Correlation between biological responses in vitro and in vivo to Ca-doped sol-gel coatings assessed using proteomic analysis.

Poor correlation between the results of in vitro testing and the subsequent in vivo experiments hinders the design of biomaterials. Thus, new characterisation methods are needed. This study used proteomic and histological techniques to analyse the effects of Ca-doped biomaterials in vitro and in vivo and verify the correlation between the two systems. The sol-gel route was employed to synthesise coatings functionalised with 0.5 and 5 wt% of CaCl2. Morphology of the coatings was examined using SEM; the Ca2+ ion release from the materials was analysed by means of ICP-AES spectroscopy. The osteogenic and inflammatory responses were inspected in vitro in human osteoblasts (HOb) and TPH-1 monocytes. The in vivo experiments used a rabbit model. The nLC-MS/MS-based proteomic methods were utilised to analyse the proteins adhering to the material samples incubated with human serum or examine protein expression in the tissues close to the implants. Ca-doped biomaterials caused a remarkable increase in the adsorption of coagulation-related proteins, both in vitro (PLMN, THRB, FIBA and VTNC) and in vivo (FBLN1, G1U978). Enhanced affinity to these materials was also observed for proteins involved in inflammation (CO5, C4BPA, IGHM and KV302 in vitro; CARD6, DDOST and CD14 in vivo) and osteogenic functions (TETN, PEDF in vitro; FBN1, AHSG, MYOC in vivo). The results obtained using different techniques were well matched, with a good correlation between the in vitro and in vivo experiments. Thus, the proteomic analysis of biological responses to biomaterials in vitro is a useful tool for predicting their impact in vivo.

Bioactive zinc-doped sol-gel coating modulates protein adsorption patterns and in vitro cell responses.

Zinc is an essential element with an important role in stimulating the osteogenesis and mineralization and suppressing osteoclast differentiation. In this study, new bioactive ZnCl2-doped sol-gel materials were designed to be applied as coatings onto titanium. The biomaterials were physicochemically characterized and the cellular responses evaluated in vitro using MC3T3-E1 osteoblasts and RAW264.7 macrophages. The effect of Zn on the adsorption of human serum proteins onto the material surface was evaluated through nLC-MS/MS. The incorporation of Zn did not affect the crosslinking of the sol-gel network. A controlled Zn2+ release was obtained, reaching values below 10 ppm after 21 days. The materials were no cytotoxic and lead to increased gene expression of ALP, TGF-ß, and RUNX2 in the osteoblasts. In macrophages, an increase of IL-1ß, TGF-ß, and IL-4 gene expression was accompanied by a reduced TNF-α liberation. Proteomic results showed changes in the adsorption patterns of proteins associated with immunological, coagulative, and regenerative functions, in a Zn dose-dependent manner. The variations in protein adsorption might lead to the downregulation of the NF-κB pathway, thus explain the observed biological effects of Zn incorporation into biomaterials. Overall, these coatings demonstrated their potential to promote bone tissue regeneration.

Lippia gracilis Schauer essential oil as a growth promoter for Japanese quail.

The use of antibiotics as performance enhancers in animal feeding is declining, so Lippia gracilis Schauer essential oil (LGSEO) could be used as a potential substitute for the conventionally used growth promoters. The LGSEO contains components such as carvacrol and thymol, which kill and/or control pathogenic bacteria, increase population of beneficial organisms, act against oxidative processes and onto nutrient digestibility and absorption. The aim of this study was to investigate the action and the effects of LGSEO as a growth promoter in the diet of Japanese quail by examining their productive performance, intestinal microbiology, blood biochemical parameters, hepatic thiobarbituric acid reactive substances (TBARS) content and intestinal gene expression. A total of 252 two-day-old quail (Coturnix coturnix japonica) were assigned to 3 treatments in 7 replicates, using 12 birds per experimental unit. The treatments consisted of a basal diet, basal diet + LGSEO at 400 mg/kg of diet and basal diet + chemical antimicrobial (bacitracin methylene disalicylate) at 500 mg/kg of diet. The experimental period was 34 days. The highest feed intake (P < 0.01) was found in the group receiving the conventional antimicrobial, whereas the best feed conversion (P < 0.01) was shown by the animals receiving LGSEO. Escherichia coli growth was restricted in the quail receiving the growth promoters. Salmonella spp. growth was controlled by the treatment containing the conventional antimicrobial. There was no difference between the treatments (P > 0.05) for the concentration of aspartate aminotransferase and alanine aminotransferase enzymes in the blood or hepatic TBARS content. Birds receiving negative-control treatment exhibited a higher expression of sodium-glucose cotransporter (SGLT1), while those receiving the treatment with essential oil showed lower catalase (CAT) and glutathione peroxidase (GPX7) expressions compared to the conventional antimicrobial and control groups, respectively. Lippia gracilis Schauer essential oil is a powerful performance enhancer for Japanese quail by virtue of its abilities to improve their intestinal environment, balance the microbial population and reduce energy expenditure for oxidative processes.

Validation of Anthropometric Measures Self-Reported in a Randomized Controlled Trial of a Web-Based Platform for Weight Loss.

INTRODUCTION: A great number of weight loss interventions have been delivered through digital solutions. Analysis of the effectiveness in terms of weight loss is fundamental to understand the real potential of digital technologies as tools for delivery of weight loss interventions. For this, we need accurate and reliable anthropometric data. For reasons of convenience, self-reported weight and height often replace actual measurements in these interventions. This might lead to misclassification of BMI status during selection of participants and to bias in the assessment of the outcomes. Therefore, it is fundamental to have validation studies of self-reported web-based data. OBJECTIVES: We aimed to validate online self-reported height, weight and BMI in a POEmaS trial subsample. METHODS: We included 12.5% of the POEmaS' population (n=159). Anthropometric data reported on the web-platform were compared to measured data by paired T-tests. Agreement was assessed by Bland-Altman plots. Multinomial regression was used to investigate factors associated with self-reported weight validity. RESULTS: There was no significant difference between reported and measured weight (0.4 kg, SD 1.7; p=0.13) and BMI (0.03 kg/m2, SD 0.87; p=0.06). Reported height was on average 0.4 cm (SD 1.2) higher than the measured ones (p<0.001). For all anthropometric data, >=95% of the cases were within the limits of agreement. Higher measured BMI was the only factor associated with low accuracy of weight report. Each unit increase in BMI increased the odds that the reported weight was lower than the one measured (OR 1.13; 95%CI 1.01-1.26). DISCUSSION: Self-reported weight and BMI change showed good agreement with measured ones. Since these are the primary outcomes of the POEmaS trial, the findings of the validation study suggest that the outcomes' accuracy is high and that it does not vary across gender, age, study group. These findings are relevant to digital health researchers and assessors and suggest that digital health interventions for weight loss might rely on self-reported assessment of outcomes. This might be particularly useful when other modes of assessment, such as anthropometry and e-scales, are not feasible or not available. However, we acknowledge that these results might not be applicable to low educated populations.

Proteomic analysis of calcium-enriched sol-gel biomaterials.

Calcium is an element widely used in the development of biomaterials for bone tissue engineering as it plays important roles in bone metabolism and blood coagulation. The Ca ions can condition the microenvironment at the tissue-material interface, affecting the protein deposition process and cell responses. The aim of this study was to analyze the changes in the patterns of protein adsorption on the silica hybrid biomaterials supplemented with different amounts of CaCl2, which can function as release vehicles. This characterization was carried out by incubating the Ca-biomaterials with human serum. LC-MS/MS analysis was used to characterize the adsorbed protein layers and compile a list of proteins whose affinity for the surfaces might depend on the CaCl2 content. The attachment of pro- and anti-clotting proteins, such as THRB, ANT3, and PROC, increased significantly on the Ca-materials. Similarly, VTNC and APOE, proteins directly involved on osteogenic processes, attached preferentially to these surfaces. To assess correlations with the proteomic data, these formulations were tested in vitro regarding their osteogenic and inflammatory potential, employing MC3T3-E1 and RAW 264.7 cell lines, respectively. The results confirmed a Ca dose-dependent osteogenic and inflammatory behavior of the materials employed, in accordance with the protein attachment patterns.

Virulence, resistance, and genetic relatedness of Escherichia coli and Klebsiella sp. isolated from mule foals / Virulência, resistência e relação genética de Escherichia coli e Klebsiella sp. isoladas de potros de muar

Respiratory diseases are common in young horses but little is known about such infections in mule foals. This study aimed to characterize Escherichia coli and Klebsiella sp. isolated from tracheal wash (TW) and fecal samples (FS) of mule foals, with or without cytological evidence of respiratory disease. Strains were analyzed against 13 antimicrobials, for presence of Extended spectrum beta-lactamase (ESBL), and virulence genes. Phylogrouping and Randomic (RAPD)-PCR profiles were used to evaluate their genetic relatedness. E. coli strains from TW and FS showed greatest resistance to tetracycline, while Klebsiella strains were mainly resistant to ampicillin; multidrug resistance and ESBL production were also detected. The blaCTX gene prevailed among the E. coli isolates, while the blaSHV gene was more frequently found in K. pneumoniae. The fimH gene was detected in most of the isolates and multiple virulence factors were identified in three E. coli isolates. Most of the E. coli isolates belonged to the B1 phylogroup, but B2 strains displayed more virulence genes. The RAPD assay revealed genetic diversity among strains and was able to distinguish FS isolates from TW isolates. Knowledge of the bacteria associated with the respiratory tract of mule foals is important in the treatment of sick animals.(AU)

Doenças respiratórias são comuns em potros de equinos, porém pouco se sabe sobre tais infecções em potros de muar. Este estudo buscou caracterizar Escherichia coli e Klebsiella sp. isolados de lavados traqueais (TW) e amostras fecais (FS) de potros de muar com e sem evidências citológicas de doença respiratória. As amostras bacterianas foram testadas contra 13 antimicrobianos, para a presença de genes de resistência estendida às betalactamases (ESBL) e de virulência. Filogrupagem e perfis de PCR randômicos (RAPD) foram usados para avaliar sua relação genética. As amostras de E. coli de TW e FS mostraram maior resistência à tetraciclina, enquanto as amostras de Klebsiella foram mais resistentes à ampicilina; multirresistência e produção de ESBL também foram detectadas. O gene blaCTX foi mais frequente entre E. coli, enquanto o gene blaSHV foi mais encontrado entre K. pneumoniae. O gene fimH foi detectado na maioria dos isolados de E. coli, enquanto múltiplos genes de virulência foram identificados em três isolados de E. coli. A maioria dos isolados de E. coli pertenceu ao filogrupo B1, porém somente isolados do filogrupo B2 apresentaram mais genes de virulência. Os ensaios de RAPD demonstraram a diversidade genética entre as amostras e distinguiram amostras TW e FS. O conhecimento de bactérias associadas a infecções de trato respiratório de potros de muar é importante no tratamento de animais doentes.(AU)

Injectable MnSr-doped brushite bone cements with improved biological performance.

Good mechanical properties and high injectability are the major requirements to ensure widespread application of calcium phosphate cements (CPCs) as bone substitutes in minimally invasive surgeries. However, obtaining CPCs that exhibit a good compromise between these two properties as well as good biological performance is still a great challenge. This study presents novel solutions to improve these properties, which include (i) co-doping ß-tricalcium phosphate (ß-TCP) powder with Sr and Mn, and (ii) adding small amounts of saccharides (sucrose or fructose) to the setting-liquid solution. The combination of these two strategies enabled full injectability and significantly increased the wet compressive strength of CPCs in comparison to undoped or solely Sr-doped CPCs. Furthermore, the proliferative response of human MG63 osteoblastic cells, their rate of collagen-I secretion, and particularly their growth behaviour on the cement surfaces were also enhanced. The overall improved relevant properties of Mn/Sr co-doped CPCs with added sucrose, including in vitro biological performance, renders them very promising materials for bone regeneration and tissue engineering.

Protective effects of exogenous and endogenous hydrogen sulfide in mast cell-mediated pruritus and cutaneous acute inflammation in mice.

The recently described 'gasomediator' hydrogen sulfide (H2S) has been involved in pain mechanisms, but its effect on pruritus, a sensory modality that similarly to pain acts as a protective mechanism, is poorly known and controversial. The effects of the slow-releasing (GYY4137) and spontaneous H2S donors (Na2S and Lawesson's reagent, LR) were evaluated in histamine and compound 48/80 (C48/80)-dependent dorsal skin pruritus and inflammation in male BALB/c mice. Animals were intradermally (i.d.) injected with C48/80 (3µg/site) or histamine (1µmol/site) alone or co-injected with Na2S, LR or GYY4137 (within the 0.3-100nmol range). The involvement of endogenous H2S and KATP channel-dependent mechanism were also evaluated. Pruritus was assessed by the number of scratching bouts, whilst skin inflammation was evaluated by the extravascular accumulation of intravenously injected 125I-albumin (plasma extravasation) and myeloperoxidase (MPO) activity (neutrophil recruitment). Histamine or C48/80 significantly evoked itching behavior paralleled by plasma extravasation and increased MPO activity. Na2S and LR significantly ameliorated histamine or C48/80-induced pruritus and inflammation, although these effects were less pronounced or absent with GYY4137. Inhibition of endogenous H2S synthesis increased both Tyrode and C48/80-induced responses in the skin, whereas the blockade of KATP channels by glibenclamide did not. H2S-releasing donors significantly attenuate C48/80-induced mast cell degranulation either in vivo or in vitro. We provide first evidences that H2S donors confer protective effect against histamine-mediated acute pruritus and cutaneous inflammation. These effects can be mediated, at least in part, by stabilizing mast cells, known to contain multiple mediators and to be primary initiators of allergic processes, thus making of H2S donors a potential alternative/complementary therapy for treating inflammatory allergic skin diseases and related pruritus.

Capsaicin and Its Role in Chronic Diseases.

A significant number of experimental and clinical studies published in peer-reviewed journals have demonstrated promising pharmacological properties of capsaicin in relieving signs and symptoms of non-communicable diseases (chronic diseases). This chapter provides an overview made from basic and clinical research studies of the potential therapeutic effects of capsaicin, loaded in different application forms, such as solution and cream, on chronic diseases (e.g. arthritis, chronic pain, functional gastrointestinal disorders and cancer). In addition to the anti-inflammatory and analgesic properties of capsaicin largely recognized via, mainly, interaction with the TRPV1, the effects of capsaicin on different cell signalling pathways will be further discussed here. The analgesic, anti-inflammatory or apoptotic effects of capsaicin show promising results in arthritis, neuropathic pain, gastrointestinal disorders or cancer, since evidence demonstrates that the oral or local application of capsaicin reduce inflammation and pain in rheumatoid arthritis, promotes gastric protection against ulcer and induces apoptosis of the tumour cells. Sadly, these results have been paralleled by conflicting studies, which indicate that high concentrations of capsaicin are likely to evoke deleterious effects, thus suggesting that capsaicin activates different pathways at different concentrations in both human and rodent tissues. Thus, to establish effective capsaicin doses for chronic conditions, which can be benefited from capsaicin therapeutic effects, is a real challenge that must be pursued.

Serotypes, virulence markers and cell invasion ability of Shiga toxin-producing Escherichia coli strains isolated from healthy dairy cattle.

AIM: The occurrence of virulence markers, serotypes and invasive ability were investigated in Shiga toxin-producing Escherichia coli (STEC) isolated from faecal samples of healthy dairy cattle at Rio de Janeiro State, Brazil. METHODS AND RESULTS: From 1562 stx-positive faecal samples, 105 STEC strains were isolated by immuno-magnetic separation (IMS) or plating onto MacConkey agar (MC) followed by colony hybridisation. Fifty (47·6%) strains belonged to nine serotypes (O8:H19, O22:H8, O22:H16, O74:H42, O113:H21, O141:H21, O157:H7, O171:H2 and ONT:H21). The prevalent serotypes were O157:H7 (12·4%), O113:H21 (6·7%) and O8:H19 (5·7%). Virulence genes were identified by polymerase chain reaction (PCR). E-hlyA (77·1%) was the more prevalent virulence marker, followed by espP (64·8%), saa (39%), eae (24·8%) and astA (21·9%). All O157:H7 strains carried the γ (gamma) variant of the locus of enterocyte effacement (LEE) genes and the stx2c gene, while the stx1/stx2 genotype prevailed among the eae-negative strains. None of the eae-positive STEC produced the localized adherence (LA) phenotype in HEp-2 or Caco-2 cells. However, intimate attachment (judged by the fluorescent actin staining test) was detected in some eae-positive strains, both in HEp-2 (23·1%) and in Caco-2 cells (11·5%). Most strains (87·5%) showed 'peripheral association' (PA) adherence phenotype to undifferentiated Caco-2 cells. Twenty-five (92·6%) of 27 strains invaded Caco-2 cells. The highest average value of invasion (9·6%) was observed among the eae-negative bovine strains from serotypes described in human disease. CONCLUSION: Healthy dairy cattle is a reservoir of STEC carrying virulence genes and properties associated with human disease. SIGNIFICANCE AND IMPACT OF THE STUDY: Although reports of human disease associated with STEC are scarce in Brazil, the colonization of the animal reservoir by potentially pathogenic strains offers a significant risk to our population.

Resistance patterns, ESBL genes, and genetic relatedness of Escherichia coli from dogs and owners

Abstract Antimicrobial resistance in Escherichia coli isolated from pet dogs can be considered a potential threat of infection for the human population. Our objective was to characterize the resistance pattern, extended spectrum beta-lactamase production and genetic relatedness of multiresistant E. coli strains isolated from dogs (n = 134), their owners (n = 134), and humans who claim to have no contact with dogs (n = 44, control), searching for sharing of strains. The strains were assessed for their genetic relatedness by phylogenetic grouping and pulsed-field gel electrophoresis. Multiresistant E. coli strains were isolated from 42 (31.3%) fecal samples from pairs of dogs and owners, totaling 84 isolates, and from 19 (43.1%) control group subjects. The strains showed high levels of resistance to ampicillin, streptomycin, tetracycline, trimethoprim and sulfamethoxazole regardless of host species or group of origin. The blaTEM, blaCTX-M, and blaSHV genes were detected in similar proportions in all groups. All isolates positive for bla genes were ESBL producers. The phylogenetic group A was the most prevalent, irrespective of the host species. None of the strains belonging to the B2 group contained bla genes. Similar resistance patterns were found for strains from dogs, owners and controls; furthermore, identical PFGE profiles were detected in four (9.5%) isolate pairs from dogs and owners, denoting the sharing of strains. Pet dogs were shown to be a potential household source of multiresistant E. coli strains.

Integration of biofiltration and advanced oxidation processes for tertiary treatment of an oil refinery wastewater aiming at water reuse.

The combination of biological and chemical oxidation processes is an interesting approach to remove ready, poor, and non-biodegradable compounds from complex industrial wastewaters. In this study, biofiltration followed by H2O2/UV oxidation (or microfiltration) and final reverse osmosis (RO) step was employed for tertiary treatment of an oil refinery wastewater. Biofiltration alone allowed obtaining total organic carbon (TOC), chemical oxygen demand (COD), UV absorbance at 254 nm (UV254), ammonium, and turbidity removal of around 46, 46, 23, 50, and 61 %, respectively. After the combined biological-chemical oxidation treatment, TOC and UV254 removal amounted to 88 and 79 %, respectively. Whereas, the treatment performance achieved with different UV lamp powers (55 and 95 W) and therefore distinct irradiance levels (26.8 and 46.3 mW/cm(2), respectively) were very similar and TOC and UV254 removal rates were highly affected by the applied C/H2O2 ratio. Silt density index (SDI) was effectively reduced by H2O2/UV oxidation, favoring further RO application. C/H2O2 ratio of 1:4, 55 W UV lamp, and 20-min oxidation reaction corresponded to the experimental condition which provided the best cost/benefit ratio for TOC, UV254, and SDI reduction from the biofilter effluent. The array of treatment processes proposed in this study has shown to be adequate for tertiary treatment of the oil refinery wastewater, ensuring the mitigation of membrane fouling problems and producing a final effluent which is suitable for reuse applications.

Resistance patterns, ESBL genes, and genetic relatedness of Escherichia coli from dogs and owners.

Antimicrobial resistance in Escherichia coli isolated from pet dogs can be considered a potential threat of infection for the human population. Our objective was to characterize the resistance pattern, extended spectrum beta-lactamase production and genetic relatedness of multiresistant E. coli strains isolated from dogs (n=134), their owners (n=134), and humans who claim to have no contact with dogs (n=44, control), searching for sharing of strains. The strains were assessed for their genetic relatedness by phylogenetic grouping and pulsed-field gel electrophoresis. Multiresistant E. coli strains were isolated from 42 (31.3%) fecal samples from pairs of dogs and owners, totaling 84 isolates, and from 19 (43.1%) control group subjects. The strains showed high levels of resistance to ampicillin, streptomycin, tetracycline, trimethoprim and sulfamethoxazole regardless of host species or group of origin. The blaTEM, blaCTX-M, and blaSHV genes were detected in similar proportions in all groups. All isolates positive for bla genes were ESBL producers. The phylogenetic group A was the most prevalent, irrespective of the host species. None of the strains belonging to the B2 group contained bla genes. Similar resistance patterns were found for strains from dogs, owners and controls; furthermore, identical PFGE profiles were detected in four (9.5%) isolate pairs from dogs and owners, denoting the sharing of strains. Pet dogs were shown to be a potential household source of multiresistant E. coli strains.

Feelings, perceptions, and expectations of patients during the process of oral cancer diagnosis.

PURPOSE: Communication barriers during the process of cancer diagnosis can adversely affect how patients understand their health, and understanding patient's perceptions and expectations can favor adherence to professional recommendations and their prognosis. This study aimed to describe the sociodemographic characteristics, perceptions, expectations, and psychological symptoms of patients during the process of oral cancer diagnosis. METHODS: Patients were assessed at two time points: pre-biopsy (T0) and post-diagnosis (T1). At T0, 49 patients answered a sociodemographic, perceived social support, lifestyle questionnaire and inventories (anxiety-State-Trait Anxiety Inventory (STAI-S/T), depression-Beck Depression Inventory (BDI), risk drinkers-Alcohol Use Disorders Identification Test (AUDIT)). They also were interviewed concerning their perceptions, feelings, and expectations. At T1, the 29 patients diagnosed with oral cancer were reassessed for anxiety (STAI-E) and depression symptoms. An interview investigated the same aspects at T0. RESULTS: Patient mean age was 59 years old (±13.7) and 73.5 % were men. At T0, depression symptoms were more frequent among cancer patients (83.3 %) than those without the disease (p < 0.05), with a prevalence of 36.7 %. Patients presented anxiety (40.8 %) and alcohol abuse behavior (32.6 %). Associations between characteristics and categories at T0 showed that patients with lower income and living with a partner reported negative feelings (p < 0.05) more frequently, together with negative expectations concerning the diagnosis among older patients (p < 0.05) and those with depressive symptoms (p < 0.05). At T1, negative feelings were more frequently reported among patients with anxiety (p < 0.01) and depression symptoms (p < 0.05). CONCLUSION: Professional support and care regarding the psychological aspects and characteristics of patients is required during the diagnosis process in order to promote patient adhesion and favorable prognosis.

Wolbachia spp. interfere na detecção molecular de Ehrlichia canis e Anaplasma platys em cães com microfilaremia / Wolbachia spp. interferes with molecular detection of Ehrlichia canis and Anaplasma platys in microfilaremic dogs

This report aimed to study the interference in molecular testing for Ehrlichia canis and Anaplasma platys in blood of 155 dogs from the coastal region of Rio de Janeiro. Five Anaplasmataceae positive samples but negative for E. canis and A. platys, from microfilaremic animals, were chosen for sequencing. These sequences, when compared to Gen et Bank database, showed 88% to 100% similarity with Wolbachia spp. denoting an interference in the detection of DNA from other members of Anaplasmataceae, possibly due to a high concentration of Wolbachia spp. DNA.(AU)

Renal transplantation in human immunodeficiency virus-positive patients: a report of four cases.

INTRODUCTION: Renal transplantation (RT) in patients infected with human immunodeficiency virus (HIV) has significantly improved under the advent of combined antiretroviral therapy (cART). The authors describe their experience in RT in patients with HIV from September 2010 to June 2013. CASES REPORT: Four patients underwent transplantation (3 with HIV-1 and 1 with HIV-2), three patients were male, and one was black. None were coinfected with hepatitis B virus (HBV) or hepatitis C virus (HCV). Etiology of kidney disease was HIV-associated nephropathy (2 patients), immunoglobulin (Ig)A nephropathy, and unknown. Average age at RT was 51 (range, 41-63) years. No patient was of high immunologic risk. Immunosuppression consisted of basiliximab for induction and prednisolone, tacrolimus (TAC), and mycophenolate mofetil for maintenance. TAC levels varied considerably in the early days (8.5-46 ng/mL), requiring major adjustments in TAC dose. Only the HIV-2 patient had delayed graft function. The follow-up of patients with HIV-1 was 37, 19, and 16 months, and 3 months for the HIV-2 patient. CD4+ T cells decreased in the early days after transplantation with subsequent improvement, along with persistent virological suppression. In the HIV-1 group there were no major infectious, cardiovascular, or neoplastic complications. Nevertheless, the HIV-2 patient died 3 months after RT due to H1N1 pneumonia complicated by pulmonary aspergillosis. Average estimated (CKD- EPI) glomerular filtration rate (eGFR) at 6 months was 85.6 mL/min/1.73 m(2). CONCLUSION: Besides the difficulty in adjusting calcineurin inhibitors levels due to its interaction with antiretroviral therapy, namely with protease inhibitors, no patient had acute rejection. Furthermore, all patients presented an excellent control of viro-immunologic parameters. At the last follow-up neither cardiovascular events nor neoplastic complications were observed. Our results highlight the favorable outcome of RT in HIV-1-infected patients. The HIV-2 patient died due to severe infection, and the clinical management and potential benefit of RT in HIV-2-infected patients needs further study.

Milder course in Duchenne patients with nonsense mutations and no muscle dystrophin.

Duchenne muscular dystrophy (DMD), a severe and lethal condition, is caused by the absence of muscle dystrophin. Therapeutic trials aiming at the amelioration of muscle function have been targeting the production of muscle dystrophin in affected Duchenne patients. However, how much dystrophin is required to rescue the DMD phenotype remains an open question. We have previously identified two exceptional golden retriever muscular dystrophy (GRMD) dogs with a milder course despite the total absence of muscle dystrophin. Here we report two unusual patients carrying nonsense mutations in the DMD gene and dystrophin deficiency but with an unexpectedly mild phenotype. Three reported polymorphisms, respectively in genes LTBP4, SPP1 and ACTN3 were excluded as possible DMD genetic modifiers in our patients. Finding the mechanisms that protect some rare patients and dogs from the deleterious effect of absent muscle dystrophin is of utmost importance and may lead to new avenues for treatment. Importantly, these observations indicate that it is possible to have a functional large muscle even without dystrophin.

Effects of Mn-doping on the structure and biological properties of ß-tricalcium phosphate.

Doping calcium phosphates with trace elements that exist in bone tissues is beneficial in terms of cell-material interactions and in vivo performance of the bone grafts made thereof. Manganese (Mn) is an essential element for normal growth and metabolism of bone tissues, but studies reporting the effects of Mn-doping calcium phosphates are scarce. The present study investigated the influence of Mn-doping on the structure, morphology and biological properties of ß-tricalcium phosphate [ß-Ca3(PO4)2] (ß-TCP). Mn-doped (MnTCP) powders, with Mn contents varying from 0 to 10 mol%, were obtained through an aqueous precipitation method followed by heat treatment at 800 °C. The successful incorporation of Mn into ß-TCP structure was proved through quantitative X-ray diffraction (XRD) phase analysis coupled with structural Rietveld refinement. Increasing Mn concentrations led to decreasing trends of a- and c-axis lattice parameters, and Mn-doping also significantly affected the morphology of ß-TCP powders. In vitro proliferation and differentiation assays of MC3T3-E1 osteoblastic-like cells, grown in the presence of the powders, revealed that the biological benefits of Mn-doped ß-TCP are limited to lower Mn incorporation levels and potentially related to their surface microstructure. The Mn1-ßTCP composition revealed the best set of bioactivity properties, potentially a good candidate for future applications of ß-TCP materials in osteoregeneration.

Construction and validation of metagenomic DNA libraries from landfarm soil microorganisms.

Landfarming biodegradation is a strategy used by the petrochemical industry to reduce pollutants in petroleum-contaminated soil. We constructed 2 metagenomic libraries from landfarming soil in order to determine the pathway used for mineralization of benzene and to examine protein expression of the bacteria in these soils. The DNA of landfarm soil, collected from Ilhéus, BA, Brazil, was extracted and a metagenomic library was constructed with the Copy Control(TM) Fosmid Library Production Kit, which clones 25-45-kb DNA fragments. The clones were selected for their ability to express enzymes capable of cleaving aromatic compounds. These clones were grown in Luria-Bertani broth plus L-arabinose, benzene, and chloramphenicol as induction substances; they were tested for activity in the catechol cleavage pathway, an intermediate step in benzene degradation. Nine clones were positive for ortho-cleavage and one was positive for meta-cleavage. Protein band patterns determined by SDS-polyacrylamide gel electrophoresis differed in bacteria grown on induced versus non-induced media (Luria-Bertani broth). We concluded that the DNA of landfarm soil is an important source of genes involved in mineralization of xenobiotic compounds, which are common in gasoline and oil spills. Metagenomic library allows identification of non-culturable microorganisms that have potential in the bioremediation of contaminated sites.