No description available.

No description available.

No description available.

The present study aims to elucidate the mechanisms underlying locomotor recovery following spinal cord injury (SCI) through the investigation of Brain-Derived Neurotrophic Factor (BDNF) delivery, and inflammatory responses associated with different spinal transection methods.Chapter 2 focuses on characterizing lumbar interneurons' activity during air- stepping following chronic intrathecal BDNF delivery to the lumbar spinal cord. BDNF has demonstrated the potential to elicit full locomotor recovery in untrained spinal animals, suggesting therapeutic benefits for SCI patients. However, the effects of BDNF on large populations of neurons responsible for this recovery are not well understood. The hypothesis is that intrathecal BDNF delivery will result in significantly increased neuronal activity in the L3-L4 segments during air-stepping. A programmable, implantable mini- pump was used to deliver BDNF at 50 ng/day for 35 days post-transection. Kinematic data was collected before and after BDNF delivery, and multiunit extracellular recordings were be obtained using 64-channel microelectrode arrays. Results from analysis suggest that while BDNF evidently increases neuronal excitability in treated cats, development of locomotor recovery seems to be achieved through subtle changes in neuronal activity. Chapter 3 investigates the mechanisms behind instances of spontaneous locomotor recovery observed in the literature, which could involve endogenous BDNF or other beneficial mechanisms. It compares locomotor recovery between open-dura and closed- dura spinal transection methods in cats. Previous studies have reported inconsistent outcomes regarding spontaneous recovery and the need for treadmill training. The ii hypothesis is that an open-dura transection will lead to better recovery during treadmill locomotion in untrained spinal cats. Kinematic data and ground reaction forces were measured to assess locomotor parameters and weight-bearing abilities, providing a quantitative analysis of recovery. The results show that an open-dura transection is associated with the development of spontaneous locomotor recovery in untrained spinal cats. Chapter 3 also examines differences in the inflammatory response at the lower thoracic cord between the two spinal transection methods, given the significant role of inflammation in CNS repair and recovery. The hypothesis is that the open-dura method will result in a higher inflammatory response, characterized by increased macrophages, microglia, and BDNF levels caudal to the transection site. Immunohistochemistry (IHC) and RNA in-situ hybridization assays were used to analyze the cellular and molecular environment near the injury site. Open dura animals show a decrease inflammatory response to injury and show no evidence of endogenous BDNF caudal to the injury. These results suggest the development of spontaneous locomotor recovery associated with a transected dura can be elicited through inflammatory mechanisms alone without the need for neurotrophic intervention. / Bioengineering

The anatomy of the cranial peripheral nervous system has been studied for over a century, yet surprisingly little is known about how the nerves are guided to their targets. The study of the development of these nerves has important implications for our understanding of craniofacial anomalies and possible treatments for both injury and genetic disorders of nerve development such as Goldenhar-Gorlin syndrome. We have discovered that retinoic acid (RA) may play a role in the development of the trigeminal nerve. Inhibition of retinoic acid receptors (RAR) results in trigeminal nerves that become unbundled or defasciculated in the eye region. To further understand how RA is affecting trigeminal development we searched for genes downregulated in response to RAR inhibition by the inhibitor BMS-453 and have identified neurotrophin-3 (NT-3), activated leukocyte cell adhesion molecule (ALCAM) and Semaphorin 4B (Sema4B). We have analyzed the expression patterns of Sema4B and NT-3 by in situ hybridization and have found NT-3 expression in the eye and Sema4B in the embryonic target of the trigeminal nerve, lens of the eye and in the pharyngeal arches. ALCAM has been analyzed via qRT-PCR and its transcription is downregulated just prior to the observed defasciculation phenotype. The pattern of expression of these genes combined with known expression of NT-3 receptors allows us to suggest a model whereby RA signaling regulates Sema4B, ALCAM and NT-3, which support the survival, guidance and fasciculation of the trigeminal nerve. This work has the potential to better understanding of the complex nature of cranial nervous system development.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 4 uppsatser.

As leucemias correspondem a 30% dos tumores pediátricos, e constituem as neoplasias mais frequentes em indivíduos com menos de 15 anos. Apesar da elevada taxa de cura, frequentemente a ela está associada resistência à quimioterapia e efeitos colaterais tardios. Por isso, novas estratégias de tratamento, diagnóstico e prognóstico são necessárias. O fator neurotrófico derivado do cérebro (BDNF) e seus receptores de quinase relacionados à tropomiosina (tropomyosin related kinase, ou Trk) estão envolvidos com muitos processos na medula óssea (MO). Entretanto, o papel do BDNF em leucemias agudas (LA) pediátricas ainda não é bem conhecido. O objetivo desse estudo foi analisar os níveis de BDNF em amostras de MO ou sangue periférico (SP) de crianças com LA, e iniciar a caracterização dos efeitos de agonistas e antagonistas de neurotrofinas sobre culturas primárias de leucemias linfóides agudas em diferentes momentos terapêuticos Foram coletadas amostras de MO ou SP de crianças e adolescentes com leucemia linfóide aguda (LLA), crianças e adolescentes com leucemia mielóide aguda (LMA), e indivíduos saudáveis (IS) da mesma faixa etária. Para análise dos níveis séricos de BDNF utilizou-se kit de imuno-ensaio enzimático tipo sanduíche. Quando comparados aos IS os níveis de BDNF de pacientes com LA, ao diagnóstico, foram significativamente menores. Resultados similares foram observados nos pacientes durante indução, consolidação, diagnóstico e tratamento de recidiva. Da mesma forma, os níveis de BDNF foram inferiores em pacientes que receberam transfusão de plaquetas e, ao diagnóstico naqueles pacientes que foram a óbito. Para a caracterização dos efeitos de agonistas e antagonistas de neurotrofinas em cultura de células, amostras de pacientes ao momento do diagnóstico e no momento de indução do tratamento foram utilizadas. Os linfócitos foram extraídos e, após plaqueamento, as células foram tratadas com BDNF (Sigma, B3795), NGF (Sigma, SRP3015) e K252a (Sigma, 05288) por 72 horas. A viabilidade foi avaliada pelo método de exclusão por azul de Tripan. Devido às dificuldades no cultivo das células, esses dados ainda estão em análise. / Leukemias account for 30% of pediatric tumors and are the most frequent cancers in people under 15 years. Despite the high cure rate, often it is associated with resistance to chemotherapy and late side effects. Therefore, new strategies for treatment, diagnosis and prognosis are necessary. The brain-derived neurotrophic factor (BDNF) and their kinase receptor related tropomyosin (tropomyosin related kinase, and Trk) are involved in many processes in bone marrow (BM), however, the role of BDNF in acute leukemias (AL) pediatric it is not well known. The aim of this study was to analyze the BDNF levels in BM samples or peripheral blood (PB) of children with AL, and start the characterization of the effects of agonists and antagonists on neurotrophin primary cultures of acute lymphoblastic leukemias in different therapeutic moments. BM or PB samples were collected from children and adolescents with acute lymphoblastic leukemia (ALL), children and adolescents with acute myeloid leukemia (AML), and healthy individuals (HI) of the same age. For analysis of serum levels of BDNF was used sandwich enzyme immunoassay kit. When compared to HI, BDNF levels in patients with AL at diagnosis were significantly lower. Similar results were observed in patients during induction, consolidation, diagnosis and treatment of relapse. Similarly, BDNF levels were lower in patients receiving platelet transfusion and at diagnosis in patients that died. To characterize the effects of agonists and antagonists for neurotrophin in cell culture, samples of patients at diagnosis and at the time of induction treatment were used. Lymphocytes were extracted and, after plating, cells were treated with BDNF (Sigma B3795), NGF (Sigma, SRP3015) and K252a (Sigma, 05288) for 72 hours. Viability was assessed by exclusion of trypan blue method. Due to difficulties in cell culture, these data are still under analysis.

No description available.

The survival of several neuron populations during development, including sympathetic neurons, is strictly regulated by neurotrophins such as nerve growth factor (NGF) released from innervation targets. NGF activates its receptor, TrkA, at axon terminals, to generate signals that are transmitted retrogradely to cell bodies to induce signaling cascades regulating survival. A general view of this process is that NGF generates retrograde survival signals that, when delivered to cell bodies, induce downstream survival signaling that prevents apoptosis. A retrograde survival signal proposed to be necessary for sympathetic neuron survival consists of endosomes containing NGF and phosphorylated TrkA. For this signal, phosphorylated TrkA arriving at cell bodies is required to initiate survival signaling. Studies have tested the necessity of TrkA phosphorylation in the cell bodies for survival: results from different studies contradict each other. Moreover, the Trk inhibitor, K252a, used in these studies, has reported non-specific effects. Using an alternate Trk inhibitor, Gö6976, data presented in this thesis demonstrates that NGF can promote survival by retrograde signaling that does not require TrkA phosphorylation in the cell bodies. These retrograde signals may be composed of signaling molecules activated downstream of TrkA in axons since pro-survival molecules downstream of TrkA, Akt and CREB, were found activated in the cell bodies/proximal axons. Data presented in this thesis also reveals a fundamentally different mechanism for how NGF promotes sympathetic neuron survival: a retrograde apoptotic signal that is suppressed by NGF. NGF withdrawal from axons induced the “axon apoptotic signal” that was retrogradely transmitted to cell bodies to activate a key pro-apoptotic molecule, c-jun. The axon apoptotic signal, which was blocked by the kinase inhibitors rottlerin and chelerythrine, was necessary for apoptosis in response to NGF deprivation. Evidence GSK3 is involved in generation or transmission of the axon apoptotic signal was provided by experiments with GSK3 inhibitors and siRNA. The axon apoptotic signal discovery refutes the previous view that NGF acting on axon terminals supports survival exclusively by generating retrograde survival signals. The axon apoptotic signal has broad implications for understanding nervous system development and other conditions where neuronal apoptosis occurs, such as neurotrauma and neurodegenerative diseases.

As leucemias correspondem a 30% dos tumores pediátricos, e constituem as neoplasias mais frequentes em indivíduos com menos de 15 anos. Apesar da elevada taxa de cura, frequentemente a ela está associada resistência à quimioterapia e efeitos colaterais tardios. Por isso, novas estratégias de tratamento, diagnóstico e prognóstico são necessárias. O fator neurotrófico derivado do cérebro (BDNF) e seus receptores de quinase relacionados à tropomiosina (tropomyosin related kinase, ou Trk) estão envolvidos com muitos processos na medula óssea (MO). Entretanto, o papel do BDNF em leucemias agudas (LA) pediátricas ainda não é bem conhecido. O objetivo desse estudo foi analisar os níveis de BDNF em amostras de MO ou sangue periférico (SP) de crianças com LA, e iniciar a caracterização dos efeitos de agonistas e antagonistas de neurotrofinas sobre culturas primárias de leucemias linfóides agudas em diferentes momentos terapêuticos Foram coletadas amostras de MO ou SP de crianças e adolescentes com leucemia linfóide aguda (LLA), crianças e adolescentes com leucemia mielóide aguda (LMA), e indivíduos saudáveis (IS) da mesma faixa etária. Para análise dos níveis séricos de BDNF utilizou-se kit de imuno-ensaio enzimático tipo sanduíche. Quando comparados aos IS os níveis de BDNF de pacientes com LA, ao diagnóstico, foram significativamente menores. Resultados similares foram observados nos pacientes durante indução, consolidação, diagnóstico e tratamento de recidiva. Da mesma forma, os níveis de BDNF foram inferiores em pacientes que receberam transfusão de plaquetas e, ao diagnóstico naqueles pacientes que foram a óbito. Para a caracterização dos efeitos de agonistas e antagonistas de neurotrofinas em cultura de células, amostras de pacientes ao momento do diagnóstico e no momento de indução do tratamento foram utilizadas. Os linfócitos foram extraídos e, após plaqueamento, as células foram tratadas com BDNF (Sigma, B3795), NGF (Sigma, SRP3015) e K252a (Sigma, 05288) por 72 horas. A viabilidade foi avaliada pelo método de exclusão por azul de Tripan. Devido às dificuldades no cultivo das células, esses dados ainda estão em análise. / Leukemias account for 30% of pediatric tumors and are the most frequent cancers in people under 15 years. Despite the high cure rate, often it is associated with resistance to chemotherapy and late side effects. Therefore, new strategies for treatment, diagnosis and prognosis are necessary. The brain-derived neurotrophic factor (BDNF) and their kinase receptor related tropomyosin (tropomyosin related kinase, and Trk) are involved in many processes in bone marrow (BM), however, the role of BDNF in acute leukemias (AL) pediatric it is not well known. The aim of this study was to analyze the BDNF levels in BM samples or peripheral blood (PB) of children with AL, and start the characterization of the effects of agonists and antagonists on neurotrophin primary cultures of acute lymphoblastic leukemias in different therapeutic moments. BM or PB samples were collected from children and adolescents with acute lymphoblastic leukemia (ALL), children and adolescents with acute myeloid leukemia (AML), and healthy individuals (HI) of the same age. For analysis of serum levels of BDNF was used sandwich enzyme immunoassay kit. When compared to HI, BDNF levels in patients with AL at diagnosis were significantly lower. Similar results were observed in patients during induction, consolidation, diagnosis and treatment of relapse. Similarly, BDNF levels were lower in patients receiving platelet transfusion and at diagnosis in patients that died. To characterize the effects of agonists and antagonists for neurotrophin in cell culture, samples of patients at diagnosis and at the time of induction treatment were used. Lymphocytes were extracted and, after plating, cells were treated with BDNF (Sigma B3795), NGF (Sigma, SRP3015) and K252a (Sigma, 05288) for 72 hours. Viability was assessed by exclusion of trypan blue method. Due to difficulties in cell culture, these data are still under analysis.