The influence of esterification of carboxyl groups of ribonuclease-a on its structure and immunological activity

The effect of modification of carboxyl groups of Ribonuclease-Aa on the enzymatic activity and the antigenic structure of the protein has been studied. Modification of four of the eleven free carboxyl groups of the protein by esterification in anhydrous methanol/0.1 M hydrochloric acid resulted in nearly 80% loss in enzymatic activity but had very little influence on the antigenic structure of the protein. Further increases in the modification of the carboxyl groups caused a progressive loss in immunological activity, and the fully methylated RNase-A exhibited nearly 30% immunological activity. Concomitant with this change in the antigenic structure of the protein, the ability of the molecule to complement with RNase-S-protein increased, clearly indicating the unfolding of the peptide "tail" from the remainder of the molecule. The susceptibility to proteolysis, accessibility of methionine residues for orthobenzoquinone reaction and the loss in immunological activity of the more extensively esterified derivatives of RNase-A are suggestive of the more flexible conformation of these derivatives as compared with the compact native conformation. The fact that even the fully methylated RNase-A retains nearly 30% of its immunological activity suggested that the modified protein contained antibody recognizable residual native structure, which presumably accommodates some antigenic determinants.

Increased potential of a cationic liposome-based delivery system:enhancing stability and sustained immunological activity in pre-clinical development

The combination of dimethyl dioctadecyl ammonium bromide (DDA) and the synthetic cord factor trehalose dibehenate (TDB) with Ag85B-ESAT-6 (H1 fusion protein) has been found to promote strong protective immune responses against Mycobacterium tuberculosis. The development of a vaccine formulation that is able to facilitate the requirements of sterility, stability and generation of a vaccine product with acceptable composition, shelf-life and safety profile may necessitate selected alterations in vaccine formulation. This study describes the implementation of a sterilisation protocol and the use of selected lyoprotective agents in order to fulfil these requirements. Concomitantly, close analysis of any alteration in physico-chemical characteristics and parameters of immunogenicity have been examined for this promising DDA liposome-based tuberculosis vaccine. The study addresses the extensive guidelines on parameters for non-clinical assessment, suitable for liposomal vaccines and other vaccine delivery systems issued by the World Health Organisation (WHO) and the European Medicines Agency (EMEA). Physical and chemical stability was observed following alteration in formulations to include novel cryoprotectants and radiation sterilisation. Immunogenicity was maintained following these alterations and even improved by modification with lysine as the cryoprotective agent for sterilised formulations. Taken together, these results outline the successful alteration to a liposomal vaccine, representing improved formulations by rational modification, whilst maintaining biological activity.

Immunological and microbiological activity of Davilla elliptica St. Hill. (Dilleniaceae) against Mycobacterium tuberculosis

Mycobacterium tuberculosis is responsible for over 8 million cases of tuberculosis (TB) annually. Natural products may play important roles in the chemotherapy of TB. The immunological activity of Davilla elliptica chloroform extract (DECE) was evaluated in vitro by the determination of hydrogen peroxide (H2O2), nitric oxide (NO), and tumor necrosis factor-alpha (TNF-alpha) release in peritoneal macrophages cultures. DECE was also tested for its antimycobacterial activity against M. tuberculosis using the microplate alamar blue assay. DECE (50, 150, 250 µg/ml) stimulated the production of H2O2 (from 1,79 ± 0,23 to 7,27 ± 2,54; 15,02 ± 2,86; 20,5 ± 2,1 nmols) (means ± SD), NO (from 2,64 ± 1,02 to 25,59 ± 2,29; 26,68 ± 2,41; 29,45 ± 5,87 µmols) (means ± SD) and TNF-alpha (from 2,44 ± 1,46 to 30,37 ± 8,13; 38,68 ± 1,59; 41,6 ± 0,90 units/ml) (means ± SD) in a dose-dependent manner and also showed a promising antimycobacterial activity with a minimum inhibitory concentration of 62,5 µg/ml. This plant may have therapeutic potential in the immunological and microbiological control of TB.

Freeze-drying of ovalbumin loaded mesoporous silica nanoparticle vaccine formulation increases antigen stability under ambient conditions

Amino functionalised mesoporous silica nanoparticles (AM-41) have been identified as a promising vaccine delivery material. The capacity of AM-41 to stabilise vaccine components at ambient temperature (23–27 °C) was determined by adsorbing the model antigen ovalbumin (OVA) to AM-41 particles (OVA-41). The OVA-41 was successfully freeze-dried using the excipients 5% trehalose and 1% PEG8000. Both the immunological activity of OVA and the nanoparticle structure were maintained following two months storage at ambient temperature. The results of immunisation studies in mice with reconstituted OVA-41 demonstrated the induction of humoral and cell-meditated immune responses. The capacity of AM-41 particles to facilitate ambient storage of vaccine components without loss of immunological potency will underpin the further development of this promising vaccine delivery platform.

Intra-epithelial T cells in coeliac disease

Coeliac disease is an enteropathy associated with dietary gluten which occurs in individuals with a genetic predisposition. The pathogenesis remains obscure although it is clear that only certain parts of the gliadin molecule are toxic and there is considerable evidence of immunological activity, including antibody production. In this issue of European Journal of Gastroenterology and Hepatology Carton et al. present evidence in favour of an inherent depletion of CD4CD8 T cells, which could result in a loss of oral tolerance to ingested gliadin. Using flow cytometry they also demonstrated that the classic T-cell infiltration of coeliac disease is not due to an increase in T cells but is an apparent increase associated with a relative decrease in enterocytes as a result of the change in architecture of the mucosa. These could be important fundamental observations in helping to unravel the pathogenesis of coeliac disease.

A soy-based product fermented by Enterococcus faecium and Lactobacillus helveticus inhibits the development of murine breast adenocarcinoma

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Ação do extrato metanólico e etanólico de Davilla elliptica St. Hill. (Malpighiaceae) na resposta imune

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

Immunostimulatory effects of the phenolic compounds from lichens on nitric oxide and hydrogen peroxide production

The effects of isolated compounds from Brazilian lichens and their derivatives on H 2O 2 and NO production were studied using murine macrophages as a part of an attempt to understand their possible immunomodulatory properties. The compound cytotoxicity was studied using MTT assay. Macrophage stimulation was evaluated by the determination of NO (Griess assay) and H 2O 2 (horseradish peroxidase/phenol red) in supernatants of peritoneal macrophage cultures of Swiss mice. This research demonstrated stimulatory activities of some phenolic compounds isolated from lichens and their derivatives on H 2O 2 and NO production. Structure-activity relationships suggest several synthetic directions for further improvement of immunological activity.

Efeitos imunológicos da ingestão do produto de soja fermentado em ratos induzidos quimicamente com câncer de cólon

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Atividade antimicrobiana e imunológica in vitro dos extratos de Senna reticulata (Willd.) Irwin & Barneby (mata-pasto) E Vismia guianensis (Aubl.)(lacre)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Influência da resposta crônica sobre as variáveis hematológicas de indivíduos diabéticos tipo 2 submetidos a um treinamento misto de bola suíça

The main function of the immune system is the defense against infection, being composed by the leukocytes that modulate the immune response, which can be innate or adaptive. The physical exercise can causes positive or negative alterations in the total or relative number of leukocytes. When the exercise has a low or moderate intensity it`s considered beneficial for improving the function of the cells responsible for the defense and to reduce the risk of infectious illnesses. The type 2 diabetes is related to an incapacity of the body in rightly respond to insulin, associated to an resistance to its actions. The purpose of this research was to make a study of the immune system characteristics, as well as the type 2 diabetes and the relation among both and the physical exercise. So, it was analyzed a group of type 2 diabetics coming from Diabetics House of Franca -SP, treated through a program of mix physical training. Two collections of blood were pre and post training program for the pair comparable of the participants. It was found positive alterations of the subpopulations of leukocytes that show a probable improvement of the immunological state what allows us to suggest about a possible improvement of the immunological activity, what would be measured by the activation of these cells forward to an inflammatory/infectious condition. So, we suggest that future studies involving diabetes, immune system and physical exercise be encouraged.

Impatto della glomerulopatia cronica da trapianto sulla disfunzione tardiva del rene trapiantato

INTRODUCTION. Late chronic allograft disfunction (CAD) is one of the more concerning issues in the management of patients (pts) with renal transplant (tx). Humoral immune response seems to play an important role in CAD pathogenesis. AIM OF THE STUDY. To identify the causes of late chronic allograft disfunction. METHODS. This study (march 2004-august 2011) enrolled pts who underwent renal biopsy (BR) because of CAD (increase of creatininemia (s-Cr) >30% and/or proteinuria >1g/day at least one year after tx). BR were classified according to 1997/2005 Banff classification. Histological evaluation of C4d (positive if >25%), glomerulitis, tubulitis, intimal arteritis, atrophy/fibrosis and arteriolar-hyalinosis were performed. Ab anti-HLA research at BR was an inclusion criteria. Pts were divided into two groups: with or without transplant glomerulopathy (CTG). RESULTS. Evaluated BR: 93/109. BR indication: impaired s-Cr (52/93), proteinuria (23/93), both (18/93). Time Tx-BR: 7.4±6.3 yrs; s-Cr at BR: 2.7±1.4 mg/dl. CTG group(n=49) not-CTG group(n=44) p Time tx-BR (yrs) 9.3±6.7 5.3±5.2 0.002 Follow-up post-BR (yrs) 2.7±1.8 4.1±1.4 0.0001 s-Cr at BR (mg/dl) 2.9±1.3 2.4±1.5 NS Rate (%) of pts: Proteinuria at BR 61% 25% 0.0004 C4d+ 84% 25% <0.0001 Ab anti-HLA+ 71% 30% 0.0001 C4d+ and/or Ab antiHLA 92% 43% 0.0001 Glomerulitis 76% 16% <0.0001 Tubulitis 6% 32% 0.0014 Intimal arteritis 18% 0% 0.002 Arteriolar hyalinosis 65% 50% NS Atrophy/fibrosis 80% 77% NS Graft survival 45% 86% 0.00005 Histological Diagnosis: CTG group (n=49:Chronic rejection 94%;IgA recurrence + humoral activity 4%;IIA acute rejection + humoral activity 2%. Not-CTG group (n=44: GN recurrence 27%;IF/TA 23%; acute rejection 23%;BKV nephritis 9%; mild not specific alterations 18%. CONCLUSIONS: CTG is the morphological lesion mainly related to CAD. In the 92% of the cases it is associated with markers of immunological activity. It causes graft failure within five years after diagnosis in 55% of pts.

Host-specificity of monogenean (platyhelminth) parasites: a role for anterior adhesive areas?

Monogeneans (flatworms) are among the most host-specific of parasites in general and may be the most host-specific of all fish parasites. Specificity, in terms of a restricted spatial distribution within an environment, is not unique to parasites and is displayed by some fungi, insects, birds, symbionts and pelagic larvae of free-living marine invertebrates. The nature of cues, how habitats are recognised and how interactions between partners are mediated and maintained is of interest across these diverse associations. We review some experiments that demonstrate important factors that contribute to host-specificity at the level of infective stages (larvae of oviparous monogeneans; juveniles of viviparous gyrodactylids) and adult parasites. Recent research on immune responses by fish to monogenean infections is considered. We emphasise the critical importance of host epidermis to the Monogenea. Monogeneans live on host epidermis, they live in its products (e.g. mucus), monopisthocotyleans feed on it, some of its products are attractants and it may be an inhospitable surface because of its immunological activity. We focus attention on fish but reference is made to amphibian hosts. We develop the concept for a potential role in host-speciality by the anterior adhesive areas, either the specialised tegument and/or anterior secretions produced by monogeneans for temporary but firm attachment during locomotion on host epithelial surfaces. Initial contact between the anterior adhesive areas of infective stages and host epidermis may serve two important purposes. (1) Appropriate sense organs or receptors on the parasite interact with a specific chemical or chemicals or with surface structures on host epidermis. (2) A specific but instant recognition or reaction occurs between component(s) of host mucus and the adhesive(s) secreted by monogeneans. The chemical composition of fish skin is known to be species-specific and our preliminary analysis of the chemistry of some monogenean adhesives indicates they are novel proteins that display some differences between parasite families and species. (C) 2000 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Estudio de las modificaciones conformacionales y su influencia en la actividad inmunológica de péptidos derivados de las familias de las proteínas spect-1 y -2, siap-1 y -2 de plasmodium falciparum

SPECT-1 y -2 y SIAP-1 y -2 son proteínas pertenecientes al esporozoíto de Plasmodium falciparum causante de la malaria más agresiva en los humanos. Estas proteínas están involucradas en el paso del parásito a través de las células del hospedero humano y en la invasión del hepatocito, haciéndolas blancos atractivos para ser estudiadas. Péptidos conservados de alta capacidad de unión (cHABPs) a células HeLa y HepG2 derivados de estas moléculas son no inmunogénicos porque son incapaces de generar una respuesta inmunitaria, pero son claves para el parásito porque cumplen una función importante durante la infección del hospedero humano. En este trabajo se encontró que algunos cHABPs pertenecientes a las proteínas SPECT-1 y -2, están posiblemente involucrados con la unión y formación de poros sobre la membrana de las células hospederas, ayudando al esporozoíto a abrirse paso través de las células del hospedero. Por otro lado, con el fin de cambiar el comportamiento inmunológico de cHABPs derivados de SPECT-1 y -2 y SIAP-1 y -2, se obtuvieron nuevos péptidos mediante el reemplazo de aminoácidos críticos por otros residuos cuya masa molecular sea similar, pero diferente en su polaridad. En este trabajo se reporta que dichas modificaciones promovieron cambios en la estructura secundaria (determinada por DC o 1H-RMN) de los péptidos modificados (mHABPs) cuando se comparó con la estructura de los cHABPs nativos; adicionalmente, estos mHABPs invirtieron su comportamiento inmunológico convirtiéndose en péptidos inmunogénicos inductores de anticuerpos. Lo que permite establecer la existencia de una relación entre la estructura que adoptan estos mHABPs con su actividad inmunológica. Además, algunos de los mHABPs estudiados aquí, pueden ser candidatos a ser incluidos en la vacuna contra la malaria químicamente sintetizada multi-epitope y multi-estadio que se está desarrollando en la Fundación Instituto de Inmunología de Colombia (FIDIC).

Purification and physicochemical, kinetic and immunological properties of allosteric serine hydroxymethyltransferase from monkey liver.

The homogeneous serine hydroxymethyltransferase purified from monkey liver, by the use of Blue Sepharose affinity chromatography, exhibited positive homotropic co-operative interactions (h = 2.5) with tetrahydrofolate and heterotropic interactions with L-serine and nicotinamide nucleotides. The enzyme had an unusually high temperature optimum of 60 degrees C and was protected against thermal inactivation by L-serine. The allosteric effects were abolished when the monkey liver enzyme was purified by using a heat-denaturation step in the presence of L-serine, a procedure adopted by earlier workers for the purification of this enzyme from mammalian and bacterial sources. The enzyme activity was inhibited completely by N5-methyltetrahydrofolate, N5-formyltetrahydrofolate, dichloromethotrexate, aminopterin and D-cycloserine, whereas methotrexate and dihydrofolate were partial inhibitors. The insoluble monkey liver enzyme-antibody complex was catalytically active and failed to show positive homotropic co-operative interactions with tetrahydrofolate (h = 1) and heterotropic interactions with NAD+. The enzyme showed a higher heat-stability in a complex with its antibody than as the free enzyme. These results highlight the pitfalls in using a heat-denaturation step in the purification of allosteric enzymes.