Macrophage suppression following phagocytosis of apoptotic neutrophils is mediated by the S100A9 calcium-binding protein

The clearance of apoptotic cells by phagocytes is a fundamental process during tissue remodeling and resolution of inflammation. In turn, the phagocytosis of apoptotic cells generates signals that suppress pro-inflammatory activation of macrophages. These events occur during the resolution phase of inflammation and therefore the malfunctioning of this process may lead to inflammation-related tissue damage. Here, we demonstrate that the calcium-binding protein S100A9, normally abundant in the cytoplasm of neutrophils and also released by apoptotic neutrophils, is involved in the suppression of macrophages after the uptake of apoptotic neutrophils. Both, spontaneous and induced production of inflammatory species (nitric oxide, hydrogen peroxide and TNF-alpha) as well as the phagocytic activity were inhibited when macrophages were in presence of apoptotic neutrophils, conditioned medium from neutrophil cultures or a peptide corresponding to the C-terminal region of S100A9 protein. On the other hand, macrophages kept in the conditioned medium of neutrophils that was previously depleted of S100A9 were shown to resume the activated status. Finally, we demonstrate that the calcium-binding property of S100A9 might play a role in the suppression process, since the stimulation of intracellular calcium release with ionomycin significantly reversed the effects of the uptake of apoptotic neutrophils in macrophages. In conclusion, we propose that S100A9 is a novel component of the regulatory mechanisms of inflammation, acting side-by-side with other suppressor factors generated upon ingestion of apoptotic cells. (C) 2009 Elsevier GmbH. All rights reserved.

Structural stability and reversible unfolding of recombinant porcine S100A12

Porcine S100A12 is a member of the S100 proteins, family of small acidic calcium-binding proteins characterized by the presence of two EF-hand motifs. These proteins are involved in many cellular events such as the regulation of protein phosphorylation, enzymatic activity, protein-protein interaction, Ca(2+) homeostasis, inflammatory processes and intermediate filament polymerization. In addition, members of this family bind Zn(2+) or Ca(2+) with cooperative effect on binding. In this study, the gene sequence encoding porcine S100A12 was obtained by the synthetic gene approach using E. coli codon bias. Additionally, we report a thermodynamic study of the recombinant S100A12 using circular dichroism, fluorescence and isothermal titration calorimetry. The results of urea and temperature induced unfolding and refolding processes indicated a reversible two-state process. Also, the ANS fluorescence studies showed that in presence of divalent ions the protein exposes hydrophobic sites which could facilitate the interaction with other proteins and trigger the physiological responses. (c) 2008 Elsevier B.V. All rights reserved.

Age and geochemistry of mantle peridotites and diorite dykes from the Baldissero body: Insights into the Paleozoic-Mesozoic evolution of the Southern Alps

Trace element and isotopic data obtained for mantle spinel Iherzolites and diorite dykes from the Baldissero massif (Ivrea-Verbano Zone, Western Italy) provide new, valuable constraints on the petrologic and geodynamic evolution of the Southern Alps in Paleozoic to Mesozoic times. Whole rock and mineral chemistry indicates that Baldissero Iherzolites can be regarded as refractory mantle residues following limited melt extraction. In particular, the Light Rare Earth Elements (LREE)-depleted and fractionated compositions of whole rock and clinopyroxene closely match modelling results for refractory residues after low degrees (similar to 4-5%) of near-fractional melting of depleted mantle, possibly under garnet-facies conditions. Following this, the peridotite sequence experienced subsolidus re-equilibration at lithospheric spinel-facies conditions and intrusion of several generations of dykes. However, Iherzolites far from dykes show very modest metasomatic changes, as evidenced by the crystallisation of accessory titanian pargasite and the occurrence of very slight enrichments in highly incompatible trace elements (e.g. Nb). The Re-Os data for Iherzolites far from the dykes yield a 376 Ma (Upper Devonian) model age that is considered to record a partial melting event related to the Variscan orogenic cycle s.l. Dioritic dykes cutting the mantle sequence have whole rock, clinopyroxene and plagioclase characterised by high radiogenic Nd and low radiogenic Sr, which point to a depleted to slightly enriched mantle source. Whole rock and mafic phases of diorites have high Mg# values that positively correlate with the incompatible trace element concentrations. The peridotite at the dyke contact is enriched in orthopyroxene, iron and incompatible trace elements with respect to the Iherzolites far from dykes. Numerical simulations indicate that the geochemical characteristics of the diorites can be explained by flow of a hydrous, silica-saturated melt accompanied by reaction with the ambient peridotite and fractional crystallisation. The composition of the more primitive melts calculated in equilibrium with the diorite minerals show tholeiitic to transitional affinity. Internal Sm-Nd, three-point isochrons obtained for two dykes suggest an Upper Triassic-Lower Jurassic emplacement age (from 204 31 to 198 29 Ma). Mesozoic igneous events are unknown in the southern Ivrea-Verbano Zone (IVZ), but the intrusion of hydrous melts, mostly silica-saturated, have been well documented in the Finero region, i.e. the northernmost part of IVZ and Triassic magmatism with calc-alkaline to shoshonitic affinity is abundant throughout the Central-Eastern Alps. The geochemical and chronological features of the Baldissero diorites shed new light on the geodynamic evolution of the Southern Alps before the opening of the Jurassic Tethys. (C) 2010 Elsevier B.V. All rights reserved.

Electrocardiographic study on geriatric dogs undergoing general anesthesia with isoflurane

The purpose of this study was to clarify the degree of influence of anesthetic agents commonly used during anesthesia on the heart conduction systems of geriatric dogs, with or without the presence of electrocardiographic changes in the pre-anesthetic electrocardiogram and also to determine the possible causes of ST-segment and T-wave changes during anesthesia, by monitoring ventilation and oxygenation. 36 geriatric dogs were evaluated. In addition to electrocardiographic evaluation, the pre-anesthetic study included serum levels of urea, creatinine, total protein, albumin and electrolytes. The pre-anesthetic medication consisted of acepromazine (0.05mg kg-1) in association with meperidine (3.0mg kg-1) by IM injection. Anesthesia was induced with propofol (3.0 to 5.0mg kg-1) by IV injection and maintained with isoflurane in 100% oxygen. During the anesthesia, the animals were monitored by continued computerized electrocardiogram. Systemic blood pressure, heart rate, respiratory rate, end-tidal carbon dioxide, partial pressure of carbon dioxide in arterial blood, arterial oxygen saturation, partial pressure of arterial oxygen and oxygen saturation of hemoglobin were closely monitored. During maintenance anesthesia, normal sinus rhythm was more common (78%). ST-segment and T-wave changes during the anesthetic procedure were quite common and were related to hypoventilation. The use of isoflurane did not result in arrhythmia, being therefore a good choice for this type of animal; Electrocardiographic findings of ST-segment and T-wave changes during the maintenance anesthesia were evident in animals with hypercapnia, a disorder that should be promptly corrected with assisted or controlled ventilation to prevent complicated arrhythmias.

Dietas com nitrogênio não-proteico para fêmeas bovinas superovuladas sem prévia adaptação durante curto tempo e em diferentes fases do ciclo estral

Objetivou-se neste estudo avaliar o efeito do fornecimento, sem prévia adaptação, durante curto tempo e em diferentes fases do ciclo estral, de dietas contendo nitrogênio não-proteico (NNP) na produção, na qualidade e no grau de desenvolvimento de embriões recuperados em fêmeas bovinas superovuladas. Sessenta e oito vacas Nelore foram distribuídas em três grupos: um controle e dois com fornecimento de ureia antes (UA = do dia -5 ao dia 0) e após (UD = dia 0 ao dia 5) a inseminação artificial. As vacas foram mantidas em pastagem e receberam concentrado (3,0 kg/animal/dia) durante 16 dias. Foram formulados dois concentrados, e as dietas totais (concentrado e consumo estimado de pastagem) apresentaram 12,0% (dieta controle) e 14,6% (dieta NNP) de proteína bruta (PB). As vacas foram sincronizadas, superovuladas e inseminadas. Sete dias (dia 7) após a inseminação (dia 0), realizou-se a colheita e análise dos embriões. Amostras de sangue foram coletadas nos dias -5, 0 e 5 para determinação da concentração de nitrogênio ureico plasmático (NUP), glicose, insulina e progesterona. A época de fornecimento de ureia influenciou as concentrações médias de NUP nos dias -5, 0 e 5, mas não tiveram efeito nas concentrações de glicose, insulina e progesterona. O período de fornecimento da ureia teve efeito na porcentagem de mórulas compactas em relação ao total de estruturas (UA = 51,4 vs. UD = 15,3%), em relação ao total de oócitos fecundados (UA = 62,5 vs. UD = 30,6%) e em relação ao total de embriões viáveis (UA = 68,8 vs. UD = 38,6%). No grupo que recebeu ureia depois da inseminação artificial, houve redução de 70,2% da proporção de mórulas compactas por total de estruturas em relação ao grupo que recebeu ureia antes da inseminação. Dietas com nitrogênio não-proteico fornecidas imediatamente após a inseminação promovem aceleração do desenvolvimento embrionário inicial.

Parâmetros sanguíneos de vacas leiteiras suplementadas com diferentes fontes de gordura

O objetivo deste trabalho foi avaliar a utilização de diferentes fontes de gordura nas rações de vacas em lactação e seus efeitos sobre o consumo de matéria seca e as concentrações dos parâmetros sangüíneos. Foram utilizadas 12 vacas da raça Holandês, agrupadas em três quadrados latinos balanceados 4x4 e alimentadas com as seguintes rações: 1) controle; 2) óleo de soja refinado; 3) grão de soja in natura; e 4) sais de cálcio de ácidos graxos (Megalac-E). As concentrações de colesterol total, colesterol, lipoproteína de baixa densidade (LDL) e colesterol de alta densidade (HDL) foram maiores (P<0,05) para as vacas alimentadas com as rações contendo fontes de gordura em relação à ração controle. As concentrações de uréia e nitrogênio ureico no soro foram semelhantes entre as rações utilizadas, com exceção da ração contendo sais de cálcio de ácidos graxos, que apresentou concentrações menores (P<0,05). O consumo de matéria seca foi menor (P<0,05) nas vacas que receberam sais de cálcio nas rações. Os animais que receberam a ração contendo grão de soja como fonte de gordura apresentaram menor (P<0,05) produção de leite quando comparada às demais rações utilizadas. A utilização de fontes de gordura em rações de vacas em lactação alterou os parâmetros sanguíneos, dependendo da fonte de gordura utilizada.

A TRIPLE-MODE BANDPASS FILTER USING A MODIFIED CIRCULAR PATCH RESONATOR

This article presents a triple-mode bandpass filter using a modified circular patch resonator. Etched slots in the resonator split the TM(1, 1, 0)(z) degenerate fundamental modes and also perturb the TM(2, 1, 0)(z) mode, approximating their resonant frequencies to form a third-order bandpass filter. A 2.42 GHz centered filter was designed and fabricated. Experimental results showed a fractional bandwidth of 29%, return loss better than 16 dB, insertion loss of 0.5 dB, and good second harmonic band rejection. The filter exhibited a size reduction of 51% compared with a filter using an unperturbed circular patch resonator at the same frequency. (C) 2008 Wiley Periodicals, Inc. Microwave Opt Technol Lett 51: 178-182, 2009; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/mop.23950

Bladder irrigation with amphotericin B and fungal urinary tract infection-systematic review with meta-analysis

Background: Candiduria is a hospital-associated infection and a daily problem in the intensive care unit. The treatment of asymptomatic candiduria is not well established and the use of amphotericin B bladder irrigation (ABBI) is controversial. The aim of this systematic review was to determine the best place for this therapy in practice. Methods: The databases searched in this study included MEDLINE, EMBASE, Web of Science, and LILACS (January 1960-June 2007). We included manuscripts with data on the treatment of candiduria using ABBI. The studies were classified as comparative, dose-finding, or non-comparative. Results: From 213 studies, nine articles (377 patients) met our inclusion criteria. ABBI showed a higher clearance of the candiduria 24 hours after the end of therapy than fluconazole (odds ratio (OR) 0.57, 95% confidence interval (CI) 0.32-1.00). Fungal culture 5 days after the end of both therapies showed a similar response (OR 1.51, 95% CI 0.81-2.80). The evaluation of ABBI using an intermittent or continuous system of delivery showed an early candiduria clearance (24 hours after therapy) of 80% and 82%, respectively (OR 0.87, 95% CI 0.52-1.36). Candiduria clearance at >5 days after the therapy showed a superior response using continuous bladder irrigation with amphotericin B (OR 0.52, 95% CI 0.29-0.94). The use of continuous ABBI for more than 5 days showed a better result (88% vs. 78%) than ABBI for less than 5 days, but without significance (OR 0.55, 95% CI 0.34-1.04). Conclusion: Although the strength of the results in the underlying literature is not sufficient to allow the drawing of definitive conclusions, ABBI appears to be as effective as fluconazole, but it does not offer systemic antifungal therapy and should only be used for asymptomatic candiduria. (C) 2008 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Anti-Group A Streptococcal Vaccine Epitope STRUCTURE, STABILITY, AND ITS ABILITY TO INTERACT WITH HLA CLASS II MOLECULES

Streptococcus pyogenes infections remain a health problem in several countries due to poststreptococcal sequelae. We developed a vaccine epitope (StreptInCor) composed of 55 amino acids residues of the C-terminal portion of the M protein that encompasses both T and B cell protective epitopes. The nuclear magnetic resonance (NMR) structure of the StreptInCor peptide showed that the structure was composed of two microdomains linked by an 18-residue alpha-helix. A chemical stability study of the StreptInCor folding/unfolding process using far-UV circular dichroism showed that the structure was chemically stable with respect to pH and the concentration of urea. The T cell epitope is located in the first microdomain and encompasses 11 out of the 18 alpha-helix residues, whereas the B cell epitope is in the second microdomain and showed no alpha-helical structure. The prediction of StreptInCor epitope binding to different HLA class II molecules was evaluated based on an analysis of the 55 residues and the theoretical possibilities for the processed peptides to fit into the P1, P4, P6, and P9 pockets in the groove of several HLA class II molecules. We observed 7 potential sites along the amino acid sequence of StreptInCor that were capable of recognizing HLA class II molecules (DRB1*, DRB3*, DRB4*, and DRB5*). StreptInCoroverlapping peptides induced cellular and humoral immune responses of individuals bearing different HLA class II molecules and could be considered as a universal vaccine epitope.

Cationic technetium and rhenium complexes with pendant carbohydrates

Three carbohydrate conjugated dipicolylamine chelators, 2-bis(2-pyridinylmethyl)amino)ethyl 1-deoxy-1-thio-beta-D-glucopyranoside (L(1)), 2-bis(2-pyridinylmethyl)amino)ethyl-beta-D-glucopyranoside (L(2)), and 2-bis(2-pyridinylmethyl)amino)carboxamide-N-(2-amino-2-deoxy-D-glucopyranose) (L(3)) were complexed to the [M(Co)(3)](+) core (M=Tc, Re) and the properties of the resulting complexes were investigated. Synthesis and characterization of the chelator 2-bis(2-pyridinylmethyl)amino)ethyl 1-deoxy-1-thio-beta-D-glucopyranoside (L(1)) and the corresponding Re complex are reported. All chelators were radiolabeled in high yield with [(99)mTc(CO)(3)(H(2)O)(3)](+) ( > 98%) and [(186)Re(CO)(3)(H(2)O)(3)](+) ( > 80%). The chelators and Re-complexes were determined to not be substrates for the glucose metabolism enzyme hexokinase. However, the biodistribution of each of the (99m)Tc complexes demonstrated fast clearance from most background tissue, including >75% clearance of the activity in the kidneys and the liver within 2 h post-injection. (C) 2010 Elsevier Ltd. All rights reserved.

Chronic American trypanosomiasis: parasite persistence in endomyocardial biopsies is associated with high-grade myocarditis

Myocyte diameter, fractional area of collagen, intensity of myocarditis and parasite persistence (explored by immunohistochemistry and PCR) were evaluated in serial sections of endomyocardial biopsies from 29 outpatients with chronic chagasic cardiopathy. The patients, 25 males and four females with a mean (S.D.) age of 43 (9) years, were subsequently followed up for 3-2861 days (median=369 days). During this follow-up, 16 (55%) of the patients died. The biopsies revealed myocarditis in 25 (86%) of the patients and high-grade myocarditis in 14 (56%). Although immunohistochemistry failed to demonstrate Trypanosoma cruzi antigens in any of the samples, five (33%) of the 15 biopsies successfully tested in the PCR-based assay for T. cruzi DNA were found positive, indicating parasite persistence. There was a significant positive association between myocardial parasite persistence and high-grade myocarditis (P= 0.014); five (71%,) of the seven endomyocardial biopsies with high-grade myocarditis that were successfully tested in the PCR assays showed persistent T. cruzi DNA. The survival time of the patients was not, however, found to be significantly associated with myocardial parasite persistence, any of the morphometric measurements taken, or the presence or intensity of myocarditis.

Extracellular matrix components and regulators in the airway smooth muscle in asthma

There is an intimate relationship between the extracellular matrix (ECM) and smooth muscle cells within the airways. Few studies have comprehensively assessed the composition of different ECM components and its regulators within the airway smooth muscle (ASM) in asthma. With the aid of image analysis, the fractional areas of total collagen and elastic fibres were quantified within the ASM of 35 subjects with fatal asthma (FA) and compared with 10 nonfatal asthma (NFA) patients and 22 nonasthmatic control cases. Expression of collagen I and III, fibronectin, versican, matrix metalloproteinase (MMP)-1, -2, -9 and -12 and tissue inhibitor of metalloproteinase-1 and -2 was quantified within the ASM in 22 FA and 10 control cases. In the large airways of FA cases, the fractional area of elastic fibres within the ASM was increased compared with NFA and controls. Similarly, fibronectin, MMP-9 and MMP-12 were increased within the ASM in large airways of FA cases compared with controls. Elastic fibres were increased in small airways in FA only in comparison with NFA cases. There is altered extracellular matrix composition and a degradative environment within the airway smooth muscle in fatal asthma patients, which may have important consequences for the mechanical and synthetic functions of airway smooth muscle.

Pkd1 Haploinsufficiency Increases Renal Damage and Induces Microcyst Formation following Ischemia/Reperfusion

Mutations in PKD1 cause the majority of cases of autosomal dominant polycystic kidney disease (ADPKD). Because polycystin 1 modulates cell proliferation, cell differentiation, and apoptosis, its lower biologic activity observed in ADPKD might influence the degree of injury after renal ischemia/reperfusion. We induced renal ischemia/reperfusion in 10- to 12-wk-old male noncystic Pkd1(+/-) and wild-type mice. Compared with wild-type mice, heterozygous mice had higher fractional excretions of sodium and potassium and higher serum creatinine after 48 h. In addition, in heterozygous mice, also cortical damage, rates of apoptosis, and inflammatory infiltration into the interstitium at time points out to 14 d after injury all increased, as well as cell proliferation at 48 h and 7 d. The mRNA and protein expression of p21 was lower in heterozygous mice than wild-type mice at 48 h. After 6 wk, we observed dilated tubules, microcysts, and increased renal fibrosis in heterozygotes. The early mortality of heterozygotes was significantly higher than that of wild-type mice when we extended the duration of ischemia from 32 to 35 min. In conclusion, ischemia/reperfusion induces a more severe injury in kidneys of Pkd1-haploin-sufficient mice, a process that apparently depends on a relative deficiency of p2l activity, tubular dilation, and microcyst formation. These data suggest the possibility that humans with ADPKD from PKD1 mutations may be at greater risk for damage from renal ischemia/reperfusion injury.

Paracoccidioidomycosis: Cells expressing IL17 and Foxp3 in cutaneous and mucosal lesions

We demonstrated and quantified by immunohistochemistry the population of cells expressing IL17 and Foxp3 in cutaneous and mucosal paracoccidioidomycosis lesions, associating these populations of cells with different presentations of granulomatous response. For this purpose, 61 skin biopsies and 55 oral mucosal biopsies were evaluated. Cells expressing IL17 were distributed in the inflammatory infiltrate in both groups of lesions and were found in the vessels` wall too. Foxp3+ expression was limited to the nuclei of lymphocytes in the inflammatory infiltrate. The distribution of IL17 was similar among the groups; however, Foxp3+ cells were increased in mucosal lesions that displayed compact granulomas. The results suggest that IL17 seems to play a role in paracoccidioidomycosis cutaneous and mucosal lesions, probably as secondary cells in the clearance of the fungal antigens. The presence of Foxp3+ cells both in skin and mucosa corroborates some previous researches that suggest the role of this group of cells in the modulation of local immune response. (C) 2011 Elsevier Ltd. All rights reserved.

Volume-activated chloride channels in mice Leydig cells

Production and secretion of testosterone in Leydig cells are mainly controlled by the luteinizing hormone (LH). Biochemical evidences suggest that the activity of Cl(-) ions can modulate the steroidogenic process, but the specific ion channels involved are not known. Here, we extend the characterization of Cl(-) channels in mice Leydig cells (50-60 days old) by describing volume- activated Cl(-) currents (I(Cl,swell)). The amplitude of I(Cl,swell) is dependent on the osmotic gradient across the cell membrane, with an apparent EC(50) of similar to 75 mOsm. These currents display the typical biophysical signature of volume- activated anion channels (VRAC): dependence on intracellular ATP, outward rectification, inactivation at positive potentials, and selectivity sequence (I(-)>Cl(-)>F(-)). Staurosporine (200 nM) did not block the activation of I(Cl), swell. The block induced by 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB; 128 mu M), SITS (200 mu M), ATP (500 mu M), pyridoxalphosphate-6- azophenyl-2`,4`-disulfonate (PPADS; 100 mu M), and Suramin (10 mu M) were described by the permeant blocker model with apparent dissociation constant at 0 mV K(d)(0) and fractional distance of the binding site (delta) of 334 mu M and 47%, 880 mu M and 35%, 2,100 mu M and 49%, 188 mu M and 27%, and 66.5 mu M and 49%, respectively. These numbers were derived from the peak value of the currents. We conclude that ICl, swell in Leydig cells are activated independently of purinergic stimulation, that Suramin and PPADS block these currents by a direct interaction with VRAC and that ATP is able to permeate this channel.