Analysis of the TCR-mediated signaling dynamics

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2014

Estudos sobre a nutrição mineral do arroz: XIV. efeitos dos excessos de Al, Cl e Mn nas variedades IAC-25 e IAC-47

Plantas de arroz, variedades IAC-25 e IAC-47 foram cultivadas em solução nutritiva completa e com excesso de Al, Cl e Mn (25, 1750 e 25 ppm, respectivamente). As últimas não completaram o ciclo. Foram observados sintomas típicos de toxidez. A produção de matéria seca foi mais afetada pelos tratamentos na variedade IAC-47. Ambas as variedades mostraram-se mais sensíveis a toxidez de Mn que à de Al. Foram determinados os teores foliares de Al, Cl e Mn associados à toxidez correspondente.

Alterações fisiológicas do sistema nervoso central de rã pelas toxinas de Cl. perfringens, Cl. oedematiens e Cl. septicum

The authors carried on experiences in order to confirm the neurotoxic theory of gas gangrene explained by Pacheco & Costa, uning preparations of isolated cord-posterior train of Leptodactylus ocellatus as described by OZORIO DE ALMEIDA & Cols. Frogs were intoxicated 3 days before the test with parcially purified toxins of Cl. perfringens, Cl. oedematiens and Cl. septicum. The intoxication produced a shortening of spinal reflexes duration time of such preparations, showing a typical alteration of the reflex activity of the spinal cord.

Trypanosoma cruzi: antigen-receptor mediated endocytosis of antibody

Trypanomastigote forms of Trypanosoma cruzi were derived from tissue culture and incubated with immune and non-immune human sera. All immune sera showed high titers of specific humoral antibodies of the IgM or the IgG type. Agglutination and swelling of parasites were observed after incubation at 37ºC, but many trypomastigotes remained free-swimming in the sera for two to three days. The quantitiy of immune serum capable of lysing a maximum of 10 x 10 [raised to the power of 6] sensitized red cells was not capable of lysing 4 x 10 [raised to the power of 3] tripomastigotes. Typically, the parasites underwent cyclical changes with the formation of clumps of amastigotes and the appearance of epimastigote forms. Multiplication of the parasites was observed in immune sera. Further, the infectivity of the parasites to susceptible mice was not lost. All sera used produced similar general effects on the growth of the parasite. The antibody bound to T. cruzi appeard to enter cells by antigen-receptor mediated endocytosis. The ferritin-conjugated antibody was internalized and delivered to phagolysosomes where they might be completely degraded to amino-acids. This seemed to be a coupled process by which the immunoglobulin is first bound to specific parasite surface receptor and then rapidly endocytosed by the cell.

CL 64, 855, a potent do anti-Trypanosoma cruzi drug, is also mutagenic in the salmonella/microsome assay

The nitroimidazole-tiadiazole derivative CL 64,855 (2-amino-5-(1-methyl-5-nitro-2-imidazolyl)-1,3,4-thiadiazole, a potent anti-trypanosomal drug, was assayed in a short-term bacterial mutagenicity test with Salmonella typhimurium strains TA 98, TA 100 and TA 102. Results indicate that CL 64,855 is a potent frameshift mutagen detected by strains TA 98 and TA 102. CL 64,855 was able to revert the indicators strains at concentrations as low as 0.1 µg/plate. Metabolic activation experiments with rat liver microsomal fractions did not increase the mutagenic action of Cl 64,855.

Insulin secretion is regulated by the glucose-dependent production of islet beta cell macrophage migration inhibitory factor.

Macrophage migration inhibitory factor (MIF), originally identified as a cytokine secreted by T lymphocytes, was found recently to be both a pituitary hormone and a mediator released by immune cells in response to glucocorticoid stimulation. We report here that the insulin-secreting beta cell of the islets of Langerhans expresses MIF and that its production is regulated by glucose in a time- and concentration-dependent manner. MIF and insulin colocalize by immunocytochemistry within the secretory granules of the pancreatic islet beta cells, and once released, MIF appears to regulate insulin release in an autocrine fashion. In perifusion studies performed with isolated rat islets, immunoneutralization of MIF reduced the first and second phase of the glucose-induced insulin secretion response by 39% and 31%, respectively. Conversely, exogenously added recombinant MIF was found to potentiate insulin release. Constitutive expression of MIF antisense RNA in the insulin-secreting INS-1 cell line inhibited MIF protein synthesis and decreased significantly glucose-induced insulin release. MIF is therefore a glucose-dependent, islet cell product that regulates insulin secretion in a positive manner and may play an important role in carbohydrate metabolism.

Flippase (FLP) recombinase-mediated marker recycling in the dermatophyte Arthroderma vanbreuseghemii.

Biological processes can be elucidated by investigating complex networks of relevant factors and genes. However, this is not possible in species for which dominant selectable markers for genetic studies are unavailable. To overcome the limitation in selectable markers for the dermatophyte Arthroderma vanbreuseghemii (anamorph: Trichophyton mentagrophytes), we adapted the flippase (FLP) recombinase-recombination target (FRT) site-specific recombination system from the yeast Saccharomyces cerevisiae as a selectable marker recycling system for this fungus. Taking into account practical applicability, we designed FLP/FRT modules carrying two FRT sequences as well as the flp gene adapted to the pathogenic yeast Candida albicans (caflp) or a synthetic codon-optimized flp (avflp) gene with neomycin resistance (nptII) cassette for one-step marker excision. Both flp genes were under control of the Trichophyton rubrum copper-repressible promoter (PCTR4). Molecular analyses of resultant transformants showed that only the avflp-harbouring module was functional in A. vanbreuseghemii. Applying this system, we successfully produced the Ku80 recessive mutant strain devoid of any selectable markers. This strain was subsequently used as the recipient for sequential multiple disruptions of secreted metalloprotease (fungalysin) (MEP) or serine protease (SUB) genes, producing mutant strains with double MEP or triple SUB gene deletions. These results confirmed the feasibility of this system for broad-scale genetic manipulation of dermatophytes, advancing our understanding of functions and networks of individual genes in these fungi.

Cell-matrix interactions in Schistosomal portal fibrosis: a dynamic event

In recent years, one of the most significant progress in the understanding of liver diseases was the demonstration that liver fibrosis is a dynamic process resulting from a balance between synthesis and degradation of several matrix components, collagen in particular. Thus, fibrosis has been found to be a very early event during liver diseases, be it of toxic, viral or parasitic origin, and to be spontaneously reversible, either partially or totally. In liver fibrosis cell matrix interactions are dependent on the existence of the many factors (sometimes acting in combination) which produce the same events at the cellular and molecular levels. These events are: (i) the recruitment of fiber-producing cells, (ii) their proliferation, (iii) the secretion of matrix constituents of the extracellular matrix, and (iv) the remodeling and degradation of the newly formed matrix. All these events represent, at least in principle, a target for a therapeutic intervention aimed at influencing the experimentally induced hepatic fibrosis. In this context, hepatosplenic schistosomiasis is of particular interest, being an immune cell-mediated granulomatous disease and a model of liver fibrosis allowing extensive studies in human and animals as well as providing original in vitro models.

MC1R-dependent, melanin-based colour polymorphism is associated with cell-mediated response in the Eleonora's falcon.

Colour polymorphism in vertebrates is usually under genetic control and may be associated with variation in physiological traits. The melanocortin 1 receptor (Mc1r) has been involved repeatedly in melanin-based pigmentation but it was thought to have few other physiological effects. However, recent pharmacological studies suggest that MC1R could regulate the aspects of immunity. We investigated whether variation at Mc1r underpins plumage colouration in the Eleonora's falcon. We also examined whether nestlings of the different morphs differed in their inflammatory response induced by phytohemagglutinin (PHA). Variation in colouration was due to a deletion of four amino acids at the Mc1r gene. Cellular immune response was morph specific. In males, but not in females, dark nestling mounted a lower PHA response than pale ones. Although correlative, our results raise the neglected possibility that MC1R has pleiotropic effects, suggesting a potential role of immune capacity and pathogen pressure on the maintenance of colour polymorphism in this species.

Immunoregulation in human Schistosomiasis by idiotypic interactions and lymphokine-mediated mechanisms

Anti-idiotypic (anti-Id) T cells from schistosomiasis patients or former patients proliferate upon exposure to polyclonal or monoclonal anti-soluble egg antigen (SEA) antibodies. Chloroquine does not inhibit, the response, which is induced by F(ab')2 (but not soluble Fab) fragments of these antibodies. Purified T cells from former patients require macrophages or exogenous IL-1 to respond to anti-SEA Ids and can respond to matrix-bound Fab fragments in the presence of IL-1. These anti-Id T cells recognize the Ids directly. Chronic schistosomiasis patients immunoregulate the production of a non-IL-2 lymphokine that stimulates IL-2 receptor expression on resting T cells. This regulation is reversed upon chemotherapeutic cure.

Prospects for a nonliving vaccine against Schistosomiasis based on cell-mediated immune resistance mechanisms

We have designed a vaccine model based on induction of cell-mediated immunity and shown that it protects mice against Schistosoma mansoni infection. Mice are immunized by intradermal injection with schistosome antigens plus BCG. Resistance is dependent on the route of antigen presentation and the adjuvant chosen. The pattern of resistance correlates with sensitization of T lymphocytes for production of gamma interferon, a macrophage activating lymphokine that stimulates the cellular effector mechanism of protection. Purified schistosome paramyosin, a muscle cell component present in soluble parasite antigenic preparations, is immunogenic for T lymphocytes and induces resistance when given intradermally with BCG. It is likely that this protein, and possibly other soluble molecules that are released by the parasites of a challenge infection, induce a cellular inflammatory response resulting in larval trapping and/or killing by activated macrophages. These results verify the feasibility of a vaccine against schistosomiasis based on induction of cell-mediated immune resistance mechanisms.

GLUT2 and the incretin receptors are involved in glucose-induced incretin secretion.

Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are incretins secreted in response to oral glucose ingestion by intestinal L and K cells, respectively. The molecular mechanisms responsible for intestinal cell glucose sensing are unknown but could be related to those described for beta-cells, brain and hepatoportal sensors. We determined the role of GLUT2, GLP-1 or GIP receptors in glucose-induced incretins secretion, in the corresponding knockout mice. GLP-1 secretion was reduced in all mutant mice, while GIP secretion did not require GLUT2. Intestinal GLP-1 content was reduced only in GIP and GLUT2 receptors knockout mice suggesting that this impairment could contribute to the phenotype. Intestinal GIP content was similar in all mice studied. Furthermore, the impaired incretins secretion was associated with a reduced glucose-stimulated insulin secretion and an impaired glucose tolerance in all mice. In conclusion, both incretins secretion depends on mechanisms involving their own receptors and GLP-1 further requires GLUT2.

In vivo and in vitro effects of somatostatin and insulin on glucagon release in a human glucagonoma.

Inhibition of pancreatic glucagon secretion has been reported to be mediated by glucose, insulin and somatostatin. As no human pancreatic alpha-cell lines are available to study in vitro the relative importance of insulin and glucose in the control of pancreatic glucagon release, we investigated a patient presenting with a malignant glucagonoma who underwent surgical resection of the tumour. Functional somatostatin receptors were present as octreotide administration decreased basal glucagon and insulin secretion by 52 and 74%, respectively. The removed tumour was immunohistochemically positive for glucagon, chromogranin A and pancreatic polypeptide but negative for insulin, gastrin and somatostatin. The glucagonoma cells were also isolated and cultured in vitro. Incubation experiments revealed that change from high (10 mM) to low (1 mM) glucose concentration was unable to stimulate glucagon secretion. A dose-dependent inhibition of glucagon release by insulin was however, observed at low glucose concentration. These findings demonstrate that insulin could inhibit glucagon secretion in vitro in the absence of elevated glucose concentrations. These data suggest, as observed in vivo and in vitro in several animal studies, that glucopenia-induced glucagon secretion in humans is not mediated by a direct effect of low glucose on alpha-cells but possibly by a reduction of insulin-mediated alpha-cell suppression and/or an indirect neuronal stimulation of glucagon release.

Nest distribution varies with dispersal method and familiarity-mediated aggression for two sympatric ants

Dispersal mechanisms and competition together play a key role in the spatial distribution of a population. Species that disperse via fission are likely to experience high levels of localized competitive pressure from conspecifics relative to species that disperse in other ways. Although fission dispersal occurs in many species, its ecological and behavioural effects remain unclear. We compared foraging effort, nest spatial distribution and aggression of two sympatric ant species that differ in reproductive dispersal: Streblognathus peetersi, which disperse by group fission, and Plectroctena mandibularis, which disperse by solitary wingless queens. We found that although both species share space and have similar foraging strategies, they differ in nest distribution and aggressive behaviour. The spatial distribution of S. peetersi nests was extremely aggregated, and workers were less aggressive towards conspecifics from nearby nests than towards distant conspecifics and all heterospecific workers. By contrast, the spatial distribution of P. mandibularis nests was overdispersed, and workers were equally aggressive towards conspecific and heterospecific competitors regardless of nest distance. Finally, laboratory experiments showed that familiarity led to the positive relationship between aggression and nest distance in S. peetersi. While unfamiliar individuals were initially aggressive, the level of aggression decreased within 1 h of contact, and continued to decrease over 24 h. Furthermore, individuals from near nests that were not aggressive could be induced to aggression after prolonged isolation. Overall, these results suggest that low aggression mediated by familiarity could provide benefits for a species with fission reproduction and an aggregated spatial distribution.