Os procedimentos são realizados utilizando o sistema de Lipogems.

Todos os procedimentos realizados nos estudos envolvendo participantes humanos estavam em conformidade com os padrões éticos do Comitê de pesquisa institucional e/ou nacional e com a declaração de Helsinque 1964 e suas alterações posteriores ou ético comparável padrões.
1. o histórico
<ol><li>Inicie o exame clínico com um histórico detalhado do paciente. Nota: Um OLT sempre deve ser suspeitado em caso de instabilidade do tornozelo com entorses repetidas associadas com inchaço, rigidez, dor e bloqueio comum. Além disso, em muitos casos pode ser associado com uma história de doença sistêmica, como doença inflamatória ou vascular, neuropatia ou doença neurológica e diabetes OLT. O uso de drogas ou problemas médicos que possam influenciar a cura deve ser avaliado e levado em conta.</li>
</ol>2. clínica exame
<ol><li>Avalie o paciente em posição ortostática para destacar o tornozelo ou pés hind deformidade. Avalie a escala de função e tornozelo músculo e tendão de movimento (ROM). Uma ternura difusa está frequentemente presente, especialmente durante a extensão e flexão máxima, e não é raro encontrar uma área sensível ao toque, ao nível da articulação articular.</li>
	<li>Efectue um teste de gaveta anterior e posterior para identificar uma instabilidade do tornozelo lateral concomitante.</li>
	<li>Durante a consulta pré-operatória, gravar os resultados clínicos e funcionais seguintes: ortopédica americana do pé & tornozelo Society (AOFAS) do tornozelo e pé hind escores15, escala Visual analógica (VAS) dor Pontuação16e 12-Item forma abreviada Saúde Survey (SF-12)17.</li>
</ol>3. Avaliação radiológica
<ol><li>Execute um peso bilateral, tendo a radiografia do pé e tornozelo. Isto consiste em convencional do peso-rolamento ântero-posterior (AP) do mortise e vistas de peso-rolamento laterais do tornozelo. Execute do mortise AP em posição neutra e com 15 graus de rotação interna para uma melhor visualização do tálus. Nota: Apenas 50% de OLT pode ser diagnosticada com radiografia; em caso de lesões grandes, uma área de moradia osso rodeado de radiolucência pode ser notável18.</li>
	<li>Execute um convencional computadorizado tomografia computadorizada-scan (tomografia) do tornozelo. Uma tomografia permite uma localização precisa e o tamanho da lesão, identificando também fragmentos ósseos em caso de desprendimento. O ponto fraco de CT é a capacidade de mostrar o status da cartilagem. Um estudo anterior mostrou uma sensibilidade e especificidade de 0,81 e 0,99, respectivamente, para a detecção de OLTs no CT19,20.</li>
	<li>Realize a ressonância magnética (RM) do tornozelo. Ressonância magnética é fundamental na avaliação de osso de cartilagem e subcondral. Além disso, a ressonância magnética não envolve radiação ionizante e permite uma melhor visualização do tecido mole. A literatura relata uma sensibilidade e especificidade de 96% na detecção de OLTs21,22.</li>
</ol>4. técnica cirúrgica
<ol><li>Colheita e processamento do tecido adiposo
	<ol>
<li>Preparar a solução de Klein: 1 L de solução salina de 9 g/L, 50 mL de lidocaína a 1%, 1 mL de epinefrina 1: 1000, 10 mL de bicarbonato de sódio 8,4% e 0,1 mL de 10 mg/mL de triancinolona acetonina.</li>
		<li>Crie dois pará-umbilical incisões de cerca de 0,5 cm com uma lâmina de bisturi. Injete aproximadamente 300 mL da solução de Klein do tecido adiposo subcutâneo do abdômen através das incisões utilizando seringas de 60 mL com uma agulha de 18G (Figura 1).</li>
		<li>Colheita de 40-45 mL de ADSCs usando uma cânula romba 13G anexado a uma seringa de 20 mL e introduzido o kit de transformação (Figura 2). Normalmente, execute a colheita na área peri-umbilical.</li>
		<li>Inserir 100-130 mL de lipoaspirate no sistema fechado. Empurrar o lipoaspirate para o dispositivo através de um filtro grande para obter uma primeira redução de cluster; ao mesmo tempo, uma quantidade correspondente de solução salina sai em direção a bolsa desperdiçando. Um papel fundamental é desempenhado por bolinhas de aço inoxidável para obter uma emulsão temporária entre óleo, sangue e soro fisiológico. Remova resíduos de óleo e sangue contaminado por uma contracorrente de gravidade de solução salina.</li>
		<li>Após esta etapa de lavagem (a solução fluindo aparece clara e o amarelo de lipoaspirate), parar o fluxo de solução salino e reverter o dispositivo (cap cinza acima), levando à redução do segunda cluster adiposa. Obter a redução, empurrando os clusters adiposos flutuantes através do filtro de corte segundo hexagonal, empurrando o fluido de baixo com uma seringa de 10 mL. Colete o produto final em uma seringa de 10 mL, conectada à abertura superior do dispositivo. Nota: O kit de transformação para o tecido adiposo melhora a técnica padrão lipofilling: na verdade, o sistema consiste de um sistema cilíndrico fechado, imersão completa, baixa pressão, a fim de obter um produto uniforme e fluido contendo um grande número de pericitos. Este procedimento permite a transformação de células adiposas exclusivamente através de forças mecânicas suaves e preservando a integridade do nicho vascular do estroma. O processo é o menos traumático possível e faz com que o produto final disponível em um curto período de tempo (15-20 min), sem tratamentos enzimáticos ou expansão. Sem danos vasculostromal nichos ajudam o processo de cicatrização.</li>
		<li>Uma vez que o ADSCs foram colhidas, aplica uma bandagem de compressão no abdômen.</li>
	</ol></li>
	<li>Procedimento cirúrgico e injeção de tecido adiposo
	<ol>
<li>Coloca o paciente em posição supina sob raquianestesia com um torniquete, a uma pressão de 250 mmHg, no nível da coxa para diminuir o sangramento e permitir uma melhor visualização artroscópica1.</li>
		<li>Marca os pontos anatômicos na pele com uma caneta de dermographic. Marcos são essenciais para evitar lesões iatrogênicas. Marcar o seguinte (Figura 3): ambos os maléolos (lateral (LM) e medial (MM)) a interlinha articular anterior (JL), identificada com dorsi - e -flexão plantar da articulação do tornozelo o tendão tibial anterior que (TAT) e do tendão de Aquiles veia safena o grande, que corre à frente do maléolo medial o nervo fibular superficial (SPN)</li>
		<li>Primeiro, execute o portal ântero-medial só medial ao tendão anterior tibial, coincidindo com um ponto fraco. Este portal representa o portal de abordagem. Na maioria dos casos, uma depressão com o tornozelo em dorsiflexão é visível e palpável.<ol>
<li>Cortar somente a pele com uma lâmina e então perfurar a cápsula por dissecção romba. Tome cuidado para evitar o nervo safeno e da veia grande safena. A veia safena é 9mm localizado lateralmente ao portal, enquanto o nervo é cerca de 7,4 mm lateral para o portal. Isto constitui um dos dois portais de visualização principal.</li>
		</ol>
</li>
		<li>Verifique a linha comum, coloque o portal ântero-lateral, medial para o maléolo lateral e lateral ao tendão extensor dos dedos. Nota: Ao executar o portal ântero-lateral, evitar danos ao nervo cutâneo dorsal intermédio (ramo lateral do nervo fibular superficial); por esta razão, após o corte, a pele deve ser seguida por dissecção romba.</li>
		<li>Inspecione a cartilagem articular para avaliar o tamanho e posição. Avalie a condição e a qualidade da cartilagem com o palpator. Lesões osteocondrais do tálus estão normalmente localizadas ou posteromedially ou anterolateralmente.<ol>
<li>Realize o tratamento artroscópico, usando arthroscopes de 2,7 mm grande angular com um ângulo de 30°, embora alguns cirurgiões usará um artroscópio maior de 4 mm e manter o instrumento no recesso anterior da articulação. Técnicas de distração conjunta não-invasivos e flexão plantar hiper podem ser usados para acessar a maioria da cúpula do tálus.</li>
			<li>No caso de uma lesão posterior, por via percutânea, coloque um propagador Hintermann para distrair a articulação e permitir a exposição da lesão. O propagador Hintermann tem uma alavanca de abertura do braço aplicado em dois fios de Kirschner previamente posicionados na tíbia e osso do tálus medialmente ou lateralmente, de acordo com o lado da lesão. No caso de uma lesão lateral, tome cuidado para inserir o fio de Kirschner proximal no osso da tíbia, evitando a fíbula, para alcançar melhor distração da articulação.</li>
		</ol>
</li>
		<li>Preparar a lesão com uma cureta, remover a cartilagem danificada e instável, a camada calcificada e osso necrótico e esclerótico, a fim de criar uma forma de regular continham lesão com bordas de ombro. Para esta etapa, use uma padrão cureta artroscópica (Figura 4).</li>
		<li>Estimula as células-tronco da medula óssea do osso subcondral realizando microperfurações redondamente do exterior para o interior da lesão.<ol>
<li>Realize perfurações com intervalos de cerca de 3 mm entre eles. Induzi a microfraturas usando uma picareta condral no osso subcondral saudável por baixo do defeito (Figura 5). Observação: Uma hemorragia resulta na formação de um coágulo de fibrina. Coagulação de produtos, através da ativação da cascata inflamatória, liberar mediadores vasoativas, fatores de crescimento e citocinas. Esses fatores têm o poder de estimular a invasão vascular e migração de células-tronco mesenquimais à porção condral da lesão. Estas células pluripotentes são estimuladas a se diferenciar em fibroblastos, condrócitos e osteoblastos e desempenham um papel importante na estimulação de reparação da lesão. Fatores de crescimento parácrina no ambiente articular promover a formação de matriz extracelular e a produção de fibrocartilagem. Ao mesmo tempo, as células da parte da lesão óssea produzem um tecido ósseo imaturo que é progressivamente substituído por um osso maduro.</li>
		</ol>
</li>
		<li>Remova a água intra-articulares, usando a máquina de barbear em aspiração e o líquido restante com uma esponja de algodão até a articulação esteja completamente seca.</li>
	</ol></li>
	<li>Injeção de células-tronco derivadas adiposo
	<ol>
<li>Injete 5-7 mL de células-tronco derivadas adiposo, preparado anteriormente na etapa 4.1.3, na articulação do tornozelo usando um dos dois portais (ântero-medial ou ântero-lateral).</li>
		<li>Libere o torniquete.</li>
	</ol></li>
</ol>5. pós-operatório cuidados
<ol><li>Evite movimentos de tornozelo durante 15 dias, com o membro operado em descarga completa.</li>
	<li>15 dias após a cirurgia, permita movimentos ativos e passivos do tornozelo, até a completa recuperação e amplitude de movimento é alcançado.</li>
	<li>Permita a carga 30 dias após a cirurgia.</li>
</ol>6. clínico e radiográfico de acompanhamento
<ol><li>Avalie pacientes clinicamente e radiograficamente aos seis e doze meses e, posteriormente, anualmente. O protocolo de acompanhamento consiste AOFAS tornozelo e pés hind partituras, pontuação de dor VAS e SF-12 em cada ponto de extremidade15,16,17. Exame radiográfico inclui uma ressonância e tomografia do tornozelo operado.</li>
</ol>

<ol>
	<li>D'Ambrosi, R., Maccario, C., Serra, N., Liuni, F., Usuelli, F. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteochondral+Lesions+of+the+Talus+and+Autologous+Matrix-Induced+Chondrogenesis:+Is+Age+a+Negative+Predictor+Outcome?.">Osteochondral Lesions of the Talus and Autologous Matrix-Induced Chondrogenesis: Is Age a Negative Predictor Outcome?.</a> Arthroscopy. 33 (2), 428-435 (2017).</li><li>Becher, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=T2-mapping+at+3+T+after+microfracture+in+the+treatment+of+osteochondral+defects+of+the+talus+at+an+average+follow-up+of+8+years.">T2-mapping at 3 T after microfracture in the treatment of osteochondral defects of the talus at an average follow-up of 8 years.</a> Knee Surg. SportsTraumatol. Arthrosc. 23 (8), 2406-2412 (2015).</li><li>Polat, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+results+of+microfracture+in+the+treatment+of+talus+osteochondral+lesions.">Long-term results of microfracture in the treatment of talus osteochondral lesions.</a> Knee Surg. Sports Traumatol. Arthrosc. 24 (4), 1299-1303 (2016).</li><li>van Bergen, C. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arthroscopic+treatment+of+osteochondral+defects+of+the+talus:+outcomes+at+eight+to+twenty+years+of+follow-up.">Arthroscopic treatment of osteochondral defects of the talus: outcomes at eight to twenty years of follow-up.</a> J. Bone Joint Surg. Am. 95 (6), 519-525 (2013).</li><li>van Eekeren, I. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Return+to+sports+after+arthroscopic+debridement+and+bone+marrow+stimulation+of+osteochondral+talar+defects:+a+5-+to+24-year+follow-up+study.">Return to sports after arthroscopic debridement and bone marrow stimulation of osteochondral talar defects: a 5- to 24-year follow-up study.</a> Knee Surg Sports Traumatol Arthrosc. 24 (4), 1311-1315 (2016).</li><li>D'Ambrosi, R., Maccario, C., Ursino, C., Serra, N., Usuelli, F. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Combining+Microfractures,+Autologous+Bone+Graft,+and+Autologous+Matrix-Induced+Chondrogenesis+for+the+Treatment+of+Juvenile+Osteochondral+Talar+Lesions.">Combining Microfractures, Autologous Bone Graft, and Autologous Matrix-Induced Chondrogenesis for the Treatment of Juvenile Osteochondral Talar Lesions.</a> Foot Ankle Int. 38 (5), 485-495 (2017).</li><li>Usuelli, F. G., D'Ambrosi, R., Maccario, C., Indino, C., Manzi, L., Maffulli, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adipose-derived+stem+cells+in+orthopaedic+pathologies.">Adipose-derived stem cells in orthopaedic pathologies.</a> British Medical Bulletin. , (2017).</li><li>Kim, Y. S., et al. <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+clinical+and+MRI+outcomes+after+mesenchymal+stem+cell+implantation+in+patients+with+knee+osteoarthritis:+a+prospective+study.">Assessment of clinical and MRI outcomes after mesenchymal stem cell implantation in patients with knee osteoarthritis: a prospective study.</a> Osteoarthr Cartilage. 24 (2), 237-245 (2016).</li><li>Koh, Y. G., Choi, Y. J., Kwon, S. K., Kim, Y. S., Yeo, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+results+and+second-look+arthroscopic+findings+after+treatment+with+adipose-derived+stem+cells+for+knee+osteoarthritis.">Clinical results and second-look arthroscopic findings after treatment with adipose-derived stem cells for knee osteoarthritis.</a> Knee Surg. Sports Traumatol. Arthrosc. 23 (5), 1308-1316 (2015).</li><li>Zuk, P. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+adipose+tissue+is+a+source+of+multipotent+stem+cells.">Human adipose tissue is a source of multipotent stem cells.</a> Mol. Biol. Cell. 13 (12), 4279-4295 (2002).</li><li>Tal&#233;ns-Visconti, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+mesenchymal+stem+cells+from+adipose+tissue:+Differentiation+into+hepatic+lineage.">Human mesenchymal stem cells from adipose tissue: Differentiation into hepatic lineage.</a> Toxicol. In Vitro. 21 (2), 324-329 (2007).</li><li>Timper, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+adipose+tissue-derived+mesenchymal+stem+cells+differentiate+into+insulin,+somatostatin,+and+glucagon+expressing+cells.">Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells.</a> Biochem. Biophys. Res. Commun. 341 (4), 1135-1140 (2006).</li><li>Tremolada, C., Palmieri, G., Ricordi, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adipocyte+transplantation+and+stem+cells:+plastic+surgery+meets+regenerative+medicine.">Adipocyte transplantation and stem cells: plastic surgery meets regenerative medicine.</a> Cell. Transplant. 19 (10), 1217-1223 (2010).</li><li>Keramaris, N. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endothelial+progenitor+cells+(EPCs)+and+mesenchymal+stem+cells+(MSCs)+in+bone+healing.">Endothelial progenitor cells (EPCs) and mesenchymal stem cells (MSCs) in bone healing.</a> Curr. Stem Cell. Res. Ther. 7 (4), 293-301 (2012).</li><li>Leigheb, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Italian+translation,+cultural+adaptation+and+validation+of+the+American+Orthopaedic+Foot+and+Ankle+Society's+(AOFAS)+ankle-hindfoot+scale.">Italian translation, cultural adaptation and validation of the American Orthopaedic Foot and Ankle Society's (AOFAS) ankle-hindfoot scale.</a> Acta Biomed. 87 (1), 38-45 (2016).</li><li>Ware, J., Kosinski, M., Keller, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+12-Item+Short-Form+Health+Survey:+construction+of+scales+and+preliminary+tests+of+reliability+and+validity.">A 12-Item Short-Form Health Survey: construction of scales and preliminary tests of reliability and validity.</a> Med. Care. 34 (3), 220-233 (1996).</li><li>Hawker, G. A., Mian, S., Kendzerska, T., French, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measures+of+adult+pain:+Visual+Analog+Scale+for+Pain+(VAS+Pain),+Numeric+Rating+Scale+for+Pain+(NRS+Pain),+McGill+Pain+Questionnaire+(MPQ),+Short-Form+McGill+Pain+Questionnaire+(SF-MPQ),+Chronic+Pain+Grade+Scale+(CPGS),+Short+Form-36+Bodily+Pain+Scale+(SF-36+BPS),+and+Measure+of+Intermittent+and+Constant+Osteoarthritis+Pain+(ICOAP).">Measures of adult pain: Visual Analog Scale for Pain (VAS Pain), Numeric Rating Scale for Pain (NRS Pain), McGill Pain Questionnaire (MPQ), Short-Form McGill Pain Questionnaire (SF-MPQ), Chronic Pain Grade Scale (CPGS), Short Form-36 Bodily Pain Scale (SF-36 BPS), and Measure of Intermittent and Constant Osteoarthritis Pain (ICOAP).</a> Arthritis Care (Hoboken). 63, S240-S252 (2011).</li><li>Bergen, C. J., Gerards, R. M., Opdam, K. T., Terra, M. P., Kerkhoffs, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diagnosing,+planning+and+evaluating+osteochondral+ankle+defects+with+imaging+modalities.">Diagnosing, planning and evaluating osteochondral ankle defects with imaging modalities.</a> World. J. Orthop. 6 (11), 944-953 (2015).</li><li>van Dijk, C. N., Reilingh, M. L., Zengerink, M., van Bergen, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteochondral+defects+in+the+ankle:+why+painful?.">Osteochondral defects in the ankle: why painful?.</a> Knee Surg. Sports Traumatol. Arthrosc. 18 (5), 570-580 (2010).</li><li>Madry, H., van Dijk, C. N., Mueller-Gerbl, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+basic+science+of+the+subchondral+bone.">The basic science of the subchondral bone.</a> Knee Surg. Sports Traumatol. Arthrosc. 18 (4), 419-433 (2010).</li><li>Mintz, D. N., Tashjian, G. S., Connell, D. A., Deland, J. T., O'Malley, M., Potter, H. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteochondral+lesions+of+the+talus:+a+new+magnetic+resonance+grading+system+with+arthroscopic+correlation.">Osteochondral lesions of the talus: a new magnetic resonance grading system with arthroscopic correlation.</a> Arthroscopy. 19 (4), 353-359 (2003).</li><li>Leumann, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+imaging+method+for+osteochondral+lesions+of+the+talus--comparison+of+SPECT-CT+with+MRI.">A novel imaging method for osteochondral lesions of the talus--comparison of SPECT-CT with MRI.</a> Am. J. Sports Med. 39 (5), 1095-1101 (2011).</li><li>Kim, Y. S., Park, E. H., Kim, Y. C., Koh, Y. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+outcomes+of+mesenchymal+stem+cell+injection+with+arthroscopic+treatment+in+older+patients+with+osteochondral+lesions+of+the+talus.">Clinical outcomes of mesenchymal stem cell injection with arthroscopic treatment in older patients with osteochondral lesions of the talus.</a> Am. J. Sports Med. 41 (5), 1090-1099 (2013).</li><li>Kim, Y. S., Lee, H. J., Choi, Y. J., Kim, Y. I., Koh, Y. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+an+injection+of+a+stromal+vascular+fraction+containing+adipose-derived+mesenchymal+stem+cells+influence+the+outcomes+of+marrow+stimulation+in+osteochondral+lesions+of+the+talus?+A+clinical+and+magnetic+resonance+imaging+study.">Does an injection of a stromal vascular fraction containing adipose-derived mesenchymal stem cells influence the outcomes of marrow stimulation in osteochondral lesions of the talus? A clinical and magnetic resonance imaging study.</a> Am. J. Sports Med. 42 (10), 2424-2434 (2014).</li><li>Kim, Y. S., Koh, Y. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Injection+of+Mesenchymal+Stem+Cells+as+a+Supplementary+Strategy+of+Marrow+Stimulation+Improves+Cartilage+Regeneration+After+Lateral+Sliding+Calcaneal+Osteotomy+for+Varus+Ankle+Osteoarthritis:+Clinical+and+Second-Look+Arthroscopic+Results.">Injection of Mesenchymal Stem Cells as a Supplementary Strategy of Marrow Stimulation Improves Cartilage Regeneration After Lateral Sliding Calcaneal Osteotomy for Varus Ankle Osteoarthritis: Clinical and Second-Look Arthroscopic Results.</a> Arthroscopy. 32 (5), 878-889 (2016).</li><li>Kim, Y. S., Lee, M., Koh, Y. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Additional+mesenchymal+stem+cell+injection+improves+the+outcomes+of+marrow+stimulation+combined+with+supramalleolar+osteotomy+in+varus+ankle+osteoarthritis:+short-term+clinical+results+with+second-look+arthroscopic+evaluation.">Additional mesenchymal stem cell injection improves the outcomes of marrow stimulation combined with supramalleolar osteotomy in varus ankle osteoarthritis: short-term clinical results with second-look arthroscopic evaluation.</a> J. Exp. Orthop. 3 (1), 12 (2016).</li><li>Hanke, C. W., Bernstein, G., Bullock, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Safety+of+tumescent+liposuction+in+15,336+patients.+National+survey+results.">Safety of tumescent liposuction in 15,336 patients. National survey results.</a> Dermatol Surg. 21 (5), 459-462 (1995).</li><li>Illouz, Y. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complications+of+liposuction.">Complications of liposuction.</a> Clin Plast Surg. 33 (1), 129-163 (2006).</li><li>Dixit, V. V., Wagh, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unfavourable+outcomes+of+liposuction+and+their+management.">Unfavourable outcomes of liposuction and their management.</a> Indian J Plast Surg. 46 (2), 377-392 (2013).</li><li>Lehnhardt, M., Homann, H. H., Daigeler, A., Hauser, J., Palka, P., Steinau, H. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Major+and+lethal+complications+of+liposuction:+review+of+72+cases+in+Germany+between+1998+and+2002.">Major and lethal complications of liposuction: review of 72 cases in Germany between 1998 and 2002.</a> Plast Reconstr Surg. 121 (6), 396e-403e (2008).</li><li>Usuelli, F. G., de Girolamo, L., Grassi, M., D'Ambrosi, R., Montrasio, U. A., Boga, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=All-Arthroscopic+Autologous+Matrix-Induced+Chondrogenesis+for+the+Treatment+of+Osteochondral+Lesions+of+the+Talus.">All-Arthroscopic Autologous Matrix-Induced Chondrogenesis for the Treatment of Osteochondral Lesions of the Talus.</a> Arthrosc Tech. 4 (3), e255-e259 (2015).</li><li>Simonson, D. C., Roukis, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Safety+of+ankle+arthroscopy+for+the+treatment+of+anterolateral+soft-tissue+impingement.">Safety of ankle arthroscopy for the treatment of anterolateral soft-tissue impingement.</a> Arthroscopy. 30 (2), 256-259 (2014).</li><li>Suzangar, M., Rosenfeld, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ankle+arthroscopy:+is+preoperative+marking+of+the+superficial+peroneal+nerve+important?.">Ankle arthroscopy: is preoperative marking of the superficial peroneal nerve important?.</a> J. Foot. Ankle Surg. 51 (2), 179-181 (2012).</li><li>Kraeutler, M. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+Concepts+Review+Update:+Osteochondral+Lesions+of+the+Talus.">Current Concepts Review Update: Osteochondral Lesions of the Talus.</a> Foot Ankle Int. 38 (3), 331-342 (2017).</li><li>Looze, C. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+and+Management+of+Osteochondral+Lesions+of+the+Talus.">Evaluation and Management of Osteochondral Lesions of the Talus.</a> Cartilage. 8 (1), 19-30 (2017).</li><li>Dragoo, J. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Healing+full-thickness+cartilage+defects+using+adipose-derived+stem+cells.">Healing full-thickness cartilage defects using adipose-derived stem cells.</a> Tissue Eng. 13 (7), 1615-1621 (2007).</li><li>Lee, S. Y., Kim, W., Lim, C., Chung, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment+of+Lateral+Epicondylosis+by+Using+Allogeneic+Adipose-Derived+Mesenchymal+Stem+Cells:+A+Pilot+Study.">Treatment of Lateral Epicondylosis by Using Allogeneic Adipose-Derived Mesenchymal Stem Cells: A Pilot Study.</a> Stem Cells. 33 (10), 2995-3005 (2015).</li><li>Feisst, V., Meidinger, S., Locke, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=From+bench+to+bedside:+use+of+human+adipose-derived+stem+cells.">From bench to bedside: use of human adipose-derived stem cells.</a> Stem Cells Cloning. 8, 149-162 (2015).</li></ol>

Federico Giuseppe Usuelli, MD, informa taxas pessoais de Integra e Geistlich e subvenções e taxas pessoais de Zimmer, fora o trabalho enviado.

Nos últimos anos, ensaios pré-clínicos e clínicos têm focado sua atenção o efeito da ADSCs para tratar diferentes patologias músculo-esqueléticas. O objetivo deste artigo é descrever o protocolo para o tratamento das lesões osteocondrais do tálus usando microfractured e purificado de tecido adiposo em associação com microperfurações artroscópica. O protocolo envolve várias etapas críticas com alto risco de complicações. Durante a colheita de gordura, as complicações podem ser divididas em complica?...

Artroscopia é o padrão ouro para o tratamento de lesões osteocondrais do tálus (OLTs) com o objetivo de alívio da dor, restaurando a funcionalidade e melhorando a qualidade de vida, especialmente em pacientes jovens e ativos.
Atualmente, técnicas artroscópicas podem ser classificadas de três maneiras. A técnica reparativa estimula células derivadas da medula óssea através de um desbridamento e microperfurações ao nível da lesão. A técnica reconstrutiva substitui a lesão utilizando um enxerto autólogo ou heterólogo ostechondral. A técnica regenerativa explora a capacidade das células multipotentes diferenciar e replicar para reconstruir o tecido danificado1,2,3,4,5,6 .
Nos últimos anos, técnicas regenerativas têm sido objecto de numerosos estudos in vitro e in vivo para o tratamento de OLTs e particularmente mesenquimais células-tronco derivadas de tecido adiposo (ADSCs)7,8 , 9. essas células-tronco mesenquimais apresentam características morfológicas e funcionais semelhantes a outras células multipotentes, isoladas de outros tecidos; Eles também têm a capacidade de se diferenciarem em vários e celulares diferentes linhas em vitro e em vivo10,11,12,13. O foco na pesquisa sobre essas células é principalmente devido à sua localização, na verdade, eles representam de 10% a 30% do peso corporal normal, com uma concentração de 5.000 células por grama de tecido13,14. Por outro lado, um fator que limita o uso dessas células está relacionado ao seu manuseamento durante os procedimentos de laboratório. O lipoaspirate contendo agregados de adipócitos, fibras de colágeno e componentes vasculares normais é enzimaticamente processada com colágeno A tipo I e submetido a hemólise antes de cultura. O objetivo aqui é descrever o protocolo para o tratamento das lesões osteocondrais do tálus usando microfractured e purificado de tecido adiposo.

<table>
	<tbody>
		<tr>
			<td>PROCESS KIT - PROCESSING KIT FOR FAT TISSUE</td>
			<td>LIPOGEMS</td>
			<td>LG PK 60</td>
			<td>Lipogems Kit to obtain microfractured and purified ADSCs</td>
		</tr>
		<tr>
			<td>HINTERMANN SPREADER</td>
			<td>INTEGRA</td>
			<td>119654</td>
			<td>The spreader allow to access most of the talar dome, in particular in case of posterior lesion</td>
		</tr>
		<tr>
			<td>CUP CURETTE</td>
			<td>ARTHREX</td>
			<td>AR-8655-02</td>
			<td>To remove the damaged cartilage and necrotic and sclerotic bone</td>
		</tr>
		<tr>
			<td>CHONDRAL PICK 30&#176; TIP / 60&#176; TIP</td>
			<td>ARTHREX</td>
			<td>AR-8655-05 
			AR-8655-06</td>
			<td>To perfrom microperforation at the level of the lesion, stimulating bleeding from the subchondral bone</td>
		</tr>
		<tr>
			<td>SHAVER</td>
			<td>ARTHREX</td>
			<td>AR-7300SR</td>
			<td>To clean the joint and aspirate water</td>
		</tr>
	</tbody>
</table>

autologous microfractured and purified adipose tissue for arthroscopic management of osteochondral lesions of the talus

Nos últimos anos, técnicas regenerativas foram cada vez mais estudadas e usadas para tratar lesões osteocondrais do tálus. Em particular, vários estudos têm focado sua atenção as células-tronco mesenquimais derivadas de tecido adiposo. Células-tronco adiposo-derivado (ADSCs) exibem características morfológicas e propriedades semelhantes a outras células mesenquimais e são capazes de se diferenciar em várias linhas celulares. Além disso, essas células também são amplamente disponíveis no tecido subcutâneo, representando 10-30% do peso corporal normal, com uma concentração de 5.000 células por grama de tecido.
A técnica apresentada, a primeira etapa envolve a colheita ADSCs do abdômen e um processo de microfratura e purificação; em seguida, o procedimento cirúrgico é realizado inteiramente artroscopicamente, com menos dissecção de tecidos moles, melhor visualização conjunta e uma recuperação mais rápida em comparação com os procedimentos padrão de abrir. Artroscopia é caracterizada por uma primeira fase em que a lesão é identificada, isolada e preparada com microperfurações; a segunda etapa, realizada a seco, envolve a injeção de tecido adiposo a nível da lesão.
Entre janeiro de 2016 e setembro de 2016, quatro pacientes foram submetidos a tratamento artroscópico da lesão osteocondral do tálus com microfractured e purificado de tecido adiposo. Todos os pacientes relataram melhora clínica, seis meses após a cirurgia sem complicações relatadas. Resultados funcionais no seguimento mais recente são encorajadores e confirmam que a técnica proporciona alívio da dor confiável e melhorias em pacientes com lesão osteocondral do tálus.

Entre janeiro de 2016 e setembro de 2016, quatro pacientes foram submetidos a tratamento artroscópico da lesão osteocondral do tálus com microfractured e purificado de tecido adiposo. Todos os pacientes relataram melhora clínica, seis meses após a cirurgia. Os resultados clínicos preliminares são relatados na tabela 1. Sem complicações foram relatadas.
Nos últimos anos, aumentou-se o uso de ADSCs para ...

Watch this Scientific Journal Video about Autologous Microfractured and Purified Adipose Tissue for Arthroscopic Management of Osteochondral Lesions of the Talus at JoVE.com

Autologous Microfractured and Purified Adipose Tissue for Arthroscopic Management of Osteochondral Lesions of the Talus

O objetivo deste estudo é relatar um protocolo para o tratamento artroscópico das lesões osteocondrais do tálus usando microfractured e células-tronco adiposo-derivado purificadas.

Microfractured autólogo e purificado tecido adiposo para gestão artroscópica das lesões osteocondrais do tálus

In recent years, regenerative techniques have been increasingly studied and used to treat osteochondral lesions of the talus. In particular, several ...

autologous-microfractured-purified-adipose-tissue-for-arthroscopic

Research

JoVE Journal

Medicine

10.2K visualizações.  IRCCS Istituto Ortopedico Galeazzi - C.A.S.C.O. Piede e Caviglia. O objetivo deste estudo é relatar um protocolo para o tratamento artroscópico das lesões osteocondrais do tálus usando microfractured e células-tronco adiposo-derivado purificadas.

Vídeo: Autologous Microfractured and Purified Adipose Tissue for Arthroscopic Management of Osteochondral Lesions of the Talus

Breathing-controlled Electrical Stimulation (BreEStim) for Management of Neuropathic Pain and Spasticity

Electrical stimulation (EStim) refers to the application of electrical current to muscles or nerves in order to achieve functional and therapeutic goals. It has been extensively used in various clinical settings. Based upon recent discoveries related to the systemic effects of voluntary breathing and intrinsic physiological interactions among systems during voluntary breathing, a new EStim protocol, Breathing-controlled Electrical Stimulation (BreEStim), has been developed to augment the effects of electrical stimulation. In BreEStim, a single-pulse electrical stimulus is triggered and delivered to the target area when the airflow rate of an isolated voluntary inspiration reaches the threshold. BreEStim integrates intrinsic physiological interactions that are activated during voluntary breathing and has demonstrated excellent clinical efficacy. Two representative applications of BreEStim are reported with detailed protocols: management of post-stroke finger flexor spasticity and neuropathic pain in spinal cord injury.

Breathing-controlled Electrical Stimulation BreEStim for Management of Neuropathic Pain and Spasticity

The purpose is to present a new method, breathing-control electrical stimulation (BreEStim) for management of neuropathic pain and spasticity.

Respiração controlada Estimulação Elétrica (BreEStim) para tratamento de dor neuropática e espasticidade

Electrical stimulation (EStim) refers to the application of electrical current to muscles or nerves in order to achieve functional and therapeutic goals. ...

Medicina

Steps for the Autologous Ex vivo Perfused Porcine Liver-kidney Experiment

The use of ex vivo perfused models can mimic the physiological conditions of the liver for short periods, but to maintain normal homeostasis for an extended perfusion period is challenging. We have added the kidney to our previous ex vivo perfused liver experiment model to reproduce a more accurate physiological state for prolonged experiments without using live animals. Five intact livers and kidneys were retrieved post-mortem from sacrificed pigs on different days and perfused for a minimum of 6 hr. Hourly arterial blood gases were obtained to analyze pH, lactate, glucose and renal parameters. The primary endpoint was to investigate the effect of adding one kidney to the model on the acid base balance, glucose, and electrolyte levels. The result of this liver-kidney experiment was compared to the results of five previous liver only perfusion models. In summary, with the addition of one kidney to the ex vivo liver circuit, hyperglycemia and metabolic acidosis were improved. In addition this model reproduces the physiological and metabolic responses of the liver sufficiently accurately to obviate the need for the use of live animals. The ex vivo liver-kidney perfusion model can be used as an alternative method in organ specific studies. It provides a disconnection from numerous systemic influences and allows specific and accurate adjustments of arterial and venous pressures and flow.

Steps for the Autologous Ex vivo Perfused Porcine Liver-kidney Experiment

Study of the animal organ physiology can be achieved by performing an experimental ex vivo perfusion system. The addition of a porcine kidney, as a homeostatic organ to our previously developed ex vivo liver perfusion model can be a principal step to achieve a better physiological environment.

The use of ex vivo perfused models can mimic the physiological conditions of the liver for short periods, but to maintain normal homeostasis for an ...

Generation and 3-Dimensional Quantitation of Arterial Lesions in Mice Using Optical Projection Tomography

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating important information on the pathogenesis of lesion formation and the action of novel therapies. Use of atherosclerosis-prone mice, surgical methods of lesion induction, and dietary modification has dramatically improved understanding of the mechanisms that contribute to disease development and the potential of new treatments.
Classically, analysis of lesions is performed ex vivo using 2-dimensional histological techniques. This article describes application of optical projection tomography (OPT) to 3-dimensional quantitation of arterial lesions. As this technique is non-destructive, it can be used as an adjunct to standard histological and immunohistochemical analyses.
Neointimal lesions were induced by wire-insertion or ligation of the mouse femoral artery whilst atherosclerotic lesions were generated by administration of an atherogenic diet to apoE-deficient mice.
Lesions were examined using OPT imaging of autofluorescent emission followed by complementary histological and immunohistochemical analysis. OPT clearly distinguished lesions from the underlying vascular wall. Lesion size was calculated in 2-dimensional sections using planimetry, enabling calculation of lesion volume and maximal cross-sectional area. Data generated using OPT were consistent with measurements obtained using histology, confirming the accuracy of the technique and its potential as a complement (rather than alternative) to traditional methods of analysis.
This work demonstrates the potential of OPT for imaging atherosclerotic and neointimal lesions. It provides a rapid, much needed ex vivo technique for the routine 3-dimensional quantification of vascular remodelling.

Ex vivo analysis of arterial lesions from animal models of cardiovascular disease classically relies on histological and immunohistochemical techniques. These provide 2-dimensional measurements in 3-dimensional lesions. This manuscript describes the generation of arterial lesions for quantitative analysis in 3-dimensions using optical projection tomography.

Geração e quantificação 3-Dimensional de Arterial lesões em ratos por meio de tomografia óptica de Projeção

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating ...

Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely related to metabolism and energy expenditure, and its dysfunction is one important contributor for obesity and diabetes. Contrary to previous belief, recent PET/CT imaging studies indicated the BAT depots are still present in human adults. PET imaging clearly shows that BAT has considerably high uptake of 18F-FDG under certain conditions. In this video report, we demonstrate that Cerenkov luminescence imaging (CLI) with 18F-FDG can be used to optically image BAT in small animals. BAT activation is observed after intraperitoneal injection of norepinephrine (NE) and cold treatment, and depression of BAT is induced by long anesthesia. Using multiple-filter Cerenkov luminescence imaging, spectral unmixing and 3D imaging reconstruction are demonstrated. Our results suggest that CLI with 18F-FDG is a practical technique for imaging BAT in small animals, and this technique can be used as a cheap, fast, and alternative imaging tool for BAT research.

In this video report, we show the application of Cerenkov Luminescence Imaging (CLI) for interscapular brown adipose tissue in mice under activated and depressed conditions.

Cerenkov luminescência imagem de Interscapular Brown tecido adiposo

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely ...

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular weight metabolites in tissues of the treated host to achieve long-term survival. The semipermeable membrane allows engrafted encapsulated cells to avoid rejection by the immune system. The encapsulation procedure was designed to enable a controlled release of bioactive compounds, such as insulin, other hormones, and cytokines. Here we describe a method for encapsulation of catabolic cells, which consume lipids for heat production and energy dissipation (thermogenesis) in the intra-abdominal adipose tissue of obese mice. Encapsulation of thermogenic catabolic cells may be potentially applicable to the prevention and treatment of obesity and type 2 diabetes. Another potential application of catabolic cells may include detoxification from alcohols or other toxic metabolites and environmental pollutants.

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

Encapsulation termogênico pré-adipócitos para transplante em adiposo Depots Tissue

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

A Technique for Subcutaneous Abdominal Adipose Tissue Biopsy via a Non-diathermy Method

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To obtain subcutaneous adipose tissue biopsies in the abdominal area, researchers and physicians use either a surgical or a needle-based technique. However, surgical subcutaneous fat biopsies can offer tissue samples that may provide a more comprehensive overview of the complexities of biological indices in white adipose tissue. Usually, a surgical adipose tissue biopsy includes a diathermy treatment for cauterizing blood vessels to prevent excessive bleeding. Nevertheless, side effects, such as flash fires and skin lesions in the tissue, have been reported after diathermy. Therefore, we aimed to standardize a surgical abdominal adipose tissue biopsy performed under local anesthesia using a non-diathermy method. We conducted 115 subcutaneous adipose tissue biopsies in healthy men using a non-diathermy abdominal surgical biopsy method. Our results showed three cases of excessive post-operation bleeding out of 115 operations (2.61%).In conclusion, our standardized subcutaneous abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men at the bedside, with minimal side effects.

We standardized an abdominal adipose tissue biopsy using a non-diathermy method performed under local anesthesia. Three cases of excessive post-operation bleeding out of 115 operations (2.61%) occurred.We conclude that an abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men.

Uma técnica para biopsia de tecido adiposo Abdominal subcutâneo através de um método não-diatermia

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To ...

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 &#181;m internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Videomicroscopy systems are used to examine functional properties of isolated adipose tissue arterioles in response to physiological and pharmacological stimuli. This technique can be used to examine microvascular phenotypes in different adipose tissue domains in obese humans.

Avaliação da função Microvascular de tecido adiposo humano usando tele-microscopia

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. ...

Clinical Protocol of Producing Adipose Tissue-Derived Stromal Vascular Fraction for Potential Cartilage Regeneration

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the knees by using different forms of mesenchymal stem cells (MSCs). In Korea, bone marrow concentrates and cord blood-derived stem cells have been approved by the Korean Food and Drug Administration (KFDA) for cartilage regeneration. In addition, an adipose tissue-derived stromal vascular fraction (SVF) has been allowed by the KFDA for joint injections in human patients. Autologous adipose tissue-derived SVF contains extracellular matrix (ECM) in addition to mesenchymal stem cells. ECM excretes various cytokines that, along with hyaluronic acid (HA) and platelet-rich plasma (PRP) activated by calcium chloride, may help MSCs to regenerate cartilage and improve knee functions. In this article, we presented a protocol to improve knee functions by regenerating cartilage-like tissue in human patients with OA. The result of the protocol was first reported in 2011 followed by a few additional publications. The protocol involves liposuction to obtain autologous lipoaspirates that are mixed with collagenase. This lipoaspirates-collagenase mixture is then cut and homogenized to remove large fibrous tissue that may clog up the needle during the injection. Afterwards, the mixture is incubated to obtain adipose tissue-derived SVF. The resulting adipose tissue-derived SVF, containing both adipose tissue-derived MSCs and remnants of ECM, is injected into knees of patients, combined with HA and calcium chloride activated PRP. Included are three cases of patients who were treated with our protocol resulting in improvement of knee pain, swelling, and range of motion along with MRI evidence of hyaline cartilage-like tissue.

Here, we present a protocol to produce an adipose tissue-derived stromal vascular fraction and its application to improve knee functions by regenerating cartilage-like tissue in human patients with osteoarthritis.

Protocolo clínico de produzir tecido adiposo-derivado do estroma Vascular fração para a regeneração da cartilagem potenciais

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the ...

Inguinal Subcutaneous White Adipose Tissue (ISWAT) Transplantation Model of Murine Islets

Pancreatic islet transplantation is a well-established therapeutic treatment for type 1 diabetes. The kidney capsule is the most commonly used site for islet transplantation in rodent models. However, the tight kidney capsule limits the transplantation of sufficient islets in large animals and humans. The inguinal subcutaneous white adipose tissue (ISWAT), a new subcutaneous space, was found to be a potentially valuable site for islet transplantation. This site has better blood supply than other subcutaneous spaces. Moreover, the ISWAT accommodates a larger islet mass than the kidney capsule, and transplantation into it is simple. This manuscript describes the procedure of mouse islet isolation and transplantation in the ISWAT site of syngeneic diabetic mouse recipients. Using this protocol, murine pancreatic islets were isolated by standard collagenase digestion and a basement membrane matrix hydrogel was used for fixing the purified islets in the ISWAT site. The blood glucose levels of the recipient mice were monitored for more than 100 days. Islet grafts were retrieved at day 100 after transplantation for histological analysis. The protocol for islet transplantation in the ISWAT site described in this manuscript is simple and effective.

Inguinal Subcutaneous White Adipose Tissue ISWAT Transplantation Model of Murine Islets

In this protocol, a method of murine islet isolation and transplantation into the inguinal subcutaneous white adipose tissue is described. Isolated syngeneic murine islets are transplanted into a murine recipient using a basement membrane hydrogel. The blood glucose level of the recipients is monitored, and histology analysis of the islet grafts is performed.

Modelo de transplante de tecido adiposo branco subcutâneo inguinal (ISWAT) de Isletas murina

Pancreatic islet transplantation is a well-established therapeutic treatment for type 1 diabetes. The kidney capsule is the most commonly used site for ...

Fat-Covered Islet Transplantation Using Epididymal White Adipose Tissue

Islet transplantation is a cellular replacement therapy for severe diabetes mellitus. The intraperitoneal cavity is typically the transplant site for this procedure. However, intraperitoneal islet transplantation has some limitations, including poor transplant efficacy, difficult graft detection ability, and a lack of graftectomy capability for post-transplant analysis. In this paper, "fat-covered islet transplantation", an intraperitoneal islet transplantation method that utilizes epididymal white adipose tissue, is used to assess the therapeutic effects of bioengineered islets. The simplicity of the method lies in the seeding of islets onto epididymal white adipose tissue and using the tissue to cover the islets. While this method can be categorized as an intraperitoneal islet transplantation technique, it shares characteristics with intra-adipose tissue islet transplantation. The fat-covered islet transplantation method demonstrates more robust therapeutic effects than intra-adipose tissue islet transplantation, however, including the improvement of blood glucose and plasma insulin levels and the potential for graft removal. We recommend the adoption of this method for assessing the mechanisms of islet engraftment into white adipose tissue and the therapeutic effects of bioengineered islets.

This fat-covered islet transplantation method is suitable for the detection of engrafted islets in the intraperitoneal cavity. Notably, it does not require the use of biobinding agents or suturing.

Transplante de ilhotas cobertas de gordura usando tecido adiposo branco epididimal

Islet transplantation is a cellular replacement therapy for severe diabetes mellitus. The intraperitoneal cavity is typically the transplant site for this ...