Post-acute delivery of erythropoietin induces stroke recovery by promoting perilesional tissue remodelling and contralesional pyramidal tract plasticity

The promotion of post-ischaemic motor recovery remains a major challenge in clinical neurology. Recently, plasticity-promoting effects have been described for the growth factor erythropoietin in animal models of neurodegenerative diseases. To elucidate erythropoietin's effects in the post-acute ischaemic brain, we examined how this growth factor influences functional neurological recovery, perilesional tissue remodelling and axonal sprouting of the corticorubral and corticobulbar tracts, when administered intra-cerebroventricularly starting 3 days after 30 min of middle cerebral artery occlusion. Erythropoietin administered at 10 IU/day (but not at 1 IU/day), increased grip strength of the contralesional paretic forelimb and improved motor coordination without influencing spontaneous locomotor activity and exploration behaviour. Neurological recovery by erythropoietin was associated with structural remodelling of ischaemic brain tissue, reflected by enhanced neuronal survival, increased angiogenesis and decreased reactive astrogliosis that resulted in reduced scar formation. Enhanced axonal sprouting from the ipsilesional pyramidal tract into the brainstem was observed in vehicle-treated ischaemic compared with non-ischaemic animals, as shown by injection of dextran amines into both motor cortices. Despite successful remodelling of the perilesional tissue, erythropoietin enhanced axonal sprouting of the contralesional, but not ipsilesional pyramidal tract at the level of the red and facial nuclei. Moreover, molecular biological and histochemical studies revealed broad anti-inflammatory effects of erythropoietin in both hemispheres together with expression changes of plasticity-related molecules that facilitated contralesional axonal growth. Our study establishes a plasticity-promoting effect of erythropoietin after stroke, indicating that erythropoietin acts via recruitment of contralesional rather than of ipsilesional pyramidal tract projection

Functional kinomics establishes a critical node of volume-sensitive cation-Cl^- cotransporter regulation in the mammalian brain

This is the final version of the article. Available from the publisher via the DOI in this record.There is another record in ORE for this publication: http://hdl.handle.net/10871/33424Cell volume homeostasis requires the dynamically regulated transport of ions across the plasmalemma. While the ensemble of ion transport proteins involved in cell volume regulation is well established, the molecular coordinators of their activities remain poorly characterized. We utilized a functional kinomics approach including a kinome-wide siRNA-phosphoproteomic screen, a high-content kinase inhibitor screen, and a kinase trapping-Orbitrap mass spectroscopy screen to systematically identify essential kinase regulators of KCC3 Thr991/Thr1048 phosphorylation – a key signaling event in cell swelling-induced regulatory volume decrease (RVD). In the mammalian brain, we found the Cl−-sensitive WNK3-SPAK kinase complex, required for cell shrinkage-induced regulatory volume decrease (RVI) via the stimulatory phosphorylation of NKCC1 (Thr203/Thr207/Thr212), is also essential for the inhibitory phosphorylation of KCC3 (Thr991/Thr1048). This is mediated in vivo by an interaction between the CCT domain in SPAK and RFXV/I domains in WNK3 and NKCC1/KCC3. Accordingly, genetic or pharmacologic WNK3-SPAK inhibition prevents cell swelling in response to osmotic stress and ameliorates post-ischemic brain swelling through a simultaneous inhibition of NKCC1-mediated Cl− uptake and stimulation of KCC3-mediated Cl− extrusion. We conclude that WNK3-SPAK is an integral component of the long-sought “Cl−/volume-sensitive kinase” of the cation-Cl− cotransporters, and functions as a molecular rheostat of cell volume in the mammalian brain.We thank the excellent technical support of the MRC-Protein Phosphorylation and Ubiquitylation Unit (PPU) DNA Sequencing Service (coordinated by Nicholas Helps), the MRC-PPU tissue culture team (coordinated by Laura Fin), the Division of Signal Transduction Therapy (DSTT) antibody purification teams (coordinated by Hilary McLauchlan and James Hastie). We are grateful to the MRC PPU Proteomics facility (coordinated by David Campbell, Robert Gourlay and Joby Varghese). We thank for support the Medical Research Council (MC_UU_12016/2; DRA) and the pharmaceutical companies supporting the Division of Signal Transduction Therapy Unit (AstraZeneca, Boehringer-Ingelheim, GlaxoSmithKline, Merck KGaA, Janssen Pharmaceutica and Pfizer; DRA). We thank Thomas J. Jentsch (Max-Delbrück-Centrum für Molekulare Medizin) for providing the KCC1/3 double KO mice and his reading of this manuscript. We thank Nathaniel Grey (Harvard) for providing the kinase inhibitor library used in this study (NIH LINCS Program grant U54HL127365). This work was also supported by a Harvard-MIT Neuroscience Grant (to KTK/SJE)

The Immune System in Stroke

Stroke represents an unresolved challenge for both developed and developing countries and has a huge socio-economic impact. Although considerable effort has been made to limit stroke incidence and improve outcome, strategies aimed at protecting injured neurons in the brain have all failed. This failure is likely to be due to both the incompleteness of modelling the disease and its causes in experimental research, and also the lack of understanding of how systemic mechanisms lead to an acute cerebrovascular event or contribute to outcome. Inflammation has been implicated in all forms of brain injury and it is now clear that immune mechanisms profoundly influence (and are responsible for the development of) risk and causation of stroke, and the outcome following the onset of cerebral ischemia. Until very recently, systemic inflammatory mechanisms, with respect to common comorbidities in stroke, have largely been ignored in experimental studies. The main aim is therefore to understand interactions between the immune system and brain injury in order to develop novel therapeutic approaches. Recent data from clinical and experimental research clearly show that systemic inflammatory diseases -such as atherosclerosis, obesity, diabetes or infection - similar to stress and advanced age, are associated with dysregulated immune responses which can profoundly contribute to cerebrovascular inflammation and injury in the central nervous system. In this review, we summarize recent advances in the field of inflammation and stroke, focusing on the challenges of translation between pre-clinical and clinical studies, and potential anti-inflammatory/immunomodulatory therapeutic approaches

A adoção de um sistema pré-fabricado e seu rebatimento no processo de projeto: um estudo de caso

Trabalho apresentado no IV Simpósio Brasileiro de Qualidade do Projeto no Ambiente Construído, Viçosa - UFV, 2015O foco do estudo foi uma investigação no campo do projeto arquitetônico de espaços produzidos a partir de sistemas construtivos pré-fabricados portáveis e o seu processo projetivo. Buscou-se investigar o modo como os arquitetos analisam o processo projetivo a partir da adoção de um sistema construtivo pré-fabricado portável, sobretudo no que se refere às potencialidades e limitações inerentes ao uso de um sistema pré-definido e seu rebatimento no projeto de arquitetura. Um dos objetivos desta pesquisa foi identificar qual a avaliação feita do sistema adotado na investigação, a partir da visão dos profissionais de arquitetura, que o analisaram com base em dados disponíveis nos catálogos técnicos, em seu aporte teórico-prático e na expertise de cada avaliador. Para tanto a pesquisa se desenvolveu através da aplicação de questionários junto aos arquitetos, seguida da análise dos dados, a tabulação das informações quantitativas, da categorização dos dados qualitativos obtidos e seleção das principais ideias que afloraram do discurso dos participantes. A análise feita pelos arquitetos à luz de suas experiências e de sua capacidade crítica apontou para a compreensão da aplicabilidade do sistema. Embora eles não tenham utilizado diretamente um sistema pré-fabricado em um projeto, houve consenso quanto ao fato de que a aplicabilidade de um sistema portável pré-definido impõe limitações ao projetista, sendo aconselhável ter cautela ao adotá-lo.The focus of the study was an investigation into the architectural design of the field spaces made from portable prefabricated building systems and this projective process. We sought to investigate how the architects analyze the projective process with the adoption of a portable prefabricated building system, especially with regard to the potential and limitations inherent in using a predefined system and its repercussion in the project architecture. One objective of this research was to identify which assessment of the system adopted in the research, from the perspective of architecture professionals, who evaluated based on data available in the technical catalogs, in its theoretical and practical contribution and expertise of each evaluator. Therefore the research was developed through the application of questionnaires to the architects, followed by data analysis, tabulation of quantitative information, the categorization of qualitative data obtained and selection of the main ideas that came out upon the speech of participants. The analysis made by the architects in light of their experience and their critical capacity pointed to the understanding of the applicability of the system. Although they have not used directly a prefabricated system in a project, there was agreement on the fact that the applicability of a predefined portable system imposes limitations to the designer, it is advisable to exercise caution in adopting it

O uso de sistema pré-fabricado e o processo de projeto: uma experiência didática

Trabalho apresentado no IV Simpósio Brasileiro de Qualidade do Projeto no Ambiente Construído, Viçosa - UFV, 2015O foco do estudo foi um experimento didático no campo do projeto arquitetônico de espaços produzidos a partir de sistemas construtivos pré-fabricados desmontáveis e o processo de sua concepção. Por meio da observação dos espaços gerados, buscou- se investigar o modo como os estudantes do 6o período do curso de Arquitetura e Urbanismo lidam com projetos concebidos a partir da adoção de um sistema construtivo pré-fabricado desmontável, de modo a contribuir para a compreensão dos elementos essenciais ao ensino desse tipo de projeto. Para tanto foi desenvolvida uma pesquisa exploratória com base em multimétodos que abrangeu: análise documental, visitas técnicas e exercício acadêmico, todos documentados por imagens. Embora o estudo não seja conclusivo, notou-se quem apesar de terem obtido um desempenho adequado, os estudantes apresentaram alguma dificuldade para acessar os recursos a eles disponibilizados. Além disso, o exercício evidenciou que o uso de um sistema pré-fabricado desmontável/portável alterou a valorização/hierarquização dos condicionantes projetuais durante o processo projetual, uma vez que, diferentemente do que acontece em disciplinas tradicionais, os ‘estudantes-projetistas’ demonstraram maior preocupação com as questões construtivas, sobretudo os elementos estruturais (apoios e cobertura), em detrimento de funcionalidade, estética e características físicas do local.The focus of the study was a teaching experiment in architectural design of spaces produced from portable prefabricated systems and the process of its design. By observing the spaces generated, we sought to investigate how architecture students of the 6th Architecture and Urbanism course of the period deal with projects designed from the adoption of a prefabricated demountable building system, in order to contribute to the understanding of essential elements to teaching this kind of project. Thus, we developed an exploratory research based on multimethod covering: document analysis, technical visits and academic exercise, all documented by pictures. Although the study is not conclusive, it was noted who despite having obtained adequate performance, the students presented some difficulty to access the resources available to them. In addition, the exercise showed that the use of a prefabricated removable / portable system changed the recovery / hierarchy of projective constraints during the design process, since, unlike what happens in traditional disciplines, the 'student-designers' show greater focus on constructive issues, especially the structural elements (support and coverage), at the expense of functionality, aesthetics and physical characteristics of the site

Synthesis of Bis-Lactones Catalysed by Pd-dppb System

Palladium acetate [Pd(OAc )2] and 1,4-bis(diphenylphosphino)butane (dppb) catalyze regioselective cyclocarbonylation of bisallyl derivs. of bisphenols affording seven-membered ring bislactones in good yields. Double cyclocarbonylation reactions carried out using different conditions afforded bislactones with two different ring sizes (7-6 and 7-5-membered)