No / Hair follicle (HF) morphogenesis is governed by a series of signals exchanged between the epidermal keratinocytes committed to HF-specific differentiation and the mesenchymal cells forming the follicular papilla (Schmidt-Ullrich and Paus, 2005). These interactions lead to the construction of the hair bulb, in which keratinocytes rapidly proliferate and differentiate into several cell populations forming the hair shaft and the inner root sheath. During the final steps of development, the HF elongates up to its maximal length and the hair shaft emerges through the epidermis through the hair canal that is formed at the distal portion of the HF epithelium (Schmidt-Ullrich and Paus, 2005). ...To elucidate a role for MMP-9 in the control of HF development and hair canal formation, its expression was studied by immunohistochemistry in cryosections of embryonic and postnatal skin of C57BL/6 mice (Sharov et al., 2003), and was correlated to the micro-anatomy of the developing hair canal depicted by high-resolution light microscopy, as described previously (Magerl et al., 2001). MMP-9 expression was not observed in the epidermis and developing HF placodes.

Yes / The membrane-type matrix metalloproteinases (MT-MMPs), an important subgroup of the wider MMP family, demonstrate widespread expression in multiple tumor types, and play key roles in cancer growth, migration, invasion and metastasis. Despite a large body of published research, relatively little information exists regarding evidence for MT-MMP expression and function in metastatic prostate cancer. This review provides an appraisal of the literature describing gene and protein expression in prostate cancer cells and clinical tissue, summarises the evidence for roles in prostate cancer progression, and examines the data relating to MT-MMP function in the development of bone metastases. Finally, the therapeutic potential of targeting MT-MMPs is considered. While MT-MMP inhibition presents a significant challenge, utilisation of MT-MMP expression and proteolytic capacity in prostate tumors is an attractive drug development opportunity.

Balance between matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) is required for normal wound healing. Chronic inflammation, such as that seen in cardiovascular and periodontal diseases, may upset this balance. The aim of this study was to determine whether initial periodontal therapy would have an effect systemically on the levels of MMP-9 and TIMP-1. Twenty-one patients with generalized chronic periodontitis were enrolled in the study. Clinical examinations were conducted and parameters measured. Scaling and root planing was performed and blood analysis done to determine the plasma concentrations of MMP-9 and serum concentrations of TIMP-1. Initial periodontal therapy resulted in improvements in gingival inflammation and plaque levels. No effect on the plasma concentrations of MMP-9 and serum concentrations of TIMP-1 could be found following therapy.

A MMP-2 é uma protease que está envolvida em muitos eventos fisiológicos e patológicos e que compartilha similaridades estruturais com a enzima conversora de angiotensina (ECA), de modo que os inibidores da ECA passaram a ser estudados com relação ao efeito inibitório também sobre a MMP-2. No entanto, este potencial inibitório não foi ainda testado na MMP-2 altamente purificada. Este estudo teve como objetivo investigar o potencial inibitório do captopril sobre a atividade da MMP- 2. Primeiramente, supôs-se que a dissolução do captopril poderia induzir a mudanças no pH de soluções tampão. Em segundo lugar, avaliou-se o efeito direto do captopril sobre a MMP-2 presente no plasma humano e a MMP-2 recombinante humana (rhMMP-2) produzida e purificada de E. coli. As análises de atividade in vitro incluíram zimogramas com gelatina e ensaios fluorimétricos com DQ gelatin. A solubilização do captopril reduziu significativamente o pH da solução tampão 50 mM (p<0,01) mas não alterou o pH da solução tampão 200 mM (p>0,05). Resultados de zimografia do plasma e da rhMMP-2 mostraram inibição da atividade gelatinolítica com significância estatística somente em concentrações iguais ou maiores que 4 e 1 mM de captopril, respectivamente (p<0,05). A presença de captopril nos ensaios de fluorimetria resultaram na inibição significante da atividade de rhMMP-2 somente em concentrações iguais ou maiores que 2 mM (p<0,01), enquanto a rhMMP-2 ativada com APMA apresentou inibição significativa diante de 0,5 mM de captopril (p<0,01). As concentrações de captopril efetivas em inibir a MMP-2 in vitro foram muito superiores àquelas referentes à concentração plasmática máxima encontrada no plasma humano após a administração de uma dose de 50 mg de captopril. Em conjunto nossos resultados sugerem que o captopril não parece promover inibição significativa da MMP-2 nas concentrações relatadas in vivo. Além disso, o pH das soluções tamponantes é um aspecto que requer mais atenção durante ensaios de inibição de protease in vitro. / MMP-2 is involved in many physiological and pathological processes. This protease shares structural similarities with the angiotensin-converting enzyme (ACE), and ACE inhibitors have been described to inhibit MMP-2. However, this inhibitory potential has not been tested using a highly purified MM-2 so far. This study aimed at investigating the inhibitory potential of captopril on MMP-2 activity. First it was tested whether the dissolution of captopril would induce changes in the pH of the solutions. Secondly, the direct inhibitory effect of captopril on plasma MMP-2 and on a recombinant human MMP-2 (rhMMP-2) produced and purified from E. coli was tested. The in vitro activity assays included gelatin zymography and a fluorimetric assay with DQ gelatin. Captopril solubilization significantly decreased the pH of the 50 mM Tris buffer solution (p<0.01) but did not decreased the pH of the 200 mM Tris Buffer solution (p>0.05). Zymography results of plasma and rhMMP-2 showed that inhibition of the activity only reached statistical significance >= 4 and 1 mM of captopril, respectively (p<0,05). The presence of captopril in the fluorimetric assay resulted in a significant inhibition of the rhMMP-2 activity only at concentrations >= 2 mM (p<0.01), whereas APMA-activated rhMMP-2 was inhibited by 0.5 mM of captopril (p<0.01). The captopril concentrations found to inhibit MMP-2 are several times of magnitude higher than the maximum plasma concentration after a dose of 50 mg of captopril. In conclusion, captopril does not seem to cause significant inhibition of MMP-2 in the concentrations found in vivo, and more attention has to be given to the pH of the solutions when testing protease inhibition in vitro.

A MMP-2 é uma protease que está envolvida em muitos eventos fisiológicos e patológicos e que compartilha similaridades estruturais com a enzima conversora de angiotensina (ECA), de modo que os inibidores da ECA passaram a ser estudados com relação ao efeito inibitório também sobre a MMP-2. No entanto, este potencial inibitório não foi ainda testado na MMP-2 altamente purificada. Este estudo teve como objetivo investigar o potencial inibitório do captopril sobre a atividade da MMP- 2. Primeiramente, supôs-se que a dissolução do captopril poderia induzir a mudanças no pH de soluções tampão. Em segundo lugar, avaliou-se o efeito direto do captopril sobre a MMP-2 presente no plasma humano e a MMP-2 recombinante humana (rhMMP-2) produzida e purificada de E. coli. As análises de atividade in vitro incluíram zimogramas com gelatina e ensaios fluorimétricos com DQ gelatin. A solubilização do captopril reduziu significativamente o pH da solução tampão 50 mM (p<0,01) mas não alterou o pH da solução tampão 200 mM (p>0,05). Resultados de zimografia do plasma e da rhMMP-2 mostraram inibição da atividade gelatinolítica com significância estatística somente em concentrações iguais ou maiores que 4 e 1 mM de captopril, respectivamente (p<0,05). A presença de captopril nos ensaios de fluorimetria resultaram na inibição significante da atividade de rhMMP-2 somente em concentrações iguais ou maiores que 2 mM (p<0,01), enquanto a rhMMP-2 ativada com APMA apresentou inibição significativa diante de 0,5 mM de captopril (p<0,01). As concentrações de captopril efetivas em inibir a MMP-2 in vitro foram muito superiores àquelas referentes à concentração plasmática máxima encontrada no plasma humano após a administração de uma dose de 50 mg de captopril. Em conjunto nossos resultados sugerem que o captopril não parece promover inibição significativa da MMP-2 nas concentrações relatadas in vivo. Além disso, o pH das soluções tamponantes é um aspecto que requer mais atenção durante ensaios de inibição de protease in vitro. / MMP-2 is involved in many physiological and pathological processes. This protease shares structural similarities with the angiotensin-converting enzyme (ACE), and ACE inhibitors have been described to inhibit MMP-2. However, this inhibitory potential has not been tested using a highly purified MM-2 so far. This study aimed at investigating the inhibitory potential of captopril on MMP-2 activity. First it was tested whether the dissolution of captopril would induce changes in the pH of the solutions. Secondly, the direct inhibitory effect of captopril on plasma MMP-2 and on a recombinant human MMP-2 (rhMMP-2) produced and purified from E. coli was tested. The in vitro activity assays included gelatin zymography and a fluorimetric assay with DQ gelatin. Captopril solubilization significantly decreased the pH of the 50 mM Tris buffer solution (p<0.01) but did not decreased the pH of the 200 mM Tris Buffer solution (p>0.05). Zymography results of plasma and rhMMP-2 showed that inhibition of the activity only reached statistical significance >= 4 and 1 mM of captopril, respectively (p<0,05). The presence of captopril in the fluorimetric assay resulted in a significant inhibition of the rhMMP-2 activity only at concentrations >= 2 mM (p<0.01), whereas APMA-activated rhMMP-2 was inhibited by 0.5 mM of captopril (p<0.01). The captopril concentrations found to inhibit MMP-2 are several times of magnitude higher than the maximum plasma concentration after a dose of 50 mg of captopril. In conclusion, captopril does not seem to cause significant inhibition of MMP-2 in the concentrations found in vivo, and more attention has to be given to the pH of the solutions when testing protease inhibition in vitro.

Systemic inflammation is associated with impaired lung function. Inflammation is part of asthma and chronic obstructive pulmonary disease (COPD), but the local and systemic inflammatory pattern differs. The overall aim was to evaluate systemic inflammatory markers in obstructive lung diseases and more specifically: To determine if CRP is related to respiratory symptoms, asthma, atopy and bronchial responsiveness (paper I), in a population sample from three countries (paper I and II); to evaluate if CRP is related to COPD, lung function and rate of lung function decline (paper II); to investigate the association of serum MMP-9 and TIMP-1 with lung function in a cross-sectional population based study (paper III); and finally, to study possible gender differences in the longitudinal association between CRP and lung function in a prospective population based study (paper IV). In the first study we reported that CRP was related to non-allergic asthma but not allergic asthma, and that CRP was related to respiratory symptoms such as wheeze, nocturnal cough and breathlessness after effort, but not associated with atopy or bronchial responsiveness. In the second study we found that COPD was more common in subjects in the highest CRP quartiles and higher CRP levels were associated with lower FEV1 values in both men and women, but the negative association between CRP and FEV1 was larger in men than women. The FEV1 decline was larger in men with high CRP levels, whereas no such association was found for women. In the third study we reported that lower FEV1 was associated with higher levels of MMP-9, TIMP-1 and their ratio MMP-9/TIMP-1. After stratification for gender this association was significant in men but not women. In the fourth study we found that CRP levels were associated with change in both FEV1 and FVC in men but not women. This association was found for both baseline CRP and change in CRP, confirming a stronger association between systemic inflammation and lung function decline in men than women. In conclusion, systemic inflammation is associated with non-allergic asthma but not allergic asthma. Our findings of a stronger association between the systemic inflammation and lung function impairment in men, but not women, may indicate a gender difference in the mechanisms of lung function decline.

Thesis (Ph. D.)--University of Oklahoma. / Bibliography: leaves 164-207.

Made available in DSpace on 2014-06-11T19:27:18Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-11-09Bitstream added on 2014-06-13T20:16:24Z : No. of bitstreams: 1 marangoni_nr_me_araca.pdf: 326349 bytes, checksum: 7dc73435330e73a5c923b91991a9e1f1 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / A leishmaniose visceral canina, causada pelo protozoário Leishmania (Leishmania) chagasi, é uma doença de grande ocorrência principalmente na América Latina. A caracterização das lesões sistêmicas associadas à infecção pelo parasita tem sido amplamente estudada, entretanto, poucos autores elucidam a patogenia na forma nervosa. Com o objetivo de compreender melhor os mecanismos envolvidos na inflamação do sistema nervoso central de cães naturalmente infectados por L. chagasi, amostras de liquor e plexo coróide foram colhidas e submetidas à zimografia para a detecção de metaloproteinases (MMPs). Amostras do plexo coróide e liquor de cães sadios foram avaliadas como controle. Os géis de zimograma foram analisados quanto à presença e atividade proteolítica das metaloproteinases -2 e -9. Formas inativas das proteases foram detectadas no plexo coróide, sendo que o Grupo de animais positivos não diferiu do negativo. No liquor foram encontradas formas ativas e inativas das MMPs-2 e -9 e a atividade proteolítica das mesmas diferiu entre os Grupos positivo e negativo. A MMP-2 teve maior detecção nos animais negativos e a MMP-9 nos positivos. O aumento da MMP- 9 no liquor dos cães doentes representa seu possível envolvimento na patogenia das lesões encefálicas ao ocasionarem o rompimento das barreiras hematoencefálica e/ou hematoliquórica, permitindo a passagem de células e proteínas envolvidas no processo inflamatório / Canine visceral leishmaniasis, caused by the protozoan Leishmania (Leishmania) chagasi, is a disease with high occurrence in Latin America. The characteristics of the systemic lesions related to the infection have been widely studied, but few studies clarify the disease on a neurological aspect. With the aim of a better understanding of the inflammation mechanisms within the central nervous system of dogs naturally infected by L. chagasi, some samples of cerebrospinal fluid and choroid plexus were collected and submitted to zymography to detect metalloproteinases (MMPs). Samples of choroid plexus and cerebrospinal fluid from healthy dogs were evaluated as control. The zymogram gels were analysed taking into account the presence and the proteolytic activity of metalloproteinase -2 and -9. Inactive forms of the proteases were detected in the choroid plexus, and the group of positive animals did not differ from negative ones. In the cerebrospinal fluid, active and inactive forms of MMP-2 and -9 were found, and their proteolytic activity differed between negative and positive groups. MMP-2 had higher detection in the negative animals and MMP-9 in the positive ones. The increasing of MMP-9 in the cerebrospinal fluid of infected dogs represents its possible involvement in the brain injuries, by causing the disruption of blood-cerebrospinal fluid barrier and/or blood-brain-barrier, allowing the passage of cells and proteins involved in inflammation process

Orientador: Sergio Luis Felisbino / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-17T13:08:44Z (GMT). No. of bitstreams: 1 Amorim_ElaineManoelaPorto_D.pdf: 2896871 bytes, checksum: d7581f6efdb5920a39d589672ca2a350 (MD5) Previous issue date: 2010 / Resumo: Diabetes mellitus é uma síndrome clínica heterogênea, causada pela falta de produção de insulina pelas células beta (?) pancreáticas ou pelo defeito nos receptores para insulina nas células-alvo, resultando em uma doença metabólica hiperglicêmica. Estudos têm demonstrado que as disfunções sexuais e reprodutivas incluindo impotência, redução da libido, prejuízo da espermatogênese, infertilidade e diminuição da qualidade espermática são frequentemente associadas com o diabete no homem e em animais de experimentação. Apesar dos efeitos do diabete sobre as funções testiculares e fertilidade de machos estarem bem esclarecidos, o impacto da doença sobre morfogênese e crescimento da próstata ainda não foi completamente descrito. Assim, o presente estudo teve por objetivo analisar os possíveis efeitos do diabete e da reposição com insulina durante o crescimento prostático na puberdade, com especial atenção para a atividade das metaloproteinases de matriz 2 e 9 (MMP-2 e MMP-9), enzimas envolvidas na remodelação dos componentes da matriz extracelular. Para tanto, foram realizadas análises morfológicas, morfométricas, imunoistoquímicas e bioquímicas de zimografia, nos lobos ventral, dorsolateral e anterior da próstata de ratos Wistar, nos quais o diabete foi induzido quimicamente aos 40 dias de idade por estreptozotocina. Nossos resultados mostraram que o diabete experimental prejudicou o crescimento glandular, caracterizado por um menor ganho de peso absoluto dos diferentes lobos da próstata e diminuição da relação parênquima/estroma, principalmente do lobo dorsolateral. O diabete causou diminuição nas concentrações plasmáticas de testosterona, mas não alterou a concentração de diidrotestosterona. As atividades das MMP-2 e MMP-9 foram menores no grupo diabético. O tratamento simultâneo e/ou tardio com insulina exógena além de reverter os parâmetros alterados nos animais diabéticos também foi efetivo em restaurar características histológicas e morfométricas dos lobos prostáticos em padrões semelhantes aos observados nos animais controles. Estes resultados permitem concluir que o diabete prejudica, mas não impede, o crescimento da próstata e causa importantes alterações estromais, evidenciadas pela diminuição da atividade das MMPs -2 e -9, reforçando o papel destas enzimas na próstata. Por outro lado, o tratamento com insulina reverteu os parâmetros alterados sugerindo que as alterações prostáticas causadas pelo diabete na próstata, nesta fase do desenvolvimento pós-natal, são aparentemente reversíveis. / Abstract: The Diabetes mellitus is a heterogeneous clinical syndrome caused by lack of insulin production by beta cells (?) or the pancreatic defect in insulin receptors on target cells, resulting in hyperglycemic metabolic disease. Studies have shown that the reproductive and sexual dysfunctions including impotence, decreased libido, impaired spermatogenesis and infertility, decreased sperm quality, are often associated with diabetes in humans and experimental animals. Although vast publications have checked the effects of diabetes on testicular function and fertility in males, the impact of disease on growth and morphogenesis of the prostate is still poor understood. Thus, this study aimed to examine the possible effects of diabetes and insulin replacement, during the maturation process prostate, with special attention to the matrix metalloproteinases 2 and 9 (MMP2 and MMP9), enzymes involved in the extracellular matrix components remodeling during development and growth of prostate. For both, analyze morphological, morphometric, immunocytochemical and biochemical zymography were done in the ventral, dorsolateral and anterior prostatic lobes of Wistar rats, which diabetes was chemically induced at 40 days old by streptozotocin. Our results showed that diabetes impaired th prostatic lobe growth, characterized by a low absolute weight gain and reduction in parenchyma/stroma ratio, mainly in the dorsolateral lobe. Diabetes caused a reduction in the testosterone plasma levels, but not in the dihydrotestosterone. The activities of MMP2 and MMP9 were reduced in the prostatic lobes from diabetic animals. Both simultaneous and late insulin treatment maintain and restored, respectively, the age-matched control morphological and morphometrical parameters. These results allow to conclude that diabetes interfere negatively in the prostate growth during puberty with important stromal changes, including reduction in MMP2 and MMP9 activities, reinforcing the role of these enzymes in the normal prostate morphogenesis. On the other hand, even a late insulin replacement can restore the deleterious effects of diabetes in the prostate during puberty. / Doutorado / Biologia Celular / Doutor em Biologia Celular e Estrutural

Abstract The invasion of a tumour through tissue limiting basement membranes is the critical step in malignant growth. Gelatinases (MMP-2 and MMP-9) are endopeptidases capable of degrading extracellular and pericellular matrix proteins such as collagen IV, the major component of basement membranes. An over-expression of these gelatinases is generally found in malignant tumours and is linked to impaired prognosis in many cancer types. Tissue inhibitors of metalloproteinases (TIMPs), endogenous regulators of the MMP activity, have recently been introduced as multifunctional proteins, which have paradoxical roles in tumour growth. Little data exists on the clinical significance of the gelatinases and TIMPs in gynaecological cancers. In this study the clinical significance of the gelatinases was studied in endometrial and uterine cervical cancers by using immunohistochemical staining with specific antibodies. In epithelial ovarian cancer (EOC) these enzymes and their TIMPs were studied in the preoperative serum samples using ELISA assay. Additionally, sequential serum measurements were performed during chemotherapy to evaluate them as treatment response indicators. In endometrial cancer, MMP-9 positivity correlated to a poor histological differentiation and an advanced clinical stage. High MMP-2 expression correlated to a poor differentiation, and unfavourable survival in stage I cancers, with mortality rates of 5% and 19% in patients with MMP-2 negative versus intensively MMP-2 positive tumours, respectively. In cervical cancers high MMP-2 expression correlated to an increased mortality risk. High MMP-9 expression was connected to a good differentiation of a tumour. In EOC, a high circulating TIMP-1 value correlated to all the examined aggressive features of EOC, including poor survival. The serum measurements of TIMP-1 were uninformative about response evaluation during chemotherapy but paradoxically, an increase in gelatinases and TIMP-2 seemed to reflect a good response to treatment. In conclusion, the data from this study show that high MMP-2 expression in tumour tissue could be prognostic in endometrial and cervical cancer, and preoperative circulating TIMP-1 could serve as an additional prognostic marker in EOC. Studies with larger patient cohorts would be necessary to further explore the value of these enzymes in clinical practice in gynaecological cancers.