Le procedure vengono eseguite utilizzando il sistema di Lipogems.

Tutte le procedure eseguite negli studi che coinvolgono i partecipanti umani erano in conformità con gli standard etici del Comitato di ricerca istituzionali e/o nazionali e con la dichiarazione di Helsinki del 1964 e sue successive modifiche o etici comparabili standard.
1. anamnesi
<ol><li>Iniziare l'esame clinico con una storia dettagliata del paziente. Nota: Un OLT deve sempre essere sospettato in caso di instabilità della caviglia con ripetute distorsioni associati con gonfiore, rigidità, dolore e blocco comune. Inoltre, in molti casi OLT può essere associato con una storia di malattia sistemica, come malattia infiammatoria o vascolare, neuropatia o malattia neurologica ed il diabete. L'uso di droghe o problemi medici che possono influenzare la guarigione deve essere valutato e preso in considerazione.</li>
</ol>2. esame obiettivo
<ol><li>Valutare il paziente in posizione ortostatica per evidenziare deformità hind-piede o della caviglia. Valutare muscoli e dei tendini della caviglia e funzione gamma di movimento (ROM). Una tenerezza diffusa è spesso presente, specialmente durante la massima flessione ed estensione, e non è raro incontrare una zona sensibile al tocco a livello del giunto articolare.</li>
	<li>Eseguire un test del cassetto anteriore e posteriore per identificare un'instabilità laterale concomitante della caviglia.</li>
	<li>Durante la consultazione pre-operatoria, registrare i seguenti punteggi clinici e funzionali: piede ortopedico americano & Ankle Society (AOFAS) hind-piede e caviglia punteggi15, scala analogica visiva (VAS) dolore Punteggio16e 12-elemento forma breve Health Survey (SF-12)17.</li>
</ol>3. valutazione radiologica
<ol><li>Eseguire una radiografia del piede e della caviglia di appoggio bilaterale. Si tratta di convenzionale del peso-cuscinetto antero-posteriore (AP) mortasa e viste laterali del peso-cuscinetto della caviglia. Eseguire da infilare AP in posizione neutra e con 15 gradi di rotazione interna per una migliore visualizzazione del talus. Nota: Solo il 50% di OLT può essere diagnosticato con radiografia; in caso di grandi lesioni, un'area di osso indipendente circondato da radiotrasparenza può essere notato18.</li>
	<li>Eseguire una convenzionale computata-esplorazione di tomografia (TAC) della caviglia. La TAC permette una precisa posizione e le dimensioni della lesione, individuando anche frammenti ossei in caso di distacco. Il punto debole del CT è la capacità di mostrare lo stato della cartilagine. Uno studio precedente ha mostrato una sensibilità e una specificità di 0,81 e 0,99, rispettivamente, per la rilevazione OLTs CT19,20.</li>
	<li>Eseguire la formazione immagine a risonanza magnetica (MRI) della caviglia. MRI è fondamentale nella valutazione dell'osso subcondrale e della cartilagine. Inoltre, la MRI non riguarda le radiazioni ionizzanti e permette una migliore visualizzazione dei tessuti molli. La letteratura segnala una sensibilità e specificità del 96% nella rilevazione OLTs21,22.</li>
</ol>4. tecnica chirurgica
<ol><li>Raccolta e lavorazione del tessuto adiposo
	<ol>
<li>Preparare la soluzione di Klein: 1 L di soluzione salina 9 g/L, 50 mL di lidocaina 1%, 1 mL di epinefrina 1: 1000, 10 mL di bicarbonato di sodio 8,4% e 0,1 mL di 10 mg/mL di acetonide del triamcinolone.</li>
		<li>Creare due para-ombelicale incisioni di circa 0,5 cm con un bisturi. Iniettare circa 300 mL di soluzione di Klein nel tessuto grasso sottocutaneo dell'addome attraverso le incisioni utilizzando siringhe da 60 mL con un ago da 18G (Figura 1).</li>
		<li>Raccogli 40-45 mL di CSTA utilizzando una cannula smussa 13G collegato a una siringa da 20 mL e introdusse il kit di trasformazione (Figura 2). In genere, di eseguire la raccolta nella zona peri-ombelicale.</li>
		<li>Inserire 100-130 mL di lipoaspirato nel sistema chiuso. Spingere il lipoaspirato nel dispositivo attraverso un filtro di grandi dimensioni per ottenere una prima riduzione di cluster; allo stesso tempo, una quantità corrispondente di soluzione salina esce verso la borsa lo spreco. Un ruolo chiave è giocato da biglie di acciaio inossidabile per ottenere un'emulsione temporanea tra olio, sangue e soluzione salina. Rimuovere residui di olio e sangue contaminato da un contro-flusso gravità di soluzione salina.</li>
		<li>Dopo questa fase di lavaggio (la soluzione scorrevole appare chiaro e il giallo di lipoaspirato), interrompere il flusso di soluzione salino e invertire il dispositivo (cappuccio grigio fino), che conduce la seconda riduzione adiposa cluster. Ottenere la riduzione spingendo i cluster adiposi galleggianti attraverso il secondo filtro esagonale taglio, spingendo da sotto liquido con una siringa da 10 mL. Raccogliere il prodotto finale in una siringa 10 mL collegata all'apertura superiore del dispositivo. Nota: Il kit di elaborazione per il tessuto grasso migliora la tecnica di lipofilling standard: infatti, il sistema consiste di un sistema cilindrico chiuso, full immersion, a bassa pressione, al fine di ottenere un prodotto fluido e uniforme che contiene un gran numero di periciti. Questa procedura consente il trattamento delle cellule adipose esclusivamente tramite forze meccaniche mite e preservando l'integrità della nicchia vascolare stromal. Il processo è il meno traumatico possibile e rende il prodotto finale disponibile in breve tempo (15-20 min.), senza trattamenti enzimatici o espansione. Vasculostromal intatto nicchie aiutano il processo di guarigione.</li>
		<li>Una volta che è state raccolte il CSTA, applicare un bendaggio di compressione sull'addome.</li>
	</ol></li>
	<li>Procedura chirurgica e l'iniezione del tessuto adiposo
	<ol>
<li>Posizionare il paziente in posizione supina in anestesia spinale con un laccio emostatico, ad una pressione di 250 mmHg, a livello della coscia per diminuire l'emorragia e consentire una migliore visualizzazione artroscopica1.</li>
		<li>Marcare i reperi anatomici sulla pelle con una penna dermografica. Luoghi d'interesse sono essenziali per evitare lesioni iatrogeniche. Contrassegnare i seguenti (Figura 3): entrambi i malleoli (laterale (LM) e mediale (MM)) la linea comune anteriore (JL), identificata con dorsi - e flessione plantare dell'articolazione della caviglia il tibiale anteriore (TAT) e il tendine di Achille della vena grande safena, che corre davanti al malleolo mediale il nervo peroneo superficiale (SPN)</li>
		<li>Innanzitutto, eseguire il portale anteromedial appena mediale per la tibiale anteriore, in concomitanza con un debole. Questo portale rappresenta il portale di approccio. Nella maggior parte dei casi, una depressione con la caviglia in dorsiflessione è visibile e palpabile.<ol>
<li>Tagliare solo la pelle con una lama e poi forare la capsula di scollamento. Fare attenzione a evitare il nervo safeno e della vena grande safena. La vena safena è situato a 9 mm lateralmente al portale, mentre il nervo è circa 7,4 mm laterale al portale. Questo costituisce uno dei due portali di visualizzazione primaria.</li>
		</ol>
</li>
		<li>Controllare la linea comune, posto sul portale anterolateral, mediale al malleolo laterale e laterale al tendine di digitorum dell'estensore. Nota: Quando si esegue il portale anterolateral, prevenire eventuali lesioni al nervo cutaneo dorsale intermedio (il ramo laterale del nervo peroneo superficiale); per questo motivo, dopo il taglio, la pelle deve essere seguita da dissezione smussa.</li>
		<li>Ispezionare la cartilagine articolare per valutare la dimensione e la posizione. Valutare la condizione e la qualità della cartilagine con il palpator. Lesioni osteocondrali dell'astragalo sono situate solitamente posteromedially o anterolaterally.<ol>
<li>Eseguire il trattamento artroscopico usando grandangolare Artroscopio 2,7-mm con un angolo di 30°, anche se alcuni chirurghi verranno utilizzato un Artroscopio 4 mm più grande e tenere l'apparecchio nel vano anteriore del giunto. Tecniche di distrazione congiunta non invasiva e iper flessione plantare può essere utilizzati per accedere alla maggior parte della cupola talar.</li>
			<li>In caso di una lesione posteriore, in modo percutaneo posizionare un divaricatore Hintermann per distrarre il giunto e consentire l'esposizione della lesione. Il divaricatore Hintermann ha una leva di apertura braccio applicato su due fili di Kirschner precedentemente posizionati nella tibia e dell'astragalo osso medialmente o lateralmente, secondo il lato della lesione. Nel caso di una lesione laterale, fare attenzione a inserire il filo di K prossimale nell'osso tibia, evitando il perone, per ottenere la migliore distrazione del giunto.</li>
		</ol>
</li>
		<li>Preparare la lesione con un curette, rimuovendo la cartilagine danneggiata e instabile, lo strato calcificato e necrotica e sclerotica dell'osso al fine di creare una forma regolare contenuti lesione con bordi dalle spalle. Per questo passaggio, è necessario utilizzare una curette standard artroscopica (Figura 4).</li>
		<li>Stimolare le cellule staminali di midollo osseo da osso subcondrale eseguire microperforazioni sonoramente dall'esterno all'interno della lesione.<ol>
<li>Eseguire perforazioni con intervalli di circa 3 mm tra di loro. Indurre microfratture usando un plettro Chondral sull'osso subcondrale sano sotto il difetto (Figura 5). Nota: Un'emorragia provoca la formazione di un coagulo di fibrina. Coagulazione prodotti, attraverso l'attivazione della cascata infiammatoria, rilascio di mediatori vasoattivi, fattori di crescita e citochine. Questi fattori hanno il potere di stimolare l'invasione vascolare e migrazione di cellule staminali mesenchimali nella porzione chondral della lesione. Queste cellule pluripotenti sono stimolate a differenziarsi in osteoblasti, condrociti e fibroblasti e svolgono un ruolo importante nella stimolazione della riparazione della lesione. Fattori di crescita paracrini nell'ambiente articolare promuovere la formazione della matrice extracellulare e la produzione di fibrocartilagine. Allo stesso tempo, le cellule della parte ossea della lesione producono un tessuto osseo immaturo che viene progressivamente sostituito da un osso maturo.</li>
		</ol>
</li>
		<li>Rimuovere l'acqua intra-articular utilizzando il rasoio in aspirazione e il liquido restante utilizzando una spugna di cotone fino a quando il giunto è completamente asciutto.</li>
	</ol></li>
	<li>Iniezione di cellule staminali di derivazione adiposa
	<ol>
<li>Iniettare 5-7 mL di cellule staminali adiposo-derivate, precedentemente preparata al punto 4.1.3, nell'articolazione della caviglia utilizzando uno dei due portali (anteromedial o anterolateral).</li>
		<li>Rilasciare il laccio emostatico.</li>
	</ol></li>
</ol>5. postoperatoria
<ol><li>Evitare movimenti della caviglia per 15 giorni, con l'arto operato in scarico completo.</li>
	<li>15 giorni dopo la chirurgia, consentono movimenti attivi e passivi della caviglia, fino al completo recupero e gamma di movimento è raggiunto.</li>
	<li>Consentire la carica 30 giorni dopo la chirurgia.</li>
</ol>6. clinico e radiografico follow-up
<ol><li>Valutare i pazienti clinicamente e radiologicamente a sei e dodici mesi e, successivamente, ogni anno. Il protocollo di follow-up è costituito da caviglia AOFAS e punteggi di hind-piede, Punteggio del dolore VAS e SF-12 a ogni endpoint15,16,17. L'esame radiografico comprende un MRI e CT-Esplora della caviglia operata.</li>
</ol>

<ol>
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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, rapporti personali da Integra e Geistlich e borse di studio e le spese personali da Zimmer, di fuori del lavoro presentato.

Negli ultimi anni, studi clinici e preclinici hanno concentrato la loro attenzione sull'effetto di CSTA per trattare diverse patologie muscolo-scheletriche. Lo scopo di questo articolo è di descrivere il protocollo per il trattamento delle lesioni osteocondrali dell'astragalo utilizzando microfratturate e tessuto adiposo purificato in associazione con microperforazioni artroscopica. Il protocollo prevede diversi passaggi critici con elevati rischi di complicazioni. Durante la raccolta del grasso, complicazioni possono e...

L'artroscopia è il gold standard per il trattamento delle lesioni osteocondrali dell'astragalo (OLTs) con l'obiettivo di alleviare il dolore, ripristino della funzionalità e migliorare la qualità della vita, soprattutto in pazienti giovani e attivi.
Attualmente, le tecniche artroscopiche possono essere classificate in tre modi. La tecnica riparativa stimola le cellule derivate dal midollo osseo attraverso un microperforazioni a livello della lesione e di sbrigliamento. La tecnica ricostruttiva sostituisce la lesione usando un innesto autologo o eterologo ostechondral. La tecnica rigenerativa sfrutta la capacità delle cellule multipotenti per differenziare e replicare per ricostruire il tessuto danneggiato1,2,3,4,5,6 .
Negli ultimi anni, tecniche rigenerative sono stati oggetto di numerosi studi in vitro e in vivo per il trattamento di OLTs e particolarmente mesenchymal cellule staminali derivate da tessuto adiposo (CSTA)7,8 , 9. queste cellule staminali mesenchimali esibiscono caratteristiche morfologiche e funzionali simili alle altre cellule multipotenti, isolati da altri tessuti; Essi hanno anche la capacità di differenziarsi in diversi e linee cellulari differenti sia in vitro che in vivo10,11,12,13. Il focus sulla ricerca per quanto riguarda queste cellule è principalmente a causa della loro localizzazione, infatti essi rappresentano dal 10% al 30% del peso corporeo normale con una concentrazione di 5000 cellule per grammo di tessuto13,14. D'altra parte, un fattore che limita l'utilizzo di queste cellule è relativo alla loro manipolazione durante le procedure di laboratorio. Il lipoaspirato contenente aggregati di adipociti, fibre di collagene e componenti vascolari normali enzimaticamente viene elaborato con collagene di tipo I A e sottoposti ad emolisi prima cultura. L'obiettivo qui è quello di descrivere il protocollo per il trattamento delle lesioni osteocondrali dell'astragalo utilizzando microfratturate e tessuto adiposo purificato.

<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

Negli ultimi anni, tecniche rigenerative sono stati sempre più studiati e usati per trattare le lesioni osteochondral del talus. In particolare, diversi studi hanno focalizzato la loro attenzione su cellule staminali mesenchimali derivate dal tessuto adiposo. Le cellule staminali adiposo-derivate (CSTA) esibiscono caratteristiche morfologiche e proprietà simili alle altre cellule mesenchimali e sono in grado di differenziarsi in diverse linee cellulari. Inoltre, queste cellule sono anche ampiamente disponibili nel tessuto sottocutaneo, che rappresentano il 10-30% del peso corporeo normale, con una concentrazione di 5000 cellule per grammo di tessuto.
La tecnica presentata, il primo passo prevede raccolta CSTA dall'addome e un processo di microfrattura e purificazione; Successivamente, la procedura chirurgica viene eseguita interamente arthroscopically, con meno dissezione dei tessuti molli, migliore visualizzazione congiunta e un recupero più veloce rispetto a procedure standard aperte. L'artroscopia è caratterizzato da una prima fase in cui la lesione è identificata, isolata e preparata con microperforazioni; la seconda fase, eseguita a secco, prevede l'iniezione di tessuto adiposo a livello della lesione.
Tra gennaio 2016 e settembre 2016, quattro pazienti hanno subito il trattamento artroscopico di lesione osteochondral del talus con microfratturate e tessuto adiposo purificato. Tutti pazienti hanno segnalato il miglioramento clinico sei mesi dopo la chirurgia senza le complicazioni segnalate. I punteggi funzionali all'ultimo follow-up sono incoraggianti e confermano che la tecnica fornisce sollievo dal dolore affidabile e miglioramenti in pazienti con lesione osteochondral del talus.

Tra gennaio 2016 e settembre 2016, quattro pazienti hanno subito il trattamento artroscopico di lesione osteochondral del talus con microfratturate e tessuto adiposo purificato. Tutti i pazienti hanno riferito il miglioramento clinico sei mesi dopo la chirurgia. Risultati clinici preliminari sono riportati in tabella 1. Nessuna complicazione sono stata segnalata.
Negli ultimi anni, ha aumentato l'uso di CSTA per...

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

Lo scopo di questo studio è di segnalare un protocollo per il trattamento artroscopico di lesioni osteocondrali dell'astragalo utilizzando microfratturate e purificato le cellule staminali adiposo-derivate.

Microfratturate autologo e nel tessuto adiposo purificato per l'amministrazione artroscopica di lesioni osteocondrali dell'astragalo

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 Views.  IRCCS Istituto Ortopedico Galeazzi - C.A.S.C.O. Piede e Caviglia. Lo scopo di questo studio è di segnalare un protocollo per il trattamento artroscopico di lesioni osteocondrali dell'astragalo utilizzando microfratturate e purificato le cellule staminali adiposo-derivate.

Video: 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.

Respirazione controllata stimolazione elettrica (BreEStim) per la gestione del dolore neuropatico e spasticità

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

Ricerca

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.

Generazione e quantificazione 3-Dimensional di arteriosa lesioni nei topi mediante tomografia ottica di proiezione

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 Luminescenza Imaging di interscapolare Brown Tessuto 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.

Incapsulamento termogenico preadipocytes al trapianto nel tessuto adiposo Depositi tessuti

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.

Una tecnica per la biopsia del tessuto adiposo sottocutaneo addominale tramite un metodo Non-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.

Valutazione della funzione microvascolare di tessuto adiposo umano mediante videomicroscopia

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.

Protocollo clinico di produrre tessuto adiposo-derivato Stromal frazione vascolare per potenziale rigenerazione della cartilagine

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.

Modello di trapianto di Murine (ISWAT) del tessuto adiposo sottocutaneo inguinale (ISWAT)

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.

Trapianto di isole ricoperte di grasso utilizzando tessuto adiposo bianco epididimo

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