Orientador: Fernando Cendes / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-19T22:33:32Z (GMT). No. of bitstreams: 1 Fernandes_DanielaAlves_M.pdf: 5738934 bytes, checksum: 206ca01256f59acd22d595142eadef05 (MD5) Previous issue date: 2012 / Resumo: A epilepsia é a segunda patologia neurológica que mais afeta a população mundial e a epilepsia do lobo temporal é a mais comum das epilepsias focais com maior frequência, associada à refratariedade e a distúrbios de memória. A lesão epileptogênica pode ser estudada através de imagens de ressonância magnética (RM), EEG entre outros exames, facilitando a indicação da cirurgia que tem como objetivo reduzir as crises, entretanto a função cognitiva pode ser afetada. Nosso propósito foi avaliar a eficácia da cirurgia quanto ao controle das crises, desempenho de memória nos pacientes e comparar os resultados de avaliações neuropsicológicas pré e pós-operatórias quanto à frequência de crises, abordagem cirúrgica e volumetria da estrutura e lacuna. Foram analisados 55 pacientes submetidos à cirurgia de epilepsia e 29 indivíduos controles saudáveis. Todos foram submetidos à aquisição de imagens em um aparelho de RM 2T em dois tempos com intervalos médios de 4,4±2,8 anos. A volumetria do hipocampo e lacuna cirúrgica foi realizada com o software Display, utilizando o protocolo de Bonilha et at., 2004. Os pacientes foram avaliados de acordo com a classificação pós-operatória de Engel Jr., 1997 e testes de avaliações neuropsicológicas que incluiu subtestes: da Wechsler Memory Scale-Revised; Wechsler Adult Intelligence Scale-Revised e Rey Auditory Verbal Learning Test, realizados com um intervalo médio de 8,5 anos após a cirurgia. Para análise dos dados pré/pós-operatórios foi utilizado o teste T de Student, de Wilcoxon ou Man-Whitney, correlação de Pearson e Spearman de acordo com as características das variáveis. Observamos diferença significativa nas avaliações neuropsicológicas, quando comparadas entre os próprios indivíduos nos grupos pré e pós-operatório e nos grupos de acordo com a Escala de Engel e lado da cirurgia, com piora no desempenho cognitivo no período pós-operatório. Mostraram melhora no desempenho de memória os que foram classificados em Engel IA quando comparados aos indivíduos que continuaram com crises. Os grupos submetidos à amigdalohipocampectomia seletiva e ressecção cortical associada à amigdalohipocampectomia não apresentaram diferenças significativas quanto ao desempenho de memória. O grau de escolaridade influenciou negativamente no resultado das avaliações. Nossos resultados mostraram que 80% foram classificados em Engel I, 18,2% em Engel II e 1,8% em Engel III. A volumetria dos hipocampos direito e esquerdo ipsilaterais à cirurgia apresentaram valores menores que dos controles. Os hipocampos esquerdos e direitos pré/pósoperatórios contralaterais à atrofia não apresentaram resultado significativo quando comparado as volumetrias dos controles e nem quando comparados entre si. Os dados de volumetria associados à memória apresentaram grau razoável e moderado de correlação. Nossos resultados mostraram que após a ressecção cirúrgica existe um declínio no desempenho de memória, porém a frequência de crises desses pacientes diminui significativamente e que o tipo de abordagem cirúrgica não interfere no desempenho de memória, mesmo com o maior volume da ressecção de pacientes submetidos à lobectomia temporal anterior / Abstract: Epilepsy is the second most frequent neurological disease and temporal lobe epilepsy is the most common form of focal epilepsy and is more frequently associated to refractoriness and memory decline. The epileptogenic lesion can be studied through magnetic resonance imaging, EEG and other tests, facilitating the indication of surgery that aims to reduce seizure frequencies, however cognitive function may be affected. We proposed to evaluate the effectiveness of surgery on the management of seizures, memory performance and compare the results of neuropsychological assessments pre/post-operative and the frequency of seizures, surgical approach and volume of hippocampi and amount of resection. We analyzed 55 patients who underwent epilepsy surgery and 29 healthy control subjects. All patients underwent imaging in a 2T MR scanner in two steps with intervals of 4.4 ± 2.8 years. The volumetry of the hippocampus and surgery gap was done with the software display, using the protocol described by Bonilha et al, 2004. Patients were evaluated according to the postoperative classification of Engel Jr., 1997 and neuropsychological evaluation that included the subtests, Wechsler Memory Scale-Revised, Wechsler Adult Intelligence Scale-Revised and Rey Auditory Verbal Learning Test, performed with an average interval 8.2 years after surgery. For data analysis, pre/post-operative we used the Student t test, Wilcoxon or Man-Whitney, Pearson and Spearman correlation test according to the characteristics of variables. We observed significant differences in neuropsychological evaluations, when we compared the groups pre/post-operative and the groups according to the Engel Scale and side of surgery, with worsening of cognitive performance in the postoperative period. There was a relative improvement in memory performance in those who were classified as Engel IA compared to individuals who continued to have seizures. The comparisons between the groups of selective amygdalohippocampectomy and anterior temporal lobe resection showed no significant differences in memory performance. The level of education had a negative influence on the outcome of neuropsychological evaluations. Our results showed that 80% were classified as Engel I, 18.2% in Engel II and 1.8% in Engel III. The volumetry of the left and right hippocampi ipsilateral to the surgery showed lower values than controls. The left and right hippocampus pre/post-operative contralateral to the side of surgery did not show significant differences when compared to controls. There was a correlation between hipocampal volumes and memory performance. Our results showed that after surgical resection there is a decline in memory performance, but the frequency of seizures in these patients decreased significantly and that the types of surgical approaches do not differ in terms of post-operative nor the total volume of the surgical lacunae / Mestrado / Neurociencias / Mestre em Ciências

Orientador: Luciana Asprino / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-20T21:58:02Z (GMT). No. of bitstreams: 1 Freire_SimeiAndredaSilvaRodrigues_D.pdf: 1346412 bytes, checksum: 35f4f1e8f5181ebc1bf60768170c3f2c (MD5) Previous issue date: 2012 / Resumo: O objetivo desse trabalho foi avaliar a epidemiologia e características do tratamento de fraturas de mandíbula atrófica; comparar a resistência mecânica e a distribuição de tensões de três técnicas de fixação interna aplicadas em fraturas de mandíbulas atróficas. CAPÍTULO I: Dados foram coletados de pacientes vítimas de fraturas em mandíbulas atróficas em um período de dez anos (1999-2009). Os dados analisados continham informações demográficas e socioeconômicas, etiologia dos traumas, diagnóstico, tipos de traumas, deslocamento das fraturas, o método de fixação utilizado, região das fraturas, traumas associados, tempo decorrido entre o trauma e tratamento. A principal causa das fraturas em mandíbulas atróficas foi a queda da própria altura, acometendo principalmente pacientes do gênero masculino desempregados. A faixa etária mais acometida foi a de trinta a sessenta anos ocorrendo predominantemente fraturas bilaterais na região de corpo mandibular. CAPÍTULO II: Avaliou-se a resistência, in vitro, por meio de testes de carregamento linear a fixação de fraturas de mandíbulas atróficas com defeito de continuidade por meio de três sistemas de fixação. Foram utilizadas réplicas de mandíbulas humanas atróficas de poliuretano submetidas a simulação de fratura com defeito de continuidade de 15mm em corpo direito, fixadas pelos seguintes sistemas: Grupo 1 - sistema 2,4mm conventional, Grupo 2 - sistema 2,4 mm com travamento e Grupo 3 - sistema 2,0mm com travamento. Pelos resultados obtidos o sistema com travamento aumentou a resistência da fixação pela melhor e mais favorável distribuição de cargas, os sistemas 2,4mm de fixação interna estável testados apresentaram adequada resistência mecânica para tratamento de fraturas de mandíbulas atróficas com defeito de continuidade. CAPÍTULO III: Avaliou-se, in sílico, pelo método de elementos finitos a fixação de fraturas de mandíbulas atróficas por meio de três sistemas de fixação submetidas a testes de carregamento linear. Os modelos criados em elementos finitos de fratura de mandíbula atrófica foram fixados pelos seguintes sistemas, sistema 2,4mm convencional, sistema 2,4mm com travamento e sistema 2,0mm com travamento. Pelos resultados obtidos o sistema 2,4mm convencional demonstrou suportar toda carga aplicada nesta simulação, os sistemas com travamento apresentaram dissipação das tensões para região anterior e posterior da mandíbula e no sistema de fixação convencional as tensões se localizaram na porção entre os furos do sistema de fixação assim como no sistema com travamento, porém este ainda apresentou dissipação para os parafusos, com concentração crescente para região apical dos parafusos próximos a simulação do traço de fratura / Abstract: The aim of this study was to evaluate the epidemiology and treatment characteristics of atrophic mandibular fractures; to compare the mechanical strength and stress distribution of three internal fixation techniques applied in atrophic mandibles fractures treatment. CHAPTER I: Data were collected from patients suffering from atrophic mandibles fractures in a period of ten years (1999-2009). The data analyzed contained demographic and socioeconomic characteristics, etiology of trauma, diagnosis, types of trauma, displacement of the fractures, the region of fracture, associated trauma, time elapsed between trauma and treatment. The main cause of fractures in atrophic mandibles was fall accidents, affecting mainly male unemployed patients. The age group most affected was between thirty to sixty years occurring bilateral fractures predominantly in the mandibular body region. CHAPTER II: Resistance was evaluated in vitro by linear loading test in the atrophic mandible fracture simulation with continuity defect by three stable internal fixation systems. It was used replicas of human atrophic polyurethane mandible subjected to simulated fracture defect continuity of 15mm in right body region, fixed by the following systems: Group 1 - 2.4mm conventional system, Group 2 - 2.4 mm locking system and Group 3 - 2.0mm locking system. The results obtained with the locking system increased the resistance setting for the better and more favorable load distribution, the 2.4mm stable internal fixation systems tested had adequate mechanical strength for the treatment of fractures of atrophic mandibles with continuity defects. CHAPTER III: It was evaluated ,in silico, by the method of finite elements with linear force three stable internal fixation systems for the treatment of atrophic mandible fractures. The finite element models created in the atrophic mandible fracture were fixed by the following systems, the conventional system 2.4mm, 2.4mm locking system and 2.0mm locking system. The results obtained demonstrated that the locking system increased the resistance by a favorable and better distribution of the stresses during the loading test when applied in atrophic mandible fractures with continuity defects. The three internal fixation systems tested in this study showed adequate mechanical efficiency to be applied in atrophic mandible fractures with continuity defects treatment / Doutorado / Cirurgia e Traumatologia Buco-Maxilo-Faciais / Doutor em Clínica Odontológica

Spinal Muscular Atrophy (SMA) is an autosomal, recessive form of childhood motor neuron disease and the most common genetic cause of infant mortality in the western world. SMA displays the characteristic hallmarks of a motor neuron disease, including loss of motor neurons in the spinal cord and atrophy of skeletal muscles. However, mounting evidence suggests that multiple tissues and body systems, beyond the neuromuscular system, are affected in SMA. Previous studies have highlighted alterations in the vascular system in both SMA patients and in a variety of mouse models of the disease, reporting alterations in vessel structure and perfusion abnormalities in peripheral tissues. In this project a detailed morphological investigation of the capillary beds of skeletal muscle and the spinal cord, two of the key pathological tissues in SMA was undertaken. This work was conducted in the Smn-/-;SMN2, Smn-/-;SMN2tg/+ and Smn-/-;SMN2;Δ7 mouse models of SMA. Significant alterations in the form and extent of the skeletal muscle and spinal cord capillary bed in SMA mice were identified, the most striking of which being a reduction in capillary density in SMA tissue when compared to control littermate tissue. In skeletal muscle, this reduction in capillary density was found to be a postnatal phenomenon, which occurred independently of denervation, in a variety of phenotypically distinct muscles and in all three SMA mouse models investigated. In the spinal cord, the capillary defect was seen to develop in a similar postnatal pattern to that observed in skeletal muscle. Importantly, a reduction in capillary density was observed in the ventral horn of the spinal cord, which houses motor neuron cell bodies, a known pathological target in SMA. These motor neurons were seen to be surrounded by fewer capillaries than their control counterparts. Using an injectable marker of hypoxia, it was determined that the cells of the ventral horn of SMA spinal cords are hypoxic. This suggests that the capillary defect identified has a functional impact on the tissues it is observed in. Having established the presence of capillary defect in SMA tissue, the effect of potential SMA therapeutics on the capillary defect was then investigated. The effect of HDAC inhibitors, which have been successfully shown to increase the levels of the disease causing Smn protein, was investigated. Treatment with the HDAC inhibitor SAHA was found to ameliorate the capillary defect, significantly improving capillary density in SMA skeletal muscle. This implies that the capillary defect is related to Smn levels in tissue and is amenable to therapeutics which increase Smn levels. Having characterised the capillary defect in SMA tissues in detail, a selection of tools were then used to investigate the underlying mechanisms resulting in the defect. First, using primary cell cultures, the growth and morphology of the key cellular component of capillaries, the endothelial cell, was examined. While displaying reduced levels of the Smn protein, endothelial cells isolated from SMA tissues showed no difference in growth rate, morphology or endothelial cell marker expression when compared to endothelial cells isolated from control tissue. This suggests that the defects seen in SMA capillary beds are not the result of defects in the structure and growth of endothelial cells. Second, retinas from SMA mice were found to exhibit similar capillary defects to those observed in SMA skeletal muscle and spinal cord. Given the entirely postnatal development of the retinal capillary network, the retina was identified as a useful experimental preparation for the further investigation of the mechanisms underlying the capillary defect in SMA. In summary, this work highlights the incidence and importance of capillary defects in mouse models of spinal muscular atrophy.

The childhood neuromuscular disease spinal muscular atrophy (SMA) is caused by low levels of survival motor neuron (SMN) protein. Historically, SMA has been characterised as a disease primarily affecting lower motor neurons. However, recent breakthroughs have revealed defects in other non-neuronal cells and tissues. In vivo analysis of peripheral nerve showed defects in Schwann cells, manifesting as abnormal myelination and delayed maturation of axo-glia interactions. The experiments in this thesis were designed to build on these observations and examine whether Schwann cell defects are intrinsic and occur as a primary result of low levels of SMN in that cell type, or rather represent a secondary consequence of pathology in neighbouring motor neurons. I initially developed a protocol to allow isolation of high-yields of purified, myelination-competent Schwann cells from ‘Taiwanese’ SMA mice. SMA-derived Schwann cells had significantly reduced SMN levels and failed to respond normally to differentiation cues. Increasing SMN levels restored myelin protein expression in Schwann cells from SMA mice. Perturbations in expression of key myelin proteins were likely due to failure of protein translation and/or stability rather than transcriptional defects. Co-cultures of healthy neurons with SMA Schwann cells revealed a significant reduction in myelination compared to cultures where wild-type Schwann cells were used. The presence of SMA Schwann cells also disrupted neurite stability. Perturbations in the expression of key extracellular matrix proteins, such as laminin α2, in SMA-derived Schwann cells suggests that Schwann cells were influencing neurite stability by modulating the composition of the extracellular matrix. Previous studies have demonstrated that low levels of SMN lead to disruption of ubiquitin homeostasis and decreased expression of ubiquitin-like modifier activating enzyme (UBA1) in the neuromuscular system, driving neuromuscular pathology via a beta-catenin dependent pathway. Label-free proteomics analysis of SMA and control Schwann cells identified 195 proteins with modified expression profiles. Bioinformatic analysis of these proteins using Ingenuity Pathway Analysis (IPA) software confirmed that major disruption of protein ubiquitination pathways was also present in Schwann cells from SMA mice. Immunolabeling and proteomics data both revealed that UBA1 levels were significantly reduced in SMA-derived Schwann cells. However, loss of UBA1 in Schwann cells did not lead to downstream modifications in beta-catenin pathways. Pharmacological inhibition of UBA1 in healthy Schwann cells was sufficient to induce defects in myelin protein expression, suggesting that UBA1 defects contribute directly to Schwann cell disruption in SMA. I conclude that low levels of SMN induce intrinsic defects in Schwann cells, mediated at least in part through disruption to ubiquitination pathways.

The disruption of the survival motor neuron (SMN1) gene leads to the children’s genetic disease spinal muscular atrophy (SMA). SMA is characterized by the degeneration of α-motor neurons and skeletal muscle atrophy. Although SMA is primarily considered a motor neuron disease, the involvement of muscle in its pathophysiology has not been ruled out. To gain a better understanding of the involvement of skeletal muscle pathophysiology in SMA, we have developed three aims: to identify cell-specific Smn-interacting proteins, to characterize postnatal skeletal muscle development in mouse models of SMA, and to assess the functional capacity of muscles from SMA model mice. We have used tandem affinity purification to discover Smn interacting partners in disease relevant cell types. We have identified novel cell-specific Smn interacting proteins of which we have validated myosin regulatory light chain as a muscle-specific Smn associated protein in vivo. We have taken advantage of two different mouse models of SMA, the severe Smn-/-;SMN2 mouse and the less severe Smn2B/- mouse, to study the postnatal development of skeletal muscle. Primary myoblasts from Smn2B/- mice demonstrate delayed myotube fusion and aberrant expression of the myogenic program. In addition, the expression of myogenic proteins was delayed in muscles from severe Smn-/-;SMN2 and less severe Smn2B/- SMA model mice. Muscle denervation and degeneration, however, are not the cause of the aberrant myogenic program. At the functional level, we demonstrate a significant decrease in force production in pre-symptomatic Smn-/-;SMN2 and Smn2B/- mice indicating that muscle weakness is an early event in these mice. Immunoblot analyses from hindlimb skeletal muscle samples revealed aberrant levels of developmentally regulated proteins important for muscle function, which may impact muscle force production in skeletal muscle of SMA model mice. The present study demonstrates early and profound intrinsic muscle weakness and aberrant expression of muscle proteins in mouse models of SMA, thus demonstrating how muscle defects can contribute to the disease phenotype independently of and in addition to that caused by motor neuron pathology.

Spinal muscular atrophy (SMA) is the most commonly inherited neurodegenerative disease that leads to infant mortality worldwide. There are no known cures for SMA, but small increase in protein levels of SMN can be beneficial. We have developed adenoviral (Ad) vectors that express a human transgene of SMN and have tested their safety in vitro. We have demonstrated that these viruses can effectively express the transgene following cell entry and that the levels are relative to the virus dose. The viruses do not appear to impact the health and function of the cells, and are capable of increasing the number of Gems. We also attempted to change the tropism of the viruses through fiber protein modifications in order to target muscles and motor neurons. Our results suggest that a therapy based on an Ad-SMN fiber-modified vector may ultimately be successful in treating patients of SMA.

The autosomal recessive disorder spinal muscular atrophy (SMA) causes motor neuron degeneration and muscle wasting, progressing to paralysis and death in severe cases. The disease is caused by deficiency of survival motor neuron protein (SMN) due to deletion or mutation of the SMN1 gene. We seek to develop a protein-based therapy for SMA using an adenoviral vector which encodes a secretable form of SMN fused to a protein transduction domain (PTD) derived from the trans-acting activator of transcription (TAT) from HIV. We generated secretable GFP proteins using transient transfection in mammalian cells and determined that the secretory peptide was inefficient when paired with the native PTD. We generated TAT-GFP proteins in bacteria and observed that the variant TAT3 most reliably tranduced cells in vitro. We did not observe uptake of the therapeutic protein following infection with an adenoviral vector and subsequent secretion of the protein from infected cells.

Orientador: Fernando Cendes / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T23:40:05Z (GMT). No. of bitstreams: 1 Fernandes_DanielaAlves_D.pdf: 7319654 bytes, checksum: db061970bef101957f53339b302dc1a9 (MD5) Previous issue date: 2015 / Resumo: A quantificação da atrofia das estruturas temporais mesiais pelas imagens de RM em pacientes com ELTM permite a identificação "in vivo" das alterações anatômicas associadas à esta patologia e sua correlação com dados neuropsicológicos, permitem a definição de um bom diagnóstico. Apesar da comprovação da eficácia do tratamento cirúrgico para o controle das crises, ainda não são claros os efeitos da ressecção de estruturas mesiais do lobo temporal na performance cognitiva, a longo prazo. O objetivo deste trabalho foi realizar um estudo prospectivo de uma série de pacientes com ELTM submetidos ao tratamento cirúrgico, comparando os resultados pré/pós-cirúrgicos obtidos a partir da análise de dados de avaliação neuropsicológica (ANP) e imagens de RM. No estudo avaliamos 119 indivíduos, 88 pacientes e 31 controles saudáveis. Os pacientes foram classificados de acordo com a escala proposta por Engel para controle de crises após a cirurgia. Avaliamos o coeficiente de inteligência estimado (QIe), as memórias verbal e não verbal com testes utilizados na rotina de investigação pré-operatória do nosso serviço. Utilizamos análises longitudinais específicas realizadas com o software SPSS®22. Consideramos p?0,05. Quanto à classificação de Engel observamos que 73,7% dos pacientes foram classificados em Engel I; 17,1% foram classificados em Engel II; 8,0% foram classificados em Engel III e 1,1% classificado em Engel IV com tempo médio de seguimento de 8,8 anos após a cirurgia. Para as ANP pré/pós-operatória observamos um declínio de memória para esses pacientes relacionado ao controle de crises e lado da cirurgia (p<0,0001). Para as ANP pós-operatórias realizadas em dois tempos diferentes e separadas em grupos quanto ao controle de crises e lado da cirurgia, não observamos diferença significativa, entretanto todos os testes indicaram uma tendência de melhora no desempenho de memória e QIe. Observamos maior volume hipocampal para os controles (média=3706±842), volume reduzido do hipocampo contralateral para pacientes livre de crises (média=3602±711) e menor ainda para os pacientes com presença de crises (média=3284±521). Para a análise de intensidade de sinal no lobo temporal contralateral dos pacientes, observamos uma diferença significativa (p=0,005) entre controles e pacientes com média menor para os controles. Também observamos alterações com diferença significativa para os tratos analisados em imagens de DTI, no lado ipsilateral à cirurgia comparados aos controles. Nossos resultados mostram bom controle de crises após a cirurgia, mesmo após um longo período. Entretanto observamos que após a cirurgia existe um declínio na performance dos testes neuropsicológicos para muitos pacientes, independente do lado operado. Porém, a análise a longo prazo mostra que existe uma recuperação parcial desse declínio, que pode estar associada a interação entre fatores de aprendizado e plasticidade cerebral; ou seja, podemos inferir que de alguma forma a melhora no controle das crises permite uma "recuperação" da eficiência cognitiva a longo prazo / Abstract: The quantification of the atrophy of the mesial temporal structures by MR images in patients with TLE allows identification "in vivo" of the anatomical changes associated with this disease and its correlation with neuropsychological data, allowing for the establishment of a proper diagnosis. Despite the evidence of the effectiveness of surgical treatment for seizure control, it is not yet clear the effects of resection of the mesial temporal lobe structures in cognitive performance in the long-term follow up. The aim of this study was to evaluate prospectively of a series of patients with TLE undergoing epilepsy surgery, comparing pre/post-surgical results obtained from neuropsychological assessment (NPA). We included 119 subjects, 88 patients and 31 healthy controls. Of the 113 patients, 88 had two NPA and 60 underwent two MRIs after surgery. Patients were classified according to Engel¿s scale. We evaluated the estimated intelligence coefficient (eIQ), the verbal and non-verbal memories with tests used in preoperative routine. We use specific longitudinal analyzes with SPSS®22 software. We observed that 73.7% of patients were classified as Engel I; 17.1% were classified as Engel II; 8.0% were classified as Engel III and 1.1% classified as Engel IV after surgery with a mean follow up of 8.8 years. The NPA pre/postoperative showed a memory decline for these patients related to seizures control and side of surgery (p<0.0001). We found no significant difference between the postoperative NPAs carried out in two different times and separated in groups regarding seizure control and side of surgery; however, all tests indicated a trend towards improvement trend in memory performance and eIQ. We observed a larger hippocampal volume for the controls (mean=3706±842), in comparison with seizure free patients (mean=3602±711) and smaller hippocampal contralateral volumes for patients with persistent seizures after surgery (mean=3284±521). We observed a significant difference (p=0.005) in T2 signal between patients and controls (increase in patients). We also observed changes with a significant difference to the white matter tracts analyzed with diffusion tensor images, in the ipsilateral side of surgery compared to controls. Our results show good seizure control after surgery, even after a long period of follow up. However, our results showed that after surgery there is a decline in the performance on neuropsychological tests for most patients, regardless of the side of surgery. However, the long-term repeated analysis showed that there is partial recovery that may be associated with the interaction between learning effect and brain plasticity. We can hypothesize that the improvement in seizure control after surgery allows "recovery" of the long-term cognitive efficiency / Doutorado / Fisiopatologia Médica / Doutora em Ciências

Living cells maintain stable transcriptional programs while exhibiting plasticity that allows them to respond to environmental stimuli. The Polycomb repressive complex 2 (PRC2) is a key regulator of chromatin structure that maintains gene silencing through the methylation of histone H3 on lysine 27 (H3K27me), establishing chromatin-based memory. Two variants of PRC2 are present in mammalian cells, PRC2-EZH2 which is predominantly present in differentiating cells, and PRC2-EZH1 that predominates in post-mitotic tissues. PRC2-EZH1α/β pathway is involved in the response of muscle cells to oxidative stress. Atrophied muscle cells respond to oxidative stress by enabling the nuclear translocation of EED and its assembly with EZH1α and SUZ12. Here we prove that the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) long noncoding RNA (lncRNA) is required for the assembly of PRC2-EZH1 components. The absence of MALAT1 significantly decreased the association between EED and EZH1α proteins. Biochemical analysis shows that the presence of MALAT1 increases the enzymatic activity of PRC2-EZH1 in vitro. In addition, we show that the simultaneous expression of PRC2 core components is necessary for their solubility. The successful expression of PRC2 proteins enables the execution of several downstream experiments, which will further explain the nature of the interplay between MALAT1 and PRC2.

Indiana University-Purdue University Indianapolis (IUPUI) / The ability to preserve, protect, or grow skeletal muscle would greatly benefit patients in health and disease. Understanding the molecular pathways that regulate muscle size is necessary to develop interventions. The extracellular signal-related kinase (ERK) and Hedgehog signaling pathways each play necessary roles in skeletal muscle development. The ERK pathway has been shown to both stimulate and inhibit muscle development at different stages, while Hedgehog signaling is vital for embryonic muscle development. Thus, these pathways represent prime targets for manipulation in diseases associated with muscle loss. In prior studies, cancer patients treated with the ERK inhibitor, Selumetinib, experienced significant gains in lean body mass. To study the mechanisms responsible, we tested the potential of Selumetinib to protect against muscle wasting in muscle cell cultures and in mice with experimental lung cancer. Selumetinib was able to induce hypertrophy of cultured muscle cells. In mice, we observed a reduction in tumor mass and in circulating mediators of muscle wasting including inflammatory cytokines. However, Selumetinib treatment did not prevent cancer-induced muscle loss. Together, these data suggest a diversity in the underlying molecular mechanisms and the need for careful consideration when extrapolating results across different disease states, clinical trials, and model systems. In separate studies, we found that the Hedgehog pathway was increased in mice and patients with cancer-associated muscle wasting and inflammation. In a series of studies in muscle cell cultures, in genetically modified mice, and in mice bearing tumors, we found that inflammatory cytokines activated Hedgehog expression in muscle. Hedgehog signaling promoted the replication of muscle stem cells but reduced the expression of genes that specify mature muscle. Inhibiting Hedgehog signaling promoted muscle growth, while activating it caused muscle wasting. Furthermore, we identified unique properties of two proteins activated by Hedgehog, Gli1 and Gli2, where Gli1 appears to promote muscle stem cell proliferation and Gli2 mature muscle gene expression. These data implicate the Hedgehog pathway, GLI1 and GLI2 as targets for treatment of muscle wasting diseases. / 2 years