Enabling 3D hepatocyte spheroids for microphysiometry.

Advances in the areas of tissue engineering and microfabrication techniques have enabled promising in vitro platforms, known as Organs-on-Chips, with the aim of mimicking complex in vivo conditions for more accurate toxicology studies. To analyze the physiological change induced by chemicals or toxic substances continuously, sensors can be used in order to measure the intracellular and extracellular environment of single cells, cell constructs, or tissue, and therefore the integration of monitoring techniques into 3D tissue culture platforms provides an essential step for the next generation Organ-on-Chip platforms. However, current in vitro platforms are not capable of combining the culture of 3D models with monitoring techniques. To address this, a novel spheroid encapsulation is designed for fluidic contact between 3D models in microwells and Intelligent Mobile Lab for In Vitro Diagnostics (IMOLA-IVD) BioChip sensors while preventing spheroid fusion. In this work, spheroid culturing protocols were developed for optimized spheroid growth and an evaluation of spheroid integrity on different porous layers was performed in order to provide a defined spheroid encapsulation on BioChip sensors.

Use of computed tomography to compare two femoral head and neck excision ostectomy techniques as performed by two novice veterinarians.

OBJECTIVES: To compare the results of femoral head and neck excision (FHNE) ostectomy performed by two novice veterinarians using an osteotome and mallet or microsagittal saw. METHODS: In this ex vivo cadaveric study, hindlimbs of eight canine cadavers were randomized to FHNE with osteotome or micro sagittal saw as performed by two recently graduated veterinarians. The hindimbs were imaged by computed tomography (CT) before and after the osteotomy. Post FHNE CT images were evaluated by a board certified radiologist blinded to the ostectomy technique for assessment of the number of bone fragments, fissures, smoothness of osteotomy margination, and volume of residual femoral neck. RESULTS: Femoral head and neck excision performed with the osteotome produced more peri-ostectomy bone fragments, cortical fissures, irregular margins, and residual femoral neck volume, compared with osteotomy using a saw. CLINICAL RELEVANCE: Compared to FHNE performed with a sagittal saw, osteotome FHNE resulted in a greater bone trauma and residual neck bone volume, which would require post-ostectomy modification in a clinical setting.

Polarization influence on reflectance measurements in the spatial frequency domain.

In this work, we quantify the influence of crossed polarizers on reflectance measurements in the spatial frequency domain. The use of crossed polarizers is a very common approach for suppression of specular surface reflections. However, measurements are typically evaluated using a non-polarized scalar theory. The consequences of this discrepancy are the focus of our study, and we also quantify the related errors of the derived optical properties. We used polarized Monte Carlo simulations for forward calculation of the reflectance from different samples. The samples' scatterers are assumed to be spherical, allowing for the calculation of the scattering functions by Mie theory. From the forward calculations, the reduced scattering coefficient [Formula: see text] and the absorption coefficient µa were derived by means of a scalar theory, as commonly used. Here, we use the analytical solution of the scalar radiative transfer equation. With this evaluation approach, which does not consider polarization, we found large errors in [Formula: see text] and µa in the range of 25% and above. Furthermore, we investigated the applicability of the use of a reference measurement to reduce these errors as suggested in literature. We found that this method is not able to generally improve the accuracy of measurements in the spatial frequency domain. Our general recommendation is to apply a polarized theory when using crossed polarizers.

An automated microphysiological assay for toxicity evaluation.

Screening a newly developed drug, food additive or cosmetic ingredient for toxicity is a critical preliminary step before it can move forward in the development pipeline. Due to the sometimes dire consequences when a harmful agent is overlooked, toxicologists work under strict guidelines to effectively catalogue and classify new chemical agents. Conventional assays involve long experimental hours and many manual steps that increase the probability of user error; errors that can potentially manifest as inaccurate toxicology results. Automated assays can overcome many potential mistakes that arise due to human error. In the presented work, we created and validated a novel, automated platform for a microphysiological assay that can examine cellular attributes with sensors measuring changes in cellular metabolic rate, oxygen consumption, and vitality mediated by exposure to a potentially toxic agent. The system was validated with low buffer culture medium with varied conductivities that caused changes in the measured impedance on integrated impedance electrodes.

Label-free monitoring of whole cell vitality.

The Intelligent Mobile Lab (IMOLA) delivers metabolic and morphological parameters of living cells in a label-free and real time way. It represents a key technology for the development of new cell-based assays. Electrochemical microsensors are used to measure the extracellular acidification (pH), cellular respiration (pO2), changes in cell number and morphology (electric impedance) in a controlled environment. These parameters are closely linked to the intracellular signaling network of the living cells. They are thus likely to respond sensitively to changes in cellular vitality. A wide spectrum of cell types can be tested with the system, including adherent and suspended cells, continuous cell lines, primary cells or tissue samples. The platform is described in detail and applications in the field's oncology, toxicology and environmental monitoring are shown.

Caracterização físico-química e sensorial de kefir tradicional e derivados / Physicolchemical and sensorial characteristic of traditional kefir and derivatives

Avaliaram-se diferentes populações de grãos de kefir tradicional e de seus derivados, o kefir, o kefir leban e o soro de kefir, quanto às características físico-químicas e sensoriais, quando inoculados em diferentes concentrações de leite padronizado, levando-se em consideração as variáveis tempo/temperatura de incubação, maturação e filtração. A manipulação da população de grãos de kefir foi eficaz na obtenção de produtos com características físico-químicas semelhantes. O volume de leite utilizado na incubação influenciou significativamente as características dos derivados. A avaliação sensorial de antepastos elaborados a partir de kefir leban, condimentados, apresentou boa aceitabilidade.

It was evaluated different traditional kefir grain populations and their derivatives, kefir, kefir leban and kefir whey, regarding physicochemical and sensorial characteristics when inoculated into different standardized milk concentrations, with incubation time/temperature, maturation and filtration variables. The manipulation of the population of kefir grains was effective in obtaining products with similar physicochemical characteristics and the volume of milk used in the incubation significantly influenced the characteristics of derivatives. The sensory evaluation of kefir leban food formulations such as spiced hors d'oeuvre, showed good acceptability.

Loss of tumor progression locus 2 (tpl2) enhances tumorigenesis and inflammation in two-stage skin carcinogenesis.

Tumor progression locus 2 (Tpl2) is a serine/threonine kinase in the mitogen-activated protein kinase signal transduction cascade known to regulate inflammatory pathways. Previously identified as an oncogene, its mutation or overexpression is reported in a variety of human cancers. To address its role in skin carcinogenesis, Tpl2(-/-) or wild-type (WT) C57BL/6 mice were subjected to a two-stage dimethylbenzanthracene/12-O-tetradecanoylphorbol-13-acetate (TPA) mouse skin carcinogenesis model. Tpl2(-/-) mice developed a significantly higher incidence of tumors (80%) than WT mice (17%), as well as a reduced tumor latency and a significantly higher number of total tumors (113 vs 6). Moreover, Tpl2(-/-) mice treated with TPA experienced significantly higher nuclear factor kappaB (NF-κB) activation, edema, infiltrating neutrophils and production of proinflammatory cytokines than did WT mice. We investigated the role of the p38, JNK, MEK and NF-κB signaling pathways both in vitro and in vivo in WT and Tpl2(-/-) mice by using inhibitors for each of these pathways. We confirmed that the proinflammatory effect in Tpl2(-/-) mice was due to heightened activity of the NF-κB pathway. These studies indicate that Tpl2 may serve more as a tumor suppressor than as an oncogene in chemically induced skin carcinogenesis, with its absence contributing to both tumorigenesis and inflammation.

Atividade antibacteriana in vitro de inflorescências de Achyrocline satureioides (Lam.) DC. - Asteraceae ("macela", "marcela") sobre agentes bacterianos de interesse em alimentos / In vitro antibacterial activity of inflorescences of Achyroclines satureioides (Lam.) DC. - Asteraceae ("macela", "marcela") on bacterial agents of interest in food

Através de Testes de diluição em sistema de tubos múltiplos determinou-se in vitro, atividade antibacteriana em inflorescências de Achyrocline satureioides (Lam.) DC. - Asteraceae ("macela", "marcela"), expressa como Intensidade de Atividade de Inibição Bacteriana (IINIB/bacteriostasia) e Intensidade de Atividade de Inativação Bacteriana (IINAB/bactericidia), a partir de formas de extração etanólica (hidroalcoolaturas) e hídrica (decoctos), sobre inóculos padronizados de Enterococcus faecalis (ATCC 19433), Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 11229) e Salmonella enteritidis (ATCC 11076). E. faecalis apresentou a maior sensibilidade, seguido por Staphylococcus aureus, enquanto S. enteritidis e E. coli apresentaram-se mais resistentes. Dentre as formas de extração, a hidroalcoolatura apresentou capacidade de inibição e/ou inativação intensa e seletiva frente aos quatro inóculos bacterianos. Os decoctos mostraram-se completamente ineficazes frente às bactérias Gram-negativas, enquanto que as Gram-positivas apresentaram somente bacteriostasia/inibição.

Dilution test in multiple tube system was used to determine the in vitro antibacterial activity in inflorescences of Achyrocline satureioides (Lam.) DC. - Asteraceae ("macela", "marcela"), expressed as intensity of bacterial inhibition activity (IINIB/bacteriostasis) and intensity of bacterial inactivation activity (IINAB/bactericidie), from ethanol (hydroalcoholic) and water (decoction) extraction forms on standardized inocula of Enterococcus faecalis (ATCC 19433), Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 11229) and Salmonella enteritidis (ATCC 11076). E. faecalis had the highest sensitivity, followed by S. aureus, while S. enteritidis and E. coli were more resistant. Of the extraction forms, the hydroalcoholic one showed intense and selective inhibition and/or inactivation capacity against four bacterial inocula. Decoctions were completely ineffective against the Gram-negative bacteria, whereas Gram-positive bacteria showed only bacteriostasis/inhibition.

Diferentes condimentos vegetais: avaliação sensorial e de atividade antibacteriana em preparação alimentar com frango cozido / Different spice plants: sensorial evaluation and antibacterial activity in chicken broth

A partir da atividade antibacteriana in vitro, predeterminada em doze plantas com indicativo etnográfico condimentar, testou-se este atributo in loco no modelo caldo com frango cozido. Primeiramente, procedeu-se ao treinamento de 10 avaliadores, segundo a legislação vigente quanto ao Consentimento Livre e Esclarecido, oportunizando conhecimentos prévios sobre as plantas salsa (Petroselinum sativum), manjerona branca (Origanum X aplii), manjerona preta (Origanum majorana), manjericão (Ocimum basilicum), sálvia (Salvia officinalis), tomilho (Thymus vulgaris), anis verde (Ocimum selloi), alfavaca (Ocimum gratissimum), alho nirá (Allium tuberosum), alho poró (Allium porrum), cúrcuma (Curcuma longa) e pimenta dedo-de-moça (Capsicum baccatum). Realizou-se, através da adição individualizada desses condimentos ao caldo com frango cozido, um Teste de Aceitação tipo escala hedônica, selecionando, dentre os doze condimentos, quatro deles que se destacaram sensorialmente, a pimenta dedo-de-moça, o alho nirá, o alho poró e o tomilho. Foi feito, então, um Teste de Aceitação de concentrações denominadas pequena, média e grande destes quatro condimentos, para determinação da intensidade sensorialmente melhor aceita. As quantidades eleitas (0,5 g de pimenta dedo-de-moça, 15 g de alho nirá, 15 g de alho poró e 5 g de tomilho) foram acrescidas ao caldo com frango cozido, sendo estes desafiados frente a Escherichia coli (ATCC 11229) em concentração final de 10 UFC mL-1, limite tolerado pela legislação, tendo como grupo-controle o caldo com frango cozido sem condimentos. O crescimento bacteriano foi aferido a cada duas horas após a inoculação, até completar 24 horas de confronto, utilizando-se meio seletivo para coliformes termo-resistentes e incubação constante a 25ºC em DBO, sendo atribuídos valores arbitrários às variações logarítmicas de crescimento. Comparados ao controle, todos os tratamentos condimentados apresentaram, individualmente, atividade antibacteriana significativa, mesmo que sem significância quando comparados entre si. Contudo, em relação ao tempo de início da atividade antibacteriana, destacou-se a pimenta dedo-de-moça, enquanto que, em relação ao prolongamento dessa ação no tempo, destacou-se o alho nirá. As 12 plantas condimentares em estudo tiveram atestada a sensorialidade, sendo que as quatro plantas com destaque tiveram a atividade anti-coliforme termo-resistente comprovada in loco. Diferentes condimentos vegetais foram capazes de fornecer qualificação sensorial e sanitária em caldo com frango cozido, em condições domésticas de manuseio.

Based on the in vitro antibacterial activity predetermined for 12 spice plants with ethnographic indicator, this feature was tested in loco in the model cooked chicken broth. First, ten evaluators were trained, according to the current legislation for Free and Informed Consent, providing previous knowledge about the plants parsley (Petroselinum sativum), marjoram (Origanum X aplii and Origanum majorana), basil (Ocimum basilicum), common sage (Salvia officinalis), thyme (Thymus vulgaris), anis-like spice (Ocimum selloi), african basilicum (Ocimum gratissimum), nirá garlic (Allium tuberosum), leek (Allium porrum), turmeric (Curcuma longa) and "dedo-de-moça" chili (Capsicum baccatum). Those spices were individually added to the chicken broth to perform a Hedonic Scale-like Acceptance Test, selecting four of the twelve spices that had higher sensory acceptance, "dedo-de-moça" chili, nirá garlic, leek and thyme. A new Acceptance Test was then performed using low, medium and high concentrations of those four spices to establish the most acceptable sensory intensities. The elected quantities (0.5 g "dedo-de-moça" chili, 15 g nirá garlic, 15 g leek and 5 g thyme) were added to the chicken broth, then challenged with Escherichia coli (ATCC 11229) at a final 10 concentration of CFU/mL, the tolerated limit according to legislation. The control group was chicken broth without spices. The bacterial growth was measured at every two hours after the inoculation until 24 hours of confront were completed, using a selective medium for thermo-resistant coliforms, under constant incubation at 25ºC in DBO. Arbitrary values were assigned to the logarithmic growth variations. Compared to the control group, all the spiced treatments individually presented significant antibacterial activity, although the latter was not significant when treatments were compared with each other. However, as regards the antibacterial activity starting time, "dedo-de-moça" chili had the best results, whereas nirá garlic had the best results as regard the extension of this activity time. The 12 studied spice plants had their sensorial characteristics attested, and the four plants that had the best results had proved in loco anti-thermo-resistant coliform activity. Different spice plants were capable of providing sensory and sanitary qualification in chicken broth, under domestic conditions of manipulation.

Atividade antibacteriana in vitro de pimentas e pimentões (Capsicum sp. ) sobre quatro bactérias toxinfectivas alimentares / In vitro antibacterial activity of hot and sweet peppers (Capsicum sp. ) on four food toxinfective bacteria

Determinou-se in vitro a Intensidade de Atividade de Inibição Bacteriana (IINIB) e a Intensidade de Atividade de Inativação Bacteriana (IINAB), através de Testes de Diluição em Sistema de Tubos Múltiplos, de extratos de oito pimentas do gênero Capsicum, etnograficamente acessadas na região metropolitana de Porto Alegre/RS/BR, frente a inóculos bacterianos padronizados (American Type Culture Collection - ATCC), respectivamente Staphylococcus aureus (25923), Enterococcus faecalis (19433), Salmonella enteritidis (13076) e Escherichia coli (11229), em doses-desafio = 10(7) UFC mL-1. Quatro destas plantas, pimenta calabresa ("pool" Capsicum sp), pimenta-de-jardim (C.annuum), pimenta dedo-de-moça (C. baccatum) e pimenta malagueta (C. frutescens), apresentaram atividades de inibição e inativação seletivas, em ordem decrescente, para salmonela, coliforme fecal, enterococo e estafilococo. As demais, pimenta cambuci (C. baccatum) e os pimentões (C. annuum) amarelo, verde e vermelho, apresentaram nenhuma atividade. Discute-se a validade da ferramenta etnográfica na prospecção de fatores de proteção anti-bacteriana em plantas, bem como a influência da inibição/inativação na preditividade do diagnóstico bacteriológico.

The intensity of bacterial inhibition activity (IINIB) and the intensity of bacterial inactivation activity (IINAB) of extracts of eight peppers of the genus Capsicum, ethnographically located in the metropolitan region of Porto Alegre, Rio Grande do Sul State, Brazil, were assessed in vitro through Dilution Tests in Multiple Tube Series against standardized bacterial inocula (American Type Culture Collection - ATCC), Staphylococcus aureus (25923), Enterococcus faecalis (19433), Salmonella enteritidis (13076), and Escherichia coli (11229), respectively, at challenge doses = 10(7) CFU mL-1. Four of these species, cayenne pepper (Capsicum sp pool), garden pepper (C. annuum), ají pepper (C. baccatum), and malagueta pepper (C. frutescens), had selective inhibition and inactivation activities, in decreasing order, to salmonella, fecal coliforms, enterococcus and staphylococcus. The remaining ones, cambuci pepper (C. baccatum) and yellow, green and red sweet peppers (C. annuum) had no activity. The validity of the ethnographic tool in the exploration of antibacterial protection factors from plants, as well as the influence of inhibition/inactivation in the bacteriological diagnosis predictability, is discussed.

Inibição e inativação in vitro de Salmonella spp. com extratos de plantas com indicativo etnográfico medicinal ou condimentar / In vitro inhibition and inactivation activity of Salmonella spp. by plant extracts with spicy or medicinal ethnographic indicative

Determinou-se in vitro a intensidade de atividade de inibição bacteriana e a intensidade de atividade de inativação bacteriana, por meio de testes de diluição e suspensão em sistema de tubos múltiplos, de diferentes extratos, aquosos ou alcoólicos/hidroalcoólicos, de 86 plantas com indicativo etnográfico medicinal ou condimentar acessadas na região metropolitana de Porto Alegre, Rio Grande do Sul, frente a Salmonella spp., ou S. enteritidis ATCC nº 13076, ou S. cholera-suis ATCC nº10708, ou S. gallinarum CPVDF-SAA/RS/BR, em doses-desafio de <10(7)UFC.mL-1. Cinquenta plantas apresentaram alguma atividade seletiva antissalmonela, e 37 nenhuma atividade. Discute-se a validade da ferramenta etnográfica na prospecção de fatores de proteção antibacteriana em plantas, bem como a influência da inibição/inativação na preditividade do diagnóstico bacteriológico.

The in vitro intensity of bacterial inhibition activity of diverse extracts (aquous, alcoholic, or hydroalcoholic) from 86 plants with medicinal and spicy ethnographic indicative, from Porto Alegre City, Rio Grande do Sul State, Brazil, were determinated in Salmonella spp. (S. enteritidis ATCC n.13076, S. cholera-suis ATCC n. 10708, or S. gallinarum CPVDF-SAA/RS/BR) in challenge dose <10(7)CFU.mL-1. Extracts of fifty plants presented some anti-Salmonella selective activity, while the other 36 extracts presented no activity. It is discussed the validity of ethnographic search instruments in the prospection of anti-bacterial protection factors in plants, as well the influence of inhibition/inactivation results in the predictivity of bacteriological diagnostic.

Atividade anti-estafilócica em extratos de plantas com indicativo medicinal ou condimentar / Anti-staphylococci activity in extracts from plants presenting medicinal and spice indicative

O gênero Staphylococcus permanece em evidência no contexto das doenças transmissíveis considerando-se os portadores humanos e animais como a sua principal fonte de infecção. Por sua vez, a pesquisa de fatores de proteção antibacteriana em recursos naturais como plantas com indicativo medicinal ou condimentar vem sendo recomendado como prioridade pelas Conferências Mundiais de Saúde. A presente revisão descreve a triagem in vitro, através de testes de diluição em sistema de tubos múltiplos, de atividade anti-estafilococo de diferentes extratos de 80 plantas com indicativo etnográfico medicinal, condimentar ou aromático, acessadas na região metropolitana de Porto Alegre, RS. Demonstrou-se atividade seletiva em 39 espécies, enquanto as demais 41 não apresentaram atividade.

The genus Staphylococcus remains in evidence regarding transmissible diseases since human and animal carriers are its main infection source. Studies on antibacterial protection factors in natural resources, such as plants presenting medicinal or spice indicative, have been recommended by World Health Conferences as a priority. Dilution tests in multiple tube system were used in this review to describe the in vitro sorting of anti-staphylococci activity of different extracts from 80 plants presenting medicinal, spice or aromatic ethnographic indicatives at the metropolitan region of Porto Alegre, Rio Grande do Sul State, Brazil. Selective activity was observed for 39 species, whereas the remaining 41 species did not show activity.

Orientational distributions and nematic order of rodlike magnetic nanoparticles in dispersions.

Using small-angle neutron scattering (SANS), we have investigated the orientational order of iron nanoparticles dispersed in cyclohexanone. The particles have rodlike shape and size distributions with an average length of 200 nm and an average diameter of 25 nm. SANS shows an anisotropy, which is a measure of orientational order, in magnetic dispersions with a volume fraction of 3.2% and 3.9% iron particles in shear flow and/or magnetic field. The scattering anisotropy can be fitted by a model assuming an Onsager distribution of the orientation of the particles in shear flow. The orientational distribution of particles oriented by a magnetic field can be described by a different model assuming the Maier-Saupe orientational distribution for uniaxial ferromagnetic particles. The orientational distribution parameter m for the Maier-Saupe distribution or alpha for the Onsager distribution and the orientational order parameter S have been determined at shear rates gamma[over ] of to 0-4000 s(-1) and in magnetic fields of 0-18 mT. The S values indicate that the particles start to orient either in a shear flow of 100 s(-1) or in a magnetic field of 6 mT. Applying only shear results in an orientational order, with the dispersion returning to the disordered state when the shear rate is decreased to zero. In sharp contrast, application of magnetic fields greater than 6 mT results in orientational order in the field-increasing cycle, and two-thirds of the orientational order remains when the field is decreased to zero. This shows that the order in a magnetic field is different from the order in a shear flow, the action of magnetizing the particles along a certain direction is irreversible, and the orientational order parameter exhibits hysteresis.

Viabilidade de ovos de Fasciola hepatica de bovinos em sistema de biodigestão anaeróbia / Viability of bovine's Fasciola hepatica eggs in a system of anaerobic biodigestion

Cattle Fasciola hepatica egg-viability was studied in closed system of anaerobic biodigestion. Two thirds of a biodigestor were filled with liquefied manure from eight Hosltein cows, naturally infected. For 10 consecutive weeks of observation, the biodigestor internal temperature ranged from 24ºC to 28ºC, and external temperature from 26ºC to 33ºC. All the effluent samples showed a constant pH of 7. The samples of the effluent were weekly collected, in a total of 10 and submitted to techniques to detect the presence and viability of the trematode eggs. It was detected a 42-day egg viability. Therefore, liquefied cattle manure containing Fasciola hepatica eggs must remain under anaerobic biodigestion for at least 42 days before its safe return to the environment.

A major lung cancer susceptibility locus maps to chromosome 6q23-25.

Lung cancer is a major cause of death in the United States and other countries. The risk of lung cancer is greatly increased by cigarette smoking and by certain occupational exposures, but familial factors also clearly play a major role. To identify susceptibility genes for familial lung cancer, we conducted a genomewide linkage analysis of 52 extended pedigrees ascertained through probands with lung cancer who had several first-degree relatives with the same disease. Multipoint linkage analysis, under a simple autosomal dominant model, of all 52 families with three or more individuals affected by lung, throat, or laryngeal cancer, yielded a maximum heterogeneity LOD score (HLOD) of 2.79 at 155 cM on chromosome 6q (marker D6S2436). A subset of 38 pedigrees with four or more affected individuals yielded a multipoint HLOD of 3.47 at 155 cM. Analysis of a further subset of 23 multigenerational pedigrees with five or more affected individuals yielded a multipoint HLOD score of 4.26 at the same position. The 14 families with only three affected relatives yielded negative LOD scores in this region. A predivided samples test for heterogeneity comparing the LOD scores from the 23 multigenerational families with those from the remaining families was significant (P=.007). The 1-HLOD multipoint support interval from the multigenerational families extends from C6S1848 at 146 cM to 164 cM near D6S1035, overlapping a genomic region that is deleted in sporadic lung cancers as well as numerous other cancer types. Parametric linkage and variance-components analysis that incorporated effects of age and personal smoking also supported linkage in this region, but with somewhat diminished support. These results localize a major susceptibility locus influencing lung cancer risk to 6q23-25.

Shear- and magnetic-field-induced ordering in magnetic nanoparticle dispersion from small-angle neutron scattering.

Small-angle neutron scattering experiments have been performed to investigate orientational ordering of a dispersion of rod-shaped ferromagnetic nanoparticles under the influence of shear flow and static magnetic field. In this experiment, the flow and flow gradient directions are perpendicular to the direction of the applied magnetic field. The scattering intensity is isotropic in zero-shear-rate or zero-applied-field conditions, indicating that the particles are randomly oriented. Anisotropic scattering is observed both in a shear flow and in a static magnetic field, showing that both flow and field induce orientational order in the dispersion. The anisotropy increases with the increase of field and with the increase of shear rate. Three states of order have been observed with the application of both shear flow and magnetic field. At low shear rates, the particles are aligned in the field direction. When increasing shear rate is applied, the particles revert to random orientations at a characteristic shear rate that depends on the strength of the applied magnetic field. Above the characteristic shear rate, the particles align along the flow direction. The experimental results agree qualitatively with the predictions of a mean field model.

Colimetrias como indicadores de qualidade de pescado artesanal do Lago Guaíba, em Porto Alegre, RS / Fish colimetry as fish in Lago Guaíba, in Porto Alegre, RS

Foram avaliadas quanto a presença de coliformes totais, fecais e Escherichia coli 43 amostras de Piava Leporinus obtusidens (Valenciennes, 1847) e 43 de Pintado Pimelodus maculatus (Lacèpéde, 1803) do Lago Guaíba, P. Alegre, RS. Os níveis colimétricos totais e fecais foram determinados segundo Brasil. Ministério da Agricultura (1992), utilizando-se a técnica dos Números Mais Prováveis. A presença de E. coli foi confirmada pelo teste do Indol. A média dos coliformes fecais é significativamente maior nos Pintados quando comparado com as Piavas (p= 0,043). Em relaçäo a E. coli existe diferença significativa entre a Piava e o Pintado (p= 0,040). O Pintado é significativamente mais contaminado que a Piava. Das Piavas analisadas 4,65 por cento estavam acima dos níveis permitidos pela legislaçäo brasileira para coliformes fecais e dos Pintados, 11,62 por cento das amostras também estavam fora destes limites. Em 23,25 por cento das amostras de Piava e 44,18 por cento de Pintados foi constatada a presença de E. coli. Os valores encontrados para enumeraçäo de coliformes totais revelam a importância de um maior controle na higiene do pescado.

Intron/exon organization and polymorphisms of the PLK3/PRK gene in human lung carcinoma cell lines.

PLK3/PRK, a conserved polo family protein serine/threonine kinase, plays a significant role at the onset of mitosis and mitotic progression. Recently, PLK3/PRK has been shown to induce apoptosis when overexpressed in cell lines and is also implicated in cell proliferation and tumor development. Forty lung tumor cell lines were used for single-strand confirmation polymorphism (SSCP) analysis and DNA sequencing to examine the mutational status of PLK3/PRK. No missense or nonsense mutations were revealed in the lung carcinoma cell lines examined. However, three polymorphisms were identified as: a G to A at position 720, an A to G at 1053, and a G to C at 1275. Intron/exon boundaries were determined by amplification of genomic DNA with PLK3/PRK exon-specific primers. The amplification products with increased size relative to the cDNA were sequenced. Fifteen exons throughout the open reading frame were characterized. None of the introns were exceptionally large, typically ranging from 100-300 basepairs in length. These results suggest that although PLK3/PRK expression is downregulated in a majority of lung carcinoma samples, mutational inactivation of the coding sequence of the PLK3/PRK gene appears to be a rare event in lung cancer.

Functional genomics of oxidant-induced lung injury.

In summary, acute lung injury is a severe (>40% mortality) respiratory disease associated with numerous precipitating factors. Despite extensive research since its initial description over 30 years ago, questions remain about the basic pathophysiological mechanisms and their relationship to therapeutic strategies. Histopathology reveals surfactant disruption, epithelial perturbation and sepsis, either as initiating factors or as secondary complications, which in turn increase the expression of cytokines that sequester and activate inflammatory cells, most notably, neutrophils. Concomitant release of reactive oxygen and nitrogen species subsequently modulates endothelial function. Together these events orchestrate the principal clinical manifestations of the syndrome, pulmonary edema and atelectasis. To better understand the gene-environmental interactions controlling this complex process, we examined the relative sensitivity of inbred mouse strains to acute lung injury induced by ozone, ultrafine PTFE, or fine particulate NiSO4 (0.2 microm MMAD, 15-150 microg/m3). Measuring survival time, protein and neutrophils in bronchoalveolar lavage, lung wet: dry weight, and histology, we found that these responses varied between inbred mouse strains, and susceptibility is heritable. To assess the molecular progression of NiSO4-induced acute lung injury, temporal relationships of 8734 genes and expressed sequence tags were assessed by cDNA microarray analysis. Clustering of co-regulated genes (displaying similar temporal expression patterns) revealed the altered expression of relatively few genes. Enhanced expression occurred mainly in genes associated with oxidative stress, anti-proteolytic function, and repair of the extracellular matrix. Concomitantly, surfactant proteins and Clara cell secretory protein mRNA expression decreased. Genome wide analysis of 307 mice generated from the backcross of resistant B6xA F1 with susceptible A strain identified significant linkage to a region on chromosome 6 (proposed as Aliq4) and suggestive linkages on chromosomes 1, 8, and 12. Combining of these QTLs with two additional possible modifying loci (chromosome 9 and 16) accounted for the difference in survival time noted in the A and B6 parental strains. Combining these findings with those of the microarray analysis has enabled prioritization of candidate genes. These candidates, in turn, can be directed to the lung epithelium in transgenic mice or abated in inducible and constitutive gene-targeted mice. Initial results are encouraging and suggest that several of these mice vary in their susceptibility to oxidant-induced lung injury. Thus, these combined approaches have led to new insights into functional genomics of lung injury and diseases.

Pathogenomic mechanisms for particulate matter induction of acute lung injury and inflammation in mice.

To begin identifying genes controlling individual susceptibility to particulate matter, responses of inbred mouse strains exposed to nickel sulfate (NiSO4*) were compared with those of mice exposed to ozone (O3) or polytetrafluoroethylene (PTFE). The A strain was sensitive to NiSO4-induced lung injury (quantified by survival time), the C3H/He (C3) strain and several other strains were intermediate in their responses, and the C57BL/6 (B6) strain was resistant. The strains showed a pattern of response similar to the patterns of response to O3 and PTFE. The phenotype of A x B6 offspring (B6AF1) resembled that of the resistant B6 parental strain, with strains exhibiting sensitivity in the order A > C3 > B6 = B6AF1. Pathology was comparable for the A and B6 mice, and exposure to NiSO4 at 15 microg/m3 produced 20% mortality in A mice. Strain sensitivity for the presence of protein or neutrophils in lavage fluid differed from strain sensitivity for survival time, suggesting that they are not causally linked but are controlled by an independent gene or genes. In the B6 strain, exposure to nickel oxide (NiO) by instillation (40 to 1000 nm) or inhalation (50 nm) produced no changes, whereas inhalation of NiSO4 (60 or 250 nm) increased lavage proteins and neutrophils. Complementary DNA (cDNA) microarray analysis with 8,734 sequence-verified clones revealed a temporal pattern of increased oxidative stress, extracellular matrix repair, cell proliferation, and hypoxia, followed by a decrease in surfactant-associated proteins (SPs). Certain expressed sequence tags (ESTs), clustered with known genes, suggest possible coregulation and novel roles in pulmonary injury. Finally, locus number estimation (Wright equation) and a genomewide analysis suggested 5 genes could explain the survival time and identified significant linkage for a quantitative trait locus (QTL) on chromosome 6, Aliq4 (acute lung injury QTL4). Haplotype analysis identified an allelic combination of 5 QTLs that could explain the difference in sensitivity to acute lung injury between parental strains. Positional candidate genes for Aliq4 include aquaporin-1 (Aqp1), SP-B, and transforming growth factor-alpha (TGF-alpha). Transgenic mice expressing TGF-alpha were rescued from NiSO4 injury (that is, they had diminished SP-B loss and increased survival time). These findings suggest that NiSO4-induced acute lung injury is a complex trait controlled by at least 5 genes (all possibly involved in cell proliferation and surfactant function). Future assessment of these susceptibility genes (including evaluations of human synteny and function) could provide valuable insights into individual susceptibility to the adverse effects of particulate matter.