Introdução: Escorpionismo é um problema de saúde no Brasil e em outras regiões do mundo. Em 2015, no Brasil, foram relatados 74.598 acidentes e 119 mortes. Neste estudo, foram descritos aspectos clínicos, epidemiológicos, laboratoriais e de vulnerabilidade nestes acidentes. Métodos: Trata-se de estudo clínico prospectivo e observacional conduzido no Hospital Municipal de Santarém (HMS), de Abril de 2008 a Janeiro de 2014. Os pacientes incluídos no estudo foram admitidos pelo autor e não representam o total de pacientes internados no HMS. Além disso, foi realizado estudo qualitativo com base teórica no conceito de vulnerabilidade que inclui as dimensões individual, social e programática. Resultados: Foram descritos 58 acidentes, presumivelmente, causados por T. obscurus na Amazônia Brasileira. A maioria dos acidentes ocorreu durante o trabalho, a maioria dos pacientes incluídos era do sexo masculino 39 (67,2%). Os principais locais de picada foram as extremidades (pés e mãos) com uma frequência de 51 (90%). Os pacientes relataram sensação de \"choques elétricos\" que podem durar horas. A grande maioria dos pacientes apresenta quadro clínico compatível com disfunção cerebelar aguda, que pode ter início minutos e durar até dois dias após a ocorrência do acidente. Apresentaram ataxia cerebelar, disdiadococinesia, dismetria, disartria, dislalia, naúseas e vômitos. Além disso, alguns pacientes, aprentaram mioclonias e fasciculações que também podem ser atribuídas à disfunção cerebelar aguda ou talvez à ação direta sobre o músculo esquelético. Seis pacientes apresentaram rabdomiólise e dois injúria renal aguda. O quadro clínico observado na maioria dos pacientes consiste, principalmente, em disfunção cerebelar aguda e manifestações neuromusculares anormais, que não foram descritos em qualquer outra região do mundo. Também foram realizadas 28 entrevistas quanti-qualitativas com pacientes vítimas de acidente escorpiônico, as quais foram submetidas à técnica de análise de discurso. Pacientes eram, em sua maioria, homens que moravam na área rural e que trabalhavam como agricultores e viviam em condições sócio demográficas desfavoráveis. Discussão: As manifestações apresentadas por esses pacientes são compatíveis com disfunção cerebelar aguda que pode ser explicada, provavelmente, porque algumas toxinas de T. obscurus (da região de Santarém), não só tem a capacidade de atravessar rapidamente, em poucos minutos, a barreira hemato-encefálica, mas também devem apresentar elevada afinidade por canais iônicos presentes nas membranas de células do cerebelo. As mioclonias e fasciculações podem ser atribuídas à disfunção muscular ou cerebelar. A vulnerabilidade individual e social foi evidenciada em vários aspectos: pouco conhecimento em relação ao comportamento de escorpiões e também às medidas preventivas, baixa escolaridade (incluindo analfabetismo), baixa qualificação profissional, trabalho informal, condições precárias de vida (falta de água encanada, energia elétrica, esgoto). Em relação à dimensão programática foi constatada dificuldade de acesso aos serviços de saúde e a falta de antiveneno no centro de referência. Portanto, o estudo destaca as particularidades do escorpionismo em Santarém e os aspectos mais importantes da vulnerabilidade destes pacientes / Introduction: Scorpionism is a health problem in Brazil and in another regions of the world. In 2015, in Brazil 74,598 accidents and 119 deaths were reported. In this study, were described clinical, epidemiologic, laboratory and vulnerability aspects of these unique scorpion accidents. Methods: A prospective and observational study was conducted in the MHS, from April, 2008 to January, 2014. Patients included in the study were admitted by the authors and do not represent the total number of hospitalized patients in the MHS. In addition, a qualitative study was conducted with the theoretical basis of the vulnerability concept, that includes individual, social and programmatic dimension. Results: We described 58 accidents presumably caused by T. obscurus in Brazilian Amazonia. Most patients were stung during work activities and the majority was male 39 (67.2%). The main sites of stung were the extremities (feet and hands), with a frequency of 51 (90%).Patients reported a sensation of \"electric shocks\" which could last hours. The vast majority of patients presented a clinical picture compatible with acute cerebellar dysfunction, that started in minutes and could last up to two days after the accident. They presented cerebellar ataxia, dysdiadocokinesia, dysmetry, dysarthria, dyslalia, nausea and vomiting. Also, some patients presented myoclonus and fasciculation which can also be attributed to cerebellar dysfunction or perhaps the result of direct action on skeletal muscle. Six patients had developed rhabdomyolysis and two acute kidney injury. The clinical picture observed in most of our patients consisted mainly from an acute cerebellar dysfunction and abnormal neuromuscular manifestations which is not described in any other region of the world. Twenty-eight quantitative and qualitative interviews with scorpion sting victims. Each patient was submitted to discourse analysis technique. The majority patients were men who live in rural areas, small farmers with unfavorable socio-demographic conditions. Discussion: The manifestations presented by these patients are compatible with acute cerebellar dysfunction which could be explained probably because some toxins from T. obscurus from Santarém region, have not only the capacity to cross quickly, within minutes, the blood-brain and also must have high affinity for ionic channels present in some cerebellar cell membranes. Myoclonus and fasciculation can be attributed to cerebellar or muscle dysfunction. The individual and social vulnerability was demonstrated in several ways: little knowledge about the scorpion behavior and about the preventive measures, low education level (including illiteracy), low-skilled, informal work, precarious living conditions (lack of running water, electricity, basic sanitation). Regarding the programmatic dimension it was found difficult access to health services and lack of antivenom serum in the reference center. Therefore, the study highlights the particularities of scorpionism in Santarem and the most important aspects of vulnerability of these patients

Introdução: Escorpionismo é um problema de saúde no Brasil e em outras regiões do mundo. Em 2015, no Brasil, foram relatados 74.598 acidentes e 119 mortes. Neste estudo, foram descritos aspectos clínicos, epidemiológicos, laboratoriais e de vulnerabilidade nestes acidentes. Métodos: Trata-se de estudo clínico prospectivo e observacional conduzido no Hospital Municipal de Santarém (HMS), de Abril de 2008 a Janeiro de 2014. Os pacientes incluídos no estudo foram admitidos pelo autor e não representam o total de pacientes internados no HMS. Além disso, foi realizado estudo qualitativo com base teórica no conceito de vulnerabilidade que inclui as dimensões individual, social e programática. Resultados: Foram descritos 58 acidentes, presumivelmente, causados por T. obscurus na Amazônia Brasileira. A maioria dos acidentes ocorreu durante o trabalho, a maioria dos pacientes incluídos era do sexo masculino 39 (67,2%). Os principais locais de picada foram as extremidades (pés e mãos) com uma frequência de 51 (90%). Os pacientes relataram sensação de \"choques elétricos\" que podem durar horas. A grande maioria dos pacientes apresenta quadro clínico compatível com disfunção cerebelar aguda, que pode ter início minutos e durar até dois dias após a ocorrência do acidente. Apresentaram ataxia cerebelar, disdiadococinesia, dismetria, disartria, dislalia, naúseas e vômitos. Além disso, alguns pacientes, aprentaram mioclonias e fasciculações que também podem ser atribuídas à disfunção cerebelar aguda ou talvez à ação direta sobre o músculo esquelético. Seis pacientes apresentaram rabdomiólise e dois injúria renal aguda. O quadro clínico observado na maioria dos pacientes consiste, principalmente, em disfunção cerebelar aguda e manifestações neuromusculares anormais, que não foram descritos em qualquer outra região do mundo. Também foram realizadas 28 entrevistas quanti-qualitativas com pacientes vítimas de acidente escorpiônico, as quais foram submetidas à técnica de análise de discurso. Pacientes eram, em sua maioria, homens que moravam na área rural e que trabalhavam como agricultores e viviam em condições sócio demográficas desfavoráveis. Discussão: As manifestações apresentadas por esses pacientes são compatíveis com disfunção cerebelar aguda que pode ser explicada, provavelmente, porque algumas toxinas de T. obscurus (da região de Santarém), não só tem a capacidade de atravessar rapidamente, em poucos minutos, a barreira hemato-encefálica, mas também devem apresentar elevada afinidade por canais iônicos presentes nas membranas de células do cerebelo. As mioclonias e fasciculações podem ser atribuídas à disfunção muscular ou cerebelar. A vulnerabilidade individual e social foi evidenciada em vários aspectos: pouco conhecimento em relação ao comportamento de escorpiões e também às medidas preventivas, baixa escolaridade (incluindo analfabetismo), baixa qualificação profissional, trabalho informal, condições precárias de vida (falta de água encanada, energia elétrica, esgoto). Em relação à dimensão programática foi constatada dificuldade de acesso aos serviços de saúde e a falta de antiveneno no centro de referência. Portanto, o estudo destaca as particularidades do escorpionismo em Santarém e os aspectos mais importantes da vulnerabilidade destes pacientes / Introduction: Scorpionism is a health problem in Brazil and in another regions of the world. In 2015, in Brazil 74,598 accidents and 119 deaths were reported. In this study, were described clinical, epidemiologic, laboratory and vulnerability aspects of these unique scorpion accidents. Methods: A prospective and observational study was conducted in the MHS, from April, 2008 to January, 2014. Patients included in the study were admitted by the authors and do not represent the total number of hospitalized patients in the MHS. In addition, a qualitative study was conducted with the theoretical basis of the vulnerability concept, that includes individual, social and programmatic dimension. Results: We described 58 accidents presumably caused by T. obscurus in Brazilian Amazonia. Most patients were stung during work activities and the majority was male 39 (67.2%). The main sites of stung were the extremities (feet and hands), with a frequency of 51 (90%).Patients reported a sensation of \"electric shocks\" which could last hours. The vast majority of patients presented a clinical picture compatible with acute cerebellar dysfunction, that started in minutes and could last up to two days after the accident. They presented cerebellar ataxia, dysdiadocokinesia, dysmetry, dysarthria, dyslalia, nausea and vomiting. Also, some patients presented myoclonus and fasciculation which can also be attributed to cerebellar dysfunction or perhaps the result of direct action on skeletal muscle. Six patients had developed rhabdomyolysis and two acute kidney injury. The clinical picture observed in most of our patients consisted mainly from an acute cerebellar dysfunction and abnormal neuromuscular manifestations which is not described in any other region of the world. Twenty-eight quantitative and qualitative interviews with scorpion sting victims. Each patient was submitted to discourse analysis technique. The majority patients were men who live in rural areas, small farmers with unfavorable socio-demographic conditions. Discussion: The manifestations presented by these patients are compatible with acute cerebellar dysfunction which could be explained probably because some toxins from T. obscurus from Santarém region, have not only the capacity to cross quickly, within minutes, the blood-brain and also must have high affinity for ionic channels present in some cerebellar cell membranes. Myoclonus and fasciculation can be attributed to cerebellar or muscle dysfunction. The individual and social vulnerability was demonstrated in several ways: little knowledge about the scorpion behavior and about the preventive measures, low education level (including illiteracy), low-skilled, informal work, precarious living conditions (lack of running water, electricity, basic sanitation). Regarding the programmatic dimension it was found difficult access to health services and lack of antivenom serum in the reference center. Therefore, the study highlights the particularities of scorpionism in Santarem and the most important aspects of vulnerability of these patients

Nous présentons ici la description clinique et génétique d’un syndrome neurocutané unique. Le laboratoire du Dr Cossette a entrepris la caractérisation clinique et génétique d'une famille canadienne-française qui a été identifiée par les Drs Giroux et Barbeau en 1972 et qui comprend plus de 100 personnes sur six générations. Les membres atteints de cette famille présentent des lésions typiques d'érythrokératodermie (EK) (OMIM 133190, EKV1 et EKV2), associées à une ataxie spinocérébelleuse pure. Dans cette famille, l'ataxie est caractérisée par des troubles de la coordination et de la démarche causés par une dégénérescence du cervelet et de la moelle épinière. Cette ataxie est transmise selon un mode autosomique dominant. Une étude antérieure de cette variante d'EK avec ataxie avait suggéré une liaison sur le chromosome 1p34-p35, soit la même région que les formes EKV de type 1 et 2, causées respectivement par des mutations dans les gènes connexin-31 (GJB3; OMIM 603324) et connexin-30.3 (GJB4; OMIM 605425). Cependant, aucune mutation n'a été retrouvée dans ces gènes pour la famille canadienne-française. Nous avons récemment recontacté la famille et effectué des examens détaillés, incluant une imagerie par résonance magnétique (IRM) et un électromyogramme (EMG). Les manifestations neurologiques des individus atteints sont compatibles avec une nouvelle forme d’ataxie cérébelleuse pure à transmission autosomique dominante (ADCA de type III dans la classification de Harding) que nous avons appelée SCA34. Une cartographie complète du génome nous a permis de localiser le gène SCA34 sur le chromosome 6p12.3-q16.2. Également, en collaboration avec les Drs Alexis Brice (Hôpital Pitié-La Salpêtrière, Paris) et Alfredo Brusco (Hôpital San Giovanni Battista di Torino, Italie), nous avons confirmé que trois autres familles européennes avec SCA inexpliquée étaient également liées au locus SCA34. Notre laboratoire a récemment entrepris la recherche des mutations responsables de SCA34. Les résultats de ce criblage de gènes candidats sont présentés dans le chapitre 3 de cette thèse. / We present here the clinical and genetic description of a unique neuro-cutaneous syndrome. Dr. Cossette’s laboratory began the clinical and genetic characterization of a French-Canadian family who was identified by Drs. Giroux and Barbeau in 1972 and includes more than 100 people over six generations. The affected members of this family have typical lesions of erythrokeratodermia (EK) (OMIM 133190, and EKV1 EKV2), associated with pure spinocerebellar ataxia. In this family, the clinical phenotype is characterized by gait ataxia caused by degeneration of the cerebellum and spinal cord and the pattern of inheritance is compatible with an autosomal dominant trait. In a previous study of this variant of ataxia with EK, putative linkage was found on chromosome 1p34-p35, the same chromosomal region of EKV1 and EKV2 that are respectively caused by mutations in the connexin-31 gene (GJB3, OMIM 603324) and connexin -30.3 (GJB4, OMIM 605425). However, no mutations have been found in these latter genes for the French-Canadian family. We recently contacted the family and carried out detailed examinations, including a magnetic resonance imaging (MRI) and electromyography (EMG). Neurological manifestations of affected individuals are consistent with a new form of pure autosomal dominant cerebellar ataxia, (ADCA type III in the classification of Harding) that we named SCA34. A whole genome scan allowed us to map the gene on chromosome 6p12.3-q16.2. Interestingly, in collaboration with Dr. Alexis Brice (Hôpital Pitié-La Salpêtrière, Paris), and Alfredo Brusco (San Giovanni Battista Hospital, Turin, Italy), we found that three additional European families with unexplained SCA were also linked to the SCA34 locus. Our laboratory has recently begun the search for mutations causing SCA34. The results of this screening of candidate genes are presented in Chapter 3 of this thesis.

Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

Les ataxies (HCA) et paraparésies spastiques héréditaires constituent les deux extrémités du spectre des entités neurodégénératives spinocérébelleuses (SCE). Elles sont marquées par une forte hétérogénéité clinique, avec des signes associés variés, et génétique. Elles peuvent se transmettre sur tous les modes d'hérédité, et des mutations ont été décrites dans une myriade de gènes. Les SCE sont donc une entité qui bénéficie particulièrement des avancées technologiques de la Nouvelle Génération de Séquençage. Ce travail décrit des résultats obtenus sur de grandes cohortes, par séquençage de panel de gènes ciblés ou de l'exome entier, ainsi que des études de familles. Celles-ci nous ont permis de décrire de nouveaux modes de transmission de mutations dans des gènes déjà connus en pathologie humaine, avec un dans un cas une dysfonction similaire, dans l'autre un gain versus une perte de fonction. Nous rapportons aussi deux gènes nouvellement impliqués, dans une forme autosomique dominante de HCA (CACNA1A), et dans un sous-type autosomique récessif de dystonie avec atrophie cérébelleuse (TOR1AIP1). Nos résultats illustrent bien la refonte nosologique en marche dans les maladies génétiques complexes, qui remettent en permanence les corrélations génotype-phénotype en question. Nous discutons du pourquoi et du comment du diagnostic moléculaire dans cette nouvelle ère du séquençage. / Hereditary cerebellar ataxias (HCA) and spastic paraplegias constitute both ends of the neurodegenerative spectrum of spinocerebellar entities (SCE). Theses diseases are marked by a pronounced heterogeneity, both clinically, with various additional neurological or extraneurological signs, and genetically. They can indeed follow all transmission modes, and mutations in a myriad of genes have been described. SCE is hence a group of diseases that benefit greatly from Next-Generation Sequencing technologies. This work reports both screenings of large cohorts of patients with either panel or whole exome sequencing, as well as family studies. The latter allowed us to describe new modes of transmission for genes previously involved in human pathology, with either similar protein dysfunction, or loss- versus gain-of-function. We also describe two new genes implicated in a form of autosomal dominant HCA (CACNA1A), and an autosomal recessive subtype of dystonia and cerebellar atrophy (TOR1AIP1). Our results are illustrative of the genetic remodelling underway in complex genetic diseases, with permanent questioning of genotype-phenotype correlations. We discuss the how and the why of molecular diagnosis in this new era of sequencing.

Neurodegenerative disorders are heterogenous in nature and include a range of ataxias with oculomotor apraxia, which are characterised by a wide variety of neurological and ophthalmological features. This family includes recessive and dominant disorders. A subfamily of autosomal recessive cerebellar ataxias are characterised by defects in the cellular response to DNA damage. These include the well characterised disorders Ataxia-Telangiectasia (A-T) and Ataxia-Telangiectasia Like Disorder (A-TLD) as well as the recently identified diseases Spinocerebellar ataxia with axonal neuropathy Type 1 (SCAN1), Ataxia with Oculomotor Apraxia Type 2 (AOA2), as well as the subject of this thesis, Ataxia with Oculomotor Apraxia Type 1 (AOA1). AOA1 is caused by mutations in the APTX gene, which is located at chromosomal locus 9p13. This gene codes for the 342 amino acid protein Aprataxin. Mutations in APTX cause destabilization of Aprataxin, thus AOA1 is a result of Aprataxin deficiency. Aprataxin has three functional domains, an N-terminal Forkhead Associated (FHA) phosphoprotein interaction domain, a central Histidine Triad (HIT) nucleotide hydrolase domain and a C-terminal C2H2 zinc finger. Aprataxins FHA domain has homology to FHA domain of the DNA repair protein 5’ polynucleotide kinase 3’ phosphatase (PNKP). PNKP interacts with a range of DNA repair proteins via its FHA domain and plays a critical role in processing damaged DNA termini. The presence of this domain with a nucleotide hydrolase domain and a DNA binding motif implicated that Aprataxin may be involved in DNA repair and that AOA1 may be caused by a DNA repair deficit. This was substantiated by the interaction of Aprataxin with proteins involved in the repair of both single and double strand DNA breaks (XRay Cross-Complementing 1, XRCC4 and Poly-ADP Ribose Polymerase-1) and the hypersensitivity of AOA1 patient cell lines to single and double strand break inducing agents. At the commencement of this study little was known about the in vitro and in vivo properties of Aprataxin. Initially this study focused on generation of recombinant Aprataxin proteins to facilitate examination of the in vitro properties of Aprataxin. Using recombinant Aprataxin proteins I found that Aprataxin binds to double stranded DNA. Consistent with a role for Aprataxin as a DNA repair enzyme, this binding is not sequence specific. I also report that the HIT domain of Aprataxin hydrolyses adenosine derivatives and interestingly found that this activity is competitively inhibited by DNA. This provided initial evidence that DNA binds to the HIT domain of Aprataxin. The interaction of DNA with the nucleotide hydrolase domain of Aprataxin provided initial evidence that Aprataxin may be a DNA-processing factor. Following these studies, Aprataxin was found to hydrolyse 5’adenylated DNA, which can be generated by unscheduled ligation at DNA breaks with non-standard termini. I found that cell extracts from AOA1 patients do not have DNA-adenylate hydrolase activity indicating that Aprataxin is the only DNA-adenylate hydrolase in mammalian cells. I further characterised this activity by examining the contribution of the zinc finger and FHA domains to DNA-adenylate hydrolysis by the HIT domain. I found that deletion of the zinc finger ablated the activity of the HIT domain against adenylated DNA, indicating that the zinc finger may be required for the formation of a stable enzyme-substrate complex. Deletion of the FHA domain stimulated DNA-adenylate hydrolysis, which indicated that the activity of the HIT domain may be regulated by the FHA domain. Given that the FHA domain is involved in protein-protein interactions I propose that the activity of Aprataxins HIT domain may be regulated by proteins which interact with its FHA domain. We examined this possibility by measuring the DNA-adenylate hydrolase activity of extracts from cells deficient for the Aprataxin-interacting DNA repair proteins XRCC1 and PARP-1. XRCC1 deficiency did not affect Aprataxin activity but I found that Aprataxin is destabilized in the absence of PARP-1, resulting in a deficiency of DNA-adenylate hydrolase activity in PARP-1 knockout cells. This implies a critical role for PARP-1 in the stabilization of Aprataxin. Conversely I found that PARP-1 is destabilized in the absence of Aprataxin. PARP-1 is a central player in a number of DNA repair mechanisms and this implies that not only do AOA1 cells lack Aprataxin, they may also have defects in PARP-1 dependant cellular functions. Based on this I identified a defect in a PARP-1 dependant DNA repair mechanism in AOA1 cells. Additionally, I identified elevated levels of oxidized DNA in AOA1 cells, which is indicative of a defect in Base Excision Repair (BER). I attribute this to the reduced level of the BER protein Apurinic Endonuclease 1 (APE1) I identified in Aprataxin deficient cells. This study has identified and characterised multiple DNA repair defects in AOA1 cells, indicating that Aprataxin deficiency has far-reaching cellular consequences. Consistent with the literature, I show that Aprataxin is a nuclear protein with nucleoplasmic and nucleolar distribution. Previous studies have shown that Aprataxin interacts with the nucleolar rRNA processing factor nucleolin and that AOA1 cells appear to have a mild defect in rRNA synthesis. Given the nucleolar localization of Aprataxin I examined the protein-protein interactions of Aprataxin and found that Aprataxin interacts with a number of rRNA transcription and processing factors. Based on this and the nucleolar localization of Aprataxin I proposed that Aprataxin may have an alternative role in the nucleolus. I therefore examined the transcriptional activity of Aprataxin deficient cells using nucleotide analogue incorporation. I found that AOA1 cells do not display a defect in basal levels of RNA synthesis, however they display defective transcriptional responses to DNA damage. In summary, this thesis demonstrates that Aprataxin is a DNA repair enzyme responsible for the repair of adenylated DNA termini and that it is required for stabilization of at least two other DNA repair proteins. Thus not only do AOA1 cells have no Aprataxin protein or activity, they have additional deficiencies in PolyADP Ribose Polymerase-1 and Apurinic Endonuclease 1 dependant DNA repair mechanisms. I additionally demonstrate DNA-damage inducible transcriptional defects in AOA1 cells, indicating that Aprataxin deficiency confers a broad range of cellular defects and highlighting the complexity of the cellular response to DNA damage and the multiple defects which result from Aprataxin deficiency. My detailed characterization of the cellular consequences of Aprataxin deficiency provides an important contribution to our understanding of interlinking DNA repair processes.

Adult neural precursor cells (NPCs) are a heterogeneous population of mitotically active, self-renewing multipotent cells of both adult and developing CNS. They can be expanded in vitro in the presence of mitogens. The B05 transgenic SCA1 mice, expressing human ataxin-1 with an expanded polyglutamine tract in cerebellar Purkinje cells (PCs), recapitulate many pathological and behavioral characteristics of the neurodegenerative disease spinocerebellar ataxia type 1 (SCA1), including progressive ataxia and PC loss. We transplanted neural precursor cells (NPCs) derived from the subventricular zone of GFP-expressing adult mice into the cerebellar white matter of SCA1 mice when they showed absent (5 weeks), initial (13 weeks) and significant PC loss (24 weeks). A stereological count demonstrates that mice with significant cell loss exhibit highest survival of grafted NPCs and migration to the vicinity of PCs as compared to wt and younger grafted animals. These animals showed improved motor skills as compared to sham animals. Confocal analysis and profiling shows that many of implanted cells present in the cerebellar cortex have formed gap junctions with host PCs and express connexin43. Grafted cells did not adopt characteristics of PCs, but stereological and morphometric analysis of the cerebellar cortex revealed that grafted animals had more surviving PCs and a better preserved morphology of these cells than the control groups. Perforated patch clamp recordings revealed a normalization of the PC basal membrane potential, which was abnormally depolarized in sham-treated animals. No significant increase in levels of several neurotrophic factors was observed, suggesting, along with morphological observation, that the neuroprotective effect of grafted NPCs was mediated by direct contact with the host PCs. In this study, evidence for a neuroprotective effect came, in addition to motor behavior improvement, from stereological and electrophysiological analyses and suggest that timing of stem cell delivery is important to determine its therapeutic effect.<p>In a brain stem cell niche, NSCs reside in a complex cellular and extracellular microenvironment comprising their own progeny, ependymal cells, numerous blood vessels and various extracellular matrix molecules. Recently, it was reported that blood vessel ECs-NSCs crosstalk plays an important role in tissue homeostasis. Bloodstream offers a natural delivery vehicle especially in case of diffuse neurodegenerative diseases which require widespread distribution of exogenous cells. As NSCs are confronted with blood-brain barrier endothelial cells (BBB-ECs) before they can enter into brain parenchyma, we investigated their interaction using primary cultures in an in vitro BBB model. We isolated human fetal neural precursor cells (hfNPCs) from aborted fetal brain tissues and expanded in vitro. We showed that in an in vitro model, human BBB endothelium induces the rapid differentiation of hfNPCs and allows them to cross the endothelial monolayer, with the differentiated progeny remaining in close contact with endothelial cells. These results are not reproduced when using a non-BBB endothelium and are partly dependent on the cytokine MCP1. Our data suggest that, in the presence of attractive signals released by a damaged brain, intravascularly administered NPCs can move across an intact BBB endothelium and differentiate in its vicinity. Overall, our findings have implications for the development of cellular therapies for cerebellar degenerative diseases and understanding of the brain stem cell niche. / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished