Immunomodulatory effects of recombinant BCG expressing pertussis toxin on TNF-alpha and IL-10 in a bladder cancer model

Background: Since successful treatment of superficial bladder cancer with BCG requires proper induction of Th1 immunity, we have developed a rBCG-S1PT strain that induced a stronger cellular immune response than BCG. This preclinical study was designed to compare the modulatory effects of BCG and rBCG-S1PT on bladder TNF-alpha and IL-10 expression and to evaluate antitumour activity. Methods: For Experiment I, the MB49 bladder cancer cell line was used in C57BL/6 mice. Chemical cauterization of the bladder was performed to promote intravesical tumor implantation. Mice were treated by intravesical instillation with BCG, rBCG-S1PT or PBS once a week for four weeks. After 35 days the bladders were removed and weighed. TNF-<alpha and IL-10 cytokine responses were measured by qPCR. Experiment II was performed in the same manner as Experiment I, except the animals were not challenged with MB49 tumor cells. Results: rBCG-S1PT immunotherapy resulted in bladder weight reduction, compared to the BCG and control group. There were increases in TNF-alpha in the BCG-treated group, as well as increases in TNF-alpha and IL-10 mRNA in the rBCG-S1PT group. Conclusion: These data indicate a significant reduction of bladder tumor volume for the rBCG group, compared to the BCG and PBS groups. This suggests that rBCG could be a useful substitute for wild-type BCG and that the potential modulation between TNF-alpha and IL-10 cytokine productions may have therapeutic value

A quantitative view of the transcriptome of Schistosoma mansoni adult-worms using SAGE

<p>Abstract</p> <p>Background</p> <p>Five species of the genus Schistosoma, a parasitic trematode flatworm, are causative agents of Schistosomiasis, a disease that is endemic in a large number of developing countries, affecting millions of patients around the world. By using SAGE (Serial Analysis of Gene Expression) we describe here the first large-scale quantitative analysis of the Schistosoma mansoni transcriptome, one of the most epidemiologically relevant species of this genus.</p> <p>Results</p> <p>After extracting mRNA from pooled male and female adult-worms, a SAGE library was constructed and sequenced, generating 68,238 tags that covered more than 6,000 genes expressed in this developmental stage. An analysis of the ordered tag-list shows the genes of F10 eggshell protein, pol-polyprotein, HSP86, 14-3-3 and a transcript yet to be identified to be the five top most abundant genes in pooled adult worms. Whereas only 8% of the 100 most abundant tags found in adult worms of S. mansoni could not be assigned to transcripts of this parasite, 46.9% of the total ditags could not be mapped, demonstrating that the 3 sequence of most of the rarest transcripts are still to be identified. Mapping of our SAGE tags to S. mansoni genes suggested the occurrence of alternative-polyadenylation in at least 13 gene transcripts. Most of these events seem to shorten the 3 UTR of the mRNAs, which may have consequences over their stability and regulation.</p> <p>Conclusion</p> <p>SAGE revealed the frequency of expression of the majority of the S. mansoni genes. Transcriptome data suggests that alternative polyadenylation is likely to be used in the control of mRNA stability in this organism. When transcriptome was compared with the proteomic data available, we observed a correlation of about 50%, suggesting that both transcriptional and post-transcriptional regulation are important for determining protein abundance in S. mansoni. The generation of SAGE tags from other life-cycle stages should contribute to reveal the dynamics of gene expression in this important parasite.</p

Immunomodulatory effects of recombinant BCG expressing pertussis toxin on TNF-alpha and IL-10 in a bladder cancer model

Abstract Background Since successful treatment of superficial bladder cancer with BCG requires proper induction of Th1 immunity, we have developed a rBCG-S1PT strain that induced a stronger cellular immune response than BCG. This preclinical study was designed to compare the modulatory effects of BCG and rBCG-S1PT on bladder TNF-α and IL-10 expression and to evaluate antitumour activity. Methods For Experiment I, the MB49 bladder cancer cell line was used in C57BL/6 mice. Chemical cauterization of the bladder was performed to promote intravesical tumor implantation. Mice were treated by intravesical instillation with BCG, rBCG-S1PT or PBS once a week for four weeks. After 35 days the bladders were removed and weighed. TNF-〈 and IL-10 cytokine responses were measured by qPCR. Experiment II was performed in the same manner as Experiment I, except the animals were not challenged with MB49 tumor cells. Results: rBCG-S1PT immunotherapy resulted in bladder weight reduction, compared to the BCG and control group. There were increases in TNF-α in the BCG-treated group, as well as increases in TNF-α and IL-10 mRNA in the rBCG-S1PT group. Conclusion These data indicate a significant reduction of bladder tumor volume for the rBCG group, compared to the BCG and PBS groups. This suggests that rBCG could be a useful substitute for wild-type BCG and that the potential modulation between TNF-α and IL-10 cytokine productions may have therapeutic value.</p

Epitope Mapping of Exposed Tegument and Alimentary Tract Proteins Identifies Putative Antigenic Targets of the Attenuated Schistosome Vaccine

The radiation-attenuated cercarial vaccine remains the gold standard for the induction of protective immunity against Schistosoma mansoni. Furthermore, the protection can be passively transferred to naïve recipient mice from multiply vaccinated donors, especially IFNgR KO mice. We have used such sera versus day 28 infection serum, to screen peptide arrays and identify likely epitopes that mediate the protection. The arrays encompassed 55 secreted or exposed proteins from the alimentary tract and tegument, the principal interfaces with the host bloodstream. The proteins were printed onto glass slides as overlapping 15mer peptides, reacted with primary and secondary antibodies, and reactive regions detected using an Agilent array scanner. Pep Slide Analyzer software provided a numerical value above background for each peptide from which an aggregate score could be derived for a putative epitope. The reactive regions of 26 proteins were mapped onto crystal structures using the CCP4 molecular graphics, to aid selection of peptides with the greatest accessibility and reactivity, prioritizing vaccine over infection serum. A further eight MEG proteins were mapped to regions conserved between family members. The result is a list of priority peptides from 44 proteins for further investigation in multiepitope vaccine constructs and as targets of monoclonal antibodies

Avaliação do questionário de Boston aplicado no pós-operatório tardio da síndrome do tunel do carpo operados pela técnica de retinaculótomo de paine por via palmar Evaluation of Boston questionnaire applied at late pos-operative period of carpal tunnel syndrome operated with the paine retinaculatome through palmar port

Entre os anos de 1995 e 1998, foram realizadas 112 cirurgias para tratamento da Síndrome do Túnel do Carpo (STC) pela técnica de incisão palmar e utilização do retináculo de Paine. Com o objetivo de avaliar os resultados em longo prazo, os pacientes foram convocados. Houve o retorno de 44 pacientes. Deste total, três pacientes, por terem doenças associadas, foram excluídos, resultando, um total de 53 mãos analisadas. Apresentaremos os resultados da avaliação subjetiva, obtidos através da aplicação de um teste de auto-avaliação chamado de questionário de Boston. Este questionário consiste em perguntas que avaliam a gravidade dos sintomas e o estado funcional no momento da aplicação do mesmo. Através da aplicação do referido questionário encontramos um escore de 1,41 ± 0,57 para gravidade dos sintomas e 1,59 ± 0,93 para o estado funcional. Como este questionário não foi aplicado no pré-operatório deste grupo de pacientes analisados, comparou-se a pontuação obtida com as encontradas na literatura pertinente. Os resultados obtidos demonstraram que as pontuações pós-operatórias são similares àquelas existentes na literatura, mesmo sendo referidas a tempos diferentes de seguimento pós-operatórios, concluindo que havendo uma melhora dos sintomas, o questionário de Boston é sensível a esta mudança clínica.<br>Between the years of 1995 and 1998, 112 surgeries were performed for treating Carpal Tunnel Syndrome (CTS) using the technique of palmar incision employing the Paine retinaculum. With the objective of analyzing results in the long-term, the patients were called for review. Forty four patients returned. From these, three patients were excluded due to associated diseases, thus resulting in a total of 53 hands assessed. Here we present the results of the subjective evaluation achieved by applying a self-assessment test called Boston questionnaire. This questionnaire consists of questions evaluating symptoms severity and functional status at the moment of its application. By applying this questionnaire, we found a score of 1.41 ± 0.57 for symptoms severity and of 1.59 ± 0.93 for functional status. As this questionnaire was not applied at the pre-operative period for those patients assessed, its scores were thus compared to those found in pertinent literature. The achieved results show that post-operative scores are similar to those described in literature, even when reported in different postoperative follow-up times, thereby concluding that when symptoms are improved, the Boston questionnaire is sensitive to that clinical change

Identification of human chromosome 22 transcribed sequences with ORF expressed sequence tags

Transcribed sequences in the human genome can be identified with confidence only by alignment with sequences derived from cDNAs synthesized from naturally occurring mRNAs. We constructed a set of 250,000 cDNAs that represent partial expressed gene sequences and that are biased toward the central coding regions of the resulting transcripts. They are termed ORF expressed sequence tags (ORESTES). The 250,000 ORESTES were assembled into 81,429 contigs. Of these, 1,181 (1.45%) were found to match sequences in chromosome 22 with at least one ORESTES contig for 162 (65.6%) of the 247 known genes, for 67 (44.6%) of the 150 related genes, and for 45 of the 148 (30.4%) EST-predicted genes on this chromosome. Using a set of stringent criteria to validate our sequences, we identified a further 219 previously unannotated transcribed sequences on chromosome 22. Of these, 171 were in fact also defined by EST or full length cDNA sequences available in GenBank but not utilized in the initial annotation of the first human chromosome sequence. Thus despite representing less than 15% of all expressed human sequences in the public databases at the time of the present analysis, ORESTES sequences defined 48 transcribed sequences on chromosome 22 not defined by other sequences. All of the transcribed sequences defined by ORESTES coincided with DNA regions predicted as encoding exons by genscan. (http://genes.mit.edu/GENSCAN.html)