Mapping of B-Cell Epitopes in a Trypanosoma cruzi Immunodominant Antigen Expressed in Natural Infections

Tc40 is an immunodominant antigen present in natural Trypanosoma cruzi infections. This immunogen was thoroughly mapped by using overlapping amino acid sequences identified by gene cloning and chemical peptide synthesis. To map continuous epitopes of the Tc40 antigen, an epitope expression library was constructed and screened with sera from human chagasic patients. A major, linear B-cell epitope spanning residues 403 to 426 (PAKAAAPPAA) was identified in the central domain of Tc40. A synthetic peptide spanning this region reacted strongly with 89.8% of the serum samples from T. cruzi-infected individuals. This indicates that the main antigenic site is defined by the linear sequence of the peptide rather than a conformation-dependent structure. The major B-cell epitope of Tc40 shares a high degree of sequence identity with T. cruzi ribosomal and RNA binding proteins, suggesting the existence of cross-reactivity among these molecules.     

Immune regulatory mechanisms influence early pathology in spinal cord injury and in spontaneous autoimmune encephalomyelitis

Injuries to the central nervous system (CNS) trigger an inflammatory reaction with potentially devastating consequences. In this report we compared the characteristics of the inflammatory response on spinal cord injury (SCI) caused by a stab wound between the T7 and T9 vertebrae and spontaneous experimental autoimmune encephalomyelitis (EAE). SCI and EAE were compared in two types of myelin basic protein Ac1-11-specific T-cell receptor transgenic mice: T/R+ mice harbor regulatory T cells, and T/R- mice lack regulatory T cells. Our results show that 8 days after SCI, T/R- mice developed a strong T-cell infiltrate in the spinal cord, with remarkable down-modulation of CD4 expression that was accompanied by a local increase in Mac-3+ and F4/80+ reactivity and diffuse local and distal astrogliosis. In contrast, T/R+ mice exhibited a modest increase in CD4+ cells localized to the site of injury, without CD4 down-modulation; focal astrogliosis was restricted to the site of the lesion, although Mac-3+ and F4/80+ cells were also present. Similarly to T/R- mice that underwent SCI, T cells displaying down-modulated CD4 expression were found in the CNS of older T/R- mice afflicted by spontaneous EAE. Overall, our results suggest that common mechanisms regulate T-cell accumulation in CNS lesions of different causes, such as mechanic lesion or autoimmune-mediated damage.     

Uptake and presentation of hepatitis C virus-like particles by human dendritic cells

Hepatitis C virus (HCV) is a major cause of chronic hepatitis worldwide. Interaction of dendritic cells (DCs) with viral particles may play an important role in the immunopathogenesis of HCV infection. Since the synthesis or purification of infectious virions is limited, we used HCV-like particles (HCV-LPs) to study the interaction of HCV with human DCs. Immature DCs exhibited an envelope-specific and saturable binding of HCV-LPs, indicating receptor-mediated DC-HCV-LP interaction. Confocal microscopy revealed that HCV-LPs were rapidly taken up by DCs in a temperature-dependent manner. Competition experiments demonstrated that C-type lectins such as mannose receptor or DC-SIGN (DC-specific intercellular adhesion molecule 3-grabbing nonintegrin) were not sufficient for mediating HCV-LP binding. HCV-LP uptake was followed by DC activation. DCs pulsed with HCV-LPs stimulated HCV core-specific CD4(+) T cells, indicating that uptake of HCV-LPs by DCs leads to antigen processing and presentation on major histocompatibility complex (MHC) class II molecules. Finally, HCV-LP-derived antigens were efficiently cross-presented to HCV core-specific CD8(+) T cells. These findings demonstrate that HCV-LPs represent a novel model system to study HCV-DC interaction allowing definition of the molecular mechanisms of HCV uptake, DC activation, and antigen presentation to T cells. Furthermore, HCV-LP-mediated DC activation and efficient antigen presentation may explain the marked immunogenicity of HCV-LPs in vivo.     

Effect of photoactivation systems and resin composites on the microleakage of esthetic restorations

AIM: The aim of this study was to evaluate the influence of four photoactivation systems [quartz tungsten halogen (QTH), light-emitting diode (LED), argon ion laser (AL), and plasma arc curing PAC)] on cementum/dentin and enamel microleakage of Class II restorations using a microhybrid [Z250-3M ESPE] and two packable composites [(SureFil-Dentsply and Tetric Ceram HB-Ivoclair/Vivadent]. METHODS AND MATERIALS: Three hundred sixty "vertical-slot Class II cavities" were prepared at the mesial surface of bovine incisors using a 245 carbide bur in a highspeed handpiece. Specimens were divided into twelve groups (composite-photoactivation systems). Half of the specimens had the gingival margin placed in enamel (n=15) and the other half in cementum/dentin (n=15). Composites were inserted and cured in 2 mm increments according to manufacturers' recommended exposure times. After polishing, the samples were immersed in 2% methylene blue solution, sectioned, and evaluated at the gingival margins. Data were submitted to statistical analysis using the Kruskal-Wallis and Mann-Whitney tests. RESULTS: No significant differences were found among the photoactivation systems and among resin composites (p>0.05). Microleakage was not significantly affected by location (enamel vs. cementum/dentin, p>0.05). These findings suggested neither the photoactivation systems nor the resin composite types might have an effect on the microleakage at gingival margins Class II cavities.     

Bisphenol A Exposure Impairs Epididymal Development during the Peripubertal Period of Rats: Inflammatory Profile and Tissue Changes

Bisphenol A (BPA) is a synthetic non‐steroidal oestrogen used in the production of plastics. BPA can cause alterations in the endocrine system of human beings and animals at varied stages of development. During puberty, altered morphological, sexual behaviour and completion of the epididymal development occur. Therefore, this study aimed to evaluate the effects of BPA on epididymal development during the peripubertal period of rats. Male Wistar rats were treated with BPA via gavage at doses of 20 μg/kg or 200 μg/kg per day [post‐natal day (PND] 36–66). The control group received the vehicles under the same conditions. Feed and water were provided ad libitum. On PND 67, the epididymis was removed, weighed, divided into caput/corpus and cauda sections. It was then used for sperm count determination; histopathological and stereological evaluation; inflammatory cell enzymatic profiling (myeloperoxidase activity – MPO; N‐acetylglucosaminidase – NAG); immunohistochemistry for IL‐6; and evaluation of superoxide anion levels and malondialdehyde (MDA). Exposure to BPA at 200 μg/kg caused a significant increase of MPO activity and immunoreactivity to IL‐6 (interleukin‐6) as well as remodelling of tissue components in the caput/corpus and cauda regions of the epididymis. Under these experimental conditions, it is concluded that BPA alters post‐natal epididymal development.     

Angiotensin II-dependent chronic hypertension and cardiac hypertrophy are unaffected by gp91phox-containing NADPH oxidase

The gp91phox-containing NADPH oxidase is the major source of reactive oxygen species (ROS) in the cardiovascular system and inactivation of gp91phox has been reported to blunt hypertension and cardiac hypertrophy seen in angiotensin (Ang) II-infused animals. In the current study, we sought to determine the role of gp91phox-derived ROS on cardiovascular outcomes of chronic exposure to Ang II. The gp91phox-deficient mice were crossed with transgenic mice expressing active human renin in the liver (TTRhRen). TTRhRen mice exhibit chronic Ang II-dependent hypertension and frank cardiac hypertrophy by age 10 to 12 weeks. Four genotypes of mice were generated: control, TTRhRen trangenics (TTRhRen), gp91phox-deficient (gp91-), and TTRhRen transgenic gp91phox-deficient (TTRhRen/gp91-). Eight to 10 mice/group were studied. ROS levels were significantly reduced (P<0.05) in the heart and aorta of TTRhRen/gp91- and gp91-mice compared with control counterparts, and this was associated with reduced cardiac, aortic, and renal NADPH oxidase activity (P<0.05). Systolic blood pressure (SBP), cardiac mass, and cardiac fibrosis were increased in TTRhRen versus controls. In contrast to its action on ROS generation, gp91phox inactivation had no effect on development of hypertension or cardiac hypertrophy in TTRhRen mice, although interstitial fibrosis was reduced. Cardiac and renal expression of gp91phox homologues, Nox1 and Nox4, was not different between groups. Thus, although eliminating gp91phox-associated ROS production may be important in cardiovascular consequences in acute insult models, it does not prevent the development of hypertension and cardiac hypertrophy in a model in which the endogenous renin-angiotensin system is chronically upregulated.