Towards a Taenia solium cysticercosis vaccine: an epitope shared by Taenia crassiceps and Taenia solium protects mice against experimental cysticercosis

The Taenia crassiceps recombinant antigen KETc7 has been shown to be effective as a vaccine against experimental murine cysticercosis, a laboratory model used to test potentially promising molecules against porcine Taenia solium cysticercosis. Based on the deduced amino acid sequence of this proline-rich polypeptide, three fragments, GK-1, GK-2, and GK-3, were chemically synthesized in linear form. Of the three peptides, only GK-1 induced sterile protection against T. crassiceps cysticercosis in 40 to 70% of BALB/cAnN male mice. GK-1 is an 18-amino-acid peptide which contains at least one B-cell epitope, as demonstrated by its ability to induce an antibody response to the peptide and T. crassiceps antigen without need of a carrier protein. Immunofluorescence studies revealed that anti-GK1 antibodies strongly react with the native protein in the tegument of T. crassiceps and also with anatomical structures of T. solium eggs, oncospheres, cysticercus, and tapeworm. GK-1 also contains at least one T-cell epitope, capable of stimulating the proliferation of CD8(+) and to a lower extent CD4(+) T cells primed either with the free peptide or T. crassiceps total antigen. The supernatant of the stimulated cells contained high levels of gamma interferon and low levels of interleukin-4. Similar results were obtained with T cells tested for intracellular cytokine production, an indication of the peptide's capacity to induce an inflammatory response. The remarkable protection induced by GK-1 immunization, its physicochemical properties, and its presence in all developmental stages of T. solium point to this synthetic peptide as a strong candidate in the construction of a synthetic vaccine against T. solium pig cysticercosis.     

Construction and characterization of affibody-Fc chimeras produced in Escherichia coli

Affibody-Fc chimeras were constructed by genetic fusion between different affibody affinity proteins with prescribed specificities and an Fc fragment derived from human IgG. Using affibody ligands previously selected for binding to respiratory syncytial virus (RSV) surface protein G and Thermus aquaticus (Taq) DNA polymerase, respectively, affibody-Fc fusion proteins showing spontaneous Fc fragment-mediated homodimerization via disulfide bridges were produced in Escherichia coli and affinity purified on protein A Sepharose from bacterial periplasms at yields ranging between 1 and 6 mg/l culture. Further characterization of the chimeras using biosensor technology showed that the affibody moieties have retained high selectivities for their respective targets after fusion to the Fc fragment. Avidity effects in the target binding were observed for the affibody-Fc chimeras compared to monovalent affibody fusion proteins, indicating that both affibody moieties in the chimeras were accessible and contributed in the binding. Fusion of a head-to-tail dimeric affibody moiety to the Fc fragment resulted in tetravalent affibody constructs which showed even more pronounced avidity effects. In addition, the Fc moiety of the chimeras was demonstrated to be specifically recognized by anti-human IgG antibody enzyme conjugates. One application for this class of "artificial antibodies" was demonstrated in a western blotting experiment in which one of the anti-RSV surface protein G affibody-Fc chimeras was demonstrated to be useful for specific detection of the target protein in a complex background consisting of a total E. coli lysate. The results show that through the replacement of the Fab portion of an antibody for an alternative binding domain based on a less complicated structure, chimeric proteins compatible with bacterial production routes containing both antigen recognition domains and Fc domains can be constructed. Such "artificial antibodies" should be interesting alternatives to, for example, whole antibodies or scFv-Fc fusions as detection devices and in diagnostic or therapeutic applications.     

Significance of premature contractions of WPW type in the acute phase of myocardial infarction

Observations in 220 patients with myocardial infarction of an intensive care unit are reported. The distinctive features of premature ventricular contractions of WPW type are described. Production of premature contractions of this type in the acute stage of myocardial infarction is regarded as a premonitory sign of rhythm disorders of considerable significance. The electrophysiological and electropathological background of rhythm disorders is analyzed in close detail, particular consideration is being given to the implications of premature contractions of WPW type. Increasing electric inhomogeneity in the acute phase of myocardial infarction is illustrated by intracavitary electrocardiograms. A relationship has been found between the protracted repolarization of the hypoxic ventricular areas and the appearance of T-U complexes. Two distinct groups of premature ventricular contractions of WPW type with reference to the conductive system are described. The origin of the delta waves in the WPW syndrome is discussed.     

DC-SIGN mediates the binding of Aspergillus fumigatus and keratinophylic fungi by human dendritic cells

Human contact with fungi does not usually lead to pathological consequences, as the immune system manages to defeat the invader pathogens. Nevertheless, under immune suppression, fungi overcome immune defenses and cause diseases that range from nonserious colonizations of keratinizated tissue (Dermatophytosis) to life threatening disseminated infections (Aspergillosis). Host defenses against fungi rely on innate and adaptative responses, with dendritic cell (DC) and macrophage surface receptors having a major role in the recognition of fungal pathogens and in the orchestration of an effective immune response. DC-SIGN is a C-type lectin involved in the recognition of bacterial, viral and parasitic pathogens, as well as in interactions between cells of the immune system. Its expression is restricted to DCs and subsets of macrophages. Here we show that DC-SIGN mediates the binding and capture of Aspergillus fumigatus and keratinophylic fungi, including Chrysosporium tropicum, by human DCs, describe the requirements of these interactions and discuss their potential involvement in the onset and persistence of pulmonary fungal infections.     

A dengue-2 Envelope fragment inserted within the structure of the P64k meningococcal protein carrier enables a functional immune response against the virus in mice

A gene fragment encoding for the amino acids (aa) 286-426 from the dengue Envelope (E) protein was expressed in Escherichia coli as two forms of fusion proteins. In one case, the E fragment was fused to the first 45 aa of the P64k protein from Neisseria meningitidis (PD2) while, in the other, it was inserted within the lipoil-binding domain of the aforementioned bacterial protein (PD3). PD2 was obtained as insoluble form within the cytoplasm of the bacteria while PD3 was distributed equally as soluble and insoluble forms. The insoluble forms of each protein as well as the soluble fraction of PD3 were semipurified to test the antigenicity and the immunogenicity in mice. The forms containing the entire P64k protein exhibited the highest recognition with different polyclonal and monoclonal antibodies. Consequently, the neutralizing antibodies elicited by the recombinant proteins were higher in the case of PD3 forms than with PD2, independently of the solubility status. In addition, mice inoculated with the semipurified insoluble form of PD3 were partially protected against lethal challenge with dengue-2 virus, administered by intracerebral inoculation. The results suggested the folding and carrier capacity of the P64k protein over the E fragment, converting PD3 as an attractive vaccine candidate against dengue-2 virus.     

CD40 is required for the optimal induction of protective immunity to Mycobacterium avium

C57Bl/6 mice and mice deficient in the CD40 molecule were infected with three strains of Mycobacterium avium. Two of the M. avium strains proliferated more extensively in CD40-deficient (CD40-/-) mice than in control mice. The increased susceptibility to infection of CD40-/- mice was associated with the generation of poorer interleukin-12 (IL-12) p40 and interferon-gamma (IFN-gamma) responses as compared to the controls, suggesting a role for CD40 in the development of protective immunity. In contrast, direct triggering of CD40 on infected macrophages failed to induce any anti-mycobacterial activity in infected macrophages.     

Retroposed new genes out of the X in Drosophila

New genes that originated by various molecular mechanisms are an essential component in understanding the evolution of genetic systems. We investigated the pattern of origin of the genes created by retroposition in Drosophila. We surveyed the whole Drosophila melanogaster genome for such new retrogenes and experimentally analyzed their functionality and evolutionary process. These retrogenes, functional as revealed by the analysis of expression, substitution, and population genetics, show a surprisingly asymmetric pattern in their origin. There is a significant excess of retrogenes that originate from the X chromosome and retropose to autosomes; new genes retroposed from autosomes are scarce. Further, we found that most of these X-derived autosomal retrogenes had evolved a testis expression pattern. These observations may be explained by natural selection favoring those new retrogenes that moved to autosomes and avoided the spermatogenesis X inactivation, and suggest the important role of genome position for the origin of new genes.     

The ancillary proteins of HATs: SLC3 family of amino acid transporters

The heteromeric amino acid transporters (HATs) are composed of a light and a heavy subunit linked by a disulfide bridge. The heavy subunits are the SLC3 members (rBAT and 4F2hc), whereas the light subunits are members of the SLC7 family of amino acid transporters. SLC3 proteins are type II membrane glycoproteins (i.e., one single transmembrane domain and the C-terminus located outside the cell) with a bulky extracellular domain that shows homology with alpha-glucosidases. rBAT heterodimerizes with b(0,+)AT (SLC7A9) constituting the amino acid transport b(0,+), the main system responsible for the apical reabsorption of cystine in kidney. The defect in this system causes cystinuria, the most common primary inherited aminoaciduria. 4F2hc subserves various amino acid transport systems by dimerization with different SLC7 proteins. The main role of SLC3 proteins is to help routing of the holotransporter to the plasma membrane. A working model for the biogenesis of HATs based on recent data on the rBAT/b(0,+)AT heterodimeric complex is presented. 4F2hc is a multifunctional protein, and in addition to its role in amino acid transport, it may be involved in other cellular functions. Studies on two SLC7 members (Asc-2 and AGT1) demonstrate heterodimerization with unknown heavy subunits.