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Orientador: Carmen Sílvia Bertuzzo / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T04:59:08Z (GMT). No. of bitstreams: 1 Assis_AngelaMariade_D.pdf: 2409399 bytes, checksum: 620279a2d72b5132a10f7af956d4bebf (MD5) Previous issue date: 2007 / Resumo: A Telangiectasia Hemorrágica Hereditária (THH) é uma desordem autossômica dominante caracterizada por epistaxe recorrente, telangiectases mucocutânea, hemorragia gastrointestinal, e malformação arteriovenosa pulmonar (PAVM), cerebral e hepática. A prevalência da doença é de 1/5000 e a mortalidade relatada da doença entre pacientes jovens quando comparado com aqueles com mais de 60 anos é de 36%. Dois genes da superfamília de receptores para TGF-? têm sido relacionados com THH, o gene da endoglina e o gene da activina (activin receptor-like-kinase). Estes genes são altamente expressos em células endoteliais e outros tecidos altamente vascularizados como pulmão e placenta. Mutações no gene da endoglina causam a THH tipo 1 que é caracterizada pela alta incidência de malformação arteriovenosa pulmonar sintomática (PAVMs). Mutações no gene da activina levam a THH tipo 2 caracterizada por epistaxe recorrente e malformação arteriovenosa gastrointestinal. Foi objetivo deste trabalho realizar uma triagem de mutações na região codificadora dos genes ALK-1 e endoglina em doze pacientes portadores de THH atendidos no Hemocentro da Unicamp. A abordagem metodológica incluiu a amplificação dos exóns dos genes da activina e endoglina seguida pela técnica de PCR/CSGE, clonagem e sequenciamento. Dos doze pacientes estudados, sete apresentaram alguma alteração molecular, destes três pacientes apresentaram uma deleção de um nucleotídeo T na posição 913 no exon 7, um paciente apresentou uma inserção de um nucleotídeo G na posição 204-205 no exon 3 e três pacientes apresentavam uma mutação misense nos exons 7 e 8 na posição 976 e 1204 respectivamente. Nossos resultados mostraram que a THH tipo 1 e tipo 2 são raras no Brasil e todas as mutações citadas na tabela IV e Figura 10 são novas, somente a mutação ocorrida no exon 8 foi previamente descrita na literatura / Abstract: Background: Hereditary hemorrhagic telangiectasia in humans, also known as HHT or Osler-Rendu-Weber syndrome, is an autosomal dominant vascular disorder characterizes by recurrent epistaxis, mucocutaneous telangiectases, gastrointestinal, pulmonary, cerebral and hepatic arteriovenous malformations (HAVM). The prevalence of the illness is of 1/5000 and the reported mortality of the illness among young patients when compared with those with more than 60 years is 36%. Two genes of the receptors superfamily for TGF-? have been related with THH, the gene of the endoglin (ENG) in the chromosome 9q33-34 and the gene of the activin (receptor-like-kinase 1 ALK-1), in the chromosome 12q11q14. These genes are highly express in endothelial cells and other tissue highly vascularized as lung and placenta. Mutations in the gene of the endoglin cause the HHT type 1 which is characterized by the high incidence of symptomatic pulmonary arteriovenous malformation (PAVM). Mutations in the activin gene cause HHT were described among patients in whom the linking with HHT in the chromosome 9q33 or mutation in the endoglin gene was excluded. It was the objective of this work to carry through a selection of mutations in the coding region of gene ALK-1 and endoglin in 12 patients carrying HHT whose are treated in the Blood Center at Unicamp. Methods: Twelve patients were analyzed. Diagnosis of HHT was carried out by means of clinical history of recurrent bleeding, heredity studies, and the presence of multiple telangiectases lesions. PCR products with consistent abnormal migration patterns were cloned into the SureCloneTM ligation kit vector system and sequenced using the DYEnamic TM ET dye terminator cycle sequencing Kit, and analyzed by the MegaBace 1000 DNA automated analyzer. A panel of 252 chromosomes from unrelated individuals without the disease was used to evaluate the frequency of each mutation in the general population. Results: In six patients three novel mutations were identified in the coding sequence of the ALK-1 gene in their families, which demonstrated clinical manifestations of HHT type 2. These mutations included an insertion a deletion of single base pairs in éxons 3 (c.204-205insG) and 7 (c.913del T), as well as missense mutations in éxons 7 (c.976A>G) and 8 (c.1204G>A) of the ALK-1 gene. The mutations identified in éxons 7 and 8 affect the kinase domain and the mutation identified in éxon 3 affect the extracellular domain. These data indicate that loss-of-function mutations in a single allele of the ALK1 locus are sufficient to contribute to defects in maintaining endothelial integrity. Conclusion: We suggest the high rate of mutation detection and the small size of the ALK-1 gene make genomic sequencing a viable diagnostic test for HHT2. This was the only research about this subject made in Brazil so far / Doutorado / Genetica Animal e Evolução / Doutora em Genética e Biologia Molecular

Charles Univerzity in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Rebeka Klingová Supervisor: PharmDr. Petra Fikrová, Ph.D. Title of diploma thesis: Effect of soluble endoglin to endothelial cells in vitro Aim: We determined the effects of soluble endoglin on endothelial cells, by the means of inflammatory markers. The aim of the study was to point out the possible association of the soluble endoglin with endothelial dysfunction. Methods: For our study we have selected the human endothelial cells from umbilical vein - HUVEC. We have influenced the cells with soluble endoglin in two concentrations and at two different time intervals. The results were evaluated in the statistical program, in which we have compared the control group with influenced cells and two concentrations between each other. The expressions of inflammatory markers were analyzed on the level of mRNA, using the real-time PCR method. Results: Significant changes in the expression of markers were observed on the vascular and intracellular adhesion molecule at both concentrations compared to the control group. Increased values of transcription were available also for cyclooxygenase 2 and decreased values for cadherin 5 compared to the control group. Conclusion: Changes in...

Endoglin, a co-receptor of the TGF-β superfamily, is predominantly expressed in endothelial cells and in some myeloid cells and implicated as a potential modulator of immune responses. We previously demonstrated that Endoglin heterozygous (Eng+/-) mice subjected to the dextran sulfate sodium colitis model developed persistent inflammation and epithelial ulceration, while Eng+/+ mice recovered following the acute phase of disease. Our aim was to assess potential alterations in distribution and number of immune cells, expression of inflammatory mediators and mechanisms of oxidative burst in Eng+/- mice. While the number of overall T, B and myeloid cells was unaltered between the genotypes, changes in neutrophil regulating cytokines and angiogenesis mediating factors were observed in Eng+/- mice. In addition, downregulation of phagocyte oxidative burst enzymes point to potential defects in microbial clearance in Eng+/- mice. These findings suggest a role for endoglin in regulating immune and vascular functions during inflammation.

Endoglin, a co-receptor of the TGF-β superfamily, is predominantly expressed in endothelial cells and in some myeloid cells and implicated as a potential modulator of immune responses. We previously demonstrated that Endoglin heterozygous (Eng+/-) mice subjected to the dextran sulfate sodium colitis model developed persistent inflammation and epithelial ulceration, while Eng+/+ mice recovered following the acute phase of disease. Our aim was to assess potential alterations in distribution and number of immune cells, expression of inflammatory mediators and mechanisms of oxidative burst in Eng+/- mice. While the number of overall T, B and myeloid cells was unaltered between the genotypes, changes in neutrophil regulating cytokines and angiogenesis mediating factors were observed in Eng+/- mice. In addition, downregulation of phagocyte oxidative burst enzymes point to potential defects in microbial clearance in Eng+/- mice. These findings suggest a role for endoglin in regulating immune and vascular functions during inflammation.

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LDL-apheresis and haemorheopheresis are the most frequent methods of extracorporeal ellimination methods used for lowering the LDL cholesterol in patients with familial hypercholesterolaemia (FH). In case of failure of conservative therapy (represented by pharmacotherapy and/or dietary regime) these methods reprensent the effective and accessible solution for the clinical status characterized by high morbidity and mortality as well. The lipid components are the most frequent observed markers for the effeciveness of intervention. However, immunity with its effector molecules plays also essential role and there is suppose, that could also reflect the state and progress of atherosclerosis in FH patients. The aim of this thesis was to evaluate the levels of selected immunological markers (plasmatic glykoprotein α-2-macroglobulin, IL-10, soluble endoglin, soluble apoptotic factor sAPO-Fas and soluble form of adhesive P-selectin) and their changes after LDL-apheresis and haemorheopheresis employing enzyme immunoanalysis and immuno nephelometry. Totally, 12 patients were involved in this study, 3 were treated by haemorheopheresis, the rest 9 received LDL-apheresis. As a results, significant decreases in serum levels of α-2- macroglobulin, soluble endoglin and sAPO-1/Fas were recorded. Observed changes of...

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Background: Angiogenesis is recognised as a critical step in tumour progression. The angiogenic switch is activated by various trigger signals, such as hypoxia, low pH, and genetic mutations. Renal cell carcinoma (RCC) is often an aggressive tumour, and advanced disease has limited treatment options and bad prognosis. This study was focused on markers of angiogenesis in RCC: endoglin (CD105) and CD31 assessing microvessel density (MVD), and carbonic anhydrase (CA) IX and hypoxia-inducible factor (HIF)-2α expressed at hypoxia. Upregulation of HIF is also associated with inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene, which is common in conventional/clear cell (c)RCC. Method: A tumour bank containing 308 tumours from patients operated 1982-2003 was used. The tumours were well characterised regarding tumour type, TNM stage, nuclear grade, tumour size, and patient survival. The tumours were prepared in tissue microarrays and fresh frozen in whole sections. To analyse the expression of endoglin, CD31, CA IX, and HIF-2α mRNA, immunohistochemistry and real-time PCR were used. Results: There was a higher endoglin expression in cRCC than in papillary (p)RCC and chromophobe (ch)RCC, and a higher CD31 expression in cRCC than in pRCC. MVD correlated inversely to TNM stage and nuclear grade in cRCC. There was also an inverse correlation between tumour diameter and CD31 expression in cRCCs. Patients with cRCC with high MVD had a more favourable prognosis than patients with lower MVD. Endoglin and CD31 were not independent prognostic factors. The CA IX expression was higher in cRCC than in pRCC and chRCC. Patients with cRCC expressing low CA IX had a significantly less favourable prognosis compared with those with higher expression. CA IX is an independent prognostic factor. There was a higher HIF-2α mRNA expression in cRCC than in pRCC and chRCC. In cRCC, there was a significant inverse correlation between HIF-2α mRNA expression, and TNM stage and nuclear grade. There was also an inverse correlation between HIF-2α mRNA expression and tumour size among patients with cRCC. HIF-2α was not an independent prognostic factor. Conclusion: In these studies, the factors related to hypoxia and vascularity were all inversely correlated to tumour aggressiveness in cRCC. MVD, CA IX, and HIF-2α expression were also higher in cRCC than in pRCC and chRCC. The relationship between angiogenesis, vascularity, and hypoxia is ambiguous. A line of reasoning including mutations increasing angiogenesis in advanced disease may also be applied to RCC. Measurements of individual angiogenic factors seem to provide prognostic information, and can potentially be combined in patient monitoring and treatment.