Influence of HLA-DRB-1 alleles on the production of antibody against CSP, MSP-1, AMA-1, and DBP in Brazilian individuals naturally infected with Plasmodium vivax

We evaluated the influence of allelic frequency of the human leukocyte antigen (HLA) -DRB1 on the acquisition of antibody response against malaria sporozoite and merozoite peptides in patients with Plasmodium vivax malaria acquired in endemic areas of Brazil. IgG antibodies were detected by enzyme-linked immunosorbent assay against four peptides of circumsporozoite protein (CSP) (amino, carboxyl, and VK210 and VK247 repeats) and peptides of merozoite surface protein 1 (MSP-1), apical membrane antigen 1 (AMA-1), and Duffy-binding protein (DBP). We found an association between HLA-DR3 and HLA-DR5 alleles and lack of antibody response to CSP amino terminal, as well as an association between HILA-DR3 and the highest antibody response to MSP1 (Pv200L). In conclusion, we suggest a potential regulatory role of the H1A-DRB1 alleles in the production of antibodies to a conserved region of P. vivax CSP and MSP1 in Brazilian population exposed to malaria. (C) 2011 Elsevier B.V. All rights reserved.

Geographic differentiation of polymorphism in the Plasmodium falciparum malaria vaccine candidate gene SERA5

SERA5 is regarded as a promising malaria vaccine candidate of the most virulent human malaria parasite Plasmodium falciparum. SERA5 is a 120 kDa abundantly expressed blood-stage protein containing a papain-like protease. Since substantial polymorphism in blood-stage vaccine candidates may potentially limit their efficacy, it is imperative to fully investigate polymorphism of the SERA5 gene (sera5). In this study, we performed evolutionary and population genetic analysis of sera5. The level of inter-species divergence (kS = 0.076) between P. falciparum and Plasmodium reichenowi, a closely related chimpanzee malaria parasite is comparable to that of housekeeping protein genes. A signature of purifying selection was detected in the proenzyme and enzyme domains. Analysis of 445 near full-length P. falciparum sera5 sequences from nine countries in Africa, Southeast Asia, Oceania and South America revealed extensive variations in the number of octamer repeat (OR) and serine repeat (SR) regions as well as substantial level of single nucleotide polymorphism (SNP) in non-repeat regions (2562 bp). Remarkably, a 14 amino acid sequence of SERA5 (amino acids 59-72) that is known to be the in vitro target of parasite growth inhibitory antibodies was found to be perfectly conserved in all 445 worldwide isolates of P. falciparum evaluated. Unlike other major vaccine target antigen genes such as merozoite surface protein-1, apical membrane antigen-1 or circumsporozoite protein, no strong evidence for positive selection was detected for SNPs in the non-repeat regions of sera5. A biased geographical distribution was observed in SNPs as well as in the haplotypes of the sera5 OR and SR regions. In Africa, OR- and SR-haplotypes with low frequency (<5%) and SNPs with minor allele frequency (<5%) were abundant and were mostly continent-specific. Consistently, significant genetic differentiation, assessed by the Wright's fixation index (FST) of inter-population variance in allele frequencies, was detected for SNPs and both OR- and SR-haplotypes among almost all parasite populations. The exception was parasite populations between Tanzania and Ghana, suggesting frequent gene flow in Africa. The present study points to the importance of investigating whether biased geographical distribution for SNPs and repeat variants in the OR and SR regions affect the reactivity of human serum antibodies to variants. (C) 2011 Elsevier Ltd. All rights reserved.

Calcium Binding to Leptospira Outer Membrane Antigen LipL32 Is Not Necessary for Its Interaction with Plasma Fibronectin, Collagen Type IV, and Plasminogen

LipL32 is the most abundant outer membrane protein from pathogenic Leptospira and has been shown to bind extracellular matrix (ECM) proteins as well as Ca2+. Recent crystal structures have been obtained for the protein in the apo-and Ca2+-bound forms. In this work, we produced three LipL32 mutants (D163-168A, Q67A, and S247A) and evaluated their ability to interact with Ca2+ and with ECM glycoproteins and human plasminogen. The D163-168A mutant modifies aspartate residues involved in Ca2+ binding, whereas the other two modify residues in a cavity on the other side of the protein structure. Loss of calcium binding in the D163-D168A mutant was confirmed using intrinsic tryptophan fluorescence, circular dichroism, and thermal denaturation whereas the Q67A and S247A mutants presented the same Ca2+ affinity as the wild-type protein. We then evaluated if Ca2+ binding to LipL32 would be crucial for its interaction with collagen type IV and plasma proteins fibronectin and plasminogen. Surprisingly, the wild-type protein and all three mutants, including the D163-168A variant, bound to these ECM proteins with very similar affinities, both in the presence and absence of Ca2+ ions. In conclusion, calcium binding to LipL32 may be important to stabilize the protein, but is not necessary to mediate interaction with host extracellular matrix proteins.

Maternal LAMP/p55gagHIV-1 DNA Immunization Induces In Utero Priming and a Long-Lasting Immune Response in Vaccinated Neonates

Infants born to HIV-infected mothers are at high risk of becoming infected during gestation or the breastfeeding period. A search is thus warranted for vaccine formulations that will prevent mother-to-child HIV transmission. The LAMP/gag DNA chimeric vaccine encodes the HIV-1 p55gag fused to the lysosome-associated membrane protein-1 (LAMP-1) and has been shown to enhance anti-Gag antibody (Ab) and cellular immune responses in adult and neonatal mice; such a vaccine represents a new concept in antigen presentation. In this study, we evaluated the effect of LAMP/gag DNA immunization on neonates either before conception or during pregnancy. LAMP/gag immunization of BALB/c mice before conception by the intradermal route led to the transfer of anti-Gag IgG1 Ab through the placenta and via breastfeeding. Furthermore, there were an increased percentage of CD4+ CD25+ Foxp3+ T cells in the spleens of neonates. When offspring were immunized with LAMP/gag DNA, the anti-Gag Ab response and the Gag-specific IFN-gamma-secreting cells were decreased. Inhibition of anti-Gag Ab production and cellular responses were not observed six months after immunization, indicating that maternal immunization did not interfere with the long-lasting memory response in offspring. Injection of purified IgG in conjunction with LAMP/gag DNA immunization decreased humoral and cytotoxic T-cell responses. LAMP/gag DNA immunization by intradermal injection prior to conception promoted the transfer of Ab, leading to a diminished response to Gag without interfering with the development of anti-Gag T- and B-cell memory. Finally, we assessed responses after one intravenous injection of LAMP/gag DNA during the last five days of pregnancy. The intravenous injection led to in utero immunization. In conclusion, DNA vaccine enconding LAMP-1 with Gag and other HIV-1 antigens should be considered in the development of a protective vaccine for the maternal/fetal and newborn periods.

Neural differentiation of rat aorta pericyte cells

Pericyte perivascular cells, believed to originate mesenchymal stem cells (MSC), are characterized by their capability to differentiate into various phenotypes and participate in tissue reconstruction of different organs, including the brain. We show that these cells can be induced to differentiation into neural-like phenotypes. For these studies, pericytes were obtained from aorta ex-plants of Sprague-Dawley rats and differentiated into neural cells following induction with trans retinoic acid (RA) in serum-free defined media or differentiation media containing nerve growth and brain-derived neuronal factor, B27, N2, and IBMX. When induced to differentiation with RA, cells express the pluripotency marker protein stage-specific embryonic antigen-1, neural-specific proteins beta 3-tubulin, neurofilament-200, and glial fibrillary acidic protein, suggesting that pericytes undergo differentiation, similar to that of neuroectodermal cells. Differentiated cells respond with intracellular calcium transients to membrane depolarization by KCl indicating the presence of voltage-gated ion channels and express functional N-methyl-D-aspartate receptors, characteristic for functional neurons. The study of neural differentiation of pericytes contributes to the understanding of induction of neuroectodermal differentiation as well as providing a new possible stem-cell source for cell regeneration therapy in the brain. (C) 2011 International Society for Advancement of Cytometry

Iodide treatment acutely increases pendrin (SLC26A4) mRNA expression in the rat thyroid and the PCCl3 thyroid cell line by transcriptional mechanisms

Iodine is a critical element involved in thyroid hormone synthesis. Its efflux into the follicular lumen is thought to occur, in part, through pendrin at the apical membrane of thyrocytes. This study attempted to investigate whether iodide administration affects SLC26A4 mRNA expression in rat thyroid and in PCCl3 cells. Rats and cells were treated or not with Nal from 30 min up to 48 h. One group was concomitantly treated with sodium perchlorate. SLC26A4 mRNA expression was also investigated in PCCl3 cells treated with actinomycin D prior to Nal treatment. Iodide administration significantly increased SLC26A4 mRNA content in both models. The simultaneous administration of Nal and perchlorate, as well as the treatment of PCCl3 cells with actinomycin D prevented this effect, indicating that intracellular iodide is essential for this event, which appears to be triggered by transcriptional mechanisms. These data show that intracellular iodide rapidly upregulates SLC26A4 mRNA expression. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Diagnostic utility of immunohistochemistry in distinguishing trichoepithelioma and basal cell carcinoma: evaluation using tissue microarray samples

Trichoepithelioma is a benign neoplasm that shares both clinical and histological features with basal cell carcinoma. It is important to distinguish these neoplasms because they require different clinical behavior and therapeutic planning. Many studies have addressed the use of immunohistochemistry to improve the differential diagnosis of these tumors. These studies present conflicting results when addressing the same markers, probably owing to the small number of basaloid tumors that comprised their studies, which generally did not exceed 50 cases. We built a tissue microarray with 162 trichoepithelioma and 328 basal cell carcinoma biopsies and tested a panel of immune markers composed of CD34, CD10, epithelial membrane antigen, Bcl-2, cytokeratins 15 and 20 and D2-40. The results were analyzed using multiple linear and logistic regression models. This analysis revealed a model that could differentiate trichoepithelioma from basal cell carcinoma in 36% of the cases. The panel of immunohistochemical markers required to differentiate between these tumors was composed of CD10, cytokeratin 15, cytokeratin 20 and D2-40. The results obtained in this work were generated from a large number of biopsies and resulted in the confirmation of overlapping epithelial and stromal immunohistochemical profiles from these basaloid tumors. The results also corroborate the point of view that trichoepithelioma and basal cell carcinoma tumors represent two different points in the differentiation of a single cell type. Despite the use of panels of immune markers, histopathological criteria associated with clinical data certainly remain the best guideline for the differential diagnosis of trichoepithelioma and basal cell carcinoma. Modern Pathology (2012) 25, 1345-1353; doi: 10.1038/modpathol.2012.96; published online 8 June 2012

Placentation in the paca (Agouti paca L)

Background: The paca is a South American rodent with potential as a commercial food animal. We examined paca placenta as part of a wider effort to understand the reproductive biology of this species. Methods: Thirteen specimens between midgestation and term of pregnancy were studied by light and transmission electron microscopy. Results: The placenta is divided into several lobes separated by interlobular trophoblast. Maternal arterial channels and fetal veins are found at the centre of each lobe. In the labyrinth, maternal blood flows through trophoblast-lined lacunae in close proximity to the fetal capillaries. The interhaemal barrier is of the haemomonochorial type with a single layer of syncytiotrophoblast. Caveolae occur in the apical membrane of the syncytiotrophoblast and recesses in the basal membrane, but there is no evidence of transtrophoblastic channels. The interlobular areas consist of cords of syncytiotrophoblast defining maternal blood channels that drain the labyrinth. Yolk sac endoderm covers much of the fetal surface of the placenta. The subplacenta comprises cytotrophoblast and syncytiotrophoblast. There are dilated intercellular spaces between the cytotrophoblasts and lacunae lined by syncytiotrophoblast. In the junctional zone between subplacenta and decidua, there are nests of multinucleated giant cells with vacuolated cytoplasm. The entire placenta rests on a pedicle of maternal tissue. An inverted yolk sac placenta is also present. The presence of small vesicles and tubules in the apical membrane of the yolk sac endoderm and larger vesicles in the supranuclear region suggest that the yolk sac placenta participates in maternal-fetal transfer of protein. Conclusion: The paca placenta closely resembles that of other hystricomorph rodents. The lobulated structure allows for a larger exchange area and the development of precocial young

Surgical complications in 550 consecutive cochlear implantation

Cochlear implantation is a safe and reliable method for auditory restoration in patients with severe to profound hearing loss. Objective: To describe the surgical complications of cochlear implantation. Materials and Methods: Information from 591 consecutive multichannel cochlear implant surgeries were retrospectively analyzed. All patients were followed-up for at least one year. Forty-one patients were excluded because of missing data, follow-up loss or middle fossa approach. Results: Of 550 cochlear implantation analyzed, 341 were performed in children or adolescents, and 209 in adults. The mean hearing loss time was 6.3 +/- 6.7 years for prelingual loss and 12.1 +/- 11.6 years for postlingual. Mean follow-up was 3.9 +/- 2.8 years. Major complications occurred in 8.9% and minor in 7.8%. Problems during electrode insertion (3.8%) were the most frequent major complication followed by flap dehiscence (1.4%). Temporary facial palsy (2.2%), canal-wall lesion (2.2%) and tympanic membrane lesion (1.8%) were the more frequent minor complications. No death occurred. Conclusion: There was a low rate of surgical complications, most of them been successfully managed. These results confirm that cochlear implant is a safe surgery and most surgical complications can be managed with conservative measures or minimal intervention.

Hyaluronidase recruits mesenchymal-like cells to the lung and ameliorates fibrosis

Hyaluronidases (HYALs) comprise a group of enzymes that degrade hyaluronic acid (HA). In this report, we reveal that a single intranasal inoculation of HYAL induces an increase in mononuclear cells within the bronchoalveolar space demonstrating a mesenchymal-like phenotype, expressing stem cell antigen-1 (SCA-1), CD44 and CD73 but not CD34, CD45, CD3, CD4, CD8 or CD19. This influx of mesenchymal stem cell (MSC)-like cells was dependent on leukotriene production within the lung parenchyma. These findings prompted experiments demonstrating that HYAL treatment potently blocked bleomycin-induced lung injury and fibrosis while decreasing transforming growth factor (TGF)-β production and collagen deposition. These data suggest that HYAL is a novel and promising tool to use autologous MSC-like cells in the treatment of pulmonary fibrosis.

Upregulation of soluble and membrane-bound human leukocyte antigen G expression is primarily observed in the milder histopathological stages of chronic hepatitis C virus infection

Chronic hepatitis C virus (HCV) infection is a worldwide health problem that may evolve to cirrhosis and hepatocellular carcinoma. Incompletely understood immune system mechanisms have been associated with impaired viral clearance. The nonclassical class I human leukocyte antigen G (HLA-G) molecule may downregulate immune system cell functions exhibiting well-recognized tolerogenic properties. HCV genotype was analyzed in chronic HCV-infected patients. Because HLA-G expression may be induced by certain viruses, we evaluated the presence of HLA-G in the liver microenvironment obtained from 89 biopsies of patients harboring chronic HCV infection and stratified according to clinical and histopathological features. Overall, data indicated that HCV genotype 1 was predominant, especially subgenotype 1a, with a prevalence of 87%. HLA-G expression was observed in 45(51%) liver specimens, and it was more frequent in milder stages of chronic hepatitis (67.4%) than in moderate (27.8%; p = 0.009) and severe (36.0%; p = 0.021) stages of the disease. Altogether, these results suggest that the expression of HLA-G in the context of HCV is a complex process modulated by many factors, which may contribute to an immunologic environment favoring viral persistence. However, because the milder forms predominantly expressed HLA-G, a protective role of this molecule may not be excluded. (C) 2012 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

Cutting Edge: Brazilian Pemphigus Foliaceus Anti-Desmoglein 1 Autoantibodies Cross-React with Sand Fly Salivary LJM11 Antigen

The environmental factors that contribute to the development of autoimmune diseases are largely unknown. Endemic pemphigus foliaceus in humans, known as Fogo Selvagem (FS) in Brazil, is mediated by pathogenic IgG4 autoantibodies against desmoglein 1 (Dsg1). Clusters of FS overlap with those of leishmaniasis, a disease transmitted by sand fly (Lutzomyia longipalpis) bites. In this study, we show that salivary Ags from the sand fly, and specifically the LJM11 salivary protein, are recognized by FS Abs. Anti-Dsg1 monoclonal autoantibodies derived from FS patients also cross-react with LJM11. Mice immunized with LJM11 generate anti-Dsg1 Abs. Thus, insect bites may deliver salivary Ags that initiate a cross-reactive IgG4 Ab response in genetically susceptible individuals and lead to subsequent FS. Our findings establish a clear relationship between an environmental, noninfectious Ag and the development of potentially pathogenic autoantibodies in an autoimmune disease. The Journal of Immunology, 2012, 189: 1535-1539.

MMP-7 and TIMP-1, news targets in predicting poor wound healing in apical periodontitis

Introduction: Matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) are strongly associated with tissue destruction because of inflammation. In this study, we investigated the expression of MMPs and TIMPs messenger RNA and protein levels in apical periodontitis lesions. Methods: Tissue samples from patients presenting clinical signs of chronic apical abscess (CAA) or asymptomatic apical periodontitis (AAP) were collected postoperatively and used for gene expression analysis of MMP-2, -3, -7, -9, -14, -16, and -25; TIMP-1; and TIMP-2 in real-time polymerase chain reaction. Immunohistochemistry was also performed to detect the expression of MMP-7 and TIMP-1 proteins. Lastly, U-937 cells were induced to terminal differentiation into macrophages, infected with purified Escherichia coli lipopolysaccharide, and assessed for the expression of MMP-7 and TIMP-1 using immunocytochemistry and confocal microscopy. Results: Significantly higher messenger RNA levels were found for all genes in AAP and CAA samples when compared with healthy control samples (P < .001). AAP cases exhibited significantly higher TIMP-1 when compared with CAA cases, whereas CAA cases showed higher MMP-2, MMP-7, and MMP-9 messenger RNA levels (P < .05). We also detected positive the expression of MMP-7 and TIMP-1 proteins in the tissue samples. The expression of both MMP-7 and TIMP-1 were increased in lipopolysaccharide-stimulated cells compared with nonstimulated cells and appear to colocalize in the Golgi apparatus. Conclusions: MMPs appear to have an influential role in CAA cases in which ongoing tissue destruction is observed. TIMPs are preferentially associated with AAP, perhaps as a subsequent defense mechanism against excessive destruction. Taken together, our findings implicate MMP and TIMP molecules in the dynamics of inflammatory periapical lesion development