Cyclosporin A pretreatment in a rat model of warm ischaemia/reperfusion injury

Background/Aims: These studies investigated the role of apoptosis following ischaemia/reperfusion (I/R) injury to the liver and the effect of pretreatment with Cyclosporin A. Methods: Male Sprague-Dawley rats received 30 min of warm ischaemia followed by a period of reperfusion of 6 h. Rats were given olive oil or Cyclosporin A (30 mg/kg p.o.) the day before surgery. Neutrophil numbers were assessed in haematoxylin-eosin-stained sections of liver. In situ staining of sections using TdT-mediated dUTP-fluoreseein nick-end labelling was carried out to determine the extent of apoptosis, followed by electron microscopy. Semi-quantitative polymerase chain reaction (PCR) analysis of the transcript for Fas antigen was performed. Results and Conclusions: High levels of apoptosis were observed in I/R injury, which were greatly ameliorated in Cyclosporin A-pretreated groups. PCR analysis indicated a reduction in the level of expression of Fas transcript in Cyclosporin A-treated rats. Histological analysis showed a significant increase in the number of neutrophils infiltrating I/R-injured tissue (62 +/- 10.69, it = 16), which was markedly reduced by Cyclosporin A pretreatment (16 +/- 7, n = 6, P < 0.05). These results indicate a role of parenchymal apoptosis in the pathogenesis of I/R injury, which occurs in association with neutrophil infiltration, both of which can be significantly reduced by Cyclosporin A pretreatment. (C) 2002 European Association for the Study of the Liver. Published by Elsevier Science B.V. All rights reserved.

Treatment of non-resectable hepatocellular carcinoma with autologous tumor-pulsed dendritic cells

Background: The response of hepatocellular carcinoma (HCC) to therapy is often disappointing and new modalities of treatment are clearly needed. Active immunotherapy based on the injection of autologous dendritic cells (DC) co-cultured ex vivo with tumor antigens has been used in pilot studies in various malignancies such as melanoma and lymphoma with encouraging results. Methods: In the present paper, the preparation and exposure of patient DC to autologous HCC antigens and re-injection in an attempt to elicit antitumor immune responses are described. Results: Therapy was given to two patients, one with hepatitis C and one with hepatitis B, who had large, multiple HCC and for whom no other therapy was available. No significant side-effects were observed. The clinical course was unchanged in one patient, who died a few months later. The other patient, whose initial prognosis was considered poor, is still alive and well more than 3 years later with evidence of slowing of tumor growth based on organ imaging. Conclusions: It is concluded that HCC may be a malignancy worthy of DC trials and sufficient details in the present paper are given for the protocol to be copied or modified. (C) 2002 Blackwell Publishing Asia Pty Ltd.

A new species of Phyllurus (Lacertilia : Gekkonidae) and a revised phylogeny and key for the Australian leaf-tailed geckos

Phyllurus gulbaru, sp. nov., is a highly distinct species of leaf-tailed gecko restricted to rocky rainforest of Pattersons Gorge, north-west of Townsville. The possession of a cylindrical, non-depressed, tapering original and regenerated tail separates P. gulbaru from all congeners except P. caudiannulatus. From this species P. gulbaru is separated by having a partially divided, as opposed to fully divided, rostral scale. Furthermore, the very small spinose body tubercles of P. gulbaru are in marked contrast to the large spinose body scales of P. caudiannulatus. An analysis of 729 bp of mitochondrial 12S rRNA and cytochrome b genes reveals P. gulbaru to be a deeply divergent lineage with closer affinities to mid-east Queensland congeners than the geographically neighbouring P. amnicola on Mt Elliot. In conservation terms, P. gulbaru is clearly at risk. Field surveys of Pattersons Gorge and the adjacent ranges indicate that this species is restricted to a very small area of highly fragmented habitat, of which only a small proportion receives a degree of protection in State forest. Further, there is ongoing, unchecked destruction of dry rainforest habitat by fire. Under current IUCN criteria, P. gulbaru warrants an Endangered ( B1, 2) listing.

Predicting the HER2 status of breast cancer from basic histopathology data: an analysis of 1500 breast cancers as part of the HER2000 International Study

The tests that are currently available for the measurement of overexpression of the human epidermal growth factor-2 (HER2) in breast cancer have shown considerable problems in accuracy and interlaboratory reproducibility. Although these problems are partly alleviated by the use of validated, standardised 'kits', there may be considerable cost involved in their use. Prior to testing it may therefore be an advantage to be able to predict from basic pathology data whether a cancer is likely to overexpress HER2. In this study, we have correlated pathology features of cancers with the frequency of HER2 overexpression assessed by immunohistochemistry (IHC) using HercepTest (Dako). In addition, fluorescence in situ hybridisation (FISH) has been used to re-test the equivocal cancers and interobserver variation in assessing HER2 overexpression has been examined by a slide circulation scheme. Of the 1536 cancers, 1144 (74.5%) did not overexpress HER2. Unequivocal overexpression (3+ by IHC) was seen in 186 cancers (12%) and an equivocal result (2+ by IHC) was seen in 206 cancers (13%). Of the 156 IHC 3+ cancers for which complete data was available, 149 (95.5%) were ductal NST and 152 (97%) were histological grade 2 or 3. Only 1 of 124 infiltrating lobular carcinomas (0.8%) showed HER2 overexpression. None of the 49 'special types' of carcinoma showed HER2 overexpression. Re-testing by FISH of a proportion of the IHC 2+ cancers showed that only 25 (23%) of those assessable exhibited HER2 gene amplification, but 46 of the 47 IHC 3+ cancers (98%) were confirmed as showing gene amplification. Circulating slides for the assessment of HER2 score showed a moderate level of agreement between pathologists (kappa 0.4). As a result of this study we would advocate consideration of a triage approach to HER-2 testing. Infiltrating lobular and special types of carcinoma may not need to be routinely tested at presentation nor may grade 1 NST carcinomas in which only 1.4% have been shown to overexpress HER2. Testing of these carcinomas may be performed when HER2 status is required to assist in therapeutic or other clinical/prognostic decision-making. The highest yield of HER2 overexpressing carcinomas is seen in the grade 3 NST subgroup in which 24% are positive by IHC. (C) 2003 Elsevier Science Ltd. All rights reserved.

Fas ligand and tumour counter-attack in colorectal cancer stratified according to microsatellite instability status

Expression of membrane-bound Fas ligand (FasL) by colorectal cancer cells may allow the development of an immune-privileged site by eliminating incoming tumour-infiltrating lymphocytes (TILs) in a Fas-mediated counter-attack. Sporadic colorectal cancer can be subdivided into three groups based on the level of DNA microsatellite instability (NISI). High-level NISI (NISI-High) is characterized by the presence of TILs and a favourable prognosis, while microsatellite-stable (MSS) cancers are TIL-deficient and low-level MSI (MSI-Low) is associated with an intermediate TIL density. The purpose of this study was to establish the relationship between MSI status and FasL expression in primary colorectal adenocarcinoma. Using immunohistochemistry and a selected series of 101 cancers previously classified as 31 MSI-High, 30 NISI-Low, and 40 MISS, the present study sought to confirm the hypothesis that increased TIL density in MSI-High cancers is associated with low or absent membrane-bound FasL expression, while increased FasL in MSS cancers allows the killing of host TILs. TUNEL/CD3 double staining was also used to determine whether MSS cancers contain higher numbers of apoptotic TILs in vivo than MSI-High or MSI-Low cancers. Contrary to the initial hypothesis, it was found that MSI-High cancers were associated with higher FasL expression (p = 0.04) and a stronger intensity of FasL staining (p = 0.007). In addition, mucinous carcinomas were independently characterized by increased FasL expression (p = 0.03) and staining intensity (p = 0.0005). Higher FasL expression and staining intensity did not correlate with reduced TIL density or increased numbers of apoptotic TILs. However, consistent with the hypothesis that curtailment of the host anti-tumour immune response contributes to the poor prognosis in MSS cancers, it was found that apoptotic TILs were most abundant in MSS carcinomas and metastatic Dukes' stage C or D tumours (p = 0.004; p = 0.046 respectively). This study therefore suggests that MSS colorectal cancers are killing incoming TILs in an effective tumour counter-attack, but apparently not via membrane-bound FasL. Copyright (C) 2003 John Wiley Sons, Ltd.

Differences in mouse strain influence leukocyte and immunoglobulin phenotype response to Porphyromonas gingivalis

This study examined the nature of the infiltrating cells in Porphyromonas gingivalis-induced lesions and immunoglobulins in the serum samples of BALB/c (H-2(d)), C57BL6 (H-2(b)), DBA/2J (H-2(d)) and CBA/CaH (H-2(k)) mice. Mice were immunized intraperitoneally with P. gingivalis outer membrane antigens or sham-immunized with phosphate-buffered saline followed by subcutaneous challenge with live organisms 1 week after the final immunization. The resulting skin abscesses were excised 7 days later, cryostat sections cut and an immunoperoxidase method used to detect the presence of CD4(+) and CD8(+) T-cell subsets, CD14(+) macrophages and CD19(+) B cells. Peroxidase positive neutrophils and IgG1- and IgG2a-producing plasma cells were also identified. Anti P. gingivalis IgG1 and IgG2a subclass antibodies were determined in serum obtained by cardiac puncture. Very few CD8(+) T cells and CD19(+) B cells were found in any of the lesions. The percentages of CD4(+) cells, CD14(+) cells and neutrophils were similar in lesions of immunized BALB/c and C57BL6 mice, with a trend towards a higher percentage of CD14(+) cells in sham-immunized mice. The percentage of CD14(+) cells was higher than that of CD4(+) cells in immunized compared with sham-immunized DBA/2J mice. The percentages of CD4(+) and CD14(+) cells predominated in immunized CBA/CaH mice and CD4(+) cells in sham-immunized CBA/CaH mice. The percentage of neutrophils in immunized CBA/CaH mice was significantly lower than that of CD14(+) cells and CD4(+) cells in sham-immunized mice. IgG1(+) plasma cells were more dominant than IgG2a(+) cells in immunized BALB/c, C57BL6 and DBA/2J mice, whereas IgG2a(+) plasma cells were more obvious in sham-immunized mice. IgG2a(+) plasma cells were predominant in immunized and sham-immunized CBA/CaH mice. In the serum, specific anti-P. gingivalis IgG2a antibody levels (Th1 response) were higher than IgG1 levels (Th2 response) in sham-immunized CBA/CaH and DBA/2J mice. In immunized BALB/c mice, IgG2a levels were lower than IgG1 levels, while IgG2a levels were higher in immunized C57BL6 mice. In conclusion, this study has shown differences in the proportion of infiltrating leukocytes and in the subclasses of immunoglobulin produced locally and systemically in response to P. gingivalis in different strains of mice, suggesting a degree of genetic control over the response to P. gingivalis.

Candidatus "Scalindua brodae", spec. nov., Candidatus "Scalindua wagneri", spec. nov., two new species of anaerobic ammonium oxidizing bacteria

Anaerobic ammonium oxidation (anammox) is both a promising process in wastewater treatment and a long overlooked microbial physiology that can contribute significantly to biological nitrogen cycling in the world's oceans. Anammox is mediated by a monophyletic group of bacteria that branches deeply in the Planctomycetales. Here we describe a new genus and species of anaerobic ammonium oxidizing planctomycetes, discovered in a wastewater treatment plant (wwtp) treating landfill leachate in Pitsea, UK. The biomass from this wwtp showed high anammox activity (5.0 +/- 0.5 nmol/mg protein/min) and produced hydrazine from hydroxylamine, one of the unique features of anammox bacteria. Eight new planctomycete 16S rRNA gene sequences were present in the 16S rRNA gene clone library generated from the biomass. Four of these were affiliated to known anammox 16S rRNA gene sequences, but branched much closer to the root of the planctomycete line of descent. Fluorescence in situ hybridization (FISH) with oligonucleotide probes specific for these new sequences showed that two species (belonging to the same genus) together made up > 99% of the planctomycete population which constituted 20% of the total microbial community. The identification of these organisms as typical anammox bacteria was confirmed with electron microscopy and lipid analysis. The new species, provisionally named Candidatus Scalindua brodae and Scalindua wagneri considerably extend the biodiversity of the anammox lineage on the 16S rRNA gene level, but otherwise resemble known anammox bacteria. Simultaneously, another new species of the same genus, Candidatus Scalindua sorokinii, was detected in the water column of the Black Sea, making this genus the most widespread of all anammox bacteria described so far.

Localization of the Epstein-Barr virus protein LMP 1 to exosomes

The Epstein-Barr virus latent membrane protein (LMP 1) functions as a constitutively active signalling molecule and associates in lipid rafts clustered with other signalling molecules. Using immunofluorescent confocal microscopy, LMP 1 was shown to have an heterogeneous distribution among individual cells which was not related to the cell cycle stage. LMP 1 was shown to localize to intracellular compartments in cells other than the plasma membrane, Co-labelling of cells with both an LIMP 1 antibody and an antibody to the Golgi protein GS15 revealed that the intracellular LMP 1 partly co-localized with the Golgi apparatus. Further confirmation of intracellular LMP 1 localization was obtained by immunoelectron microscopy with rabbit polyclonal LIMP 1 antibodies and cryosectioning. As well as being present in intracellular foci, LMP 1 co-localized in part with MHC-II and was present on exosomes derived from a lymphoblastoid cell line. Preparations of LMP 1 containing exosomes were shown to inhibit the proliferation of peripheral blood mononuclear cells, suggesting that LIMP 1 could be involved in immune regulation. This may be of particular relevance in EBV-associated tumours such as nasopharyngeal carcinoma and Hodgkin's disease, as LMP 1-containing exosomes may be taken up by infiltrating T-lymphocytes, where LMP 1 could exert an anti-proliferative effect, allowing the tumour cells to evade the immune system.

Early pregnancy factor treatment suppresses the inflammatory response and adhesion molecule expression in the spinal cord of SJL/J mice with experimental autoimmune encephalomyelitis and the delayed-type hypersensitivity reaction to trinitrochlorobenzene in normal BALB/c mice

Early pregnancy factor (EPF) is a secreted protein, present in serum during early pregnancy and essential for maintaining viability of the embryo. It is a homologue of chaperonin 10 (Cpn10) but, unlike Cpn10, it has an extracellular role. EPF has immunosuppressive and growth regulatory properties. Previously we have reported the preparation of recombinant EPF (rEPF) and shown that treatment with rEPF will suppress clinical signs of MBP-EAE in Lewis rats and PLP-EAE in SJL/J mice. In the present study, these findings have been extended to investigate possible mechanisms involved in the action of EPF. Following treatment of mice with rEPF from the day of inoculation, there were fewer infiltrating CD3+ and CD4+ cells in the parenchyma of the spinal cord during the onset of disease and after the initial episode, compared with mice treated with vehicle. Expression of the integrins LFA-1, VLA-4 and Mac-1 and of members of the immunoglobulin superfamily of adhesion molecules ICAM-1 and VCAM-1 was suppressed in the central nervous system (CNS) following rEPF treatment. The expression of PECAM-1 was not affected. To determine if rEPF suppressed T cell activation in the periphery, the delayed-type hypersensitivity (DTH) reaction of normal BALB/c mice to trinitrochlorobenzene (TNCB) following treatment with rEPF was studied. The results showed that treatment with rEPF suppressed the DTH reaction, demonstrating the ability of EPF to downregulate the cell-mediated immune response. These results indicate that suppression of immunological mechanisms by rEPF plays a major role in the reduction of clinical signs of disease in experimental autoimmune encephalomyelitis (EAE). (C) 2003 Elsevier Science B.V. All rights reserved.

Early pregnancy factor suppresses the infiltration of lymphocytes and macrophages in the spinal cord of rats during experimental autoimmune encephalomyelitis but has no effect on apoptosis

Early pregnancy factor (EPF) is a secreted protein with immunosuppressive and growth factor properties that has been shown to suppress acute experimental autoimmune encephalomyelitis (EAE) induced with myelin basic protein (MBP) in Lewis rats. EAE is associated with infiltration of the central nervous system (CNS) with inflammatory cells. Spontaneous recovery involves the loss of T lymphocytes from the CNS and the selective apoptosis of Vbeta8.2(+) cells. In the present study, T cell, macrophage (CD11b/c(+)) and B cell (CD45RA(+)) populations in spinal cord and popliteal lymph nodes (LN) of Lewis rats with EAE were quantitated and apoptosis was studied. Rats were treated with EPF or vehicle. Following treatment on day 14 after inoculation with MBP, neither 1 x 100 mug nor 2 x 100 mug doses of EPF affected the total number of cells infiltrating the spinal cord on day 15, although the higher dose caused a decrease in the number of CD5(+) and CD11b/c(+) cells. Treatment with 2 x 100 mug/day from days 10 to 14 decreased the total number of infiltrating cells, and the numbers of CD5(+), CD11b/c(+) and CD45RA(+) cells. Apoptosis was unaffected. No alteration on the number or type of inflammatory cells in the popliteal LN was observed after treatment on days 10-14. However, treatment with EPF from days 0 to 11 increased the total number of T and B cells and CD5(+) T cells found on day 12 in the LN. Similarly, there was an increase in the frequency of MBP-reactive cells in the LN as determined by limiting dilution analysis. These results suggest that EPF treatment reduces the numbers of lymphocytes and macrophages in the CNS, possibly through an effect on cell trafficking. (C) 2003 Elsevier B.V. All rights reserved.

Suppressor of cytokine signalling gene expression is elevated in breast carcinoma

Cytokines are important for breast cell function, both as trophic hormones and as mediators of host defense mechanisms against breast cancer. Recently, inducible feedback suppressors of cytokine signalling (SOCS/JAB/SSI) have been identified, which decrease cell sensitivity to cytokines. We examined the expression of SOCS genes in 17 breast carcinomas and 10 breast cancer lines, in comparison with normal tissue and breast lines. We report elevated expression of SOCS-1-3 and CIS immunoreactive proteins within in situ ductal carcinomas and infiltrating ductal carcinomas relative to normal breast tissue. Significantly increased expression of SOCS-1-3 and CIS transcripts was also shown by quantitative in situ hybridisation within both tumour tissue and reactive stroma. CIS transcript expression was elevated in all 10 cancer lines, but not in control lines. However, there was no consistent elevation of other SOCS transcripts. CIS protein was shown by immunoblot to be present in all cancer lines at increased levels, mainly as the 47 kDa ubiquitinylated form. A potential proliferative role for CIS overexpression is supported by reports that CIS activates ERK kinases, and by strong induction in transient reporter assays with an ERK-responsive promoter. The in vivo elevation of SOCS gene expression may be part of the host/tumour response or a response to autocrine/paracrine GH and prolactin. However, increased CIS expression in breast cancer lines appears to be a specific lesion, and could simultaneously shut down STAT 5 signalling by trophic hormones, confer resistance to host cytokines and increase proliferation through ERK kinases.

Anomalous water absorption in porous materials

The absorption of fluid by unsaturated, rigid porous materials may be characterized by the sorptivity. This is a simple parameter to determine and is increasingly being used as a measure of a material's resistance to exposure to fluids (especially moisture and reactive solutes) in aggressive environments. The complete isothermal absorption process is described by a nonlinear diffusion equation, with the hydraulic diffusivity being a strongly nonlinear function of the degree of saturation of the material. This diffusivity can be estimated from the sorptivity test. In a typical test the cumulative absorption is proportional to the square root of time. However, a number of researchers have observed deviation from this behaviour when the infiltrating fluid is water and there is some potential for chemo-mechanical interaction with the material. In that case the current interpretation of the test and estimation of the hydraulic diffusivity is no longer appropriate. Kuntz and Lavallee (2001) discuss the anomalous behaviour and propose a non-Darcian model as a more appropriate physical description. We present an alternative Darcian explanation and theory that retrieves the earlier advantages of the simple sorptivity test in providing parametric information about the material's hydraulic properties and allowing simple predictive formulae for the wetting profile to be generated.

Avalia????o das pol??ticas p??blicas na Fran??a

Ap??s um longo per??odo de discuss??o sobre a reforma do Estado, a Fran??a, em 1990, implementou um sistema interdepartamental para a avalia????o das pol??ticas p??blicas. O aspecto interessante nesta reforma e o meio pelo qual um pequeno grupo de pessoas, de diferentes institui????es e com experi??ncias pol??ticas distintas, convenceu-se da import??ncia da avalia????o das pol??ticas p??blicas e se conscientizou dos diversos problemas relativos a esta quest??o. Finalmente, com uma atitude bipartid??ria, este grupo foi capaz de criar uma realidade que modifica profundamente o sistema legislativo, tanto no que se refere ao processo decis??rio quanto ao sistema de implementa????o das pol??ticas. Neste trabalho, o autor descreve os passos do debate e os recursos das diferentes propostas que acabaram por se transformar no projeto de reforma e na sua implementa????o.

Crítica da razão excludente : desvelando novas racionalidades a partir das pesquisas em educação especial numa perspectiva inclusiva

O objetivo desta tese é produzir uma reflexão hermenêutica e fenomenológica acerca dos saberes e fazeres que vêm sido produzidos a partir de pesquisas que tratam da inclusão de alunos com necessidades especiais nas escolas de ensino regular. Buscou-se uma pesquisa de natureza bibliográfica que debruçou-se na produção de conhecimento relativa a dissertações e teses que foram produzidas no PPGE-UFES entre os anos de 2000 e 2010 que buscaram analisar os saberes-fazeres inclusivos que vêm se constituindo a partir da inserção de alunos com necessidades especiais nos espaços-tempos das escolas comuns. Procurou-se num primeiro momento traçar o estado da arte de tais trabalhos de pesquisa para, posteriormente, compreendê-los numa dimensão filosófica. Como resultado concluiu-se que estas pesquisas apontam movimentos profundamente ambíguos e paradoxais, revelando uma profunda crise de um modelo de racionalidade excludente que encontra-se enraizado em nossa cultura e no cotidiano das escolas. Frente a esse quadro, procura-se compreender, a partir das pistas e rastros deixados pelas dissertações e teses pesquisadas, como vêm se configurando outras formas de racionalidades que instauram novas formas de ensinar-aprender conjugadas aos diferentes modos de ser-sentir-conhecer o mundo.

A formação de educadores e a travessia de cercas invisíveis de acesso/produção de conhecimentos : experiências do MST nas inter-relações com universidades brasileiras /

Este estudo sistematiza e aborda a temática da formação de educadores desenvolvida nas inter-relações entre universidades brasileiras e o Movimento dos Trabalhadores Rurais Sem Terra, especificamente nos cursos de graduação em História na Universidade Federal da Paraíba e de Engenharia Agronômica na Universidade Federal de Sergipe, no período de 2004 a 2008. Tal processo desenvolve-se em um contexto difícil e complexo da luta pela reforma agrária no Brasil, principalmente pelas transformações ocorridas nos últimos anos oriundas da ampliação das lógicas de produção do agronegócio. Esta condição leva o MST a discutir e propor uma nova concepção de reforma agrária que a designa de popular em substituição à proposta de reforma agrária clássica. Por outro lado, apresenta uma visão das universidades brasileiras e dos projetos que são construídos e implementados nos últimos anos, inclusive, observando coincidências com as políticas econômicas gerais para a sociedade. Apresenta uma concepção de formação que vem sendo construída no interior das práticas do MST que também procura as universidades para firmar convênios e desenvolver processos de escolarização/formação de seus militantes educadores. Dentre as dimensões desse processo educativo/formativo, destacam-se: vínculo permanente com os processos orgânicos; a formação como um processo ético, estético, místico que trata das atitudes/comportamentos e como um processo dialógico, crítico e articulado que contempla saberes, experiências, em uma interação que busca superar as monoculturas. Captura por intermédio da pesquisa de campo: estranhamentos, entraves, sentidos da ocupação pedagógica e coletiva, alternativas, legados que permanecem tanto no MST como na universidade e apresenta o resultado do envolvimento e atuação dos egressos de ambos os cursos na atualidade. Aponta também para desafios, possibilidades outras de enfrentar a difícil mas necessária tarefa de formar educadores, militantes capazes de coletivamente levar adiante a luta por um mundo mais justo, solidário e democrático, em que a terra e o conhecimento, juntamente com os demais bens econômicos e culturais sejam profundamente democratizados. Pretende ser uma contribuição para o debate acerca da relevância dessas inter-relações entre universidade e movimentos sociais, em que essas experiências demarcam novas possibilidades de abertura e avanços democráticos e menos elitista da universidade e novos patamares de escolarização/formação para integrantes do Movimento dos Sem Terra.