無資金源。

1。出産時の胎盤の準備<ol><li>もの（長子の場合）または2（セカンド生まれ用）クランプを持つ双子のへその緒にラベルを付けます。 </li><li>完全性や混乱のための胎盤の母体と胎児の表面を点検。 </li><li>コー​​ドの挿入（、中央の偏心、限界や卵膜）、臍帯の血管の数（通常は1静脈と時には2動脈、唯一の動脈）との両方胎盤株式の間に色差のタイプ：次のデータを記録する。割っ膜のセクションは、chorionicityの種類を確認するために病理に送信することができます。 </li><li>胎盤は、プラスチック製のボウルに入れて最終試験（最高の一週間以内）と色の染料を注入するまで冷蔵することができます。 </li><li>胎盤は、（ホルマリンは使用しないでください）​​凍結または固定されてはならない。 </li></ol> 2。臍血管のカテーテル<ol><li>暖かい水または生理食塩水で胎盤を洗ってください。 </li><li>末梢膜をトリミング、amnions（血管吻合と胎盤の写真のより良い品質の良い視覚化のための）オフ間ツイン割っ膜と皮を取り除く。 </li><li>コー​​ドの挿入から約5 cmの距離でそれぞれのへその緒を横に切断する。 </li><li>ゆっくりと臍帯血管と胎盤の血管から血の塊を絞り出す。 </li><li>臍静脈は2つの臍動脈のより小さい直径に比べて、その大きな直径のために識別するために、通常は簡単です。 </li><li>適切なサイズのカテーテルと臍静脈のカニューレを挿入する。偽の通路を避けてください。 </li><li>小さなカテーテルを使用すると、ワン臍動脈をカニューレを挿入する。臍動脈の内腔を広げるためにピンセットを使用してください。偽の通路を避けてください。わずか2臍動脈の一つが吻合ので、カテーテルを挿入する必要がある（ヒルトルの）コードの挿入近くに2動脈を接続します。 </li><li>他の臍帯のための手順の両方を繰り返します。 </li><li>カテーテルの配置は、臍帯血管の穏やか前後にマッサージによって促進することができます。カテーテルの任意のタイプは、この手順のために使用することができます。我々は（とリサイクル）新生児の臍カテーテル挿入のための私達の新生児病棟で使用されるカテーテルを使用することを選択。 </li><li>色素注入中に着色された染料の流れをバック避けるために、両方のコードの周りにテープの一部を接続します。 </li></ol> 3。色の染料と注入<ol><li>それぞれのカテーテルへの着色された染料を充填した20 mlの注射器を接続してください。 </li><li>任意の粘性色染料は血管アーキテクチャ胎盤を視覚化するために使用することができます。吻合（動脈のための暗い色、静脈のための明るい色）の良好な可視化を可能にするために対照的な色を使用してください。 </li><li>アシスタントが優しくも、すべて胎盤血管、最小のものを埋めるために色の染料を可能にするために染料を押している間は静かに（低気圧による）静脈の色の色素を注入する。 </li><li>血管の赤道近くにある小さな血管（血管赤道のいずれかツインの吻合が相互に接続する場所です）に特に注意してください。 </li><li>動脈に着色された染料を注入する前の手順を繰り返します。注目すべきは：動脈はより多くの忍耐を注入し、必要とするより難しいかもしれません。 </li><li>他の臍帯のための手順の両方を繰り返します。 </li></ol> 4。着色された染料の注入後の胎盤の評価および文書<ol><li>慎重に血管赤道を調査し、吻合の数と種類を記録。 </li><li>デジタル画像上で直径と胎盤株式を測定するため胎盤に巻き尺を置きます。 </li><li>高解像度のデジタルカメラを使用し、注入された胎盤の写真を撮る。写真は胎盤に垂直に取られていることを確認します。 </li></ol> 5。代表的な結果： 単一絨毛膜性の双子の血管アーキテクチャ胎盤は、単一絨毛膜性の双胎妊娠の種類によって異なります。注入の研究では、AA、AVおよびVV吻合がそれぞれ80％、95％、合併症のない単一絨毛膜性の双子妊娠1（図1）の20％に存在することを明らかにした。 AAの吻合は、TTTSとTAPS 6の開発から保護するために考えられている。注射の研究では、AAの吻合がTTTS胎盤のわずか20％とTAPS胎盤1,2,7（図2）の10％で発生することが示されている。 fetoscopicレーザー手術で治療TTTSの胎盤では、血管吻合のレーザーの凝固によって引き起こされる傷跡は胎盤表面（図3および4）で見ることができます。 TAPSの胎盤は、わずか数極小のAVの吻合2（図5）の存在によって特徴付けられる。ドナーの胎盤シェアは（図6）を淡であるのに対し、TAPSの受信者の胎盤シェアは、しばしば多血です。出生時体重の不一致による単一絨毛膜性の双子妊娠、成長制限された胎児の多くの場合、被膜のコードの挿入とはるかに小さい胎盤シェア（図7）があります。 AAの吻合は、出生時体重の不一致8の双生児から単一絨毛膜性の胎盤でしばしば存在する。単一絨毛膜性の- monoamniotic胎盤は、両方のコードの挿入（図8）との間の短い距離で特徴的な血管のアーキテクチャを持っている。 monoamniotic -単一絨毛膜性の胎盤におけるAA吻合の発生率はほぼ100％であるとmonoamniotic双子9 TTTSの発症を予防する。この記事の図では、色の染料で私達の中心に注入された単一絨毛膜性の胎盤の特徴的所見を示す。我々は日常的に動脈と静脈のための明るい色（黄色、ピンクまたはオレンジ色）のために暗い色（青または緑）を使用してください。 <img alt="図1" src="/files/ftp_upload/3208/3208fig1.jpg" /> 図1。青動脈から黄色の静脈への緑の動脈からピンク静脈（白い星）にいくつかのAV吻合を示す通常の、合併症を伴わない単一絨毛膜性の双胎妊娠から単一絨毛膜性の胎盤、いくつかのVA吻合（緑の星）とで識別される1大規模なAA吻合（青と​​緑の色素の混合、ブルースター）。 <img alt="図2" src="/files/ftp_upload/3208/3208fig2.jpg" /> AAの吻合なしだけのAV（白い星）とVA（緑の星）吻合の存在を示すシリアルamnioreductionで治療TTTSの妊娠で2単一絨毛膜性の胎盤図 。 <img alt="図3" src="/files/ftp_upload/3208/3208fig3.jpg" /> 小さ ​​な残留吻合が誤って特許（ホワイトスター）残された選択的なレーザー技術を使用して血管吻合のfetoscopicレーザー凝固後の図3。TTTS胎盤を。選択的レーザー技術で、血管吻合は、最初に同定され、その後、一つずつを凝固されています。 <img alt="図4" src="/files/ftp_upload/3208/3208fig4.jpg" /> 図4。TTTS胎盤、個々吻合の同定と凝固した後、完全な血管の赤道は、片方の胎盤マージンから凝固されているソロモンの技法を使用してfetoscopicレーザー凝固の後に。 <img alt="図5" src="/files/ftp_upload/3208/3208fig5.jpg" /> 図5。わずか、ほんのわずかなVAの吻合（白い星）は血管の赤道に沿って表示されているTAPS胎盤。 <img alt="図6" src="/files/ftp_upload/3208/3208fig6.jpg" /> 図6。TAPS胎盤の母体側（図5に示すように）両方胎盤株式の間に特徴的な色の違いを示しています。 <img alt="図7" src="/files/ftp_upload/3208/3208fig7.jpg" /> 図7。ツインサイズの選択的子宮内発育制限と双胎妊娠の単一絨毛膜性の胎盤。成長制限された胎児は、被膜のコードの挿入とはるかに小さい胎盤を共有しています。白い星は、AA吻合を示している。 <img alt="図8" src="/files/ftp_upload/3208/3208fig8.jpg" /> 図8 Monoamniotic胎盤：様々なAV（グリーンスター）、VA（ブルースター）と2 AA吻合（白い星）、および両方のコードの挿入との間の短い距離に注意してください。

<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>

利害の衝突は宣言されません。

TTTSとTAPSは、単一絨毛膜性の双子で発生することができる2つの深刻な疾患である。両疾患は、アンバランスintertwinの血流を引き起こすAV吻合によるものです。最後の20年間で、fetoscopicレーザー手術が可能となり、TTTS 10の治療のための最善の方法であることが示された。 fetoscopicレーザー治療の目的は、胎盤表面の血管吻合の凝固を通じて相互双子の循環を中断することです。しかし?...

<table border="1"><tbody><tr><th>試薬の名前</th><th>会社</th><th>カタログ番号</th><th>コメント</th></tr><tr><td>臍カテーテル（様々な大きさ、長さ40センチ） </td><td> Vygon </td><td> 1270.08（8 F）1270.08（5 F）1270.04（4 F）1270.03（3.5 F）1270.02（2.5 F） </td><td>カテーテルの任意のタイプを使用することができます。 </td></tr><tr><td> 20ミリリットル注射器</td><td> BD Plastipak </td><td> 300613 </td><td></td></tr><tr><td>色染料（異なる色） </td><td>ロイヤルTalens Schoolverf </td><td> 36716010（青）36715010（緑）36712350（黄）36713570（ピンク） </td><td>任意の粘性カラー染料を使用することができます。 </td></tr></tbody></table>

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

胎盤血管吻合の存在は、conditio必須条件双胎児間輸血症候群（TTTS）と双子の貧血の赤血球増加症のシーケンス（TAPS）1,2の開発のためです。双子の胎盤の注入の研究では、このような吻合がdichorionic双子1でほとんど常に単一絨毛膜性の双子に存在すると極めて稀であることが示されている。吻合の3つのタイプが文書化されています：動脈から動脈に、静脈から静脈へと動脈から静脈へ。動静脈（AV）吻合は単方向であり、動静脈（AA）と静脈静脈（VV）吻合が双方向であると呼ばれるのに対し、彼らは、共有胎盤葉を進めますので、&quot;深い&quot;吻合と呼ばれています彼らは絨毛膜板上にあるので、として、&quot;表面的&quot;。 TTTSおよびTAPSの両方がAV吻合による双子の間に血流の正味の不均衡によって引き起こされます。ツイン（ドナー）からの血液は、共有胎盤葉に動脈を通ってポンプで、その後他の双子（受信者）の循環に静脈を通じて排出される。血液が逆向きの深いAV吻合を介して、または表面の吻合を介して受信者からドナーに戻って励起されていない限り、血液量の不均衡は徐々にTTTSまたはTAPSの開発につながる、発生します。 AAの吻合の存在は、単方向のAV吻合の1,2によって引き起こされる循環の不均衡を補償することによってTTTSとTAPSの開発から保護することが示されている。単一絨毛膜性の胎盤の注射は、生後間もなく単一絨毛膜性の双子の様々な（血液学）合併症の病因を理解するために有用な平均値であり、TAPS 2の診断に到達するために必要なテストです。さらに、fetoscopicレーザー手術で治療TTTSの胎盤の注入が可能残留吻合3-5の識別が可能になります。この追加情報は、TTTSやTAPSを持つ単一絨毛膜性の双子の管理とケアに関わるすべてのperinatologistsのための極めて重要である。いくつかの胎盤注射の技術は、現在使用されています。我々は、正確に着色された染料の注入を使用して（残留）血管吻合の存在を評価するための単純なプロトコルを提供します。

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

双胎児間輸血症候群とツイン貧血の赤血球増加症のシーケンスは、周産期医療の2つの潜在的に壊滅的な問題です。両疾患は、単一絨毛膜性の双子や胎盤の血管吻合を介して不平衡血流からの結果で発生する。我々は正確に出産後の胎盤血管の色の色素注入を用いた血管吻合の存在を評価するための単純なプロトコルを提供します。

カラー染料を使用して単一絨毛膜性の胎盤における血管吻合の正確かつ簡単に評価

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 ビュー.  Leiden University Medical Center. 双胎児間輸血症候群とツイン貧血の赤血球増加症のシーケンスは、周産期医療の2つの潜在的に壊滅的な問題です。両疾患は、単一絨毛膜性の双子や胎盤の血管吻合を介して不平衡血流からの結果で発生する。我々は正確に出産後の胎盤血管の色の色素注入を用いた血管吻合の存在を評価するための単純なプロトコルを提供します。

ビデオ: 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.

全血刺激のためのアッセイを用いて炎症性サイトカインIL -1βの正確かつ簡単に測定

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

医学

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.

単一絨毛膜性のツイン胎盤の血管のマッピングのためのガイドワイヤ補助カテーテルと有色色素注入

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.

マウス肺挿管の簡単な方法

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.

マウスAortocaval瘻の技術的な側面

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.

高速かつ正確な呼気アンモニア測定

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.

皮下腫瘍モデルにおける腫瘍浸潤白血球サブセットの評価

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.

大動脈弁移植 - 教則ビデオのマウスモデルでスリーブを用いた

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.

リアルなシミュレータを使用した Monochorionic Diamniotic ツイン胎盤の胎児レーザー光凝固でモデル手術トレーニング: スキルの習得

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.

心臓手術中に経食道心エコーを用いた渦形成時間の非侵襲的定量

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;

経カテーテル閉鎖前の心房通信の心エコー検査

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