Nessun fonti di finanziamento.

1. Preparazione della placenta al momento del parto <ol><li> Etichettare il cordone ombelicale dei gemelli con una sola (per i primogeniti) o due (per il secondo nato) morsetti. </li><li> Ispezionare la superficie materna e fetale della placenta per completezza o interruzioni. </li><li> Registrare i seguenti dati: tipo di inserzione del cordone (centrale, eccentrica, marginale o velamentous), numero di vasi sanguigni del cordone ombelicale (in genere una vena e due arterie, a volte solo una delle arterie) e la differenza di colore tra le due parti della placenta. Una sezione della membrana divisoria possono essere inviati alla patologia per confermare il tipo di corialità. </li><li> La placenta può essere inserito in una ciotola di plastica e refrigerati fino alla prova finale (al meglio in una settimana) e colorante iniezione di colore. </li><li> La placenta non devono essere congelati o fissi (non usare formalina). </li></ol> 2. Cateterismo dei vasi ombelicali <ol><li> Lavare la placenta con acqua tiepida o soluzione salina. </li><li> Tagliare le membrane periferiche, rimuovere l&#39;inter-twin membrana che divide e staccare la amnions (per una migliore visualizzazione delle anastomosi vascolari e una migliore qualità delle immagini placentare). </li><li> Transetto ogni cordone ombelicale a circa 5 cm di distanza dalla inserzione del cordone. </li><li> Premere delicatamente i coaguli di sangue dai vasi ombelicali e dei vasi placentari. </li><li> La vena ombelicale è di solito facile identificare grazie al suo diametro maggiore, rispetto al diametro più piccolo dei due arterie ombelicali. </li><li> Cannulate la vena ombelicale con un catetere di dimensioni appropriate. Evitare passaggi false. </li><li> Cannulate un&#39;arteria ombelicale con un catetere più piccolo. Utilizzare una pinzetta per allargare il lume dell&#39;arteria ombelicale. Evitare passaggi false. Solo uno dei due arterie ombelicali deve essere cateterizzati poiché un anastomosi (di Hyrtl) collega le 2 arterie nei pressi della inserzione del cordone. </li><li> Ripetere i due passaggi per il cordone ombelicale altri. </li><li> Il posizionamento del catetere può essere facilitata da tornare indietro massaggio dolce e dei vasi ombelicali. Qualsiasi tipo di catetere può essere utilizzato per questa procedura. Abbiamo scelto di usare (e riciclare), i cateteri utilizzati presso il nostro reparto di neonatologia per cateterizzazione ombelicale nei neonati. </li><li> Legare un pezzo di nastro adesivo intorno ad entrambi i cavi per evitare di nuovo il flusso del colorante durante l&#39;iniezione colorante. </li></ol> 3. Iniezione con colorante <ol><li> Collegare una siringa da 20 ml piena di colorante ad ogni catetere. </li><li> Ogni colorante viscoso di colore può essere utilizzato per visualizzare le placentare angio-architettura. Usa colori a contrasto per consentire una buona visualizzazione delle anastomosi (colori scuri per le arterie, colori vivaci per le vene). </li><li> Iniettare delicatamente (a bassa pressione) il colorante nella vena, mentre un assistente spinge delicatamente il colorante per permettere al colorante di riempire tutti i vasi placentari, anche i più piccoli. </li><li> Prestare particolare attenzione ai piccoli vasi vicino all&#39;equatore vascolare (l&#39;equatore vascolare è il luogo dove l&#39;anastomosi da entrambi i gemelli collegare tra di loro). </li><li> Ripetere i passaggi precedenti per iniettare colorante nell&#39;arteria. Di nota: le arterie può essere più difficile da iniettare e richiedono più pazienza. </li><li> Ripetere i due passaggi per il cordone ombelicale altri. </li></ol> 4. Valutazione e documentazione della placenta dopo l&#39;iniezione di colore colorante <ol><li> Esaminare attentamente l&#39;equatore vascolare e registrare il numero e il tipo di anastomosi. </li><li> Posizionare un metro sulla placenta per misurare i diametri e condivide placentare sull&#39;immagine digitale. </li><li> Utilizzare una fotocamera ad alta risoluzione digitale e scattare foto della placenta iniettato. Assicurarsi che le immagini sono prese perpendicolarmente alla placenta. </li></ol> 5. Rappresentante dei risultati: La placenta angio-architettura nei gemelli monocoriali varia a seconda del tipo di monocoriali gravidanza gemellare. Gli studi hanno dimostrato che l&#39;iniezione anastomosi AA, AV e VV sono presenti rispettivamente nel 80%, 95% e 20% di gravidanze senza complicazioni gemelli monocoriali 1 (Figura 1). Anastomosi AA sono considerati per la protezione contro lo sviluppo di TTTS e TAPS 6. Gli studi hanno dimostrato che iniezioni di anastomosi AA si verificano solo nel 20% dei TTTS placente e il 10% dei rubinetti placente 1,2,7 (Figura 2). In placente TTTS trattati con chirurgia laser fetoscopic, le cicatrici causate da coagulazione laser delle anastomosi vascolari possono essere visti in superficie placentare (Figura 3 e 4). Placente RUBINETTI sono caratterizzate dalla presenza di anastomosi minuscoli solo pochi AV 2 (Figura 5). La quota placentare del destinatario TAPS è spesso pletorica, mentre la quota di placentare del donatore è pallido (Figura 6). In gravidanze gemellari monocoriali discordanza con peso alla nascita, la crescita limitata fetoha spesso un inserimento velamentous cavo e una quota molto più piccola della placenta (Figura 7). Anastomosi AA sono spesso presenti nelle placente di gemelli monocoriali discordanza con peso alla nascita 8. Monocoriali-monoamniotici placente hanno una caratteristica angio-architettura, con una breve distanza tra i due inserimenti cavo (Figura 8). L&#39;incidenza di anastomosi AA in monoamniotici-placente monocoriali è praticamente il 100% e impedisce lo sviluppo di TTTS in monoamniotici gemelli 9. I dati in questo articolo mostrano i risultati caratteristici di placente monocoriali iniettato nel nostro centro con tintura a colori. Noi abitualmente utilizzare colori più scuri (blu o verde) per arterie e colori più chiari (giallo, rosa o arancione) per le vene. <img alt="Figura 1" src="/files/ftp_upload/3208/3208fig1.jpg" /> Figura 1. Placenta monocoriali da un normale, non complicata gravidanza gemelli monocoriali che mostra diverse anastomosi AV dalle arterie alle vene verde rosa (stelle bianche), diverse anastomosi VA dalle arterie alle vene blu, giallo (stelle verdi) e 1 grande AA anastomosi (identificato dal miscelazione del colorante blu e verde, blu stella). <img alt="Figura 2" src="/files/ftp_upload/3208/3208fig2.jpg" /> Figura 2. Placenta monocoriali in una gravidanza TTTS trattati con amnioreduction seriale che mostra la presenza di un solo AV (stelle bianche) e VA (stelle verdi) anastomosi senza anastomosi AA. <img alt="Figura 3" src="/files/ftp_upload/3208/3208fig3.jpg" /> Figura 3. TTTS placenta dopo la coagulazione laser fetoscopic delle anastomosi vascolari con la tecnica laser selettiva in cui una anastomosi piccolo residuo è stato inavvertitamente lasciato brevetto (White Star). Con la tecnica laser selettiva, le anastomosi vascolari vengono prima identificati e poi coagulato uno per uno. <img alt="Figura 4" src="/files/ftp_upload/3208/3208fig4.jpg" /> Figura 4. TTTS placenta dopo la coagulazione laser fetoscopic con la tecnica Salomone in cui, dopo l&#39;identificazione e la coagulazione di anastomosi ogni individuo, è coagulato l&#39;equatore completa vascolare da un margine placentare all&#39;altro. <img alt="Figura 5" src="/files/ftp_upload/3208/3208fig5.jpg" /> Figura 5. RUBINETTI placenta in cui solo pochi, minuscoli anastomosi VA (stelle bianche) sono visibili lungo l&#39;equatore vascolare. <img alt="Figura 6" src="/files/ftp_upload/3208/3208fig6.jpg" /> Figura 6. Il lato materno della placenta TAPS (Figura 5) mostra la differenza caratteristico colore tra le due parti della placenta. <img alt="Figura 7" src="/files/ftp_upload/3208/3208fig7.jpg" /> Figura 7. Placenta monocoriali di una gravidanza gemellare con selettivo restrizione della crescita intrauterina di un gemello. Il feto ha una crescita limitata inserimento velamentous cavo e una quota molto più piccola della placenta. La stella bianca indica una anastomosi AA. <img alt="Figura 8" src="/files/ftp_upload/3208/3208fig8.jpg" /> Figura 8 monoamniotici placenta:. Noti i vari AV (verde stella), VA (stella blu) e l&#39;AA 2 anastomosi (bianco stelle), e la breve distanza tra i due inserimenti cavo.

<ol>
	<li>Lopriore, E., Middeldorp, J. M., Sueters, M., Vandenbussche, F. P., Walther, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Twin-to-twin+transfusion+syndrome:+from+placental+anastomoses+to+long-term+neurodevelopmental+outcome.">Twin-to-twin transfusion syndrome: from placental anastomoses to long-term neurodevelopmental outcome.</a> Curr. Pediatr. Rev. 1, 191-203 (2005).</li><li>Slaghekke, F., Kist, W. J., Oepkes, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Twin+anemia-polycythemia+sequence:+diagnostic+criteria,+classification,+perinatal+management+and+outcome.">Twin anemia-polycythemia sequence: diagnostic criteria, classification, perinatal management and outcome.</a> Fetal Diagn. Ther. 27, 181-190 (2010).</li><li>Lopriore, E., Middeldorp, J. M., Oepkes, D., Klumper, F. J., Walther, F. J., Vandenbussche, F. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Residual+anastomoses+after+fetoscopic+laser+surgery+in+twin-to-twin+transfusion+syndrome:+frequency,+associated+risks+and+outcome.">Residual anastomoses after fetoscopic laser surgery in twin-to-twin transfusion syndrome: frequency, associated risks and outcome.</a> Placenta. 28, 204-208 (2007).</li><li>Lopriore, E., Slaghekke, F., Middeldorp, J. M., Klumper, F. J., Oepkes, D., Vandenbussche, F. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Residual+anastomoses+in+twin-to-twin+transfusion+syndrome+treated+with+selective+fetoscopic+laser+surgery:+localization,+size,+and+consequences.">Residual anastomoses in twin-to-twin transfusion syndrome treated with selective fetoscopic laser surgery: localization, size, and consequences.</a> Am. J. Obstet. Gynecol. , (2009).</li><li>Lewi, L., Jani, J., Cannie, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intertwin+anastomoses+in+monochorionic+placentas+after+fetoscopic+laser+coagulation+for+twin-to-twin+transfusion+syndrome:+is+there+more+than+meets+the+eye.">Intertwin anastomoses in monochorionic placentas after fetoscopic laser coagulation for twin-to-twin transfusion syndrome: is there more than meets the eye.</a> Am. J. Obstet. Gynecol. 194, 790-795 (2006).</li><li>Denbow, M. L., Cox, P., Taylor, M., Hammal, D. M., Fisk, N. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placental+angioarchitecture+in+monochorionic+twin+pregnancies:+relationship+to+fetal+growth,+fetofetal+transfusion+syndrome,+and+pregnancy+outcome.">Placental angioarchitecture in monochorionic twin pregnancies: relationship to fetal growth, fetofetal transfusion syndrome, and pregnancy outcome.</a> Am. J. Obstet. Gynecol. 182, 417-426 (2000).</li><li>van Meir, H., Slaghekke, F., Lopriore, E., van Wijngaarden, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arterio-arterial+anastomoses+do+not+prevent+the+development+of+twin+anemia-polycythemia+sequence.">Arterio-arterial anastomoses do not prevent the development of twin anemia-polycythemia sequence.</a> Placenta. 31, 163-165 (2010).</li><li>De Paepe, M. E., Shapiro, S., Young, L., Luks, F. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placental+characteristics+of+selective+birth+weight+discordance+in+diamniotic-monochorionic+twin+gestations.">Placental characteristics of selective birth weight discordance in diamniotic-monochorionic twin gestations.</a> Placenta. 31, 380-386 (2010).</li><li>Umur, A., van Gemert, M. J., Nikkels, P. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monoamniotic-versus+diamniotic-monochorionic+twin+placentas:+anastomoses+and+twin-twin+transfusion+syndrome.">Monoamniotic-versus diamniotic-monochorionic twin placentas: anastomoses and twin-twin transfusion syndrome.</a> Am. J. Obstet. Gynecol. 189, 1325-1329 (2003).</li><li>Senat, M. V., Deprest, J., Boulvain, M., Paupe, A., Winer, N., Ville, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endoscopic+laser+surgery+versus+serial+amnioreduction+for+severe+twin-to-twin+transfusion+syndrome.">Endoscopic laser surgery versus serial amnioreduction for severe twin-to-twin transfusion syndrome.</a> N. Engl. J. Med. 351, 136-144 (2004).</li><li>Lopriore, E., Sueters, M., Middeldorp, J. M., Vandenbussche, F. P., Walther, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Haemoglobin+differences+at+birth+in+monochorionic+twins+without+chronic+twin-to-twin+transfusion+syndrome.">Haemoglobin differences at birth in monochorionic twins without chronic twin-to-twin transfusion syndrome.</a> Prenat. Diagn. 25, 844-850 (2005).</li><li>Papathanasiou, D., Witlox, R., Oepkes, D., Walther, F. J., Bloemenkamp, K. W., Lopriore, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monochorionic+twins+with+ruptured+vasa+previa:+double+trouble!.">Monochorionic twins with ruptured vasa previa: double trouble!.</a> Fetal Diagn. Ther. 28, 48-50 (2010).</li><li>Weingertner, A. S., Kohler, A., Kohler, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+and+placental+characteristics+in+four+new+cases+of+twin+anemia-polycythemia+sequence.">Clinical and placental characteristics in four new cases of twin anemia-polycythemia sequence.</a> Ultrasound Obstet. Gynecol. 35, 490-494 (2010).</li><li>Lewi, L., Cannie, M., Blickstein, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placental+sharing,+birthweight+discordance,+and+vascular+anastomoses+in+monochorionic+diamniotic+twin+placentas.">Placental sharing, birthweight discordance, and vascular anastomoses in monochorionic diamniotic twin placentas.</a> Am. J. Obstet. Gynecol. 197, 587-588 (2007).</li><li>Lewi, L., Gucciardo, L., Huber, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+outcome+and+placental+characteristics+of+monochorionic+diamniotic+twin+pairs+with+early-+and+late-onset+discordant+growth.">Clinical outcome and placental characteristics of monochorionic diamniotic twin pairs with early- and late-onset discordant growth.</a> Am. J. Obstet. Gynecol. , (2008).</li><li>De Paepe, M. E., Friedman, R. M., Poch, M., Hansen, K., Carr, S. R., Luks, F. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placental+findings+after+laser+ablation+of+communicating+vessels+in+twin-to-twin+transfusion+syndrome.">Placental findings after laser ablation of communicating vessels in twin-to-twin transfusion syndrome.</a> Pediatr. Dev. Pathol. 7, 159-165 (2004).</li><li>De Paepe, M. E., DeKoninck, P., Friedman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascular+distribution+patterns+in+monochorionic+twin+placentas.">Vascular distribution patterns in monochorionic twin placentas.</a> Placenta. 26, 471-475 (2005).</li><li>Nikkels, P. G., Hack, K. E., vanGemert, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathology+of+twin+placentas+with+special+attention+to+monochorionic+twin+placentas.">Pathology of twin placentas with special attention to monochorionic twin placentas.</a> J. Clin. Pathol. 61, 1247-1253 (2008).</li><li>Quintero, R. A., Martinez, J. M., Lopez, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Individual+placental+territories+after+selective+laser+photocoagulation+of+communicating+vessels+in+twin-twin+transfusion+syndrome.">Individual placental territories after selective laser photocoagulation of communicating vessels in twin-twin transfusion syndrome.</a> Am. J. Obstet. Gynecol. 192, 1112-1118 (2005).</li><li>Quintero, R. A., Comas, C., Bornick, P. W., Allen, M. H., Kruger, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selective+versus+non-selective+laser+photocoagulation+of+placental+vessels+in+twin-to-twin+transfusion+syndrome.">Selective versus non-selective laser photocoagulation of placental vessels in twin-to-twin transfusion syndrome.</a> Ultrasound Obstet. Gynecol. 16, 230-236 (2000).</li></ol>

Nessun conflitto di interessi dichiarati.

TTTS e RUBINETTI sono due patologie gravi che possono verificarsi nei gemelli monocoriali. Entrambi i disturbi sono dovuti ad anastomosi AV causando squilibri del flusso sanguigno intertwin. Negli ultimi 2 decenni, la chirurgia laser fetoscopic si sono resi disponibili e ha dimostrato di essere il metodo migliore per il trattamento di TTTS 10. Lo scopo del trattamento laser fetoscopic è quello di interrompere l&#39;inter-doppia circolazione attraverso la coagulazione delle anastomosi vascolari sulla superfic...

<table border="1"><tbody><tr><th> Nome del reagente </th><th> Azienda </th><th> Numero di catalogo </th><th> Commenti </th></tr><tr><td> Cateteri ombelicali (varie dimensioni, lunghezza 40 cm) </td><td> Vygon </td><td> 1.270,08 (8 F) 1.270,08 (5 F) 1.270,04 (4 F) 1.270,03 (3,5 F) 1.270,02 (2,5 F) </td><td> Qualsiasi tipo di catetere può essere utilizzato </td></tr><tr><td> Siringhe da 20 ml </td><td> BD Plastipak </td><td> 300613 </td><td></td></tr><tr><td> A colori dye (colori diversi) </td><td> Reale Talens Schoolverf </td><td> 36716010 (blu) 36715010 (verde) 36712350 (giallo) 36.713.570 (rosa) </td><td> Qualsiasi tintura colore viscoso può essere utilizzato </td></tr></tbody></table>

accurate and simple evaluation of vascular anastomoses in monochorionic placenta using colored dye

La presenza di anastomosi vascolare placentare è una conditio sine qua non per lo sviluppo del gemello-gemello sindrome di trasfusione (TTTS) e doppia sequenza policitemia anemia (TAPS) 1,2. Studi iniezione di placente gemelli hanno dimostrato che tali anastomosi sono quasi sempre presenti nei gemelli monocoriali ed estremamente raro in dicoriale gemelli 1. Tre tipi di anastomosi sono stati documentati: da arteria per arteria, dalla vena alla vena e dalla arteria alla vena. Artero-venose (AV) anastomosi sono unidirezionali e sono indicati come &quot;profonda&quot; anastomosi dal momento che procedere attraverso una condivisione cotiledone placentare, mentre artero-arteriosa (AA) e veno-venoso (VV) anastomosi sono bi-direzionali e di cui come &quot;superficiale&quot; dal momento che si trovano sul piatto coriali. Sia TTTS e RUBINETTI sono causati da squilibrio netto del flusso di sangue tra i due gemelli a causa di anastomosi AV. Sangue da uno dei gemelli (il donatore) viene pompato attraverso un&#39;arteria nel cotiledone condiviso placentare e poi drenato attraverso una vena nella circolazione degli altri fratelli gemelli (il destinatario). A meno che il sangue viene pompato dal destinatario al donatore attraverso dirette in verso opposto anastomosi profonda AV o tramite anastomosi superficiali, uno squilibrio di volumi di sangue si verifica, via via che porta allo sviluppo di TTTS o TAPS. La presenza di una anastomosi AA ha dimostrato di proteggere contro lo sviluppo di TTTS e rubinetti, compensando lo squilibrio circolatorio causato dalla uni-direzionale 1,2 anastomosi AV. Iniezione di placente monocoriali subito dopo la nascita è un mezzo utile per comprendere l&#39;eziologia delle varie (ematologiche) complicazioni nei gemelli monocoriali ed è una prova necessaria per raggiungere la diagnosi di TAPS 2. Inoltre, l&#39;iniezione di placente TTTS trattati con la chirurgia laser fetoscopic consente l&#39;identificazione di possibili anastomosi residuo 3-5. Queste informazioni aggiuntive è di fondamentale importanza per tutti perinatologists coinvolti nella gestione e cura dei gemelli monocoriali con TTTS o TAPS. Diverse tecniche di iniezione placentare sono attualmente utilizzate. Forniamo un semplice protocollo per valutare con precisione la presenza di (residua) anastomosi vascolari mediante iniezione di colore colorante.

Watch this Scientific Journal Video about Accurate and Simple Evaluation of Vascular Anastomoses in Monochorionic Placenta using Colored Dye at JoVE.com

Accurate and Simple Evaluation of Vascular Anastomoses in Monochorionic Placenta using Colored Dye

Gemello-gemello sindrome di trasfusione e doppia sequenza policitemia anemia sono due problemi potenzialmente devastanti in medicina perinatale. Entrambi i disturbi si verificano solo in gemelli monocoriali e il risultato di flusso di sangue placentare sbilanciato attraverso anastomosi vascolari. Forniamo un semplice protocollo per valutare con precisione la presenza di anastomosi vascolari mediante iniezione di colore tintura dei vasi placentari dopo la nascita.

La valutazione accurata e semplice delle anastomosi vascolari in placenta monocoriale con colorante

The presence of placental vascular anastomoses is a conditio sine qua non for the development of twin-to-twin transfusion syndrome (TTTS) and twin anemia ...

accurate-simple-evaluation-vascular-anastomoses-monochorionic

Research

JoVE Journal

Medicine

26.6K Views.  Leiden University Medical Center. Gemello-gemello sindrome di trasfusione e doppia sequenza policitemia anemia sono due problemi potenzialmente devastanti in medicina perinatale. Entrambi i disturbi si verificano solo in gemelli monocoriali e il risultato di flusso di sangue placentare sbilanciato attraverso anastomosi vascolari. Forniamo un semplice protocollo per valutare con precisione la presenza di anastomosi vascolari mediante iniezione di colore tintura dei vasi placentari dopo la nascita.

Video: Accurate and Simple Evaluation of Vascular Anastomoses in Monochorionic Placenta using Colored Dye

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&beta; using a Whole Blood Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues as well as altered cytokine homeostasis. The innate immune system plays a crucial role in recognizing pathogens and other endogenous danger stimuli. One of the major cytokines released by innate immune cells is Interleukin (IL)-1. Therefore, we utilize a whole blood stimulation assay in order to measure the secretion of inflammatory cytokines and specifically of the pro-inflammatory cytokine IL-1&beta; 1, 2, 3.
Patients with genetic dysfunctions of the innate immune system causing autoinflammatory syndromes show an exaggerated release of mature IL-1&beta; upon stimulation with LPS alone. In order to evaluate the innate immune component of patients who present with inflammatory-associated pathologies, we use a specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns (PAMPs), such as the gram-negative bacterial endotoxin, lipopolysaccharide (LPS). These PAMPs are recognized by pathogen recognition receptors (PRRs), which are found on the cells of the innate immune system 4, 5, 6, 7. A primary signal, LPS, in conjunction with a secondary signal, ATP, is necessary for the activation of the inflammasome, a multiprotein complex that processes pro-IL-1&beta; to its mature, bioactive form 4, 5, 6, 8, 9, 10.
The whole blood assay requires minimal sample manipulation to assess cytokine production when compared to other methods that require labor intensive isolation and culturing of specific cell populations. This method differs from other whole blood stimulation assays; rather than diluting samples with a ratio of RPMI media, we perform a white blood cell count directly from diluted whole blood and therefore, stimulate a known number of white blood cells in culture 2. The results of this particular whole blood assay demonstrate a novel technique useful in elucidating patient cohorts presenting with autoinflammatory pathophysiologies.

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&amp;#946; using a Whole Blood Stimulation Assay

We describe a simple immunoassay to measure the production of pro-inflammatory cytokines, such as IL-1 beta production, in patients presenting with autoinflammatory phenotypes. By activating cells in whole blood cultures with pathogen-associated molecular patterns, specifically with lipopolysaccharide, cytokine secretion can be conveniently evaluated in whole blood supernatants.

Misurazione accurata e semplice della citochina pro-infiammatoria IL-1β utilizzando un intero saggio stimolazione del sangue

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues ...

Ricerca

Medicina

Guide Wire Assisted Catheterization and Colored Dye Injection for Vascular Mapping of Monochorionic Twin Placentas

Monochorionic (MC) twin pregnancies are associated with significantly higher morbidity and mortality rates than dichorionic twins. Approximately 50% of MC twin pregnancies develop complications arising from the shared placenta and associated vascular connections1. Severe twin-to-twin syndrome (TTTS) is reported to account for approximately 20% of these complications2,3. Inter-twin vascular connections occur in almost all MC placentas and are related to the prognosis and outcome of these high-risk twin pregnancies. The number, size and type of connections have been implicated in the development of TTTS and other MC twin conditions. Three types of inter-twin vascular connections occur: 1) artery to vein connections (AVs) in which a branch artery carrying deoxygenated blood from one twin courses along the fetal surface of the placenta and dives into a placental cotyledon. Blood flows via a deep intraparenchymal capillary network into a draining vein that emerges at the fetal surface of the placenta and brings oxygenated blood toward the other twin. There is unidirectional flow from the twin supplying the afferent artery toward the twin receiving the efferent vein; 2) artery to artery connections (AAs) in which a branch artery from each twin meets directly on the superficial placental surface resulting in a vessel with pulsatile bidirectional flow, and 3) vein to vein connections (VVs) in which a branch vein from each twin meets directly on the superficial placental surface allowing low pressure bidirectional flow. In utero obstetric sonography with targeted Doppler interrogation has been used to identify the presence of AV and AA connections4. Prenatally detected AAs that have been confirmed by postnatal placental injection studies have been shown to be associated with an improved prognosis for both twins5. Furthermore, fetoscopic laser ablation of inter-twin vascular connections on the fetal surface of the shared placenta is now the preferred treatment for early, severe TTTS. 
Postnatal placental injection studies provide a valuable method to confirm the accuracy of prenatal Doppler ultrasound findings and the efficacy of fetal laser therapy6. Using colored dyes separately hand-injected into the arterial and venous circulations of each twin, the technique highlights and delineates AVs, AAs, and VVs. This definitive demonstration of MC placental vascular anatomy may then be correlated with Doppler ultrasound findings and neonatal outcome to enhance our understanding of the pathophysiology of MC twinning and its sequelae. Here we demonstrate our placental injection technique.

Vascular mapping of monochorionic (MC) twin placentas after birth provides a means for detailed demonstration of vascular connections between the twins&#x2019; circulations. 
Imbalance of these connections is thought to play a pivotal role in the development of complications of MC twinning including twin-to-twin transfusion syndrome.

Guida cateterizzazione filo assistita e colorate iniezione Colorante per la mappatura vascolare di placente monocoriali Gemelle

Monochorionic (MC) twin pregnancies are associated with significantly higher morbidity and mortality rates than dichorionic twins. Approximately 50% of MC ...

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

Un metodo semplice di intubazione mouse polmone

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Technical Aspects of the Mouse Aortocaval Fistula

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta and inferior vena cava (IVC) are exposed. The proximal infrarenal aorta and the distal aorta are dissected for clamp placement and needle puncture, respectively. Special attention is paid to avoid dissection between the aorta and the IVC. After clamping the aorta, a 25 G needle is used to puncture both walls of the aorta into the IVC. The surrounding connective tissue is used for hemostatic compression. Successful creation of the AVF will show pulsatile arterial blood flow in the IVC. Further confirmation of successful AVF can be achieved by post-operative Doppler ultrasound.

The goal is to produce an arteriovenous fistula that is simple and reproducible. This method does not use sutures or glue adhesive. Therefore the samples can be used with the least amount of foreign materials for analysis.

Aspetti tecnici della fistola aorto-cavale mouse

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta ...

Fast and Accurate Exhaled Breath Ammonia Measurement

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that uses a quantum cascade based laser. The monitor is coupled to a sampler that measures mouth pressure and carbon dioxide. The system is temperature controlled and specifically designed to address the reactivity of this compound. The sampler provides immediate feedback to the subject and the technician on the quality of the breath effort. Together with the quick response time of the monitor, this system is capable of accurately measuring exhaled breath ammonia representative of deep lung systemic levels. 
Because the system is easy to use and produces real time results, it has enabled experiments to identify factors that influence measurements. For example, mouth rinse and oral pH reproducibly and significantly affect results and therefore must be controlled. Temperature and mode of breathing are other examples. As our understanding of these factors evolves, error is reduced, and clinical studies become more meaningful. This system is very reliable and individual measurements are inexpensive. 
The sampler is relatively inexpensive and quite portable, but the monitor is neither. This limits options for some clinical studies and provides rational for future innovations.

Ammonia is an important physiologic metabolite relevant to various disease and wellness states. It is also a difficult molecule to measure in breath, which demands particular precautions be taken to obtain accurate results. Not all factors influencing ammonia are known, but progress can be difficult without accounting for these factors.

Veloce e preciso espirata Ammoniaca misura

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that ...

Evaluation of Tumor-infiltrating Leukocyte Subsets in a Subcutaneous Tumor Model

Specialized immune cells that infiltrate the tumor microenvironment regulate the growth and survival of neoplasia.&#160; Malignant cells must elude or subvert anti-tumor immune responses in order to survive and flourish. Tumors take advantage of a number of different mechanisms of immune &#8220;escape,&#8221; including the recruitment of tolerogenic DC, immunosuppressive regulatory T cells (Tregs), and myeloid-derived suppressor cells (MDSC) that inhibit cytotoxic anti-tumor responses. Conversely, anti-tumor effector immune cells can slow the growth and expansion of malignancies: immunostimulatory dendritic cells, natural killer cells which harbor innate anti-tumor immunity, and cytotoxic T cells all can participate in tumor suppression. The balance between pro- and anti-tumor leukocytes ultimately determines the behavior and fate of transformed cells; a multitude of human clinical studies have borne this out. Thus, detailed analysis of leukocyte subsets within the tumor microenvironment has become increasingly important. Here, we describe a method for analyzing infiltrating leukocyte subsets present in the tumor microenvironment in a mouse tumor model. Mouse B16 melanoma tumor cells were inoculated subcutaneously in C57BL/6 mice. At a specified time, tumors and surrounding skin were resected en bloc and processed into single cell suspensions, which were then stained for multi-color flow cytometry. Using a variety of leukocyte subset markers, we were able to compare the relative percentages of infiltrating leukocyte subsets between control and chemerin-expressing tumors. Investigators may use such a tool to study the immune presence in the tumor microenvironment and when combined with traditional caliper size measurements of tumor growth, will potentially allow them to elucidate the impact of changes in immune composition on tumor growth. Such a technique can be applied to any tumor model in which the tumor and its microenvironment can be resected and processed.

This protocol describes a method for the detailed evaluation of leukocyte subsets within the tumor microenvironment in a mouse tumor model. Chemerin-expressing B16 melanoma cells were implanted subcutaneously into syngeneic mice. Cells from the tumor microenvironment were then stained and analyzed by flow cytometry, allowing for detailed leukocyte subset analyses.

Valutazione del tumore infiltrante leucociti sottoinsiemi in un tumore sottocutaneo modello

Specialized immune cells that infiltrate the tumor microenvironment regulate the growth and survival of neoplasia.&#160; Malignant cells must elude or ...

Using the Sleeve Technique in a Mouse Model of Aortic Transplantation - An Instructional Video

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the aorta in mice using the sleeve method was short and easy averaging 20 min, permitting studies of iso/allo grafts. The following article describes the aortic transplantation procedure used in our laboratory. The mice were anesthetized with a mixture of 1.5% volume isoflurane and 100% oxygen through a face mask. At this point, the segment of the aorta between the renal arteries and its bifurcation was separated from the vena cava, freely prepared and clampedat the proximal and distal segments with a single silk suture. Prior to the removal of the aorta, a saline solution containing heparin was injected into the inferior vena cava. Then the aorta was cut between the clamps and a saline heparin solution was used to flush the lumen. The sleeve technique with monofilament sutures was used in order to transplant the abdominal aorta in the orthotopic position.

We present an orthotopic aortic transplantation model using the sleeve technique in mice. It is a very rapid anastomosis method, which can be employed in studies of vascular disease.

Utilizzando la tecnica di manica in un modello murino di trapianto aortico - un Video didattico

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the ...

Model Surgical Training: Skills Acquisition in Fetoscopic Laser Photocoagulation of Monochorionic Diamniotic Twin Placenta Using Realistic Simulators

Fetoscopic laser coagulation of arterio-venous anastomoses (AVA) in a monochorionic placenta is the standard of care for twin-twin transfusion syndrome (TTTS), but is technically challenging and can lead to significant complications. Acquiring and maintaining the necessary surgical skills require consistent practice, a critical caseload, and time. Training on realistic surgical simulators can potentially shorten this steep learning curve and enables several proceduralists to acquire procedure-specific skills simultaneously. Here we describe realistic simulators designed to allow the user familiarity with the equipment and specific steps required in the surgical treatment of TTTS, including fetoscopic handling, approaches to anterior and posterior placenta, recognition of anastomoses, and efficient coagulation of vessels. We describe the skills that are especially important in conducting placental laser coagulation that the surgeon can practice on the model and apply in a clinical case. These models can be adapted easily depending on the availability of materials and require standard fetoscopy equipment. Such training systems are complementary to traditional surgical apprenticeships and can be useful aids for fetal medicine units that provide this clinical service.

Practicing the specific skills required for fetoscopic laser coagulation of monochorionic placental anastomoses on realistic models can aid less experienced surgeons in overcoming the steep learning curve associated with this procedure that is now regarded as the standard of care for twin-twin transfusion syndrome.

Modello di Training chirurgico: Acquisizione di competenze in photocoagulation del Laser Fetoscopic di Monochorionic Diamniotic Twin Placenta utilizzando simulatori realistici

Fetoscopic laser coagulation of arterio-venous anastomoses (AVA) in a monochorionic placenta is the standard of care for twin-twin transfusion syndrome ...

Noninvasive Determination of Vortex Formation Time Using Transesophageal Echocardiography During Cardiac Surgery

Trans-mitral blood flow produces a three-dimensional rotational body of fluid, known as a vortex ring, that enhances the efficiency of left ventricular (LV) filling compared with a continuous linear jet. Vortex ring development is most often quantified with vortex formation time (VFT), a dimensionless parameter based on fluid ejection from a rigid tube. Our group is interested in factors that affect LV filling efficiency during cardiac surgery. In this report, we describe how to use standard two-dimensional (2D) and Doppler transesophageal echocardiography (TEE) to noninvasively derive the variables needed to calculate VFT. We calculate atrial filling fraction (&#946;) from velocity-time integrals of trans-mitral early LV filling and atrial systole blood flow velocity waveforms measured in the mid-esophageal four-chamber TEE view. Stroke volume (SV) is calculated as the product of the diameter of the LV outflow track measured in the mid-esophageal long axis TEE view and the velocity-time integral of blood flow through the outflow track determined in the deep transgastric view using pulse-wave Doppler. Finally, mitral valve diameter (D) is determined as the average of major and minor axis lengths measured in orthogonal mid-esophageal bicommissural and long axis imaging planes, respectively. VFT is then calculated as 4 &#215; (1-&#946;) &#215; SV/(&#960;D3). We have used this technique to analyze VFT in several groups of patients with differing cardiac abnormalities. We discuss our application of this technique and its potential limitations and also review our results to date. Noninvasive measurement of VFT using TEE is straightforward in anesthetized patients undergoing cardiac surgery. The technique may allow cardiac anesthesiologists and surgeons to assess the impact of pathological conditions and surgical interventions on LV filling efficiency in real time.

We describe a protocol to measure vortex formation time, an index of left ventricular filling efficiency, using standard transesophageal echocardiography techniques in patients undergoing cardiac surgery. We apply this technique to analyze vortex formation time in several groups of patients with differing cardiac pathologies.

Determinazione non invasiva del vortice formazione tempo usando l'ecocardiografia Transesophageal durante l'ambulatorio cardiaco

Trans-mitral blood flow produces a three-dimensional rotational body of fluid, known as a vortex ring, that enhances the efficiency of left ventricular ...

Echocardiographic Evaluation of Atrial Communications before Transcatheter Closure

Transthoracic (TTE) and transesophageal echocardiography (TEE) is the standard imaging method for atrial septal defect (ASD) and patent foramen ovale (PFO) detection, for patient selection for transcatheter ASD/PFO closure,&#160;for intraoperative guidance and for long-term follow-up. The size, shape, location and the number of the atrial communications schould be determined. The accuracy of PFO detection can be improved by using agitated saline together with maneuvers to transiently increase the right atrial (RA) pressure. The appearance of microbubbles in the left atrium (LA) within 3 cardiac cycles after opacification of the RA is considered positive for the presence of an intracardiac shunt. Three dimensional TEE identifies further septal fenestrations and describes the dynamic morphology of ASD/PFO and atrial septal aneurysm. Follow-up evaluations with TTE is recommended at 1, 6, and 12 months after the procedure, with a subsequent evaluation every year. Previous studies showed an increased incidence of atrial arrhythmias early after device closure. Speckle tracking analysis may help to understand functional left atrial remodeling following percutaneous closure and its impact on atrial arrhythmias.

Transthoracic (TTE) and transesophageal (TEE) echocardiography represent the basic imaging tools for interatrial septum examination. Three dimensional (3D) TEE provides incremental information in the assessment of the interatrial septum. Further advanced echocardiography techniques using speckle tracking echocardiography are applied for sensitive volumetric and functional assessment of the heart chambers.&#160;

Valutazione ecocardiografica delle comunicazioni atriali prima della chiusura transcatetere

Transthoracic (TTE) and transesophageal echocardiography (TEE) is the standard imaging method for atrial septal defect (ASD) and patent foramen ovale ...