccine against BoHV-1 in cattle.

Characterization of a Nanovaccine Platform Based on an α1,2-Mannobiose Derivative Shows Species-non-specific Targeting to Human, Bovine, Mouse, and Teleost Fish Dendritic Cells.

Dendritic cells serve as the main immune cells that trigger the immune response. We developed a simple and cost-effective nanovaccine platform based on the α1',2-mannobiose derivative for dendritic cell targeting. In previous work, we have formulated the α1,2-mannobiose-based nanovaccine platform with plasmid DNA and tested it in cattle against BoHV-1 infection. There, we have shown that the dendritic cell targeting using this nanovaccine platform in vivo can boost the immunogenicity, resulting in a long-lasting immunity. In this work, we aim to characterize the α1',2-mannobiose derivative, which is key in the nanovaccine platform. This DC-targeting strategy takes advantage of the specific receptor known as DC-SIGN and exploits its capacity to bind α1,2-mannobiose that is present at terminal ends of oligosaccharides in certain viruses, bacteria, and other pathogens. The oxidative conjugation of α1',2-mannobiose to NH2-PEG2kDa-DSPE allowed us to preserve the chemical structure of the non-reducing mannose of the disaccharide and the OH groups and the stereochemistry of all carbons of the reducing mannose involved in the binding to DC-SIGN. Here, we show specific targeting to DC-SIGN of decorated micelles incubated with the Raji/DC-SIGN cell line and uptake of targeted liposomes that took place in human, bovine, mouse, and teleost fish DCs in vitro, by flow cytometry. Specific targeting was found in all cultures, demonstrating a species-non-specific avidity for this ligand, which opens up the possibility of using this nanoplatform to develop new vaccines for various species, including humans.

A plasmid encoding the extracellular domain of CD40 ligand and Montanide™ GEL01 as adjuvants enhance the immunogenicity and the protection induced by a DNA vaccine against BoHV-1.

DNA vaccines are capable of inducing humoral and cellular immunity, and are important to control bovine herpesvirus 1 (BoHV-1), an agent of the bovine respiratory disease complex. In previous work, a DNA plasmid that encodes a secreted form of BoHV-1 glycoprotein D (pCIgD) together with commercial adjuvants provided partial protection against viral challenge of bovines. In this work, we evaluate new molecules that could potentiate the DNA vaccine. We show that a plasmid encoding a soluble CD40 ligand (CD40L) and the adjuvant Montanide™ GEL01 (GEL01) activate in vitro bovine afferent lymph dendritic cells (ALDCs). CD40L is a co-stimulating molecule, expressed transiently on activated CD4+ T cells and, to a lesser extent, on activated B cells and platelets. The interaction with its receptor, CD40, exerts effects on the presenting cells, triggering responses in the immune system. GEL01 was designed to improve transfection of DNA vaccines. We vaccinated cattle with: pCIgD; pCIgD-GEL01; pCIgD with GEL01 and CD40L plasmid (named pCIgD-CD40L-GEL01) or with pCIneo vaccines. The results show that CD40L plasmid with GEL01 improved the pCIgD DNA vaccine, increasing anti-BoHV-1 total IgGs, IgG1, IgG2 subclasses, and neutralizing antibodies in serum. After viral challenge, bovines vaccinated with pCIgD-GEL01-CD40L showed a significant decrease in viral excretion and clinical score. On the other hand, 80% of animals in group pCIgD-GEL01-CD40L presented specific anti-BoHV-1 IgG1 antibodies in nasal swabs. In addition, PBMCs from pCIgD-CD40L-GEL01 had the highest percentage of animals with a positive lymphoproliferative response against the virus and significant differences in the secretion of IFNγ and IL-4 by mononuclear cells, indicating the stimulation of the cellular immune response. Overall, the results demonstrate that a plasmid expressing CD40L associated with the adjuvant GEL01 improves the efficacy of a DNA vaccine against BoHV-1.

MANα1-2MAN decorated liposomes enhance the immunogenicity induced by a DNA vaccine against BoHV-1.

New technologies in the field of vaccinology arise as a necessity for the treatment and control of many diseases. Whole virus inactivated vaccines and modified live virus ones used against Bovine Herpesvirus-1 (BoHV-1) infection have several disadvantages. Previous works on DNA vaccines against BoHV-1 have demonstrated the capability to induce humoral and cellular immune responses. Nevertheless, 'naked' DNA induces low immunogenic response. Thus, loading of antigen encoding DNA sequences in liposomal formulations targeting dendritic cell receptors could be a promising strategy to better activate these antigen-presenting cells (APC). In this work, a DNA-based vaccine encoding the truncated version of BoHV-1 glycoprotein D (pCIgD) was evaluated alone and encapsulated in a liposomal formulation containing LPS and decorated with MANα1-2MAN-PEG-DOPE (pCIgD-Man-L). The vaccinations were performed in mice and bovines. The results showed that the use of pCIgD-Man-L enhanced the immune response in both animal models. For humoral immunity, significant differences were achieved when total antibody titres and isotypes were assayed in sera. Regarding cellular immunity, a significant increase in the proliferative response against BoHV-1 was detected in animals vaccinated with pCIgD-Man-L when compared to the response induced in animals vaccinated with pCIgD. In addition, upregulation of CD40 molecules on the surface of bovine dendritic cells (DCs) was observed when cells were stimulated and activated with the vaccine formulations. When viral challenge was performed, bovines vaccinated with MANα1-2MAN-PEG-DOPE elicited better protection which was evidenced by a lower viral excretion. These results demonstrate that the dendritic cell targeting using MANα1-2MAN decorated liposomes can boost the immunogenicity resulting in a long-lasting immunity. Liposomes decorated with MANα1-2MAN-PEG-DOPE were tested for the first time as a DNA vaccine nanovehicle in cattle as a preventive treatment against BoHV-1. These results open new perspectives for the design of vaccines for the control of bovine rhinotracheitis.

An Improved DNA Vaccine Against Bovine Herpesvirus-1 Using CD40L and a Chemical Adjuvant Induces Specific Cytotoxicity in Mice.

Bovine herpesvirus-1 (BoHV-1) uses many mechanisms to elude the immune system; one of them is spreading intracellularly, even in the presence of specific antiviral antibodies. Cytotoxic T lymphocytes (CTLs) are necessary to eliminate the virus. The main preventive strategy is vaccination based on inactivated virus. These vaccines are poor inducers of cellular immune responses, and complicate serological diagnosis and determination of the real prevalence of infection. DNA vaccines are a good option because of the capacity of Differentiating Infected from Vaccinated Animals-(DIVA vaccine)-and may be the best way to induce cytotoxic responses. Although this type of vaccines leads to only weak "in vivo" expression and poor immune responses, incorporation of molecular and/or chemical adjuvants can improve the latter, both in magnitude and in direction. In this study, we have investigated the specific immune responses elicited in mice by DNA vaccines based on the BoHV-1 glycoprotein D (pCIgD) with and without two different adjuvants: a plasmid encoding for murine CD40L (pCD40L) or Montanide™ 1113101PR (101). Mice vaccinated with pCIgD+CD40L, pCIgD+101, and pCIgD+CD40L+101 developed significantly higher specific antibody titers against BoHV-1 than the pCIgD group (p < 0.01). The animals vaccinated with pCgD+pCD40L+101 raised significantly higher levels of IgG2a and IgG2b (p < 0.01 and p < 0.001, respectively) than mice vaccinated with pCIgD alone. On the contrary, when the activity of CTL against cells infected with BoHV-1 was measured, the vaccine pCgD+pCD40L+101 induced significantly higher levels of cytotoxicity activity (p < 0.001) than pCIgD alone. A significant increase in the CD4+ populations in the group receiving pCIgD+CD40L+101 in comparison with the pCIgD group was observed and, also, interferon gamma, interleukin (IL)-6, and IL-17A levels were higher. Considering the results obtained from this study for humoral and cellular responses in mice, the inclusion of pCD40L and 101 as adjuvants in a BoHV-1 DNA vaccine for cattle is highly recommendable.

Optimized Adenoviral Vector That Enhances the Assembly of FMDV O1 Virus-Like Particles in situ Increases Its Potential as Vaccine for Serotype O Viruses.

Although replication-defective human adenovirus type 5 (Ad5) vectors that express in situ the capsid-encoding region of foot-and-mouth disease virus (FMDV) have been proven to be effective as vaccines in relevant species for several viral strains, the same result was not consistently achieved for the O1/Campos/Brazil/58 strain. In the present study, an optimization of the Ad5 system was explored and was proven to enhance the expression of FMDV capsid proteins and their association into virus-like particles (VLPs). Particularly, we engineered a novel Ad5 vector (Ad5[PVP2]OP) which harbors the foreign transcription unit in a leftward orientation relative to the Ad5 genome, and drives the expression of the FMDV sequences from an optimized cytomegalovirus (CMV) enhancer-promoter as well. The Ad5[PVP2]OP vaccine candidate also contains the amino acid substitutions S93F/Y98F in the VP2 protein coding sequence, predicted to stabilize FMD virus particles. Cells infected with the optimized vector showed an ∼14-fold increase in protein expression as compared to cells infected with an unmodified Ad5 vector tested in previous works. Furthermore, amino acid substitutions in VP2 protein allowed the assembly of FMDV O1/Campos/Brazil/58 VLPs. Evaluation of several serological parameters in inoculated mice with the optimized Ad5[PVP2]OP candidate revealed an enhanced vaccine performance, characterized by significant higher titers of neutralizing antibodies, as compared to our previous unmodified Ad5 vector. Moreover, 94% of the mice vaccinated with the Ad5[PVP2]OP candidate were protected from homologous challenge. These results indicate that both the optimized protein expression and the stabilization of the in situ generated VLPs improved the performance of Ad5-vectored vaccines against the FMDV O1/Campos/Brazil/58 strain and open optimistic expectations to be tested in target animals.

Bovine Dendritic Cell Activation, T Cell Proliferation and Antibody Responses to Foot-And-Mouth Disease, Is Similar With Inactivated Virus and Virus Like Particles.

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals that causes severe economic losses in the livestock industry. Currently available vaccines are based on the inactivated FMD virus (FMDV). Although inactivated vaccines have been effective in controlling the disease, they have some disadvantages. Because of these disadvantages, investigations are being made to produce vaccines in low containment facilities. The use of recombinant empty capsids (also referred as Virus Like Particles, VLPs) has been reported to be a promising candidate as a subunit vaccine because it avoids the use of virus in the vaccine production and conserves the conformational epitopes of the virus. Mignaqui and collaborators have produced recombinant FMDV empty capsids from serotype A/ARG/2001 using a scalable technology in mammalian cells that elicited a protective immunity against viral challenge in a mouse model. However, further evaluation of the immune response elicited by these VLPs in cattle is required. In the present work we compare the effect that VLPs or inactivated FMDV has on bovine dendritic cells and the humoral response elicited in cattle after a single vaccination.

Characterization of GASA-1, a new vaccine candidate antigen of Babesia bovis.

Surface proteins bound to the cell membrane by glycosylphosphatidylinositol (GPI) anchors are considered essential for the survival of pathogenic protozoans. In the case of the tick-transmitted hemoparasite Babesia bovis, the most virulent causative agent of bovine babesiosis, the GPI-anchored proteome was recently unraveled by an in silico approach. In this work, one of the identified proteins, GASA-1 (GPI-Anchored Surface Antigen-1), was thoroughly characterized. GASA-1 is 179 aa long and has the characteristic features of a GPI-anchored protein, including a signal peptide, a hydrophilic core and a hydrophobic tail that harbors a GPI anchor signal. Transcriptomic analysis shows that it is expressed in pathogenic and attenuated B. bovis strains. Notably, the gasa-1 gene has syntenic counterparts in B. bigemina and B. ovata, which also encode GPI-anchored proteins. This is highly unusual since all piroplasmid GPI-anchored proteins described so far have been found to be species-specific. Sequencing of gasa-1 alleles from B. bovis geographical isolates originating from Argentina, USA, Brazil, Mexico and Australia showed over 98 % identity in both nucleotide and amino acid sequences. A recombinant form of GASA-1 (rGASA-1) was generated in E. coli and anti-rGASA-1 antibodies were raised in mice. Fixed and live immunofluorescence assays showed that GASA-1 is expressed in in vitro cultured B. bovis merozoites and surface-exposed. Moreover, incubation of B. bovis in vitro cultures with anti-GASA-1 antibodies partially, but significantly, reduced erythrocyte invasion, indicating that this protein bears neutralization-sensitive antibody epitopes. Splenocytes of rGASA-1-inoculated mice showed a specific proliferative response when exposed to the recombinant protein, indicating that GASA-1 bears T-cell epitopes. Finally, sera from a group of B. bovis-infected cattle reacted with the recombinant protein, demonstrating that GASA-1 is expressed during natural infection of bovines with B. bovis, and suggesting that it is immunodominant. The high degree of conservation among B. bovis isolates and the presence of syntenic genes in other Babesia species suggest a relevant role of GASA-1 and GASA-1-like proteins for parasite survival, especially considering that, due to their surface location, they are exposed to the selection pressure of the host immune system. The highlighted features of GASA-1 make it an interesting candidate for the development of vaccines against bovine babesiosis.

A New Cage-Like Particle Adjuvant Enhances Protection of Foot-and-Mouth Disease Vaccine.

Foot-and-Mouth Disease (FMD) is an acute viral disease that causes important economy losses. Vaccines with new low-cost adjuvants that stimulate protective immune responses are needed and can be assayed in a mouse model to predict their effectiveness in cattle. Immunostimulant Particle Adjuvant (ISPA), also known as cage-like particle adjuvant, consisting of lipid boxes of dipalmitoyl-phosphatidylcholine, cholesterol, sterylamine, alpha-tocopherol, and QuilA saponin, was shown to enhance protection of a recombinant vaccine against Trypanosoma cruzi in a mouse model. Thus, in the present work, we studied the effects on the magnitude and type of immunity elicited in mice and cattle in response to a vaccine based on inactivated FMD virus (iFMDV) formulated with ISPA. It was demonstrated that iFMDV-ISPA induced protection in mice against challenge and elicited a specific antibody response in sera, characterized by a balanced Th1/Th2 profile. In cattle, the antibody titers reached corresponded to an expected percentage of protection (EPP) higher than 80%. EPP calculates the probability that livestock would be protected against a 10,000 bovine infectious doses challenge after vaccination. Moreover, in comparison with the non-adjuvanted iFMDV vaccine, iFMDV-ISPA elicited an increased specific T-cell response against the virus, including higher interferon gamma (IFNγ)+/CD8+ lymphocyte production in cattle. In this work, we report for first time that an inactivated FMDV serotype A vaccine adjuvanted with ISPA is capable of inducing protection against challenge in a murine model and of improving the specific immune responses against the virus in cattle.

Mouse model as an efficacy test for foot-and-mouth disease vaccines.

Protection against foot-and-mouth disease virus (FMDV) has been linked to the development of a humoral response. In Argentina, the official control tests for assessing the potency of FMD vaccines are protection against podal generalization (PPG) and expected percentage of protection (EPP) curves built with quantitative data of antibodies determined by liquid-phase blocking ELISA (lpELISA). The results of these tests are used to accept or discard vaccines at the batch level. In this report, a mouse model was assessed as an alternative efficacy control for FMDV vaccines. To this aim, groups of cattle (n = 18) and BALB/c mice (n = 16) were inoculated with commercial FMDV vaccines and bleedings were performed 60 days post vaccination (dpv) in cattle and 21 dpv in mice. Specific FMDV antibody titres were measured in both species by a standardized lpELISA. A statistically significant association between antibody levels in cattle and mice has already been demonstrated. However, some vaccines have been misclassified since they were considered protective based on lpELISA results but did not induce good protection in cattle upon challenge. For this reason, other immunological parameters were evaluated to improve the prediction of protection in mice, without the need of using infective virus. In addition, antibody titres by lpELISA, the IgG2b/IgG1 isotype ratio and the Avidity Index were identified as good predictors, resulting in an optimal predictive model of protection. This mouse model could be a simple and economic alternative for testing FMD vaccines since the disadvantages of high costs and facility requirements associated with the use of large animals are overcome.

Immune response to Neospora caninum live tachyzoites in prepubertal female calves.

The aim of the present study was to characterize the specific immune response in prepubertal female calves inoculated with Neospora caninum. Forty-eight N. caninum-seronegative 6-month-old Angus female calves were randomly allocated into two groups: group A calves were inoculated subcutaneously (sc) with 1 × 106 tachyzoites of the low virulence NC-Argentina LP1 isolate in sterile phosphate-buffered saline (PBS); group B calves were mock inoculated sc with sterile PBS. Calves from group A developed a specific immune response characterized by the production of IgG antibodies and the expression of IFN-γ and TNF-α cytokines. Animals did not present any febrile reaction or reactions at the site of inoculation. Although chronic N. caninum infection was developed in 50% of calves of group A after inoculation, according to the presence of antibodies against rNc-SAG4, antigen characteristic of bradyzoites, N. caninum antibodies dropped below the cut-off of ELISA from day 210 post-inoculation onwards. Future trials using the same group of inoculated animals will allow the characterization of the evolution of the immune response during pregnancy and to determine whether the immunization with the local isolate is able to prevent congenital transmission and to protect against heterologous challenges.

Interleukin-8 Secreted by Glioblastoma Cells Induces Microvascular Hyperpermeability Through NO Signaling Involving S-Nitrosylation of VE-Cadherin and p120 in Endothelial Cells.

Glioblastoma is a highly aggressive brain tumor, characterized by the formation of dysfunctional blood vessels and a permeable endothelial barrier. S-nitrosylation, a post-translational modification, has been identified as a regulator of endothelial function. In this work we explored whether S-nitrosylation induced by glioblastoma tumors regulates the endothelial function. As proof of concept, we observed that S-nitrosylation is present in the tumoral microenvironment of glioblastoma in two different animal models. Subsequently, we measured S nitrosylation and microvascular permeability in EAhy296 endothelial cells and in cremaster muscle. In vitro, conditioned medium from the human glioblastoma cell line U87 activates endothelial nitric oxide synthase, causes VE-cadherin- S-nitrosylation and induces hyperpermeability. Blocking Interleukin-8 (IL-8) in the conditioned medium inhibited S-nitrosylation of VE-cadherin and hyperpermeability. Recombinant IL-8 increased endothelial permeability by activating eNOS, S-nitrosylation of VE-cadherin and p120, internalization of VE-cadherin and disassembly of adherens junctions. In vivo, IL-8 induced S-nitrosylation of VE-cadherin and p120 and conditioned medium from U87 cells caused hyperpermeability in the mouse cremaster muscle. We conclude that eNOS signaling induced by glioma cells-secreted IL-8 regulates endothelial barrier function in the context of glioblastoma involving S-nitrosylation of VE-cadherin and p120. Our results suggest that inhibiting S-nitrosylation may be an effective way to control and/or block damage to the endothelial barrier and prevent cancer progression.

Galectin-8 induces endothelial hyperpermeability through the eNOS pathway involving S-nitrosylation-mediated adherens junction disassembly.

The permeability of endothelial cells is regulated by the stability of the adherens junctions, which is highly sensitive to kinase-mediated phosphorylation and endothelial nitric oxide synthase (eNOS)-mediated S-nitrosylation of its protein components. Solid tumors can produce a variety of factors that stimulate these signaling pathways leading to endothelial cell hyperpermeability. This generates stromal conditions that facilitate tumoral growth and dissemination. Galectin-8 (Gal-8) is overexpressed in several carcinomas and has a variety of cellular effects that can contribute to tumor pathogenicity, including angiogenesis. Here we explored whether Gal-8 has also a role in endothelial permeability. We show that recombinant Gal-8 activates eNOS, induces S-nitrosylation of p120-catenin (p120) and dissociation of adherens junction, leading to hyperpermeability of the human endothelial cell line EAhy926. This pathway involves focal-adhesion kinase (FAK) activation downstream of eNOS as a requirement for eNOS-mediated p120 S-nitrosylation. This suggests a reciprocal, yet little understood, regulation of phosphorylation and S-nitrosylation events acting upon adherens junction permeability. In addition, glutathione S-transferase (GST)-Gal-8 pull-down experiments and function-blocking ß1-integrin antibodies point to ß1-integrins as cell surface components involved in Gal-8-induced hyperpermeability. Endogenous Gal-8 secreted from the breast cancer cell line MCF-7 has similar hyperpermeability and signaling effects. Furthermore, the mouse cremaster model system showed that Gal-8 also activates eNOS, induces S-nitrosylation of adherens junction components and is an effective hyperpermeability agent in vivo. These results add endothelial permeability regulation by S-nitrosylation as a new function of Gal-8 that can potentially contribute to the pathogenicity of tumors overexpressing this lectin.

Dendrimeric peptides can confer protection against foot-and-mouth disease virus in cattle.

Foot-and-mouth disease virus (FMDV) causes a highly contagious disease in cloven-hoofed animals. A synthetic vaccine candidate consisting of dendrimeric peptides harbouring two copies of a B-epitope [VP1(136-154)] linked to a T-cell epitope [3A(21-35)] of FMDV confers protection to type O FMDV challenge in pigs. Herein we show in cattle that novel dendrimeric peptides bearing a T-cell epitope [VP1(21-40] and two or four copies of a B-cell epitope [VP1(135-160)] from type O1 Campos FMDV (termed B2T and B4T, respectively) elicited FMDV specific immune responses to similar levels to a commercial vaccine. Animals were challenged with FMDV and 100% of vaccinated cattle with B2T or B4T were protected to podal generalization. Moreover, bovines immunized with B4T were completely protected (with no clinical signs) against FMDV challenge after three vaccine doses, which was associated with titers of viral neutralizing antibodies in serum higher than those of B2T group (p< 0.05) and levels of opsonic antibodies similar to those of animals immunized with one dose of FMDV commercial vaccine. Bovines vaccinated with both dendrimeric peptides presented high levels of IgG1 anti FMDV in sera and in mucosa. When IgA in nasal secretions was measured, 20% or 40% of the animals in B2T or B4T groups respectively, showed anti-FMDV IgA titers. In addition, B2T and B4T peptides evoked similar consistent T cell responses, being recognized in vitro by lymphocytes from most of the immunized cattle in the proliferation assay, and from all animals in the IFN-γ production assay. Taken together, these results support the potential of dendrimers B2T or B4T in cattle as a highly valuable, cost-effective FMDV candidate vaccine with DIVA potential.

Phylogeography and Ecological Niche Modeling of the Desert Iguana (Dipsosaurus dorsalis, Baird & Girard 1852) in the Baja California Peninsula.

Understanding the factors that explain the patterns of genetic structure or phylogeographic breaks at an intraspecific level is key to inferring the mechanisms of population differentiation in its early stages. These topics have been well studied in the Baja California region, with vicariance and the dispersal ability of individuals being the prevailing hypothesis for phylogeographic breaks. In this study, we evaluated the phylogeographic patterns in the desert iguana (Dipsosaurus dorsalis), a species with a recent history in the region and spatial variation in life history traits. We analyzed a total of 307 individuals collected throughout 19 localities across the Baja California Peninsula with 15 microsatellite DNA markers. Our data reveal the existence of 3 geographically discrete genetic populations with moderate gene flow and an isolation-by-distance pattern presumably produced by the occurrence of a refugium in the Cape region during the Pleistocene Last Glacial Maximum. Bayesian methods and ecological niche modeling were used to assess the relationship between population genetic structure and present and past climatic preferences of the desert iguana. We found that the present climatic heterogeneity of the Baja California Peninsula has a marked influence on the population genetic structure of the species, suggesting that there are alternative explanations besides vicariance. The information obtained in this study provides data allowing a better understanding of how historical population processes in the Baja California Peninsula can be understood from an ecological perspective.

Galectin-8 activates dendritic cells and stimulates antigen-specific immune response elicitation.

Galectin-8 (Gal-8) is a mammalian ß-galactoside-binding lectin, endowed with proinflammatory properties. Given its capacity to enhance antigen-specific immune responses in vivo, we investigated whether Gal-8 was also able to promote APC activation to sustain T cell activation after priming. Both endogenous [dendritic cells (DCs)] and bone marrow-derived DCs (BMDCs) treated with exogenous Gal-8 exhibited a mature phenotype characterized by increased MHC class II (MHCII), CD80, and CD86 surface expression. Moreover, Gal-8-treated BMDCs (Gal-8-BMDCs) stimulated antigen-specific T cells more efficiently than immature BMDCs (iBMDCs). Proinflammatory cytokines IL-3, IL-2, IL-6, TNF, MCP-1, and MCP-5, as well as growth factor G-CSF, were augmented in Gal-8-BMDC conditioned media, with IL-6 as the most prominent. Remarkably, BMDCs from Gal-8-deficient mice (Lgals8-/- BMDC) displayed reduced CD86 and IL-6 expression and an impaired ability to promote antigen-specific CD4 T cell activation. To test if Gal-8-induced activation correlates with the elicitation of an effective immune response, soluble Gal-8 was coadministrated with antigen during immunization of BALB/cJ mice in the experimental foot-and-mouth disease virus (FMDV) model. When a single dose of Gal-8 was added to the antigen formulation, an increased specific and neutralizing humoral response was developed, sufficient to enhance animal protection upon viral challenge. IL-6 and IFN-γ, as well as lymphoproliferative responses, were also incremented in Gal-8/antigen-immunized animals only at 48 h after immunization, suggesting that Gal-8 induces the elicitation of an inflammatory response at an early stage. Taking together, these findings argue in favor of the use of Gal-8 as an immune-stimulator molecule to enhance the adaptive immune response.

S-nitrosylation of VASP at cysteine 64 mediates the inflammation-stimulated increase in microvascular permeability.

We tested the hypothesis that platelet-activating factor (PAF) induces S-nitrosylation of vasodilator-stimulated phosphoprotein (VASP) as a mechanism to reduce microvascular endothelial barrier integrity and stimulate hyperpermeability. PAF elevated S-nitrosylation of VASP above baseline levels in different endothelial cells and caused hyperpermeability. To ascertain the importance of endothelial nitric oxide synthase (eNOS) subcellular location in this process, we used ECV-304 cells transfected with cytosolic eNOS (GFPeNOSG2A) and plasma membrane eNOS (GFPeNOSCAAX). PAF induced S-nitrosylation of VASP in cells with cytosolic eNOS but not in cells wherein eNOS is anchored to the cell membrane. Reconstitution of VASP knockout myocardial endothelial cells with cysteine mutants of VASP demonstrated that S-nitrosylation of cysteine 64 is associated with PAF-induced hyperpermeability. We propose that regulation of VASP contributes to endothelial cell barrier integrity and to the onset of hyperpermeability. S-nitrosylation of VASP inhibits its function in barrier integrity and leads to endothelial monolayer hyperpermeability in response to PAF, a representative proinflammatory agonist.NEW & NOTEWORTHY Here, we demonstrate that S-nitrosylation of vasodilator-stimulated phosphoprotein (VASP) on C64 is a mechanism for the onset of platelet-activating factor-induced hyperpermeability. Our results reveal a dual role of VASP in endothelial permeability. In addition to its well-documented function in barrier integrity, we show that S-nitrosylation of VASP contributes to the onset of endothelial permeability.