Transcriptional suppression of Sox9 expression in chondrocytes by retinoic acid

SOX9 is a transcription factor that is expressed in chondrocytes and regulates expression of chondrocyte phenotype related genes. Expression of these genes is known to be suppressed by retinoic acid (RA). We, therefore, examined whether the Sox9 gene expression is regulated by RA in chondrocytes. RA treatment suppressed Sox9 mRNA expression in primary chondrocytes prepared from newborn mouse rib cartilage within 12 h and this suppression lasted at least up to 24 h. The RA suppression of Sox9 mRNA levels was dose-dependent starting at 0.5 muM with a maximum at 1 muM. Nuclear run-on assays revealed that RA reduced the rate of transcription of Sox9 gene. Finally, Western blot analysis indicated that RA suppressed SOX9 protein revels in these chondrocytes. Furthermore, overexpression of SOX9 reversed RA suppression of Col/2a1 enhancer activity. These observations indicate that RA suppresses Sox9 gene expression in chondrocytes at least in part through transcriptional events. (C) 2001 Wiley-Liss, Inc.

A histone deacetylase inhibitor, azelaic bishydroxamic acid, shows cytotoxicity on Epstein-Barr virus-transformed B-cell lines - A potential therapy for posttransplant lymphoproliferative disease

Background. Posttransplant lymphoproliferative disease (PTLD), driven by the presence of Epstein-Barr virus (EBV), is becoming an increasingly important clinical problem after solid organ transplantation. The use of immunosuppressive therapy leads to the inhibition of the cytotoxic T cells that normally control the EBV latently infected B cells. The prognosis for many patients with PTLD is poor, and the optimal treatment strategy is not well defined. Method. This study investigates the use of a histone deacetylase inhibitor, azelaic bishydroxamic acid (ABRA), for its ability to effectively kill EBV-transformed lymphoblastoid cell lines. Results. In vitro treatment of lymphoblastoid cell lines with ABRA showed that they were effectively killed by low doses of the drug (ID50 2-5 mug/ml) within 48 hr. As well as being effective against polyclonal B-cell lines, ABHA was also shown to be toxic to seven of eight clonal Burkitt's lymphoma cell lines, indicating that the drug may also be useful in the treatment of late-occurring clonal PTLD. In addition, ABHA treatment did not induce EBV replication or affect EBV latent gene expression. Conclusion. These studies suggest that ABHA effectively kills both polyclonal and clonal B-cell lines and has potential in the treatment of PTLD.

Development of a novel titration and off-gas analysis (TOGA) sensor for study of biological processes in wastewater treatment systems

The development of the new TOGA (titration and off-gas analysis) sensor for the detailed study of biological processes in wastewater treatment systems is outlined. The main innovation of the sensor is the amalgamation of titrimetric and off-gas measurement techniques. The resulting measured signals are: hydrogen ion production rate (HPR), oxygen transfer rate (OTR), nitrogen transfer rate (NTR), and carbon dioxide transfer rate (CTR). While OTR and NTR are applicable to aerobic and anoxic conditions, respectively, HPR and CTR are useful signals under all of the conditions found in biological wastewater treatment systems, namely, aerobic, anoxic and anaerobic. The sensor is therefore a powerful tool for studying the key biological processes under all these conditions. A major benefit from the integration of the titrimetric and off-gas analysis methods is that the acid/base buffering systems, in particular the bicarbonate system, are properly accounted for. Experimental data resulting from the TOGA sensor in aerobic, anoxic, and anaerobic conditions demonstrates the strength of the new sensor. In the aerobic environment, carbon oxidation (using acetate as an example carbon source) and nitrification are studied. Both the carbon and ammonia removal rates measured by the sensor compare very well with those obtained from off-line chemical analysis. Further, the aerobic acetate removal process is examined at a fundamental level using the metabolic pathway and stoichiometry established in the literature, whereby the rate of formation of storage products is identified. Under anoxic conditions, the denitrification process is monitored and, again, the measured rate of nitrogen gas transfer (NTR) matches well with the removal of the oxidised nitrogen compounds (measured chemically). In the anaerobic environment, the enhanced biological phosphorus process was investigated. In this case, the measured sensor signals (HPR and CTR) resulting from acetate uptake were used to determine the ratio of the rates of carbon dioxide production by competing groups of microorganisms, which consequently is a measure of the activity of these organisms. The sensor involves the use of expensive equipment such as a mass spectrometer and requires special gases to operate, thus incurring significant capital and operational costs. This makes the sensor more an advanced laboratory tool than an on-line sensor. (C) 2003 Wiley Periodicals, Inc.

GH treatment in adults with chronic liver disease: A randomized, double-blind, placebo-controlled, cross-over study

Patients with chronic liver disease (CLD) are catabolic and GH-resistant. The effects of supraphysiological recombinant human GH (rhGH; 0.2 IU.kg(-1).d(-1)) treatment in adults with CLD were assessed in a randomized, double-blind, placebo-controlled cross-over trial (4-wk dietary run-in, 4-wk treatment, and 2-wk wash-out phases). Nine adults with mild- to moderate-severity CLD participated (median age, 49 yr; three males and six females; Child's classification A in six and B in three). Biopsy-proven etiologies were: alcohol (four patients), primary biliary cirrhosis (three patients), non-A, non-B, non-C hepatitis (one patient), and cryptogenic (one patient). Treatment with rhGH increased serum IGF-I (median increase over placebo, +93 mug.liter(-1); P = 0.004), IGF-binding protein-3 (+0.9 mg.liter(-1): P = 0.004), and acid labile subunit (+10.7 nM; P = 0.004). Total body potassium (+8.0 g; P = 0.023), body weight (+1.6 kg; P = 0.008), and total body water (by bioelectrical impedance; +4.9 kg; P = 0.004) increased. Resting metabolic rate (+313 ml.kg(-1).min(-1); P = 0.004) and lipid oxidation (+1072.0 kcal.d(-1); P = 0.032) increased. Metabolic changes included increased fasting plasma glucose (+1.2 mm; P = 0.008), insulin (+33.8 mU.liter(-1); P = 0.004), C-peptide (+0.7 nM; P = 0.004), and free-fatty acids (+0.1 mEq.liter(-1); P = 0.04). Clinical side effects included worsening edema and ascites. Hepatocellular function did not change. Therefore, rbGH treatment in CLD: 1) overcame hepatic GH resistance; 2) may have improved whole-body protein catabolism; 3) increased lipolysis and lipid oxidation; 4) increased insulin resistance; and 5) had potent antinatriuretic effects. Long-term safety and efficacy require further assessment.

Growth hormone receptor and insulin-like growth factor-I immunoreactivity in osteoclast-like cells during tooth eruption in the toothless (Osteopetrotic) rat following treatment with colony-stimulating factor-1

In the toothless (tl/tl) osteopetrotic rat, teeth form but fail to erupt. Treatment of tl/tl rats with colony-stimulating factor-1 (CSF-1) activates bone resorption by osteoclasts, permits tooth eruption, and up-regulates the immunoreactivity of bone marrow mononuclear cells to growth hormone receptor (GHr) and insulin-like growth factor (IGF)-I. This study examined the distribution of tartrate-resistant acid phosphatase (TRAP) and immunoreactivity for GHr and IGF-I in osteoclast-like cells located on the alveolar bone margin, adjacent to the lower first molar crown, in 14-day-old normal and tl/tl rats, following treatment with CSF-1. Osteoclast-like cells demonstrated a positive reaction for TRAP, GHr, and IGF-I in all groups. However, in tl/tl tissue, osteoclast-like cells were generally negative for GHr. There was no significant difference in the total number of TRAP, GHr, and IGF-I-positive osteoclast-like cells on the adjacent bone margin in normal, normal treated with CSF-1, and tl/tl rats. CSF-1 treatment of the tl/tl rat significantly increased the total number of osteoclast-like cells, which were positive for TRAP (p < 0.001), GHr (p < 0.05) and IGF-I (P < 0.01).

Effects of rosiglitazone and linoleic acid on human preadipocyte differentiation

Background Peroxisome proliferator activated receptor gamma (PPARgamma) is a ligand-activated transcription factor known to be central to both adipose tissue development and insulin action. Growth of adipose tissue requires differentiation of preadipocytes with acquisition of specific cellular functions including insulin sensitivity, leptin secretion and the capacity to store triglyceride. Dietary fatty acids and members of the thiazolidinedione class of compounds have been reported to influence adipogenesis at the transcriptional level. Here, we compare the effects of a dietary fatty acid, linoleic acid, and a thiazolidinedione, rosiglitazone, on biochemical and functional aspects of human preadipocyte differentiation in vitro . Materials and methods Human omental and subcutaneous preadipocytes were subcultured 2-3 times and subsequently differentiated for 21 days in the presence of either linoleic acid or rosiglitazone. Differentiation was assessed using a number of biochemical and functional parameters. Results Omental and subcutaneous preadipocytes differentiated in the presence of linoleic acid showed marked cytoplasmic triacylglycerol accumulation however, no biochemical markers of differentiation (LPL expression, G3PDH gene expression and enzyme activity and leptin expression or secretion) were detected. In contrast, treatment of these cells with rosiglitazone induced full biochemical differentiation as judged by all markers assessed, despite comparatively little lipid accumulation. The rosiglitazone effects were subcutaneous depot-specific. Cells treated with linoleic acid showed decreased glucose uptake cf rosiglitazone-treated cells. A luciferase reporter assay demonstrated that rosiglitazone potently activates h-peroxisome proliferator activated receptor gamma while linoleic acid had no effect. Conclusions These studies demonstrate that (a) human preadipocytes have the potential to accumulate triacylglycerol irrespective of their stage of biochemical differentiation; (b) while omental preadipocytes are refractory to biochemical differentiation in vitro , they are able to accumulate triacylglycerol; and (c) rosiglitazone and linoleic acid may exert their effects via different biochemical pathways.

Heat shock and salicylic acid on postharvest preservation of organic strawberries

Heat shock and salicylic acid have been studied on shelf-life extension of fruits. The benefits of these techniques have been related to their effect on inducing physiological defense responses against the oxidative stress and pathogen development. The objective of this study was to evaluate the effect of heat shock and salicylic acid on the postharvest preservation and contents of total phenolics, anthocyanins, ascorbic acid, fresh weight loss and microbiological quality of organic strawberries cv. Dover. Strawberries produced organically and stored at 5 ºC were subjected to heat shock (45 ºC ± 3 ºC for 3 h), application of salicylic acid (soaking in 2.0 mmol L-1 solution), heat shock in combination with salicylic acid and control. After treatment, the fruits were packed and stored in a climatic chamber at 5 ºC ± 2 ºC. At 1, 7 and 14 days, the experimental units were removed from refrigeration and kept at room temperature of approximately 20 ºC for two days. There was no effect of treatments on fresh weight loss, incidence of pathogens or chemical variations in strawberry fruits during the storage period. In natural conditions, organically grown strawberries remained in good condition for sale up to seven days of storage in all treatments.

Cork processing wastewater treatment/valorisation by nanofiltration

Nanofiltration process for the treatment/valorisation of cork processing wastewaters was studied. A DS-5 DK 20/40 (GE Water Technologies) nanofiltration membrane/module was used, having 2.09 m(2) of surface area. Hydraulic permeability was determined with pure water and the result was 5.2 L.h(-1).m(-2).bar(-1). The membrane presents a rejection of 51% and 99% for NaCl and MgSO4 salts, respectively. Two different types of regimes were used in the wastewaters filtration process, total recycling mode and concentration mode. The first filtration regime showed that the most favourable working transmembrane pressure was 7 bar working at 25 degrees C. For the concentration mode experiments it was observed a 30% decline of the permeate fluxes when a volumetric concentration factor of 5 was reached. The permeate COD, BOD5, colour and TOC rejection values remained well above the 90% value, which allows, therefore, the concentration of organic matter (namely the tannin fraction) in the concentrate stream that can be further used by other industries. The permeate characterization showed that it cannot be directly discharged to the environment as it does not fulfil the values of the Portuguese discharge legislation. However, the permeate stream can be recycled to the process (boiling tanks) as it presents no colour and low TOC (< 60 ppm) or if wastewater discharge is envisaged we have observed that the permeate biodegradability is higher than 0.5, which renders conventional wastewater treatments feasible.

Study on the glycerolysis reaction of high free fatty acid oils for use as biodiesel feedstock

Biodiesel is the main alternative to fossil diesel and it may be produced from different feedstocks such as semi-refined vegetable oils, waste frying oils or animal fats. However, these feedstocks usually contain significant amounts of free fatty acids (FFA) that make them inadequate for the direct base catalyzed transesterification reaction (where the FFA content should be lower than 4%). The present work describes a possible method for the pre-treatment of oils with a high content of FFA (20 to 50%) by esterification with glycerol. In order to reduce the FFA content, the reaction between these FFA and an esterification agent is carried out before the transesterification reaction. The reaction kinetics was studied in terms of its main factors such astemperature, % of glycerin excess, % of catalyst used, stirring velocity and type of catalyst used. The results showed that glycerolysis is a promising pretreatment to acidic oils or fats (> 20%) as they led to the production of an intermediary material with a low content of FFA that can be used directly in thetransesterification reaction for the production of biodiesel. (C) 2011 Elsevier B.V. All rights reserved.

Reactivity of human fetal and adult immortalized hepatocytes to potentially toxic bilirubin and bile acid species

Dissertação apresentada para a obtenção do Grau de Mestre em Genética Molecular e Biomedicina, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Recovery of acid and base from brackish water by bipolar membrane electrodialysis

Nesta dissertação pretendeu-se estudar a viabilidade do uso de eletrodiálise com membranas bipolares (BM) na recuperação de ácido clorídrico e de hidróxido de sódio a partir de um efluente industrial que contém 1.4 mol/L de cloreto de sódio. Estas membranas mostraram ser uma ferramenta eficiente para a produção de ácidos e bases a partir do respetivo sal. Foi feita uma seleção de diferentes membranas bipolares (Neosepta, Fumatech e PCA) e aniónicas (PC-SA e PC-ACID 60) na tentativa de encontrar a combinação mais adequada para o tratamento do efluente. Dependendo do critério, o melhor arranjo de membranas é o uso de PC-ACID 60 (membrana aniónica), PC-SK (membrana catiónica) e membranas bipolares do tipo Neosepta para maior pureza dos produtos; membranas bipolares Fumatech para maior eficiência de dessalinização e membranas bipolares PCA para um maior grau de dessalinização. Tecnologicamente foi possível obter uma dessalinização de 99.8% em quatro horas de funcionamento em modo batch com recirculação de todas as correntes. Independentemente da combinação usada é recomendável que o processo seja parado quando a densidade de corrente deixa de ser máxima, 781 A/m2. Assim é possível evitar o aumento de impurezas nos produtos, contra difusão, descida instantânea do pH e uma dessalinização pouco eficiente. A nível piloto o principal fornecedor de membranas e unidade de tratamento “stack” é a marca alemã PCA. Sendo assim realizaram-se ensaios de repetibilidade, contra difusão, avaliação económica e upscaling utilizando as membranas bipolares PCA. A nível económico estudou-se o uso de dois tipos de unidades de tratamento; EDQ 380 e EDQ 1600, para diferentes níveis de dessalinização (50, 75 e 80%). Tendo em conta a otimização económica, é recomendável uma dessalinização máxima de 80%, uma vez que a eficiência de processo a este ponto é de 40%. A aplicação do método com a unidade EDQ 1600 para uma dessalinização de 50% é a mais vantajosa economicamente, com custos de 16 €/m3 de efluente tratado ou 0,78 €/kg Cl- removido. O número de unidades necessárias é 4 posicionados em série.

Carbon key-properties for microcystin adsorption in drinking water treatment: structure or surface chemistry?

Dissertação para Obtenção de Grau de Mestre em Engenharia Química e Bioquímica

The effect of water inorganic matrix in ibuprofen adsorption onto activated carbon for water and wastewater treatment

Dissertação para obtenção do Grau de Mestre em Engenharia Química e Bioquímica

SHIFT IN HUMAN ROTAVIRUS DISTRIBUTION IN BELO HORIZONTE, BRAZIL DETECTED BY RIBONUCLEIC ACID ELECTROPHORESIS

Rotavirus has been considered the main agent of infectious diarrhea especially among younger children. We addressed the prevalence of rotavirus-associated diarrhea and the diversity of circulating electropherotypes by immunochromatography and RNA electrophoresis. Stool samples were taken from 391 children (267 with diarrhea) from the lower socioeconomic stratum who sought treatment in the Hospital Infantil João Paulo II/Belo Horizonte, during 2005 and 2006. Rotavirus was detected in 79/20.2% of subjects, 64/24.0% with diarrhea and 15/12.1% with no diarrhea. The virus was strongly associated with diarrhea (p = 0.003). A total of 76/19.4% and 69/17.6% rotavirus-positive children were identified by immunochromatography and electrophoresis, respectively. Rotavirus-associated diarrhea was more frequently detected in dry months (p < 0.001) and almost exclusively in children aged up to three years. Long profile strains prevailed (54/78.3%) but a shift toward short electropherotype was identified. Despite the decrease seen in 2006, rotavirus infection is still very common in our area. Although viral RNA electrophoresis is useful as a typing method, it should not be used exclusively in the diagnosis of rotavirus infection. We confirmed a shift from long to short profile strains, as already described for other South American countries.

ACUTE KIDNEY INJURY CAUSED BY Crotalus AND Bothrops SNAKE VENOM: A REVIEW OF EPIDEMIOLOGY, CLINICAL MANIFESTATIONS AND TREATMENT

SUMMARY Ophidic accidents are an important public health problem due to their incidence, morbidity and mortality. An increasing number of cases have been registered in Brazil in the last few years. Several studies point to the importance of knowing the clinical complications and adequate approach in these accidents. However, knowledge about the risk factors is not enough and there are an increasing number of deaths due to these accidents in Brazil. In this context, acute kidney injury (AKI) appears as one of the main causes of death and consequences for these victims, which are mainly young males working in rural areas. Snakes of the Bothrops and Crotalus genera are the main responsible for renal involvement in ophidic accidents in South America. The present study is a literature review of AKI caused by Bothrops and Crotalus snake venom regarding diverse characteristics, emphasizing the most appropriate therapeutic approach for these cases. Recent studies have been carried out searching for complementary therapies for the treatment of ophidic accidents, including the use of lipoic acid, simvastatin and allopurinol. Some plants, such as Apocynaceae, Lamiaceae and Rubiaceae seem to have a beneficial role in the treatment of this type of envenomation. Future studies will certainly find new therapeutic measures for ophidic accidents.