Os autores gostariam de agradecer a Koby Sheffy, PhD pela sua revisão criteriosa deste trabalho e Sotka William para apoio e assistência técnica. Este trabalho foi financiado em parte por concessões do National Institutes of Health (K12 HL087746, RC2HL103400, 1U01HL100397), eo Programa de Pesquisa do Tabaco Califórnia Related Disease da Universidade da Califórnia (18XT-0098).

I. preparar o paciente para um Estudo de Endo-PAT <ol><li> Antes do estudo, certifique-se o paciente realizar um jejum de pelo menos 4 horas, e absteve-se por pelo menos 8 horas de cafeína, tabaco, vitaminas ou medicamentos que possam afetar o tônus ​​vascular. O paciente pode querer usar o banheiro antes do estudo. </li><li> O estudo Endo-PAT deve ser realizado em um silêncio, mal iluminada sala de exame, com temperatura controlada para reduzir as oscilações do tônus ​​vascular. </li><li> Telefones celulares ou dispositivos de paginação deve ser silenciada, e roupas restritivas que possam interferir com o fluxo sanguíneo para os braços devem ser removidos. O paciente também deve remover relógios, anéis ou outras jóias nas mãos ou dedos. </li><li> Inspecionar os dedos do paciente para qualquer deformidades ou lesões que possam afetar o estudo. Não coloque as pontas de prova em um dedo que é cortado ou lesionado. Unhas não deve exceder 5 mm ou 1 / 5 de uma polegada além da ponta do tecido dedo. Aparar as unhas ou o arquivo se necessário, para evitar danificar as membranas internas das sondas PAT e deslocando o dedo da região de detecção da sonda. </li><li> O dedo indicador é recomendado para o estudo, no entanto, se esse dedo é inadequado, um dígito diferente (exceto o polegar) podem ser usadas, contanto que o mesmo dedo é usado em ambas as mãos. </li><li> O paciente deve ser confortável e supina por 15 minutos, de modo a atingir um steady-state cardiovascular. Coloque os dois apoiadores ao longo de cada braço de lados do paciente. </li><li> Medir a pressão arterial usando o braço de controle (o braço que não está obstruída durante o estudo Endo-PAT. </li><li> Coloque um manguito de pressão arterial no braço para ser obstruída durante o estudo Endo-PAT. Aplicar o manguito firmemente, mas sem excesso de pressão. Não inflar o manguito no momento. </li></ol> II.Prepare o Sistema Endo-PAT de Estudos <ol><li> Lançamento do Endo-PAT 2000 software e clique no &quot;Paciente Informação&quot; ícone na barra de ferramentas para criar um arquivo novo paciente. </li><li> Preencha a caixa de diálogo do Paciente informação, incluindo identificação do paciente, nome (opcional), idade, sexo, altura, peso, pressão arterial sistólica e diastólica. Campos opcionais permitem comentários de texto livre. Selecione o seu nome da lista pré-definida no campo Nome Patographer. Se seu nome não foi inserido adicioná-lo na lista e selecione. </li><li> Selecionar duas novas sondas PAT e conectar-se a tubulação pneumo-elétrica. Para conectar as pontas de prova, insira o conector de guia na fenda sonda e pressione suavemente o conector da sonda até que se encaixe no lugar. </li><li> Coloque as pontas de prova conectadas nas tomadas dos apoios de braço e pressione o botão &quot;Deflate&quot; na parte superior do Endo-PAT 2000 dispositivo. </li></ol> III.Conduct um Estudo de Endo-PAT <ol><li> Coloque os dedos pacientes índice completamente na sondas, confirme com o paciente que ele ou ela pode sentir o fim das sondas, e pressione o botão &quot;inflar&quot; botão na parte superior do Endo-PAT 2000 dispositivo. </li><li> Coloque um anel de espuma de âncora na base do dedo médio adjacentes. Assegurar que o anel de espuma eo sensor PAT não se tocam. Caso contrário, o anel pode interferir mecanicamente com o sensor. </li><li> Criar um loop cerca de 7-10cm com o tubo de pneumo-elétrica. O loop deve se estender a partir do sensor PAT e voltar aos ringues de espuma no dedo adjacente, enquanto o resto do tubo que se conecta ao dispositivo EndoPAT está apontando tubulação para a ponta do dedo. </li><li> Posição dos braços do paciente para os antebraços são suportados no braço e os dedos suporta balançar livremente para fora da borda do suporte. Certifique-se que as sondas não estão em contato com qualquer objeto, incluindo o apoio do braço, anel de espuma, tubulação, o colchão ou de outro dedo. </li><li> Peça ao paciente para se abster de mover os dedos, pois isso irá criar artefatos mecânicos. É importante que o paciente estar relaxado durante o estudo. Explicar ao paciente que, durante o teste que irá inflar o manguito do braço, e durante esse tempo eles podem sentir algum desconforto, dormência ou formigamento. </li><li> Clique no ícone &quot;Standby&quot; na interface do Endo-PAT de computador. Ajustar a base de tempo para 1 minuto e ajustar o ganho de sinal na tela para maximizar a clareza do sinal. Inspecionar os traçados dos sinais PAT das duas sondas para confirmar que eles estão livres de sinais artifactual. Se os sinais artifactual estão presentes, veriify que as sondagens não estão tocando nada e que o paciente não está se movendo os dedos. </li><li> Para começar o estudo, clique no ícone &quot;Go&quot; na interface do computador. Iniciar o cronômetro, clicando no botão &quot;Start / Stop Timer&quot; ícone. Isso irá iniciar uma contagem regressiva de cinco minutos para o período de gravação de linha de base. Após cinco minutos, parar o cronômetro clicando no botão &quot;Start / Stop Timer&quot; ícone. </li><li> Dizer ao paciente que você está indo noflate do manguito para a fase de oclusão e que ele ou ela deve ficar relaxada e não mover os dedos. </li><li> Rapidamente inflar o manguito de pressão para uma pressão supra-sistólica de 60mmHg acima da pressão sistólica do paciente ou 200mmHg, o que for maior e começar o cronômetro novamente. Cessação completa do fluxo sangüíneo para a mão é verificada pela ausência de um sinal PAT do braço ocluída. Para confirmar a oclusão aumentar o ganho na tela do canal do lado ocluído para 20.000, mantendo o ganho do lado contra-lateral constante. Diminuir a base de tempo de ambos os canais a 30 segundos. Verifique se você não observou sinais com uma periodicidade que corresponde ao sinal do braço de controle, pois isso indica uma oclusão incompleta. Se este for o caso, então ainda mais inflar o manguito até que não haja sinais são vistos. O manguito pode ser inflado até um máximo de 300 mmHg. </li><li> Isso irá iniciar uma contagem regressiva de cinco minutos para o período de oclusão arterial de gravação. Perto do fim do período de oclusão dizer ao paciente que você vai liberar o manguito e que eles devem continuar a abster-se de mover os dedos. Depois de exatamente cinco minutos, desinflar o manguito de forma abrupta o mais rápido possível e parar o cronômetro clicando no botão &quot;Start / Stop Timer&quot; ícone. </li><li> Clique no botão &quot;Start / Stop Timer&quot; ícone novamente para iniciar um período de cinco minutos pós-período de gravação oclusão. Parar o cronómetro após cinco minutos e clique no botão &quot;Teste de Stop&quot; ícone para completar o estudo. As sondas serão automaticamente desinflar. </li><li> Remover as sondas, fita adesiva, espuma e anéis dos dedos do paciente e desconecte as pontas de prova PAT da tubulação de pneumo-elétrica. Descartar as sondas utilizadas. </li></ol> IV.Review e Análise <ol><li> Carregar o arquivo de estudo para a tela usando o ícone de carga. Para executar a análise automática, clique no botão &quot;mágico stick&quot; ícone. O período de oclusão será destacada em azul eo resultado do teste será exibido, incluindo o Índice de hiperemia reativa (RHI) e freqüência cardíaca (FC), na coluna do lado direito da tela. </li><li> A revisão de dados adicionais, incluindo parâmetros de estudo, variáveis ​​calculadas, as informações do paciente, e medidas de qualidade de sinal, clique no botão &quot;Abrir os resultados da última Cálculo&quot; ícone. Isto irá abrir uma planilha com os parâmetros do estudo e resultados para todas as análises realizadas até agora, com a última linha na tabela contendo os dados a partir da análise mais recente. </li></ol> Representante Resultados V. Um representante da tela Endo-PAT de um estudo realizado em um indivíduo com função vasodilatadora endotelial normal, é mostrado na Figura 1. Uma tela de representante de um estudo de Endo-PAT realizada em um indivíduo com disfunção endotelial vasodilatador é mostrado na Figura 2. <img src="/files/ftp_upload/2167/2167fig1.jpg" alt="Figura 1" /> Figura 1: Função vasodilatador endotelial normal. Gravação de representante de um indivíduo com função vasodilatadora endotelial normais, caracterizado por um aumento na amplitude do sinal após a liberação do manguito em relação ao valor basal. <img src="/files/ftp_upload/2167/2167fig2.jpg" alt="Figura 2" /> Figura 2: Disfunção Endotelial vasodilatador. Gravação de representante de um indivíduo com disfunção endotelial vasodilatador.

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L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impaired+Vascular+Function+in+Obese+Adolescents+with+Insulin+Resistance.">Impaired Vascular Function in Obese Adolescents with Insulin Resistance.</a> J Pediatr. 155 (5), 678-682 (2009).</li><li>Hirata, Y., Nagata, D., Suzuki, E., Nishimatsu, H., Suzuki, J., Nagai, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diagnosis+and+treatment+of+endothelial+dysfunction+in+cardiovascular+disease.">Diagnosis and treatment of endothelial dysfunction in cardiovascular disease.</a> Int Heart J. 51 (1), 1-6 (2010).</li><li>Hamburg, N. M., Keyes, M. J., Larson, M. G., Vasan, R. S., Schnabel, R., Pryde, M. M., Mitchell, G. F., Sheffy, J., Vita, J. A., Benjamin, E. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cross-Sectional+Relations+of+Digital+Vascular+Function+to+Cardiovascular+Risk+Factors+in+the+Framingham+Heart+Study.">Cross-Sectional Relations of Digital Vascular Function to Cardiovascular Risk Factors in the Framingham Heart Study.</a> Circulation. 117 (19), 2467-2474 (2008).</li><li>Truschel, E., Jarczok, M. N., Fischer, J. E., Terris, D. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+ambulatory+assessment+of+digital+reactive+hyperemia:+Concurrent+validity+with+known+cardiovascular+risk+factors+and+potential+confounding.">High-throughput ambulatory assessment of digital reactive hyperemia: Concurrent validity with known cardiovascular risk factors and potential confounding.</a> Prev Med. 49 (6), 468-4672 (2009).</li><li>Rubinshtein, R., Kuvin, J. T., Soffler, M., Lennon, R. J., Lavi, S., Nelson, R. E., Pumper, G. M., Lerman, L. O., Lerman, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+Endothelial+Function+by+Non-invasive+Peripheral+Arterial+Tonometry+Predicts+Late+Cardiovascular+Adverse+Events.">Assessment of Endothelial Function by Non-invasive Peripheral Arterial Tonometry Predicts Late Cardiovascular Adverse Events.</a> Eur Heart. , (2010).</li><li>Yamaoka-Tojo, M., Tojo, T., Kosugi, R., Hatakeyama, Y., Yoshida, Y., Machida, Y., Aoyama, N., Masuda, T., Izumi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+ezetimibe+add-on+therapy+for+high-risk+patients+with+dyslipidemia.">Effects of ezetimibe add-on therapy for high-risk patients with dyslipidemia.</a> Lipids Health Dis. 8 (1), 41-41 (2009).</li></ol>

O desenvolvimento de vídeo foi apoiado por Itamar Medical Co. Dr Cooke é o inventor de patentes detidas pela Universidade de Stanford para aplicações de diagnóstico e terapêutica da via NOS partir do qual ele recebe royalties.

A principal causa de morbidade e mortalidade em todo o mundo é a doença aterosclerótica vascular, levando a infarto, acidente vascular cerebral do miocárdio, insuficiência cardíaca, insuficiência renal, ruptura de aneurisma ou embolia, claudicação intermitente e 1,2 gangrena. A disfunção endotelial é um dos primeiros eventos no processo fisiopatológico que conduziram a estes distúrbios ateroscleróticos 3. Disfunção endotelial, além disso, contribui para a progressão da doença, facilitando a inflamação e trombose. 4-6. Os tradicionais fatores de risco cardiovascular estão associadas a disfunção endotelial vasodilatador 7-11. Disfunção endotelial vasodilatador pode ser detectado em indivíduos aparentemente saudáveis ​​que estão em risco de desenvolver doença cardiovascular 12. Além disso, o achado de disfunção endotelial é vasodilatador preditivos dos principais eventos cardiovasculares adversos, bem como a mortalidade 13-16. Os vasodilatadores liberados pelo endotélio incluem prostanóides vasodilatadores como a prostaciclina, peptídeos, tais como adrenomedulina e peptídeo natriurético atrial, e moléculas pequenas, como fator dependente do endotélio hiperpolarizante, monóxido de carbono e óxido nítrico 17. Além disso, o efeito das correntes hiperpolarizantes gerado no endotélio pode ser transmitido para o músculo liso vascular subjacente de embarcações mais pequenas, relaxá-los 18. Uma avaliação indireta da geração dessas influências vasodilatador endotelial pode ser adquirida através do estudo de reatividade vascular. O método mais prevalente para avaliar regulação da reatividade vascular endotelial não-invasiva tem sido ultra-sonografia duplex para detectar a vasodilatação fluxo-mediada da artéria braquial 19. A força de tracção de fluxo de fluido estimula o endotélio a liberação de vasodilatadores, o óxido nítrico mais proeminente 20. Este fenômeno pode ser observado por ultra-som na artéria braquial durante aumentos no fluxo sanguíneo do antebraço reativa induzida por 19 hiperemia. Esta tecnologia tem sido amplamente utilizada para documentar a associação de disfunção endotelial vasodilatador com fatores de risco cardiovascular, a relação de disfunção endotelial vasodilatador para biomarcadores diversos, tais como peptídeo C-reativa, ou dimetilarginina assimétrica (o antagonista endógeno da óxido nítrico sintase), e a correção da função vasodilatadora endotelial com modificações nutricionais e estilo de vida, bem como com o uso de inibidores da enzima conversora da angiotensina, bloqueadores dos receptores da angiotensina, estatinas, agentes sensibilizadores de insulina ou aspirina 21-23. A avaliação da vasodilatação mediada por fluxo da artéria braquial por ultra-som requer equipamento caro, é altamente dependente do operador, a resposta tem uma gama muito pequena dinâmica ea relação sinal-ruído é baixa. Novas abordagens para resolver estes problemas com a avaliação da função endotelial vasodilatador são necessários 24. A Endo-PAT 2000 é uma nova abordagem para avaliar a função vasodilatadora endotelial de modo não-invasivo rápido. A técnica fornece valores para o cálculo de um índice de hiperemia reativa (RHI), que dá uma indicação da função vasodilatadora endotelial. A RHI é o pós-oclusão de pré-relação sinal PAT no braço ocluído, em relação à mesma proporção no grupo de controlo, e corrigidos para o tom de base vascular. Estudos utilizando o EndoPAT têm mostrado que a pontuação RHI reflete biodisponibilidade de NO-25. A RHI correlaciona-se com a medição da função vasodilatadora endotelial nas artérias coronárias e com 26 de febre aftosa braquial 27. Pacientes com um grau maior de doenças cardiovasculares apresentam uma menor pontuação 28 e valores também são inferiores em outras condições associadas com a função endotelial prejudicada e risco de doença cardiovascular 29-33. Nomeadamente, os valores RHI parecem ser preditivos de desfechos cardiovasculares 35. A RHI baixo (indicando disfunção endotelial) pode ser revertida com o tratamento 36. Em conclusão, demonstramos como realizar um teste confiável e reprodutível para a função vasodilatadora endotelial com a Endo-2000 PAT. O teste é não invasivo, de fácil execução, e é uma ferramenta de pesquisa útil. Sua utilidade na monitorização clínica da função endotelial e na adaptação de gestão da doença está sob investigação.

assessing endothelial vasodilator function with the endo-pat 2000

O endotélio é uma monocamada de células delicadas que as linhas de todos os vasos sanguíneos, e que compreende os capilares sistêmicos e linfático. Em virtude da panóplia de fatores parácrinos que segrega, o endotélio regula o estado contrátil e proliferativo do músculo liso vascular subjacente, bem como a interação da parede do vaso com elementos do sangue circulante. Por causa de seu papel central na mediação tom navio e crescimento, a sua posição como porta de entrada para as células imunológicas que circulam, e sua regulamentação local da hemostasia e coagulação, o endotélio eo bom funcionamento é a chave para a saúde cardiovascular. Por outro lado, os primeiros distúrbios na maioria das doenças vasculares é a disfunção endotelial. Na circulação arterial, o endotélio saudável geralmente exerce uma influência vasodilatador sobre o músculo liso vascular. Há uma série de métodos para avaliar a função vasodilatadora endotelial. A Endo-PAT 2000 é um novo dispositivo que é usado para avaliar a função vasodilatadora endotelial de uma forma rápida e não invasiva. Ao contrário da técnica comumente utilizada de duplex ultra-sonografia para avaliar a vasodilatação mediada pelo fluxo, é totalmente não-dependente do operador e do equipamento é uma ordem de magnitude menos caro. Os registros dispositivo mediada pelo endotélio mudanças na forma de onda de pulso digital conhecido como o sinal de PAT (Tone arterial periférica), medido com um par de novas sondas modificada pletismográfico situado no dedo indicador de cada mão. Mediada pelo endotélio as alterações do sinal PAT são extraídas através da criação de uma resposta a jusante hiperêmica. Hiperemia é induzida por oclusão do fluxo sangüíneo através da artéria braquial por 5 minutos, utilizando um manguito inflável de um lado. A resposta a hiperemia reativa é calculado automaticamente pelo sistema. Uma relação PAT é criado usando o post e valores pré oclusão. Estes valores são normalizados para medições do braço contra-lateral, que serve como controle para não-endotelial dependente efeitos sistêmicos. Mais notavelmente, esta normalização controles para flutuações na saída do nervo simpático que pode induzir alterações no tônus ​​arterial periférico que são sobrepostos na resposta hiperêmica. Neste vídeo mostramos como usar o Endo-PAT 2000 para realizar uma avaliação clínica relevante da função vasodilatadora endotelial.

Watch this Scientific Journal Video about Assessing Endothelial Vasodilator Function with the Endo-PAT 2000 at JoVE.com

Assessing Endothelial Vasodilator Function with the Endo-PAT 2000

Um procedimento não-invasivo para avaliar a função endotelial é demonstrada através dos 2000 Endo-PAT.

Avaliação da função endotelial com a Vasodilatador 2000 Endo-PAT

The endothelium is a delicate monolayer of cells that lines all blood vessels, and which comprises the systemic and lymphatic capillaries. By virtue of ...

assessing-endothelial-vasodilator-function-with-the-endo-pat-2000

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43.5K visualizações.  Stanford University. Um procedimento não-invasivo para avaliar a função endotelial é demonstrada através dos 2000 Endo-PAT.

Vídeo: Assessing Endothelial Vasodilator Function with the Endo-PAT 2000

Dual-mode Imaging of Cutaneous Tissue Oxygenation and Vascular Function

Accurate assessment of cutaneous tissue oxygenation and vascular function is important for appropriate detection, staging, and treatment of many health disorders such as chronic wounds. We report the development of a dual-mode imaging system for non-invasive and non-contact imaging of cutaneous tissue oxygenation and vascular function. The imaging system integrated an infrared camera, a CCD camera, a liquid crystal tunable filter and a high intensity fiber light source. A Labview interface was programmed for equipment control, synchronization, image acquisition, processing, and visualization. Multispectral images captured by the CCD camera were used to reconstruct the tissue oxygenation map. Dynamic thermographic images captured by the infrared camera were used to reconstruct the vascular function map. Cutaneous tissue oxygenation and vascular function images were co-registered through fiduciary markers. The performance characteristics of the dual-mode image system were tested in humans.

A dual-mode imaging system was developed for non-contact assessment of cutaneous tissue oxygenation and vascular function.

Dual-mode de imagem de oxigenação dos tecidos cutâneos e Função Vascular

Accurate assessment of cutaneous tissue oxygenation and vascular function is important for appropriate detection, staging, and treatment of many health ...

Medicina

In Vivo Canine Muscle Function Assay

We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric 
contractions. The pelvic limb of an anesthetized dog is immobilized in a stereotactic frame to align the tibia at a right angle to the femur. Adhesive wrap affixes the 
paw to a pedal mounted on the shaft of a servomotor to measure torque. Percutaneous nerve stimulation activates pelvic limb muscles of the paw to either push (extend) or 
pull (flex) against the pedal to generate isometric torque. Percutaneous tibial nerve stimulation activates tibiotarsal extensor muscles. Repeated eccentric (lengthening) 
contractions are induced in the tibiotarsal flexor muscles by percutaneous peroneal nerve stimulation. The eccentric protocol consists of an initial isometric contraction 
followed by a forced stretch imposed by the servomotor. The rotation effectively lengthens the muscle while it contracts, e.g., an eccentric contraction. During 
stimulation flexor muscles are subjected to an 800 msec isometric and 200 msec eccentric contraction. This procedure is repeated every 5 sec. To avoid fatigue, 4 min rest 
follows every 10 contractions with a total of 30 contractions performed.

In Vivo Canine Muscle Function Assay

We describe a minimally-invasive and painless method to measure canine hindlimb muscle strength and muscle response to repeated eccentric contractions.

Em função Assay Canine Muscle Vivo

We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric 
...

Assessing Replication and Beta Cell Function in Adenovirally-transduced Isolated Rodent Islets

Glucose homeostasis is primarily controlled by the endocrine hormones insulin and glucagon, secreted from the pancreatic beta and alpha cells, respectively. Functional beta cell mass is determined by the anatomical beta cell mass as well as the ability of the beta cells to respond to a nutrient load. A loss of functional beta cell mass is central to both major forms of diabetes 1-3. Whereas the declining functional beta cell mass results from an autoimmune attack in type 1 diabetes, in type 2 diabetes, this decrement develops from both an inability of beta cells to secrete insulin appropriately and the destruction of beta cells from a cadre of mechanisms. Thus, efforts to restore functional beta cell mass are paramount to the better treatment of and potential cures for diabetes.
Efforts are underway to identify molecular pathways that can be exploited to stimulate the replication and enhance the function of beta cells. Ideally, therapeutic targets would improve both beta cell growth and function. Perhaps more important though is to identify whether a strategy that stimulates beta cell growth comes at the cost of impairing beta cell function (such as with some oncogenes) and vice versa.
By systematically suppressing or overexpressing the expression of target genes in isolated rat islets, one can identify potential therapeutic targets for increasing functional beta cell mass 4-6. Adenoviral vectors can be employed to efficiently overexpress or knockdown proteins in isolated rat islets 4,7-15. Here, we present a method to manipulate gene expression utilizing adenoviral transduction and assess islet replication and beta cell function in isolated rat islets (Figure 1). This method has been used previously to identify novel targets that modulate beta cell replication or function 5,6,8,9,16,17.

This protocol allows one to identify factors that modulate functional beta cell mass to find potential therapeutic targets for the treatment of diabetes. The protocol consists of a streamlined method to assess islet replication and beta cell function in isolated rat islets following manipulation of gene expression with adenoviruses.

Avaliação de replicação e função das células beta no Adenovirally transduzidas isoladas ilhotas de roedores

Glucose homeostasis is primarily controlled by the endocrine hormones insulin and glucagon, secreted from the pancreatic beta and alpha cells, ...

Non-invasive Assessment of Microvascular and Endothelial Function

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of cardiovascular disease. Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. Percent capillary recruitment is assessed by dividing the increase in capillary density induced by postocclusive reactive hyperemia (postocclusive reactive hyperemia capillary density minus baseline capillary density), by the maximal capillary density (observed during passive venous occlusion). Percent perfused capillaries represents the proportion of all capillaries present that are perfused (functionally active), and is calculated by dividing postocclusive reactive hyperemia capillary density by the maximal capillary density. Both percent capillary recruitment and percent perfused capillaries reflect the number of functional capillaries. The forearm blood flow (FBF) technique provides accepted non-invasive measures of endothelial function: The ratio FBFmax/FBFbase is computed as an estimate of vasodilation, by dividing the mean of the four FBFmax values by the mean of the four FBFbase values. Forearm vascular resistance at maximal vasodilation (FVRmax) is calculated as the mean arterial pressure (MAP) divided by FBFmax. Both the capillaroscopy and forearm techniques are readily acceptable to patients and can be learned quickly.
The microvascular and endothelial function measures obtained using the methodologies described in this paper may have future utility in clinical patient cardiovascular risk-reduction strategies. As we have published reports demonstrating that microvascular and endothelial dysfunction are found in initial stages of hypertension including prehypertension, microvascular and endothelial function measures may eventually aid in early identification, risk-stratification and prevention of end-stage vascular pathology, with its potentially fatal consequences.

Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. The forearm blood flow technique provides accepted non-invasive measures of endothelial function.

Avaliação não invasiva da função microvascular e endotelial

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of ...

Diagnostic Ultrasound Imaging of Mouse Diaphragm Function

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive surgical procedures. Although this is an effective method of assessing in vitro diaphragm activity, it involves non-survival surgery. The application of non-invasive ultrasound imaging as an in vivo procedure is beneficial since it not only reduces the number of animals sacrificed, but is also suitable for monitoring disease progression in live mice. Thus, our ultrasound imaging method may likely assist in the development of novel therapies that alleviate muscle injury induced by various respiratory diseases. Particularly, in clinical diagnoses of obstructive lung diseases, ultrasound imaging has the potential to be used in conjunction with other standard tests to detect the early onset of diaphragm muscle fatigue. In the current protocol, we describe how to accurately evaluate diaphragm contractility in a mouse model using a diagnostic ultrasound imaging technique.

Diagnostic ultrasound imaging has proven to be effective in diagnosing various respiratory diseases in human and animal subjects. We demonstrate a comprehensive ultrasound protocol utilized by Dr. Zuo's lab to analyze diaphragm kinetics specifically in mouse models. This is also a non-invasive research technique which can provide quantitative information on mouse respiratory muscle function.

Diagnóstico Ecografia de função de mouse Diafragma

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive ...

Assessment of Vascular Function in Patients With Chronic Kidney Disease

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this is only partially explained by traditional CVD risk factors. Vascular dysfunction is an important non-traditional risk factor, characterized by vascular endothelial dysfunction (most commonly assessed as impaired endothelium-dependent dilation [EDD]) and stiffening of the large elastic arteries. While various techniques exist to assess EDD and large elastic artery stiffness, the most commonly used are brachial artery flow-mediated dilation (FMDBA) and aortic pulse-wave velocity (aPWV), respectively. Both of these noninvasive measures of vascular dysfunction are independent predictors of future cardiovascular events in patients with and without kidney disease. Patients with CKD demonstrate both impaired FMDBA, and increased aPWV. While the exact mechanisms by which vascular dysfunction develops in CKD are incompletely understood, increased oxidative stress and a subsequent reduction in nitric oxide (NO) bioavailability are important contributors. Cellular changes in oxidative stress can be assessed by collecting vascular endothelial cells from the antecubital vein and measuring protein expression of markers of oxidative stress using immunofluorescence. We provide here a discussion of these methods to measure FMDBA, aPWV, and vascular endothelial cell protein expression.

The degree of vascular dysfunction and contributing physiological mechanisms can be assessed in patients with chronic kidney disease by measuring brachial artery flow-mediated dilation, aortic pulse-wave velocity, and vascular endothelial cell protein expression.

Avaliação da função vascular em pacientes com doença renal crônica

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this ...

Isolation of Human Lymphatic Endothelial Cells by Multi-parameter Fluorescence-activated Cell Sorting

Lymphatic system disorders such as primary lymphedema, lymphatic malformations and lymphatic tumors are rare conditions that cause significant morbidity but little is known about their biology. Isolating highly pure human lymphatic endothelial cells (LECs) from diseased and healthy tissue would facilitate studies of the lymphatic endothelium at genetic, molecular and cellular levels. It is anticipated that these investigations may reveal targets for new therapies that may change the clinical management of these conditions. A protocol describing the isolation of human foreskin LECs and lymphatic malformation lymphatic endothelial cells (LM LECs) is presented. To obtain a single cell suspension tissue was minced and enzymatically treated using dispase II and collagenase II. The resulting single cell suspension was then labelled with antibodies to cluster of differentiation (CD) markers CD34, CD31, Vascular Endothelial Growth Factor-3 (VEGFR-3) and PODOPLANIN. Stained viable cells were sorted on a fluorescently activated cell sorter (FACS) to separate the CD34LowCD31PosVEGFR-3PosPODOPLANINPos LM LEC population from other endothelial and non-endothelial cells. The sorted LM LECs were cultured and expanded on fibronectin-coated flasks for further experimental use.

The goal of this protocol is to isolate lymphatic endothelial cells lining human lymphatic malformation cyst-like vessels and foreskins using fluorescence-activated cell sorting (FACS). Subsequent cell culturing and expansion of these cells permits a new level of experimental sophistication for genetic, proteomic, functional and cell differentiation studies.

Isolamento de linfático humano endotelial por multi-parâmetro activadas por fluorescência celular

Lymphatic system disorders such as primary lymphedema, lymphatic malformations and lymphatic tumors are rare conditions that cause significant morbidity ...

Using Saccadometry with Deep Brain Stimulation to Study Normal and Pathological Brain Function

The oculomotor system involves a large number of brain areas including parts of the basal ganglia, and various neurodegenerative diseases including Parkinson's and Huntington's can disrupt it. People with Parkinson's disease, for example, tend to have increased saccadic latencies. Consequently, the quantitative measurement of saccadic eye movements has received considerable attention as a potential biomarker for neurodegenerative conditions. A lot more can be learned about the brain in both health and disease by observing what happens to eye movements when the function of specific brain areas is perturbed. Deep brain stimulation is a surgical intervention used for the management of a range of neurological conditions including Parkinson's disease, in which stimulating electrodes are placed in specific brain areas including several sites in the basal ganglia. Eye movement measurements can then be made with the stimulator systems both off and on and the results compared. With suitable experimental design, this approach can be used to study the pathophysiology of the disease being treated, the mechanism by which DBS exerts it beneficial effects, and even aspects of normal neurophysiology.

This paper describes the use of quantitative measurement of eye movements in conjunction with stimulation of focal areas of the deep brain in order to study physiology, pathophysiology, and the mechanisms of deep brain stimulation.

Usando Saccadometry com Estimulação Cerebral Profunda para estudar normal e patológico função cerebral

The oculomotor system involves a large number of brain areas including parts of the basal ganglia, and various neurodegenerative diseases including ...

Ultrasound Assessment of Endothelial Function: A Technical Guideline of the Flow-mediated Dilation Test

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a pathological condition characterized mainly by a disruption in the balance between vasodilator and vasoconstrictor substances and is proposed to play an important role in the development of atherosclerotic cardiovascular disease. Therefore, a precise evaluation of endothelial function in humans represents an important tool that could help better understand the etiology of multiple cardio-centric pathologies.
Over the past twenty-five years, many methodological approaches have been developed to provide an assessment of endothelial function in humans. Introduced in 1989, the FMD test incorporates a forearm occlusion and subsequent reactive hyperemia that promotes nitric oxide production and vasodilation of the brachial artery. The FMD test is now the most widely utilized, non-invasive, ultrasonic assessment of endothelial function in humans and has been associated with future cardiovascular events.
Although the FMD test could have clinical utility, it is a physiological assessment that has inherited several confounding factors that need to be considered. This article describes a standardized protocol for determining FMD including the recommended methodology to help minimize the physiological and technical issues and improve the precision and reproducibility of the assessment.

The flow mediated dilation (FMD) test is the most commonly utilized, non-invasive, ultrasound assessment of endothelial function in humans. Although the FMD test has been related with the prediction of future cardiovascular disease and events, it is a physiological assessment with many inherent confounding factors that need to be considered.

Avaliação ultra-sons da função endotelial: A Orientação Técnica do Teste de dilatação fluxo-mediada

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a ...

Nerve-sparing Mid-urethral Obstruction (NeMO) in Female Small Rodents

Partial bladder outlet obstruction (pBOO) has a high prevalence, causes significant patient burden, and immense health care costs. The most common animal model to investigate bladder remodeling in pBOO are female rodents undergoing partial obstruction at the proximal urethra. Variability in the degree of obstruction and animal mortality are major concerns with proximal obstruction. Furthermore, dissecting around the proximal urethra and bladder neck jeopardizes bladder innervation.
We developed a nerve-sparing mid-urethral obstruction (NeMO) model for pBOO avoiding the disadvantages of the traditional model. We approached the urethra just inferior to the pubic symphysis, which obviated the need for laparotomy as well as for dissection in this area; also, the striated urethral sphincter remained untouched. We performed NeMO in female Sprague-Dawley rats (12 obstructions, 6 sham animals) as well as in female C57/bl6 mice (20 obstructions, 18 sham animals). After two weeks, we evaluated bladder function, bladder mass, and body mass.
We had no mortalities among obstructed- or sham-operated female rats; as described for the traditional proximal pBOO-method, we tied the suture around the proximal urethra and a temporarily placed 0.9 mm metal rod. NeMO induced an 85% increase in bladder mass after two weeks, average residual urine volume was 0.4 mL in partially obstructed rats while only 0.03 mL in sham animals. In mice, we tested 3 sizes of cannulas that we placed along the urethra when tying the suture. We found that using a 27-gauge cannula resulted in over 50% animal mortality; placing the 25-gauge cannula did not yield the desired response in increasing bladder mass; utilizing a 26-gauge cannula yielded favorable results with minimal animal mortality (1/8) yet a significant 2-fold increase in bladder mass.

Nerve-sparing Mid-urethral Obstruction NeMO in Female Small Rodents

Traditional modeling of partial bladder outlet obstruction in rodents is fraught with animal mortality. A denervation injury from dissection around the proximal urethra and bladder neck is also of major concern. We developed and evaluated a safe and reliable mid-urethral obstruction model, avoiding the shortcomings of the traditional model.

-Nervo poupadores Mid-obstrução uretral (Nemo) em fêmeas pequenos roedores

Partial bladder outlet obstruction (pBOO) has a high prevalence, causes significant patient burden, and immense health care costs. The most common animal ...