A Novel Nomogram to Identify Candidates for Extended Pelvic Lymph Node Dissection Among Patients with Clinically Localized Prostate Cancer Diagnosed with Magnetic Resonance Imaging-targeted and Systematic Biopsies

Background

Available models for predicting lymph node invasion (LNI) in prostate cancer (PCa) patients undergoing radical prostatectomy (RP) might not be applicable to men diagnosed via magnetic resonance imaging (MRI)-targeted biopsies.

Identification of casein as the major allergenic and antigenic protein of cow's milk

The objective of this study was to analyze both the allergenicity and immunogenicity of cow's milk proteins. To this end, 80 milk-atopic patients were selected on the basis of the presence of cow's milk-specific IgE antibodies in serum and compatible clinical history. Fifteen patients allergic to other allergens and 10 nonatopic subjects were studied as controls. The specificity of serum IgG and IgE antibodies was determined by immunoblotting, employing both cow's milk and milk components, i.e., α- and β-casein, β-lactoglobulin, and α-lactalbumin separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The experiments showed that casein-specific IgE antibodies were present in all (80/80) sera examined; 10/80 showed reactivity to α-lactoglobulin, and 5/80 showed reactivity to α-lactalbumin. None of the 25 negative control sera analyzed showed the presence of specific IgE antibodies against milk proteins. These results were similar to those corresponding to the detection, by the radioallergosorbent test, of IgE antibodies against the milk components coupled to paper disks. All sera from milk-atopic patients also showed IgE reactivity against a high-molecular-mass fraction that hardly enters the gel. This fraction, after separation by gel filtration and treatment with β-mercaptoethanol and urea, was shown by SDS-PAGE analysis to be formed by casein monomers. All sera analyzed by immunoblotting reacted against the components corresponding to casein monomers. Inhibition of immunoblotting by adsorption with different milk components confirmed that those high-molecular-mass aggregates are formed by casein components. The results presented here strongly suggest that casein is the major allergenic component of cow's milk.     

Brucella lumazine synthase elicits a mixed Th1-Th2 immune response and reduces infection in mice challenged with Brucella abortus 544 independently of the adjuvant formulation used

The immunogenicity and protective efficacy of recombinant lumazine synthase from Brucella spp. (rBLS) administered with different adjuvants was evaluated in mice. Mice were immunized with rBLS in the absence or the presence of aluminum hydroxide gel (BLS-Al), monophosphoryl lipid A (BLS-MPA), or incomplete Freund's adjuvant (BLS-IFA). rBLS per se induced a vigorous immunoglobulin G (IgG) response, with high titers of IgG1 as well as IgG2. All the adjuvants increased this response; the BLS-IFA formulation was the most effective at inducing BLS-specific IgG antibodies. In addition, after in vitro stimulation with rBLS, spleen cells from BLS-IFA-, BLS-Al-, or BLS-MPA-immunized mice proliferated and produced interleukin-2 (IL-2), gamma interferon (IFN-gamma), IL-10, and IL-4, suggesting the induction of a mixed Th1-Th2 response. Immunization with rBLS protected mice against challenge with B. abortus 544. The levels of protection in the spleen were similar for all adjuvants, but only BLS-Al and BLS-IFA were effective in the liver. Our results indicate that BLS might be a useful candidate for the development of subunit vaccines against brucellosis, since it elicits antigen-specific cellular responses, with production of IFN-gamma and protection, independently of the adjuvant formulation used.     

Development of high‐sensitive enzyme immunoassays for gliadin quantification using the streptavidin‐biotin amplification system

Optimization of three enzyme immunoassays of very high sensitivity using three antiprolamin monoclonal antibodies (MAbs) (13B4, 11C4 and 12A1) is presented here. These MAbs are specific for those prolamins toxic for coeliac patients, as determined by immunoblotting analysis. Biotinylated MAbs were used in two of the assays. In a competitive ELISA, the binding of each biotinylated MAb to a gliadin‐coated solid phase was inhibited by gliadin in the fluid phase. The best result was obtained using the biotinylated MAbl3B4 (detection limit: 20 ng ml^?1). With regard to capture ELISA, we tested the performance of the three MAbs. In this sandwich ELISA, the MAb used for antigenic capture was the same as that used as secondary biotinylated antibody. The MAbl2Al had the best performance (detection limit: 1 ng ml^?1). The use of biotin‐labelled gliadin in a quantitative immunoassay with a detection limit of 5 ng ml^?1 is also reported. This assay involves an antigenic capture using the MAbl2Al followed by a competition between biotinylated and non‐biotinylated gliadin. We have found the use of the streptavidin‐biotin interaction as signal amplification system to be very useful. This technique, as far as we know, has not been previously reported for gliadin quantification.     

Isolation of a Trypanosoma cruzi antigen by affinity chromatography with a monoclonal antibody. Preliminary evaluation of its possible applications in serological tests.

By affinity chromatography with a monoclonal antibody (163B6), obtained in our laboratory, we have isolated a T. cruzi antigen which could be useful for differential diagnosis of Chagas' disease from leishmaniasis. This antigen, a 52-kD protein, reacted with all sera from Chagas' disease patients tested but not with sera from patients with leishmania, in ELISA. The 52-kD antigen is widely distributed in the Trypanosoma genus since the 163B6 monoclonal antibody reacts with T. rangeli and 8 strains and a clone of T. cruzi epimastigotes.     

A lytic monoclonal antibody to Trypanosoma cruzi bloodstream trypomastigotes which recognizes an epitope expressed in tissues affected in Chagas' disease.

It has been suggested that molecular mimicry between the antigens of Trypanosoma cruzi and the host could have a role in the onset of the chronic stage of Chagas' disease. In this article, we report on a monoclonal antibody (MAb), CAK20.12 (immunoglobulin G2b), which reacts with a polypeptidic epitope of a 150-kDa antigen expressed on the surface of several strains of T. cruzi. This MAb also causes lysis of bloodstream trypomastigotes. Serum samples from 30 of 30 patients with chronic and 11 of 13 patients with acute Chagas' disease present specific antibodies to this antigen. MAb CAK20.12 reacts, by indirect immunofluorescence, with human and syngeneic murine striated muscle tissue, with the smooth muscle layer of cardiac arteries, with the lamina muscularis mucosae and the external striated muscle layer of the esophagus, and with the smooth muscle cells of the colon from normal syngeneic mice. Reactivity with the small intestine was very weak, and no reactivity with ventricle or atrium tissue was detected. Adsorption with an antigenic fraction from normal murine striated muscle or from T. cruzi epimastigotes confirmed that MAb CAK20.12 recognizes a common epitope present in parasites and host tissues. MAb CAK20.12, lytic for the infective form of T. cruzi, recognizes an epitope expressed in striated and smooth muscle cells of the host tissues affected in the chronic stage of Chagas' disease.     

In vitro immunomodulatory properties of tucaresol in HIV infection

The immunomodulatory properties of tucaresol (compound 589C80) were tested on in vitro antigen- and mitogen-stimulated proliferation and cytokine production by peripheral blood mononuclear cells (PBMC) of HIV-infected individuals and healthy controls (HC). Results showed that tucaresol: (1) increases influenza A virus-, gp 160 peptide-, and HLA alloantigen-stimulated proliferation as well as interleukin (IL)-2 and interferon gamma (IFN gamma) production by PBMC of HIV-infected individuals with higher CD4 counts (>500/microl) but had only a marginal immunomodulatory effect on PBMC of patients with lower CD4 counts (<500/microl); (2) did not modify IL-10 production; (3) augmented CD25 expression on mitogen-stimulated T cells of HC but not of HIV-infected individuals; and (4) marginally increased CTL activity. The immunomodulatory properties of tucaresol were confirmed by PCR analyses; additional data showed that tucaresol costimulated CD3-dependent triggering of T cells and that this stimulation was independent of CD28 costimulation. The immunomodulatory effects of tucaresol on T cell functions are characterized by a bell-shaped dose response curve; the action of the compound is optimal in the 100 to 300 microM range. Analyses of mitogen-stimulated apoptosis demonstrated that the lack of effect of tucaresol at higher doses is not the result of increased cell death, suggesting a role of functional impairment. These data confirm that tucaresol can stimulate T helper cell function and enhance the production of type 1 cytokines, thus eliciting cell-mediated immunity, and warrant its potential utility in the therapy of HIV infection.     

Occurrence and Potential Diagnostic Applications of Serological Cross-Reactivities between Brucella and Other Alpha-Proteobacteria

Agrobacterium, Sinorhizobium, and Ochrobactrum are genera closely related to Brucella but, in contrast to the latter, are not pathogenic for humans and animals. We studied by an indirect enzyme-linked immunosorbent assay (ELISA) the reactivities of brucellosis sera against cytosolic (CYT) and membrane (MA) antigens from these nonpathogenic bacteria, and we evaluated the potential usefulness of these cross-reactions for the diagnosis of brucellosis in humans, sheep, cows, and dogs. Canine infection by Brucella canis was detected with high specificity by CYT antigen-based ELISAs (96% for Agrobacterium, 96% for Sinorhizobium, and 91% for Ochrobactrum), while sensitivity was variable (58% for Agrobacterium, 88% for Sinorhizobium, and 84% for Ochrobactrum). In addition, it was possible to diagnose canine disease shortly after exposure to the pathogen (15 days). Similar results for canine brucellosis were obtained with MA antigens. In contrast, normal sera from humans, sheep, and cattle reacted strongly with all the antigens (CYT and MA antigens from the three bacteria), producing high cutoff values and, consequently, low sensitivities. While for some host species the reactivity patterns of normal sera by Western blotting were similar to those produced with sera from infected individuals, the reactivity pattern of bovine sera against Sinorhizobium meliloti antigens exhibited some differential bands for the two groups of sera. These results show that crude fractions from nonpathogenic alpha-proteobacteria can be used to diagnose canine brucellosis but may need to be further separated into simpler fractions to have diagnostic usefulness in ovine, bovine, or human infection. By reducing the biosafety requirements, the use of antigens derived from these nonpathogenic bacteria would simplify the production of diagnostic kits for brucellosis, especially in settings where biosafety level-3 facilities are scarce or absent.     

The lupus-prone BXSB strain: the Yaa gene model of systemic lupus erythematosus

Conclusion Genetic analysis of SLE in the various autoimmune mice has revealed that this disease is multigenic in nature and that several, quite distinct genetic backgrounds are compatible with this disease. Although the nature of these genetic components has not been defined, studies on New Zealand mice have indicated that multiple, unlinked genes are responsible for the expression of various disease manifestations and the production of autoantibodies. However, it is significant that single gene mutations, such as lpr, gld and Yaa, markedly influence the development of the lupus-like autoimmune syndrome. In addition, it is becoming clear, based on the results obtained from H-2 congenic mice, that the MHC class II genes play a crucial role in the development of SLE. However, the absence of severe autoimmune pathology in mice lacking the SLE background, even in the presence of the single autoimmune mutant gene and of the appropriate MHC class II genes, clearly indicates the requirement of supplementary genetic abnormalities for the fullblown manifestations of SLE.Although the abnormality of the lpr and possibly gld mutation is associated with the molecules mediating the apoptosis, the nature of the Yaa gene defect has not yet been identified. The differences in autoimmune accelerating effects between the Yaa gene and the lpr gene on autoimmune-prone and non-autoimmune mice strongly suggest that the molecular defect of the Yaa gene is likely to differ from those of the lpr and gld genes. We propose that the Yaa gene effect may result from the expression of the Yaa gene-related molecule on B cells. This molecule could behave as an intercellular adhesion-like molecule, thereby facilitating the interaction and subsequent activation of autoreactive T and B cells. Alternatively, the Yaa molecule-derived peptide could be efficiently presented in the context of MHC class II antigens by B cells, and its recognition by Yaa-specific T helper cells could activate autoantigen-specific B cells even in the absence of autoantigen-specific T helper cells. Obviously, the molecular identification of the Yaa gene product would be of paramount importance in helping answer this important and interesting question.     

A psychometric-genetic study of schizotypal disorder

This study aimed to clarify the structure and the etiological constituents of schizotypal disorder (SPD) by directly interviewing pairs of twins. A latent class analysis was applied to each individual's outcome for specified SPD items, such that each subject's phenotype could be redefined in terms of individual probabilities of class membership. Intraclass correlations were then calculated for each twin pair, and a standard univariate twin model applied. The best latent class analysis solution encompassed a model with four latent classes (three latent classes of SPD subjects, one of non-SPD). The intraclass correlations revealed a moderate to high heritability for two out of three SPD classes and for the modal class (a class composed of subjects that possessed a conditional probability of belonging to any of the SPD classes). Model fittings revealed considerable variation in the extent to which the different classes of SPD were influenced by additive genetic constituents or non-genetic factors. Although these data confirm the importance of genetic contributors in determining liability to SPD and the schizophrenia spectrum, they indicate that there is a relationship between psychometric multidimensionality and etiological heterogeneity in SPD.