A Vaccine Encoding Conserved Promiscuous HIV CD4 Epitopes Induces Broad T Cell Responses in Mice Transgenic to Multiple Common HLA Class II Molecules

Current HIV vaccine approaches are focused on immunogens encoding whole HIV antigenic proteins that mainly elicit cytotoxic CD8+ responses. Mounting evidence points toward a critical role for CD4+ T cells in the control of immunodeficiency virus replication, probably due to cognate help. Vaccine-induced CD4+ T cell responses might, therefore, have a protective effect in HIV replication. In addition, successful vaccines may have to elicit responses to multiple epitopes in a high proportion of vaccinees, to match the highly variable circulating strains of HIV. Using rational vaccine design, we developed a DNA vaccine encoding 18 algorithm-selected conserved, ""promiscuous"" ( multiple HLA-DR-binding) B-subtype HIV CD4 epitopes - previously found to be frequently recognized by HIV-infected patients. We assessed the ability of the vaccine to induce broad T cell responses in the context of multiple HLA class II molecules using different strains of HLA class II-transgenic mice (-DR2, -DR4, -DQ6 and -DQ8). Mice displayed CD4+ and CD8+ T cell responses of significant breadth and magnitude, and 16 out of the 18 encoded epitopes were recognized. By virtue of inducing broad responses against conserved CD4+ T cell epitopes that can be recognized in the context of widely diverse, common HLA class II alleles, this vaccine concept may cope both with HIV genetic variability and increased population coverage. The vaccine may thus be a source of cognate help for HIV-specific CD8+ T cells elicited by conventional immunogens, in a wide proportion of vaccinees.

Mast cell repopulation of the peritoneal cavity: contribution of mast cell progenitors versus bone marrow derived committed mast cell precursors

Background: Mast cells have recently gained new importance as immunoregulatory cells that are involved in numerous pathological processes. One result of these processes is an increase in mast cell numbers at peripheral sites. This study was undertaken to determine the mast cell response in the peritoneal cavity and bone marrow during repopulation of the peritoneal cavity in rats. Results: Two mast cell specific antibodies, mAb AA4 and mAb BGD6, were used to distinguish the committed mast cell precursor from more mature mast cells. The peritoneal cavity was depleted of mast cells using distilled water. Twelve hours after distilled water injection, very immature mast cells could be isolated from the blood and by 48 hours were present in the peritoneal cavity. At this same time the percentage of mast cells in mitosis increased fourfold. Mast cell depletion of the peritoneal cavity also reduced the total number of mast cells in the bone marrow, but increased the number of mast cell committed precursors. Conclusions: In response to mast cell depletion of the peritoneal cavity, a mast cell progenitor is released into the circulation and participates in repopulation of the peritoneal cavity, while the committed mast cell precursor is retained in the bone marrow.

The Lectin ArtinM Induces Recruitment of Rat Mast Cells from the Bone Marrow to the Peritoneal Cavity

Background: The D-mannose binding lectin ArtinM is known to recruit neutrophils, to degranulate mast cells and may have potential therapeutic applications. However, the effect of ArtinM on mast cell recruitment has not been investigated. Methodology: Male Wistar rats were injected i.p. with ArtinM or ConA (control). The ability of the lectin to degranulate peritoneal and mesenteric mast cells was examined. Recruitment of mast cells to the peritoneal cavity and mesentery after ArtinM injection was examined with or without depletion of peritoneal mast cells by distilled water. Results: ArtinM degranulated both peritoneal and mesentery mast cells in vitro. Three days after i.p. injection of the lectin there were reduced numbers of mast cells in the peritoneal lavage, while at 7 days post injection of ArtinM, the number of peritoneal mast cells was close to control values. Since immature mast cells are recruited from the bone marrow, the effect of the lectin on bone marrow mast cells was examined. Injection of ArtinM resulted in an increased number of mast cells in the bone marrow. To determine if degranulation of mast cells in the peritoneal cavity was required for the increase in bone marrow mast cells, the peritoneal cavity was depleted of mast cells with ultrapure water. Exposure to ArtinM increased the number of mast cells in the bone marrow of rats depleted of peritoneal mast cells. Conclusions: The ArtinM induced recruitment of mast cells from the bone marrow to the peritoneal cavity may partially explain the therapeutic actions of ArtinM.

Epigenetic Silencing of CRABP2 and MX1 in Head and Neck Tumors

Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous disease affecting the epithelium of the oral cavity, pharynx and larynx. Conditions of most patients are diagnosed at late stages of the disease, and no sensitive and specific predictors of aggressive behavior have been identified yet. Therefore, early detection and prognostic biomarkers are highly desirable for a more rational management of the disease. Hypermethylation of CpG islands is one of the most important epigenetic mechanisms that leads to gene silencing in tumors and has been extensively used for the identification of biomarkers. In this study, we combined rapid subtractive hybridization and microarray analysis in a hierarchical manner to select genes that are putatively reactivated by the demethylating agent 5-aza-2'-deoxycytidine (5Aza-dC) in HNSCC cell lines (FaDu, UM-SCC-14A, UM-SCC-17A, UM-SCC-38A). This combined analysis identified 78 genes, 35 of which were reactivated in at least 2 cell lines and harbored a CpG island at their 5' region. Reactivation of 3 of these 35 genes (CRABP2, MX1, and SLC15A3) was confirmed by quantitative real-time polymerase chain reaction (PCR; fold change, >= 3). Bisulfite sequencing of their CpG islands revealed that they are indeed differentially methylated in the HNSCC cell lines. Using methylation-specific PCR, we detected a higher frequency of CRABP2 (58.1% for region 1) and MX1 (46.3%) hypermethylation in primary HNSCC when compared with lymphocytes from healthy individuals. Finally, absence of the CRABP2 protein was associated with decreased disease-free survival rates, supporting a potential use of CRABP2 expression as a prognostic biomarker for HNSCC patients.

Photodynamic therapy for root canals infected with Enterococcus faecalis

Objective: The aim of this study was to investigate the effects of photodynamic therapy (PDT) on endodontic pathogens by evaluating the decrease in numbers of Enterococcus faecalis colonies in the canals of extracted human teeth. Background Data: Failure in endodontics is usually related to inadequate cleaning and disinfection of the root canal system. This is due to the establishment of microorganisms in areas where the instruments and chemical agents used during root canal preparation cannot eliminate them. PDT is a complementary therapeutic method that could be used to eliminate these remaining bacteria. PDT is a process in which radiation acts on a dye that is applied to the target organism, resulting in bacterial death. Materials and Methods: Forty-six uniradicular teeth had their canals contaminated with bacteria and were incubated for 48 h at 35 degrees C. After that, the teeth were divided into a control group (CG) and a test group (TG). The 23 CG teeth did not undergo any intervention, whereas in the TG the teeth received a solution of 0.0125% toluidine blue for 5 min followed by irradiation using a 50-mW diode laser (Ga-Al-As) at a wavelength of 660 nm. Bacterial samples were taken before and after irradiation. In each of the samples, the number of colony-forming units (CFU) was counted. Results: The mean decrease in CFU was 99.9% in the TG, whereas in the CG an increase of 2.6% was observed. Conclusion: PDT was effective as a bactericidal agent in Enterococcus faecalis-contaminated root canals.

In utero protein restriction causes growth delay and alters sperm parameters in adult male rats

Background: Recent studies have supported the concept of ""fetal programming"" which suggests that during the intrauterine development the fetus may be programmed to develop diseases in adulthood. The possible effects of in utero protein restriction on sexual development of rat male offspring were evaluated in the present study. Methods: Pregnant Wistar rats were divided into two experimental groups: one group treated with standard chow (SC, n = 8, 17% protein) and the other group treated with hypoproteic chow (HC, n = 10, 6% protein) throughout gestation. After gestation the two experimental groups received standard chow. To evaluate the possible late reproductive effects of in utero protein restriction, the male offspring of both groups were assessed at different phases of sexual development: prepubertal (30 days old); peripubertal (60 days old); adult (90 days old). Student's t test and Mann-Whitney test were utilized. Differences were considered significant when p < 0.05. Results: We found that in utero protein restriction reduced the body weight of male pups on the first postnatal day and during the different sexual development phases (prepubertal, peripubertal and adult). During adulthood, Sertoli cell number, sperm motility and sperm counts in the testis and epididymal cauda were also reduced in HC. Furthermore, the numbers of sperm presenting morphological abnormalities and cytoplasmic drop retention were higher in HC. Conclusions: In conclusion, in utero protein restriction, under these experimental conditions, causes growth delay and alters male reproductive-system programming in rats, suggesting impairment of sperm quality in adulthood.

Placentation in Sigmodontinae: a rodent taxon native to South America

Background: Sigmodontinae, known as ""New World rats and mice,"" is a large subfamily of Cricetidae for which we herein provide the first comprehensive investigation of the placenta. Methods: Placentas of various gestational ages ranging from early pregnancy to near term were obtained for five genera, i.e. Necromys, Euryoryzomys, Cerradomys, Hylaeamys, and Oligoryzomys. They were investigated by means of histology, immunohistochemistry, a proliferation marker, DBA-lectin staining and transmission electron microscopy. Results: The chorioallantoic placenta was organized in a labyrinthine zone, spongy zone and decidua and an inverted yolk sac persisted until term. The chorioallantoic placenta was hemotrichorial. The interhemal barrier comprised fetal capillary endothelium and three layers of trophoblast, an outermost, cellular layer and two syncytial ones, with interspersed trophoblast giant cells (TGC). In addition, accumulations of TGC occurred below Reichert's membrane. The junctional zone contained syncytial trophoblast, proliferative cellular trophoblast, glycogen cells and TGC that were situated near to the maternal blood channels. In three of the genera, TGC were also accumulated in distinct areas at the placental periphery. PAS-positive glycogen cells derived from the junctional zone invaded the decidua. Abundant maternal uNK cells with positive response to PAS, vimentin and DBA-lectin were found in the decidua. The visceral yolk sac was completely inverted and villous. Conclusion: The general aspect of the fetal membranes in Sigmodontinae resembled that found in other cricetid rodents. Compared to murid rodents there were larger numbers of giant cells and in some genera these were seen to congregate at the periphery of the placental disk. Glycogen cells were found to invade the decidua but we did not identify trophoblast in the walls of the deeper decidual arteries. In contrast these vessels were surrounded by large numbers of uNK cells. This survey of wild-trapped specimens from five genera is a useful starting point for the study of placentation in an important subfamily of South American rodents. We note, however, that some of these rodents can be captive bred and recommend that future studies focus on the study of time dated pregnancies.

A SEARCH FOR UNRECOGNIZED CARBON-ENHANCED METAL-POOR STARS IN THE GALAXY

We have developed a new procedure to search for carbon-enhanced metal-poor (CEMP) stars from the Hamburg/ESO (HES) prism-survey plates. This method employs an extended line index for the CH G band, which we demonstrate to have superior performance when compared to the narrower G-band index formerly employed to estimate G-band strengths for these spectra. Although CEMP stars have been found previously among candidate metal-poor stars selected from the HES, the selection on metallicity undersamples the population of intermediate-metallicity CEMP stars (-2.5 <= [Fe/H] <= -1.0); such stars are of importance for constraining the onset of the s-process in metal-deficient asymptotic giant branch stars (thought to be associated with the origin of carbon for roughly 80% of CEMP stars). The new candidates also include substantial numbers of warmer carbon-enhanced stars, which were missed in previous HES searches for carbon stars due to selection criteria that emphasized cooler stars. A first subsample, biased toward brighter stars (B < 15.5), has been extracted from the scanned HES plates. After visual inspection (to eliminate spectra compromised by plate defects, overlapping spectra, etc., and to carry out rough spectral classifications), a list of 669 previously unidentified candidate CEMP stars was compiled. Follow-up spectroscopy for a pilot sample of 132 candidates was obtained with the Goodman spectrograph on the SOAR 4.1 m telescope. Our results show that most of the observed stars lie in the targeted metallicity range, and possess prominent carbon absorption features at 4300 angstrom. The success rate for the identification of new CEMP stars is 43% (13 out of 30) for [Fe/H] < -2.0. For stars with [Fe/H] < -2.5, the ratio increases to 80% (four out of five objects), including one star with [Fe/H] < -3.0.

Fatores gravimétricos no Brasil para um modelo anelástico de Terra

Geodetic observations are affected by the disturbing potential of the luni-solar tide. Among those observations, the value of g obtained from gravimetric survey needs correction by the gravimetric factor. This correction is derived from the Numbers of Love, which depend on the adopted model of Earth. Because of this, it is necessary to update the correction since the gravimetric factor widely used in Brazil as delta = 1.20 does not consider local rheological variations and they are latitude dependent. A discrepancy of about 1% between the observed tidal gravimetric factors d of the ""Trans World Tidal Gravity Profiles"" (TWTGP), related to Brussels fundamental station, and those obtained by recent observations reported by Freitas and Ducarme ( 1991). Experiments based on inertial force effects also reveal a variation of about 0.5% in the observed d. A same order of magnitude difference is obtained for an anelastic Earth model when compared with a viscous-elastic model and even when different frequencies of tidal perturbations are considered. In this paper regression models are presented for gravimetric factors for the lunar components O(1) and M(2) in Brazil. These models were obtained from observations performed at stations belonging to the Brazilian segment of the TWTGP.

Pollen foraging in colonies of Melipona bicolor (Apidae, Meliponini): effects of season, colony size and queen number

We evaluated the ratio between the number of pollen foragers and the total number of bees entering colonies of Melipona bicolor, a facultative polygynous species of stingless bees. The variables considered in our analysis were: seasonality, colony size and the number of physogastric queens in each colony. The pollen forager ratios varied significantly between seasons; the ratio was higher in winter than in summer. However, colony size and number of queens per colony had no significant effect. We conclude that seasonal differences in pollen harvest are related to the production of sexuals and to the number of individuals and their body size.

Unsuccessful attacks dominate a drone-preying wasp's hunting performance near stingless bee nests

Bee males (drones) of stingless bees tend to congregate near entrances of conspecific nests, where they wait for virgin queens that initiate their nuptial flight. We observed that the Neotropical solitary wasp Trachypus boharti (Hymenoptera, Cabronidae) specifically preys on males of the stingless bee Scaptotrigona postica (Hymenoptera, Apidae); these wasps captured up to 50 males per day near the entrance of a single hive. Over 90% of the wasp attacks were unsuccessful; such erroneous attacks often involved conspecific wasps and worker bees. After the capture of non-male prey, wasps almost immediately released these individuals unharmed and continued hunting. A simple behavioral experiment showed that at short distances wasps were not specifically attracted to S. postica males nor were they repelled by workers of the same species. Likely, short-range prey detection near the bees' nest is achieved mainly by vision whereas close-range prey recognition is based principally on chemical and/or mechanical cues. We argue that the dependence on the wasp's visual perception during attack and the crowded and dynamic hunting conditions caused wasps to make many preying attempts that failed. Two wasp-density-related factors, wasp-prey distance and wasp-wasp encounters, may account for the fact that the highest male capture and unsuccessful wasp bee encounter rates occurred at intermediate wasp numbers.

Effects of protein deprivation and re-feeding on P2X(2) receptors in enteric neurons

AIM: To investigate the effects of malnutrition and refeeding on the P2X(2) receptor, nitric oxide synthase (NOS), calretinin, calbindin and choline acetyltransferase (ChAT) in neurons of the rat ileum. METHODS: We analyzed the co-localization, numbers and sizes of P2X(2)-expressing neurons in relation to NOS-IR (immunoreactive), calbindin-IR, ChAT-IR, and calretinin-IR neurons of the myenteric and submucosal plexus. The experimental groups consisted of: (1) rats maintained on normal feed throughout pregnancy until 42 d post-parturition (N); (2) rats deprived of protein throughout pregnancy and 42 d post-parturition (D); and (3) rats undernourished for 21 d post-parturition and then given a protein diet from days 22 to 42 (DR). The myenteric and submucosal plexuses were evaluated by double labeling by immunohistochemical methods for P2X(2) receptor, NOS, ChAT, calbindin and calretinin. RESULTS: We found similar P2X(2) receptor immunoreactivity in the cytoplasm and surface membranes of myenteric and submucosal neurons from the N, D and DR groups. Double labeling of the myenteric plexus demonstrated that approximately 100% of NOS-IR, calbindin-IR, calretinin-IR and ChAT-IR neurons in all groups also expressed the P2X(2) receptor. In the submucosal plexus, the calretinin-IR, ChAT-IR and calbindinIR neurons were nearly all immunoreactive for the P2X(2) receptor. In the myenteric plexus, there was a 19% increase in numbers per cm(2) for P2X(2) receptor-IR neurons, 64% for NOS-IR, 84% for calretinin-IR and 26% for ChAT-IR neurons in the D group. The spatial density of calbindin-IR neurons, however, did not differ among the three groups. The submucosal neuronal density increased for calbindin-IR, calretinin-IR and ChAT-IR neurons. The average size of neurons in the myenteric plexus neurons in the D group was less than that in the controls and, in the re-fed rats; there was a 34% reduction in size only for the calretinin-IR neurons. CONCLUSION: This work demonstrates that expression of the P2X(2) receptor is present in inhibitory, intrinsic primary afferent, cholinergic secretomotor and vasomotor neurons. Undernutrition affected P2X(2) receptor expression in the submucosal plexus, and neuronal and size. These changes were rescued in the re-fed rats. (C) 2010 Baishideng. All rights reserved.