EpiJen:a server for multistep T cell epitope prediction

Background - The main processing pathway for MHC class I ligands involves degradation of proteins by the proteasome, followed by transport of products by the transporter associated with antigen processing (TAP) to the endoplasmic reticulum (ER), where peptides are bound by MHC class I molecules, and then presented on the cell surface by MHCs. The whole process is modeled here using an integrated approach, which we call EpiJen. EpiJen is based on quantitative matrices, derived by the additive method, and applied successively to select epitopes. EpiJen is available free online. Results - To identify epitopes, a source protein is passed through four steps: proteasome cleavage, TAP transport, MHC binding and epitope selection. At each stage, different proportions of non-epitopes are eliminated. The final set of peptides represents no more than 5% of the whole protein sequence and will contain 85% of the true epitopes, as indicated by external validation. Compared to other integrated methods (NetCTL, WAPP and SMM), EpiJen performs best, predicting 61 of the 99 HIV epitopes used in this study. Conclusion - EpiJen is a reliable multi-step algorithm for T cell epitope prediction, which belongs to the next generation of in silico T cell epitope identification methods. These methods aim to reduce subsequent experimental work by improving the success rate of epitope prediction.

MHCPred : a server for quantitative prediction of peptide-MHC binding

Accurate T-cell epitope prediction is a principal objective of computational vaccinology. As a service to the immunology and vaccinology communities at large, we have implemented, as a server on the World Wide Web, a partial least squares-base multivariate statistical approach to the quantitative prediction of peptide binding to major histocom-patibility complexes (MHC), the key checkpoint on the antigen presentation pathway within adaptive,cellular immunity. MHCPred implements robust statistical models for both Class I alleles (HLA-A*0101, HLA-A*0201, HLA-A*0202, HLA-A*0203,HLA-A*0206, HLA-A*0301, HLA-A*1101, HLA-A*3301, HLA-A*6801, HLA-A*6802 and HLA-B*3501) and Class II alleles (HLA-DRB*0401, HLA-DRB*0401and HLA-DRB* 0701).

Histidine hydrogen bonding in MHC at pH 5 and pH 7 modeled by molecular docking and molecular dynamics simulations

Hydrogen bonds play important roles in maintaining the structure of proteins and in the formation of most biomolecular protein-ligand complexes. All amino acids can act as hydrogen bond donors and acceptors. Among amino acids, Histidine is unique, as it can exist in neutral or positively charged forms within the physiological pH range of 5.0 to 7.0. Histidine can thus interact with other aromatic residues as well as forming hydrogen bonds with polar and charged residues. The ability of His to exchange a proton lies at the heart of many important functional biomolecular interactions, including immunological ones. By using molecular docking and molecular dynamics simulation, we examine the influence of His protonation/deprotonation on peptide binding affinity to MHC class II proteins from locus HLA-DP. Peptide-MHC interaction underlies the adaptive cellular immune response, upon which the next generation of commercially-important vaccines will depend. Consistent with experiment, we find that peptides containing protonated His residues bind better to HLA-DP proteins than those with unprotonated His. Enhanced binding at pH 5.0 is due, in part, to additional hydrogen bonds formed between peptide His+ and DP proteins. In acidic endosomes, protein His79β is predominantly protonated. As a result, the peptide binding cleft narrows in the vicinity of His79β, which stabilizes the peptide - HLA-DP protein complex. © 2014 Bentham Science Publishers.

Peptide binding to HLA-DP proteins at pH 5.0 and pH 7.0:a quantitative molecular docking study

Background: HLA-DPs are class II MHC proteins mediating immune responses to many diseases. Peptides bind MHC class II proteins in the acidic environment within endosomes. Acidic pH markedly elevates association rate constants but dissociation rates are almost unchanged in the pH range 5.0 - 7.0. This pH-driven effect can be explained by the protonation/deprotonation states of Histidine, whose imidazole has a pKa of 6.0. At pH 5.0, imidazole ring is protonated, making Histidine positively charged and very hydrophilic, while at pH 7.0 imidazole is unprotonated, making Histidine less hydrophilic. We develop here a method to predict peptide binding to the four most frequent HLA-DP proteins: DP1, DP41, DP42 and DP5, using a molecular docking protocol. Dockings to virtual combinatorial peptide libraries were performed at pH 5.0 and pH 7.0. Results: The X-ray structure of the peptide - HLA-DP2 protein complex was used as a starting template to model by homology the structure of the four DP proteins. The resulting models were used to produce virtual combinatorial peptide libraries constructed using the single amino acid substitution (SAAS) principle. Peptides were docked into the DP binding site using AutoDock at pH 5.0 and pH 7.0. The resulting scores were normalized and used to generate Docking Score-based Quantitative Matrices (DS-QMs). The predictive ability of these QMs was tested using an external test set of 484 known DP binders. They were also compared to existing servers for DP binding prediction. The models derived at pH 5.0 predict better than those derived at pH 7.0 and showed significantly improved predictions for three of the four DP proteins, when compared to the existing servers. They are able to recognize 50% of the known binders in the top 5% of predicted peptides. Conclusions: The higher predictive ability of DS-QMs derived at pH 5.0 may be rationalised by the additional hydrogen bond formed between the backbone carbonyl oxygen belonging to the peptide position before p1 (p-1) and the protonated ε-nitrogen of His 79β. Additionally, protonated His residues are well accepted at most of the peptide binding core positions which is in a good agreement with the overall negatively charged peptide binding site of most MHC proteins. © 2012 Patronov et al.; licensee BioMed Central Ltd.

EPIPOX:immunoinformatic characterization of the shared T-cell epitome between variola virus and related pathogenic orthopoxviruses

Concerns that variola viruses might be used as bioweapons have renewed the interest in developing new and safer smallpox vaccines. Variola virus genomes are now widely available, allowing computational characterization of the entire T-cell epitome and the use of such information to develop safe and yet effective vaccines. To this end, we identified 124 proteins shared between various species of pathogenic orthopoxviruses including variola minor and major, monkeypox, cowpox, and vaccinia viruses, and we targeted them for T-cell epitope prediction. We recognized 8,106, and 8,483 unique class I and class II MHC-restricted T-cell epitopes that are shared by all mentioned orthopoxviruses. Subsequently, we developed an immunological resource, EPIPOX, upon the predicted T-cell epitome. EPIPOX is freely available online and it has been designed to facilitate reverse vaccinology. Thus, EPIPOX includes key epitope-focused protein annotations: time point expression, presence of leader and transmembrane signals, and known location on outer membrane structures of the infective viruses. These features can be used to select specific T-cell epitopes suitable for experimental validation restricted by single MHC alleles, as combinations thereof, or by MHC supertypes.

Ampliação do Setor de Aulas Teóricas IV - UFRN: proposta arquitetônica baseada no conceito da flexibilidade e adaptabilidade, utilizando os parâmetros de conforto ambiental

In recent years, the Universidade Federal do Rio Grande do Norte (UFRN) has come through an intense process of expansion, common for the Instituições Federais de Ensino Superior (IFES) - Higher Education Institutions – since the implantation of the Programa de Reestruturação e Expansão das Universidades Federais (REUNI), a program of restructuring and expansion of the federal universities. The incentives made at UFRN can be seen into the expansion of the fields of teaching, research and extension, besides hiring human resources and adaptation / expansion of physical spaces. The focus of the UFRN at the technological area through the project (REUNI) has reflected in a significant expansion of the physical structure of the Center of Technology, area where the section of classes IV remains being the main set of rooms that supports the demand of the courses in the technological area, in which highlights the lack of classrooms. In this sense, the work presented here, proposes an architectural blueprint for expanding the physical space of this sector, based in this main parameters: (i) the spatial flexibility that attends the varying numbers of students per class; (ii) the environmental comfort, using the parameters and guidelines constructive for the Bioclimatic Zone 8; (iii) the requirements contained in the rules of ABNT NBR 9050:2004, 9077:2001 and NBR NBR 15220-3:2003. The architectural proposal used the maximum template allowed by the UFRN Central Campus Master Plan, four floors, considering the existing building (area equal to 7413.95 m²) as the ground floor and the extension proposed distributed on three floors, with an area of around 2,535, 00 m² each, making up a total of 7602.65 m² of total area built

Tratamento de cascalho de perfuração utilizando sistemas microemulsionados

The oil industry is one of the activities that generates more waste to the environment. The drill cuttings is a waste generated in large quantities in the drilling process and that may cause environmental damage such as soil contamination and consequently the contamination of groundwater if disposed of without prior treatment. Arises the need to develop scientific activities and research ways to adapt these wastes the current environmental standards. In the case of solid wastes, the NBR 10004: 2004 of the Brazilian Association of Technical Standards (ABNT) classifies them into class I waste (hazardous) and class II (not dangerous), which determines which wastes may or may not be discarded in the environment without causing environmental impact. This study presents a novel alternative for treating drill cuttings, where this waste was classified as class I (Abreu & Souza, 2005), mainly by removing the n-paraffin present in it, since this arises when using drilling fluids base oil. Using microemulsion systems promotes the removal of this contaminant drill cuttings samples from wells located in Alto do Rodrigues - RN. Initially, we determined the concentration of paraffin using infrared method in samples were extracted with ultrasound, we obtained a paraffin concentration in the range from 36.59 to 43.52 g of paraffin per kilogram of cuttings. Used two microemulsion systems containing two nonionic surfactants from different classes, one is an alcohol ethoxylated (UNTL-90) and the other an nonylphenol ethoxylated (RNX 110). The results indicated that the system UNTL-90 surfactant has better efficiency than the system with RNX 110. The study of the influence of contact time at the extraction showed that for times greater than 25 minutes has a tendency to increase the percentage extraction with increasing contact time. It was also observed that the extraction is fast because at 1 minute contact has 22.7% extraction. The reuse of the microemulsion system without removing the paraffin extracted in previous steps, showed reduction of 29.32 in percentage of extraction by comparing the first and third extraction, but by comparing the first and second extractions reduction is 8.5 in percentage extraction, so the systems reuse optimization can be an option for economically viable removing paraffin from cuttings. The extraction with shaking is more effective in the treatment of cuttings, reaching the extraction percentage of 87.04%, that is, obtaining a drill cuttings with 0.551% paraffin. Using the percentage of paraffin employed in non-aqueous drilling fluids and fluid maximum limit on cuttings for disposal established by the Environmental Protection Agency of the United States (US EPA), one arrives at the conclusion that the level of paraffin on gravel cannot exceed 3.93%. Conclude that the amount of paraffin in the treated cuttings with the microemulsion system with shaking is below the established by US EPA, showing that the system used was efficient in removing the paraffin from the drill cuttings.

Ethics in the provision of removable partial dentures

Aim: To investigate the construction of cobalt-chromium removable partial dentures by commercial private dental laboratories. Methods: Ninety master casts for fabrication of cobalt-chromium removable partial dentures were obtained from three commercial laboratories randomly selected. Casts were assessed for dental arch treated, Kennedy classification, cast surveying, denture design information provided by the dentist, and mouth preparation (rest seat, guiding plane and retentive area). Dental technicians answered a questionnaire regarding qualification of assisted dentists, monthly number of framework castings, and use of dental surveyor. Mouth preparation was compared among laboratories using Kruskal-Wallis test (α=0.05). Results: The percentage of Kennedy class I was 16%, class II 19%, class III 56%, and class IV 9%. The majority of master cats (51%) examined was sent to dental laboratories without any design information and did not comply with ethical guidelines in the provision of RPD. Approximately half of the casts were considered “inappropriate” for guiding planes and retentive areas. One of the laboratories presented all casts “inappropriate” for rest seat distribution (p<0.001). Conclusions: Mouth preparation frequently failed for guiding planes, retentive areas and distribution of rest seats. It is necessary to provide students with adequate clinical experience at the dental school environment, which will actually be carried into the practice of dentistry.

Ethics in the provision of removable partial dentures

Aim: To investigate the construction of cobalt-chromium removable partial dentures by commercial private dental laboratories. Methods: Ninety master casts for fabrication of cobalt-chromium removable partial dentures were obtained from three commercial laboratories randomly selected. Casts were assessed for dental arch treated, Kennedy classification, cast surveying, denture design information provided by the dentist, and mouth preparation (rest seat, guiding plane and retentive area). Dental technicians answered a questionnaire regarding qualification of assisted dentists, monthly number of framework castings, and use of dental surveyor. Mouth preparation was compared among laboratories using Kruskal-Wallis test (α=0.05). Results: The percentage of Kennedy class I was 16%, class II 19%, class III 56%, and class IV 9%. The majority of master cats (51%) examined was sent to dental laboratories without any design information and did not comply with ethical guidelines in the provision of RPD. Approximately half of the casts were considered “inappropriate” for guiding planes and retentive areas. One of the laboratories presented all casts “inappropriate” for rest seat distribution (p<0.001). Conclusions: Mouth preparation frequently failed for guiding planes, retentive areas and distribution of rest seats. It is necessary to provide students with adequate clinical experience at the dental school environment, which will actually be carried into the practice of dentistry.

Prótese parcial removível retida por implantes: relato de caso

A ausência de dentes posteriores inferiores representa dificuldades no planejamento e controle de próteses parciais removíveis, pois o suporte é proporcionado por dentes e mucosa, que apresentam características anatômicas diferentes. O planejamento de próteses parciais removíveis apoiadas sobre implantes na região distal torna a prótese dento-implanto-suportada e não dento-muco-suportada sendo, uma opção aos casos de extremidade livre. Por meio deste relato de caso, um paciente portador de arco classe II de Kennedy foi reabilitado com prótese parcial removível apoiada sobre implante na região distal. A colocação do implante resultou em uma oclusão mais estável, o que melhorou a funcionalidade da prótese e proporcionou maior conforto ao paciente. Observou-se que esta resolução pode ser uma alternativa viável de tratamento, pois apresenta custo reduzido em relação à prótese fixa sobre implantes, entretanto, requer um acompanhamento em longo prazo com uma série de casos para tornar-se um método rotineiro de tratamento.

Prótese parcial removível retida por implantes: relato de caso

A ausência de dentes posteriores inferiores representa dificuldades no planejamento e controle de próteses parciais removíveis, pois o suporte é proporcionado por dentes e mucosa, que apresentam características anatômicas diferentes. O planejamento de próteses parciais removíveis apoiadas sobre implantes na região distal torna a prótese dento-implanto-suportada e não dento-muco-suportada sendo, uma opção aos casos de extremidade livre. Por meio deste relato de caso, um paciente portador de arco classe II de Kennedy foi reabilitado com prótese parcial removível apoiada sobre implante na região distal. A colocação do implante resultou em uma oclusão mais estável, o que melhorou a funcionalidade da prótese e proporcionou maior conforto ao paciente. Observou-se que esta resolução pode ser uma alternativa viável de tratamento, pois apresenta custo reduzido em relação à prótese fixa sobre implantes, entretanto, requer um acompanhamento em longo prazo com uma série de casos para tornar-se um método rotineiro de tratamento.

The Irish paradigm on the natural progression of hepatitis C virus infection: an investigation in a homogeneous patient population infected with HCV 1b (Review)

The aetiological agent of chronic hepatitis C is the hepatitis C virus. The hepatitis C virus is spread by parenteral transmission of body fluids, primarily blood or blood products. In 1989, after more than a decade of research, HCV was isolated and characterised. The hepatitis C viral genome is a positive-sense, single-stranded RNA molecule approximately 9.4 kb in length, which encodes a polyprotein of about 3100 amino acids. There are 6 main genotypes of HCV, each further stratified by subtype. In 1994, a cohort of women was identified in Ireland as having been iatrogenically exposed to the hepatitis C virus. The women were all young and exposed as a consequence of the receipt of HCV 1b contaminated anti-D immunoglobulin. The source of the infection was identified as an acutely infected female. As part of a voluntary serological screening programme involving 62,667 people, 704 individuals were identified as seropositive for exposure to the hepatitis C virus; 55.4% were found to be positive for the viral genome 17 years after exposure. Of these women 98% had evidence of inflammation, but suprisingly, a remarkable 49% showed no evidence of fibrosis. Clinicopathology and virological analysis has identified associations between viral load and the histological activity index for inflammation, and, between inflammation and levels of the liver enzyme alanine aminotransferase. Infection at a younger age appears to protect individuals from progression to advanced liver disease. Molecular analyses of host immunogenetic elements shows that particular class II human leukocyte associated antigen alleles are associated with clearance of the hepatitis C virus. Additional class II alleles have been identified that are associated with stable viraemia over an extended period of patient follow-up. Although, investigation of large untreated homogeneous cohorts is likely to become more difficult, as the efficacy of anti-viral therapy improves, further investigation of host and viral factors that influence disease progression will help provide an evidence based approach were realistic expectations regarding patient prognosis can be ascertained.

Calcium/Calmodulin-Dependent Protein Kinase Kinase 2 (CaMKK2) Regulates Dendritic Cells and Myeloid Derived Suppressor Cells Development in the Lymphoma Microenvironment

Calcium (Ca2+) is a known important second messenger. Calcium/Calmodulin (CaM) dependent protein kinase kinase 2 (CaMKK2) is a crucial kinase in the calcium signaling cascade. Activated by Ca2+/CaM, CaMKK2 can phosphorylate other CaM kinases and AMP-activated protein kinase (AMPK) to regulate cell differentiation, energy balance, metabolism and inflammation. Outside of the brain, CaMKK2 can only be detected in hematopoietic stem cells and progenitors, and in the subsets of mature myeloid cells. CaMKK2 has been noted to facilitate tumor cell proliferation in prostate cancer, breast cancer, and hepatic cancer. However, whethter CaMKK2 impacts the tumor microenvironment especially in hematopoietic malignancies remains unknown. Due to the relevance of myeloid cells in tumor growth, we hypothesized that CaMKK2 has a critical role in the tumor microenvironment, and tested this hyopothesis in murine models of hematological and solid cancer malignancies.

We found that CaMKK2 ablation in the host suppressed the growth of E.G7 murine lymphoma, Vk*Myc myeloma and E0771 mammary cancer. The selective ablation of CaMKK2 in myeloid cells was sufficient to restrain tumor growth, of which could be reversed by CD8 cell depletion. In the lymphoma microenvironment, ablating CaMKK2 generated less myeloid-derived suppressor cells (MDSCs) in vitro and in vivo. Mechanistically, CaMKK2 deficient dendritic cells showed higher Major Histocompatibility Class II (MHC II) and costimulatory factor expression, higher chemokine and IL-12 secretion when stimulated by LPS, and have higher potent in stimulating T-cell activation. AMPK, an anti-inflammatory kinase, was found as the relevant downstream target of CaMKK2 in dendritic cells. Treatment with CaMKK2 selective inhibitor STO-609 efficiently suppressed E.G7 and E0771 tumor growth, and reshaped the tumor microenvironment by attracting more immunogenic myeloid cells and infiltrated T cells.

In conclusion, we demonstrate that CaMKK2 expressed in myeloid cells is an important checkpoint in tumor microenvironment. Ablating CaMKK2 suppresses lymphoma growth by promoting myeloid cells development thereby decreasing MDSCs while enhancing the anti-tumor immune response. CaMKK2 inhibition is an innovative strategy for cancer therapy through reprogramming the tumor microenvironment.

Adult orthodontics: internet information and a national survey

Aims: 1. To investigate the reliability and readability of information on the Internet on adult orthodontics. 2. To evaluate the profile and treatment of adults by specialist orthodontists in the Republic of Ireland (ROI). Materials and methods: 1. An Internet search was conducted in May 2015 using three search engines (Google, Yahoo and Bing), with two search terms (“adult orthodontics” and “adult braces”). The first 50 websites from each engine were screened and exclusion criteria applied. Included websites were then assessed for reliability using the JAMA benchmarks, the DISCERN and LIDA tools and the presence of the HON seal. Readability was assessed using the FRES. 2. A pilot-tested questionnaire about adult orthodontics was distributed to 122 eligible specialist orthodontists in the ROI. Questions addressed general and treatment information about adult orthodontic patients, methods of information provision and respondent demographics. Results: 1. Thirteen websites met the inclusion criteria. Three websites contained all JAMA benchmarks and one displayed the HON Seal. The mean overall score for DISCERN was 3.9/5 and the mean total LIDA score was 115/120. The average FRES score was 63.1. 2. The questionnaire yielded a response rate of 83%. The typical demographic profile of adult orthodontic patients was professional females between 25-35 years. The most common incisor relationship and skeletal base was Class II, division 1 (51%) and Class II (61%) respectively. Aesthetic upper brackets and metal lower brackets were the most frequently used appliances. Only 30% of orthodontists advise their adult patients to find extra information on the Internet. Conclusions: 1. The reliability and readability of information on the Internet on adult orthodontics is of moderate quality. 2. The provision of adult orthodontic treatment is common among specialist orthodontists in the Republic of Ireland.

Three-dimensional outcomes of twin-block appliance treatment and the influence of the postured wax bite

Aims 1 To identify the 3D soft tissue volumetric and linear landmark changes following treatment with the Twin-Block Appliance TBA. 2 To estimate the TBA treatment outcome on the soft tissue facial profile volumetric and linear landmark changes from the Postured Wax Bite (PWB). 3 To identify if there is any association between certain soft tissue landmark variables and successful treatment outcome of the TBA as measured by the reduction in overjet. 4 To assess the effects of TBA treatment on facial expressions. Materials and Methods Forty-seven Caucasian subjects with Class II division 1 were recruited. 3D images captured of each subject, pre-treatment (T1), with the PWB (T2) and at the end of treatment (T3). Soft tissue volumetric and linear changes as well as the correlation between facial parameters and successful treatment were calculated. Results The mean soft tissue volumetric change from T1 to T3 was 22.24 ± 16.73 cm³. Soft tissue profile linear changes from T1-T3 for lower facial landmarks were 4-5 mm. From T1-T3, the mean soft tissue volumetric change of the total sample was 60% of the change produced by the PWB (T1 to T2). Correlations were weak for all 3D facial parameters and successful overjet reduction. Facial expression changes were only significant for the lower landmarks. Conclusions 1 TBA treatment, in growing subjects, increased the lower facial soft tissue volume and caused forward movement of the lower soft tissue facial profile landmarks.2 The PWB can be used to estimate the treatment outcome of the TBA on soft tissue profile changes.3 No association was found between soft tissue landmark variables and successful overjet reduction.4 TBA treatment had no effect on the upper facial landmarks for each facial expression but it changed the lower facial expressions significantly except for maximal smile in males.