Grade 2 gliomas are malignant brain tumours affecting otherwise healthy adults. Although the long-term prognosis is poor, many patients are well and may have a high quality of life for several years. There is, however, a large variability in the natural course of the disease which makes it essential to identify patients who might benefit from early surgery or radio-therapy. The aim of the present thesis was to define new and clinically useful prognostic markers that may assist in the initial treatment decision and in patient follow-up. A retrospective study of 189 patients with gliomas WHO grade 2 showed no advantage in survival of early tumour resection or radiotherapy, and confirmed that histological subtype and patient age are the most important predictors of survival (I). In 89 patients, the pre-treatment uptake of 11C-methionine (MET) measured with positron emission tomography (PET) was identified as a prognostic marker for survival (II). At the time of tumour progression, irradiated tumours demonstrated signs of a residual radiotherapeutic effect that correlated with the pre-treatment uptake of MET (III). Pre-treatment uptake of MET may, therefore, be important both in predicting the natural course of the disease and the response after treatment. Immunohistochemical staining of 40 tumour samples showed an inverse association between the number of tumour cells expressing platelet-derived growth factor alpha receptor (PDGFRa) and survival (IV). Also, a reduction was observed in the number of receptor-positive cells after malignant transformation, supporting the prognostic value of PDGFRa. Lumbar puncture was performed in eight patients with newly diagnosed low-grade gliomas to identify three important growth factors in tumour development. Neither PDGF nor vascular endothelial growth factor (VEGF) were detected in the cerebrospinal fluid (CSF), and fibroblast growth factor 2 (FGF-2) was measurable at extremely low concentrations in two of the patients (V). A proteome screening of the CSF, using two-dimensional gel electrophoresis and mass spectrometry, detected alpha 2-HS glycoprotein at significantly higher concentrations than in a control group (VI). This glycoprotein emerges as a novel substance in glioma research and may be of great interest because of its suggested involvement in the embryonic development of the neocortex.

Adipose tissue (AT) expansion and remodeling that maintains healthy function relies on stromal preadipocytes capable of differentiating into new adipocytes (adipogenesis). During chronic positive energy balance, a relative deficit in adipogenesis, from either a decrease in preadipocyte number or their capacity to differentiate, leads to excessive adipocyte hypertrophy and AT dysfunction. AT contains macrophages whose number and activation state is dynamically regulated with changes in AT mass. This study aims to investigate the effect of macrophage-secreted factors on preadipocyte survival. To assess the effect of macrophage-secreted factors on preadipocytes, murine 3T3-L1 preadipocytes or human primary preadipocytes were incubated with macrophage-conditioned medium (MacCM), prepared from either murine (J774A.1, RAW264.7, bone marrow-derived) or human (THP-1, monocyte-derived) macrophage models, respectively. MacCM inhibited preadipocyte apoptosis and activated pro-survival signaling in both preadipocyte models. Inhibition of PDGFR, Akt, or ERK1/2 reduced the pro-survival effect of MacCM in 3T3-L1 preadipocytes. Inhibition of reactive oxygen species (ROS) generation, or enhancement of ROS clearance, reduced MacCM-dependent 3T3-L1 preadipocyte survival. Whereas anti-inflammatory activated macrophages retained the ability to prevent preadipocyte apoptosis, pro-inflammatory activated macrophages did not. TNF-α immunoneutralization restored the survival activity of pro-inflammatory MacCM on 3T3-L1 preadipocytes. These studies reveal a novel pro-survival effect of MacCM on preadipocytes, and identify signaling molecules (PDGF, Akt, ERK1/2, and ROS) that underlie this action. Macrophage activation was found to regulate the pro-survival activity of MacCM. These in vitro cell culture studies are consistent with a model in which the extent of preadipocyte apoptosis in vivo may determine preadipocyte number and the ability of AT to expand while maintaining healthy function during chronic positive energy balance.

Activated receptor tyrosine kinases recruit many signalling proteins to initiate downstream cell proliferation and survival pathways, including phosphatidylinositol 3-kinase (PI3K), a heterodimer consisting of a p85 regulatory protein and a p110 catalytic protein. Our laboratory has previously shown the p85α protein also has in vitro GTPase activating protein (GAP) activity towards Rab5 and Rab4, small GTPases that regulate vesicle trafficking events for activated receptors. Expression of a p85α protein containing an arginine to alanine substitution at position 274 (p85R274A) that affects its GAP activity, caused sustained levels of activated platelet-derived growth factor receptors (PDGFRs), enhanced downstream signalling, and resulted in cellular transformation. Together with other data, this suggested that in p85R274A-expressing cells, PDGFRs are more rapidly trafficked through the endocytic pathway, which reduces opportunities for sorting events necessary for receptor degradation. Our laboratory has observed previously that p85 was capable of binding to both Rab5-GDP, as well as Rab5-GTP, which is an atypical characteristic of GAP proteins, whereas p110β had previously been reported to bind Rab5-GTP selectively. Based on these observations, this thesis project was designed to test the hypothesis that both proteins contributed GAP activity towards Rab5, with p85 providing a catalytic arginine residue (R274) and p110β providing switch stabilization functions specific to the GTP-bound state. To accomplish the thesis objective, cells expressing individual p85 defects (lacking GAP activity, R274A; or lacking p110-binding ability through deletion of residues 478-513, Δ110) were compared to cells expressing a double mutant missing both functions. Stable clonal NIH 3T3 cell lines were generated and selected in G418 and clones expressing similar levels of FLAG-tagged p85 wild type or mutants compared to the control cell lines (NIH 3T3, FLAG-vector control, p85 wild type, and p85R274A) were chosen for analysis. A time-course of PDGF stimulation showed that cells expressing p85R274A or p85Δ110+R274A have sustained phosphorylation levels of the PDGFR, reduced rates of PDGFR degradation and sustained MAPK/Erk signalling. Contrary to the cellular transformation previously reported for p85R274A-expressing cells, expression of p85Δ110+R274A did not lead to cellular transformation. These divergent results suggest that p85-associated p110 serves two functions. As the catalytic subunit of PI3K, one function is the localized generation of PI3,4,5P3 lipids at the plasma membrane for Akt activation, and possibly during receptor endocytosis where it could impact MAPK/Erk activation/deactivation kinetics and cell transformation. These results support a second function for p110 in the regulation of PDGFR activation/deactivation kinetics and PDGFR half-life, both strongly influenced by alterations in PDGFR trafficking. This suggests that p110β may regulate PDGFR trafficking by providing Rab5-GTP switch stabilization that complements the catalytic arginine residue (R274) within p85, and that p85α and p110β work together as a Rab5 GAP. The role of PDGFR in the localization of the RabGAP function of p85 to specific subcellular compartments was also examined. It was hypothesized that PDGFR may help localize the RabGAP function of p85 to vesicles containing Rab5 or Rab4 through the binding of p85 to phosphorylated tyrosine residues on activated PDGFR. Stable cell lines expressing individual p85 defects (lacking GAP activity, R274A; or lacking PDGFR-binding ability through site-directed mutation of residues 358 and 649 from arginine to alanine, ΔR; or a double mutant missing both functions) demonstrated that p85R274A or p85ΔR+R274A expression leads to sustained PDGFR activation and signalling, and to delayed PDGFR degradation in response to PDGF stimulation. The sustained signalling observed resulted in cellular transformation in cells expressing p85R274A or p85ΔR+R274A. The data suggests that PDGFR does not play a role in the localization of the RabGAP activity of p85. The findings of this study elucidates important non-canonical functions of the PI3K heterodimer and contributes to our understanding of how specific mutations in both p85 and p110β within regions implicated in the regulation of RabGAP activity can alter signalling events and lead to enhancement of tumour-associated phenotypes.

Adipose tissue (AT) expansion and remodeling that maintains healthy function relies on stromal preadipocytes capable of differentiating into new adipocytes (adipogenesis). During chronic positive energy balance, a relative deficit in adipogenesis, from either a decrease in preadipocyte number or their capacity to differentiate, leads to excessive adipocyte hypertrophy and AT dysfunction. AT contains macrophages whose number and activation state is dynamically regulated with changes in AT mass. This study aims to investigate the effect of macrophage-secreted factors on preadipocyte survival. To assess the effect of macrophage-secreted factors on preadipocytes, murine 3T3-L1 preadipocytes or human primary preadipocytes were incubated with macrophage-conditioned medium (MacCM), prepared from either murine (J774A.1, RAW264.7, bone marrow-derived) or human (THP-1, monocyte-derived) macrophage models, respectively. MacCM inhibited preadipocyte apoptosis and activated pro-survival signaling in both preadipocyte models. Inhibition of PDGFR, Akt, or ERK1/2 reduced the pro-survival effect of MacCM in 3T3-L1 preadipocytes. Inhibition of reactive oxygen species (ROS) generation, or enhancement of ROS clearance, reduced MacCM-dependent 3T3-L1 preadipocyte survival. Whereas anti-inflammatory activated macrophages retained the ability to prevent preadipocyte apoptosis, pro-inflammatory activated macrophages did not. TNF-α immunoneutralization restored the survival activity of pro-inflammatory MacCM on 3T3-L1 preadipocytes. These studies reveal a novel pro-survival effect of MacCM on preadipocytes, and identify signaling molecules (PDGF, Akt, ERK1/2, and ROS) that underlie this action. Macrophage activation was found to regulate the pro-survival activity of MacCM. These in vitro cell culture studies are consistent with a model in which the extent of preadipocyte apoptosis in vivo may determine preadipocyte number and the ability of AT to expand while maintaining healthy function during chronic positive energy balance.

O tratamento tópico de feridas visa favorecer um processo de cicatrização eficaz, rápido e seguro. Como uma das opções, o Plasma Rico em Plaquetas – PRP - um concentrado de plaquetas obtido por meio de centrifugação sanguínea ou aférese, vem sendo também utilizado no tratamento de feridas por conter os fatores de crescimento plaquetários. Visto que se trata de uma terapia inovadora, este estudo objetivou buscar as evidências sobre o seu uso na terapia tópica de feridas crônicas de perna. Para tanto, realizou-se revisão sistemática de literatura, seguindo-se as etapas preconizadas pela Colaboração Cochrane. Os estudos foram levantados até 2006, por meio dos descritores platelet rich plasma, platelet derived growth factor, platelet gel, platelet releasate, platelet lysate, CT-102 activated supernatant, wound healing, chronic wound, foot ulcer, diabetic foot, e varicose ulcer, utilizando diferentes combinações, conforme a base de dados consultada (Cochrane, PubMed, Lilacs, Embase e Cinahal). Para a análise da validade interna dos estudos, empregaram-se: a Escala de Jadad, Escala de Avaliação do Grau de Recomendação e Evidência e Escala de Avaliação do Controle das Variáveis. De 56 estudos pré-selecionados, chegou-se à amostra de 18 ensaios clínicos, indexados, principalmente, no PubMed/ Medline (17 / 94,5%), originários dos EUA (12 / 66,6%) e publicados em língua inglesa. Desses, sete (39%) eram ensaios clínicos randomizados, que obtiveram forte recomendação (A) e nível de evidência alto. A partir das metanálises desses ensaios randomizados, em diferentes combinações, os resultados mostraram que o PRP favorece o processo de cicatrização (IC95% 1,84 - 7,41), principalmente em úlceras diabéticas (IC95% 2,94 - 20,31), e quando utilizado como CT-102 (IC95% 2,70-41,40). Concluindo, esta revisão sistemática e metanálise mostram que há evidências científicas sobre os resultados favoráveis do uso do PRP em feridas crônicas de perna, principalmente as de etiologia diabética / The wound topical treatment stimulates an effective, fast and safe wound healing. The platelet rich plasma (PRP) – a concentrated of platelets obtained from centrifugation or single apheresis, has been used as the treatment of wounds because it contains platelet derived growth factor. Being a new therapy, the aim of this study is to show some evidence about the effectiveness of PRP on the healing of chronic wound leg. To this test, a systematic review was conducted, as the recommendation of the Cochrane Library. The studies were screened until 2006 using some key words: platelet rich plasma, platelet derived growth factor, platelet gel, platelet releasate, platelet lysate, CT-102 activated supernatant, wound healing, chronic wound, foot ulcer, diabetic foot, and varicose ulcer; with different combinations, according to data base (Cochrane, PubMed, Lilacs, Embase e Cinahal). From 56 studies, 18 were clinical trials, specially found in PubMed (17 / 94,5%), originated in USA (12 / 66,6%) and published in English. Seven (39%) were clinical trials randomized , classified as a strong recommendation (A) with high evidence level. The meta-analysis of these randomized trials, shows the PRP promotion in wound healing (CI 95% 1,84-7,41), mainly in diabetic ulcer (CI 95% 2,70-41,40). To sum up, this study provides a scientific evidence on repair of chronic wound leg, mainly diabetic ulcer, using PRP

Orientador: Marta Helena Krieger / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T15:25:30Z (GMT). No. of bitstreams: 1 Oliveira_MarianaGoncalvesde_M.pdf: 2027053 bytes, checksum: c424b2043397f6d28d8adebb3fc72bd3 (MD5) Previous issue date: 2013 / Resumo: O óxido nítrico (NO) é um multifuncional agente biológico que nas últimas décadas tem sido alvo de uma infinidade de estudos e constitui hoje um dos mais importantes mediadores de processos intra e extracelulares. Diversos estudos demonstram sua capacidade de prevenção da ativação e adesão plaquetária ou leucocitária e inibição da proliferação e migração de células musculares lisas vasculares (VSMCs), entretanto, em condições de baixa disponibilidade do NO esses processos são prejudicados. Atualmente há o grande interesse no desenvolvimento de compostos capazes de liberar NO de forma modulada e estável, e, nesse sentido, os complexos nitrosilos de rutênio têm se destacado por suas características excepcionais. É amplamente reconhecido que a modulação fenotípica de VSMCs tem papel crítico na progressão de diversas doenças vasculares proeminentes. Sabe-se que o fator de crescimento derivado de plaquetas (PDGF-BB) é um dos principais estimulantes desse processo. Este estudo se propôs a caracterizar os efeitos inibitórios do complexo de rutênio trans-[Ru(NO)Cl(cyclam)](PF6)2, nomeado Ru(cyclam)NO, na modulação fenotípica, resposta proliferativa e migratória de VSMCs induzidas por PDGF-BB. VSMCs foram obtidas por técnica de cultura primária. A citotoxicidade do complexo, na faixa de concentração de 100 ?M a 1500 ?M, foi determinada em ensaios de redução do MTT e incorporação neutral red (NR), e comparadas a do nitroprussiato de sódio (SNP). A concentração 100 ?M foi definida para os demais protocolos experimentais. Western blotting, ensaios transwell e wound healing, e incorporação de timidina triciada foram utilizados para determinação da modulação fenotípica, migração e proliferação celular, respectivamente, e níveis de nitrato no meio foram determinados por quimiluminescência para avaliação do perfil de liberação de NO. O complexo demonstrou baixa citotoxicidade, mesmo na maior concentração e após 48 horas de exposição, reduzindo ao máximo em 30% a porcentagem de células viáveis em ambos os ensaios, demonstrando ser menos tóxico que o SNP. Níveis de nitrato no meio atigiram a concentração máxima após 30 minutos (11 ?M ± 4,8), de maneira mais lenta em relação ao SNP, cuja concentração máxima foi após 5 minutos (13 ?M ± 3,7). A proliferação das VSMCs induzida por PDGF-BB foi inibida, reduzindo à metade a radioatividade incorporada, bem como a expressão do marcador de proliferação PCNA. Observou-se redução de 45% na migração induzida por PDGF-BB nos ensaios transwell, e no wound-healing, embora qualitativo, a redução é notável. A modulação fenotípica da VSMC foi observada pela redução em 60% da expressão da proteína alfa-actina, característica do fenótipo maduro, e foi quase totalmente prevenida pelo tratamento com o complexo. Tal prevenção pode ser mediada pelo fator de transcrição ELK-1, que favorece a expressão de genes de diferenciação, e cuja fosforilação foi estimulada pelo PDGF-BB, porém inibida em quase 50% pelo pré-tratamento com Ru(cyclam)NO. As respostas observadas nos tratamentos com Ru(cyclam)NO foram promissoras, e, embora seu mecanismo de ação x ainda não esteja completamente esclarecido, este complexo demonstrou atividade biológica singular, e sua aplicação em condições clínicas onde há descontrole de processos de proliferação e migração de VSMCs, como a reestenose, apresenta-se como uma proposta interessante / Abstract: Nitric oxide (NO) is a multifuctional biological agent that in the recent decades has been the subject of a plethora of studies and today is one of the most important intracellular and extracellular processes mediators. Several studies have demonstrated its ability to prevent leukocyte or platelet adhesion and activation, and inhibition of vascular smooth muscle cells (VSMCs) proliferation and migration. However, low availability of NO conditions determines impairement of these processes. There is a keen interest in the development of compounds capable of releasing NO modulated so stable, and the nitrosyl ruthenium complexes have gained prominence for its exceptional features. It is widely recognized that the phenotypic modulation of VSMCs, and its uncontrolled proliferation and migration, plays a critical role in the progression of several prominent vascular diseases. It is known that the platelet-derived growth factor (PDGF) is a primary stimulant of the process. This study aimed to determine the inhibitory effects of the ruthenium complex NO donor trans-[Ru(NO)Cl(cyclam)](PF6)2, named Ru(cyclam)NO, in the phenotypic switching, on migratory and proliferative responses of VSMCs induced by PDGF-BB, and its biological response. VSMCs were obtained from primary culture methodology. The complex cytotoxicity were determined by MTT reduction and incorporation of neutral red (NR) assays, in the range of concentration from 100 ?M to 1500 ?M, and compared to sodium nitroprusside (SNP). For the following experimental protocols the concentration of the 100 ?M was set. Western blotting, transwell and wound healing assays, and incorporation of tritiated thymidine were used for determination of phenotypic switching, cell migration and proliferation, respectively. Evaluation of the NO profile release was determined as nitrate levels in the culture medium by chemiluminescence. The complex Ru(cyclam)NO showed low cytotoxicity even at the highest concentration evaluated and after 48 hours of exposition, reducing only 30% the percentage of viable cells in both trials, showing be less toxic than the SNP. Medium nitrate levels exibhited the highest concentration after 30 min (11 ?M ± 4.8), slower when compared to SNP, which reached the maximum concentration after 5 minutes (13 ?M ± 3.7). The proliferation of VSMCs induced by PDGF-BB was inhibited by half of the radioactivity incorporated counting, as well as the reduction on the expression of the proliferation marker PCNA. Observed a reduction by 45% in the migration induced by PDGF-BB determined in transwell assays, and on the wound-healing, although a qualitative result, the reduction of migration induced by PDGF-BB. The 60% reduction by PDGF-BB treatment of the contractile protein expression ?-SMA, characteristic of mature phenotype, revealed the modulation of VSMCs phenotype, and it was almost completely prevented by treatment with the complex. Such prevention was associated with inhibition by almost 50% of phosphorylation of the transcription factor ELK-1 stimulated by PDGF-BB. The responses determined with complex treatments revealed promising for future development of cardiovascular devices. Although its xii mechanism of action is not completely understood, this complex showed singular biological activity, and its application in some clinical conditions where there is uncontrolled proliferation and migration of VSMCs presents as a substancial proposal / Mestrado / Fisiologia / Mestra em Biologia Funcional e Molecular

Adipose tissue (AT) expansion and remodeling that maintains healthy function relies on stromal preadipocytes capable of differentiating into new adipocytes (adipogenesis). During chronic positive energy balance, a relative deficit in adipogenesis, from either a decrease in preadipocyte number or their capacity to differentiate, leads to excessive adipocyte hypertrophy and AT dysfunction. AT contains macrophages whose number and activation state is dynamically regulated with changes in AT mass. This study aims to investigate the effect of macrophage-secreted factors on preadipocyte survival. To assess the effect of macrophage-secreted factors on preadipocytes, murine 3T3-L1 preadipocytes or human primary preadipocytes were incubated with macrophage-conditioned medium (MacCM), prepared from either murine (J774A.1, RAW264.7, bone marrow-derived) or human (THP-1, monocyte-derived) macrophage models, respectively. MacCM inhibited preadipocyte apoptosis and activated pro-survival signaling in both preadipocyte models. Inhibition of PDGFR, Akt, or ERK1/2 reduced the pro-survival effect of MacCM in 3T3-L1 preadipocytes. Inhibition of reactive oxygen species (ROS) generation, or enhancement of ROS clearance, reduced MacCM-dependent 3T3-L1 preadipocyte survival. Whereas anti-inflammatory activated macrophages retained the ability to prevent preadipocyte apoptosis, pro-inflammatory activated macrophages did not. TNF-α immunoneutralization restored the survival activity of pro-inflammatory MacCM on 3T3-L1 preadipocytes. These studies reveal a novel pro-survival effect of MacCM on preadipocytes, and identify signaling molecules (PDGF, Akt, ERK1/2, and ROS) that underlie this action. Macrophage activation was found to regulate the pro-survival activity of MacCM. These in vitro cell culture studies are consistent with a model in which the extent of preadipocyte apoptosis in vivo may determine preadipocyte number and the ability of AT to expand while maintaining healthy function during chronic positive energy balance.

No description available.

Nous avons découvert que TRF2 est aussi sur-exprimée au niveau des cellules endothéliales de nombreux types de cancers humains alors qu’elle n’est pas détectable dans les vaisseaux des tissus sains adjacents. Des cellules endothéliales extraites de tumeurs ex-vivo manifestent une expression supérieure de TRF2, une migration et une prolifération accrues et une aptitude à former des tubules élevée, comparées aux endotheliums isolées de tissus sains. La sur-expression de cette protéine in vitro dans des cellules endothéliales primaires et ex-vivo entraine l’augmentation de la prolifération, de la migration et de la capacité de ces dernières à former des tubules. La diminution de l’expression de TRF2 conduit à l’effet inverse. Par ailleurs, la modulation de l’expression de TRF2 n’affecte pas la proportion de cellules apoptotiques. De même, les variations des niveaux d’expression de TRF2 n’induisent aucune réponse aux dommages à l’ADN et les modifications des facultés angiogéniques sont indépendantes d’ATM. Les effets angiogéniques de TRF2 semblent donc distincts des fonctions télomériques. Etant donné que le facteur de transcription WT1 (Wilms’ tumour suppressor 1) est fortement exprimé dans les vaisseaux de tumeurs humaines et régule les propriétés angiogéniques des cellules endothéliales, nous nous sommes penché sur la régulation potentielle de TRF2 par WT1. WT1 se lie en effet sur le promoteur de TRF2 pour activer sa transcription. Enfin, nous avons démontré que l’activité angiogénique de TRF2 réside en partie dans sa capacité à se fixer sur le promoteur du gène codant pour le récepteur angiogénique à activité tyrosine kinase PDGFRβ et à activer sa transcription. / We discovered that TRF2 is expressed in endothelial cells of many human cancer types but not in the vessels of healthy adjacent tissues. Endothelial cells derived from tumours ex vivo exhibited a significantly increased TRF2 expression, and a higher migration, proliferation and tube formation potential as endothelium obtained from healthy tissues. In vitro TRF2 over-expression in primary or ex vivo endothelial cells resulted in an increased proliferation, migration and tube formation, while silencing of TRF2 led to the opposite results. No changes in apoptosis could be observed. Interestingly, modulation of TRF2 in endothelium does not induce DNA damage responses and the observed changes in the angiogenic behaviour are ATM –independent. The angiogenic effects of TRF2 seem therefore to be uncoupled from its telomeric function. Since the transcription factor WT1 (Wilms’ tumour suppressor 1) is highly expressed in human tumour vessels and mediates angiogenic properties of endothelial cells, we investigated whether TRF2 expression could be regulated by WT1. Indeed, WT1 binds the TRF2 promoter and activates its transcription. Finally, we demonstrated that TRF2 promotes angiogenesis by binding to the promoter of the gene encoding for the angiogenic tyrosine kinase receptor PDGFRβ and activating its transcription.

A neuraminidase 1 (Neu1) é a enzima que regula o catabolismo de sialoglicoconjugados nos lisossomos. A deficiência da Neu1 é a base da sialidose, doença grave associada a um amplo espectro de manifestações, incluindo hipotonia e fraqueza muscular. Camundongos com deficiência de Neu1 desenvolvem degeneração muscular caracterizada principalmente por atrofia, invasão das fibras musculares por fibroblastos e expansão da matriz extracelular. A Neu1 controla a proliferação de fibroblastos de pacientes por meio da desialilação dos receptores de PDGF e IGF. Além disso, há enzimas lisossomais que são moduladas pela Neu1, tais como as catepsinas, que são capazes de degradar componentes musculares e estariam excessivamente ou erroneamente ativas (sialiladas) em decorrência da deficiência de Neu1. O objetivo deste trabalho foi identificar se o fenótipo da musculatura esquelética de camundongos Neu1-/- poderia estar associado à atividade do IGF-1R, PDGFR e/ou à sialilação de catepsina B, através da análise histológica e proteica de músculos esqueléticos e fibroblastos de camundongos Neu1+/+ e Neu1-/- tratados com inibidores de IGF1-R e PDGFR. O estudo da expressão proteica de catepsina B foi realizado nos músculos tratados com os inibidores de IGF-1R e PDGFR, e nas frações citosólica e lisossomal de fibroblastos tratados com neuraminidase exógena. Em comparação com camundongos Neu1+/+, os músculos de animais Neu1-/- apresentam menor área de fibra, peso corporal, expressão de pAkt e maior expressão de catepsina B; e os fibroblastos Neu1-/- exibem maior proliferação e expressão de pAkt. A inibição do IGF-1R em camundongos Neu1-/- aumentou a área das fibras musculares, expressão de pAKt e diminuiu a expressão de catepsina B; em relação aos fibroblastos Neu1-/-, entretanto aumentou a proliferação celular com diminuição de pAkt. A inibição do PDGFR em músculos de camundongos Neu1-/- levou ao aumento da expressão de pAkt, da área das fibras, com diminuição de pERK e catepsina L, quando comparados com os controles Neu1-/-; a mesma inibição in vitro conduziu à diminuição da expressão de pAkt, pERK e proliferação. A catepsina B encontra-se bastante ativa na fração lisossomal e o tratamento com neuraminidase foi eficaz na correção de seu peso molecular e compartimentalização lisossomal. De forma geral, o fenótipo muscular de camundongos Neu1-/- parece estar relacionado com a atividade de IGF-1R e PDGFR, e a catepsina B hipersialilada é potencialmente deletéria para o músculo esquelético / Neuraminidase 1 (Neu1) is an enzyme that regulates the catabolism of sialoglycoconjugates in lysosomes. Neu1 deficiency is the basis of sialidosis, a severe disease associated with a broad spectrum of manifestations, including hypotonia and muscle weakness. Neu1 deficient mice develop muscular degeneration characterized by atrophy, invasion of muscle fibers by fibroblasts, and expansion of the extracellular matrix. Neu1 controls the proliferation of fibroblasts from patients through the desialylation of PDGF and IGF receptors. In addition, lysosomal enzymes are modulated by Neu1, such as cathepsins, which degrade muscle components and are excessively or erroneously active (sialylated) as a result of Neu1 deficiency. The aim of this study was to identify whether skeletal muscle phenotype of Neu1-/- mice may be associated with IGF-1R, PDGFR and/or sialylation of cathepsin B, through protein and histological analysis of skeletal muscles and fibroblast from Neu1+/+ and Neu1-/- mice treated with IGF-1R and PDGFR inhibitors. The study of cathepsin B protein expression was performed in skeletal muscles treated with IGF-1R and PDGFR inhibitors, and in the cytosolic and lysosomal fractions of fibroblasts treated with exogenous neuraminidase. Compared with Neu1+/+ animals, Neu1-/- muscles showed smaller muscle fiber area, body weight, pAkt expression and higher cathepsin B expression; and Neu1-/- fibroblasts exhibited increased proliferation and expression of pAkt. The inhibition of IGF-1R Neu1-/- mice increased the area of muscle fibers, expression of pAkt and decreased expression of cathepsin B; but, considering Neu1-/- fibroblasts, there was increased cell proliferation with reduction of pAkt. The inhibition of PDGFR in muscles of Neu1-/- mice led to increased expression of pAkt, muscle fiber area, with decreased expression of pERK and cathepsin L, when compared with the Neu1-/- controls; the same inhibition in vitro led to reduced expression of pAkt, pERK and cell proliferation. Cathepsin B presented high activity in the lysosomal fraction and the treatment with neuraminidase was effective in the correction of its molecular weight and lysosomal compartmentalization. In general, the muscular phenotype of Neu1-/- mice is possibly related to IGF-1R and PDGFR activity, and oversialylated cathepsin B is potentially deleterious for the skeletal muscle