עבודה זו נתמכה על ידי הצבא, חיל הים, NIH, חיל האוויר, VA לענייני בריאות לתמוך במאמץ AFIRM השנייה, תחת פרס מס W81XWH-13-2-0053. פעילות רכישה הצבא האמריקני למחקר רפואי, 820 צ&#39;נדלר רחוב, פורט דטריק MD 21,702-5,014 הוא משרד הרכישה הענקת וניהלה. דעות, פרשנויות, מסקנות והמלצות הן על דעת המחבר ואינם שאושרו בהכרח על ידי משרד הגנה. המחברים מבקשים להודות ג&#39;סיקה איזי, DVM, קרוליין גארט, DVM וג&#39;ולי ווטסון, DVM לתמיכת הווטרינרים המעולה שלהם במהלך המחקר.

כל הניסויים נערכו בהתאם מדריך לטיפול ושימוש בחיות מעבדה של המכון הלאומי לבריאות (NIH) ואושרו על ידי ועדת טיפול בבעלי חיים באוניברסיטת ג&#39;ונס הופקינס ושימוש (JHUACUC). הנהלים הספציפיים בוצעו תחת MO13M108 פרוטוקול ACUC אושר. מבצע התורם 1. <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> נהל שיכוך כאבים בנקודת הזמן המתאים לכל ניסוח תרופתי לפני ניתוח. על פי השימוש בפרוטוקול טיפול בבעלי חיים ושימוש אישרה 0.1 מ&quot;ג / ק&quot;ג BW של עצירות תת עורי 1 hr לפני החתך בעור. </li><li style=";text-align:right;direction:rtl"> התורם שלווה עם isoflurane מיושם באמצעות תא המצורף מאדה isoflurane ב 4%; לשמור הרגעה והרדמה ב -2% באמצעות חרטומו. בצע השתקפות נסיגת קמצוץ בוהן כדי לפקח על עומק ההרדמה לפני תחילתו של ההליך. </li><li style=";text-align:right;direction:rtl"> לובשים מסכות, שמלות בידוד הפנויה וכפפות. </li><li style=";text-align:right;direction:rtl"> לגלח את ar כירורגיתea, בפרט הגפה ובמפשעה האחוריות, ו prep עם 10% Povidone - יוד. </li><li style=";text-align:right;direction:rtl"> השתמש וילון שדה סטרילי, מכשירים autoclaved ו מיקרוסקופ בהגדלה גבוהה (40X). </li><li style=";text-align:right;direction:rtl"> בצעו חתך עור במפשעה באמצעות מספרי proximally לאזור אמצע הירכיים ואת circumferentially לחבר את החתך כדי לתחום את הגפיים אחוריים משאר גוף העכבר. </li><li style=";text-align:right;direction:rtl"> זהה לנתח את עורק, וריד עצב הירך. הפרד כל שלושת המבנים באמצעות מלקחיים ומספריים מיקרו. </li><li style=";text-align:right;direction:rtl"> לאחר pedicle וסקולרית הוא גזור לחלק את הכלי ברמה של הרצועה מפשעתי באמצעות מספרי מייקרו. </li><li style=";text-align:right;direction:rtl"> לאחר מכן, ממשיכים לחלק את גחון הפרט (שרירי ירך gracilis המדיאלי) וקבוצות שרירים הגבו 20 proximally ברמה של ירך האמצע להפריד השתל מן החיה התורמת באמצעות מספריים. </li><li style=";text-align:right;direction:rtl"> Transect הירך וגוזר באמצע פיר הירך באמצעות מספריים. </li><li style=";text-align:right;direction:rtl"> להרדים בעלי חיים על ידי fol מנת יתר isofluraneשאחריו נקע בצוואר הרחם. אשר הפסיק פעימה ונשימת לב. </li><li style=";text-align:right;direction:rtl"> שטוף את האיבר עם 2 מיליליטר heparinized (30 IE) קר (4 ° C) מלוח באמצעות מחט שטיפת G 33 רכובה על מזרק (ראה לוח חומרים). </li><li style=";text-align:right;direction:rtl"> מניח שרוול polyimide אחד על וריד, ועצם ירך, בהתאמה. </li><li style=";text-align:right;direction:rtl"> עוטפים השתל לתוך גזה צמר גפן רטוב, מקום בצלחת פטרי ולאחסן ב 4 ° C עד הבלעה. </li></ol> 2. נמען מבצע <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> הסרת הגפיים האחוריות <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> נהל שיכוך כאבים בנקודת הזמן המתאים לכל ניסוח תרופתי לפני ניתוח. על פי השימוש בפרוטוקול טיפול בבעלי חיים ושימוש אישרה 0.1 מ&quot;ג / ק&quot;ג BW של עצירות SC 1 hr לפני החתך בעור. </li><li style=";text-align:right;direction:rtl"> התורם שלווה עם isoflurane מיושם באמצעות תא המצורף מאדה isoflurane ב 4%; לשמור הרגעה והרדמה ב -2% באמצעות חרטומו. בצע השתקפות נסיגת קמצוץ בוהן כדי לפקח על המחלקהשעות של הרדמה לפני תחילתו של ההליך. </li><li style=";text-align:right;direction:rtl"> השתמש במשחה וטרינריים המשפיעים על העיניים של העכבר כדי למנוע יובש לאחר הרדמה. </li><li style=";text-align:right;direction:rtl"> לגלח את אזור הניתוח, ובמיוחד הגפיים האחוריות ובמפשעה מכין עם 10% Povidone - יוד. </li><li style=";text-align:right;direction:rtl"> בצעו חתך עור במפשעה באמצעות מספרי proximally לאזור אמצע הירכיים ואת circumferentially לחבר את החתך כדי לתחום את הגפיים אחוריים משאר גוף העכבר. </li><li style=";text-align:right;direction:rtl"> זהה לנתח את עורק הירך, וריד ועצב ולהפריד כל שלושת המבנים באמצעות מלקחיים ומספריים מיקרו. </li><li style=";text-align:right;direction:rtl"> לאחר pedicle וסקולרית הוא גזור, מהדק את כלי הירך ברמה של הרצועה מפשעתי. </li><li style=";text-align:right;direction:rtl"> חותכים את כלי דיסטלי ברמה של העורק ברום הבטן שטחית. </li><li style=";text-align:right;direction:rtl"> לאחר מכן, ממשיכים לחלק את גחון הפרט (שרירי ירך gracilis המדיאלי) וקבוצות שרירים הגבו 20 proximally ברמה של ירך האמצע להפריד את האחוריות li הילידיםMB מבעלי החיים לנמען באמצעות מספריים. </li><li style=";text-align:right;direction:rtl"> Transect הירך באמצע פיר הירך באמצעות מספריים. </li><li style=";text-align:right;direction:rtl"> לצרוב שרירי ירך transected בעבר כדי למנוע דימום של אתר לנתיחה ובכך איבוד דם נמען. </li></ol></li><li style=";text-align:right;direction:rtl"> הַשׁרָשָׁה <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> להקטין את איבוד הנוזלים על ידי להשקיית שדה הניתוח עם מלח חמים (37 מעלות צלזיוס) והזרקת 0.3 מ&quot;ל תמיסת מלח חמים לפני ואחרי הניתוח. </li><li style=";text-align:right;direction:rtl"> מניחים את השתל באופן המשקף את עמדת אנטומי מדויק של הגפיים האחוריות יליד ידי יישור עצם הירך של הנמען ואת השתל ולחבר אותם באמצעות מחט השדרה 20 G כמו מסמר תוך לשדי. </li><li style=";text-align:right;direction:rtl"> Coapt קבוצות שרירים הגחון ועל הגב באמצעות חומר תפר נספג (6-0 Polysorb). </li><li style=";text-align:right;direction:rtl"> חבר את כלי הירך באמצעות טכניקת השרוול-תפר הכללי; בפירוט, למשוך את הצד המקבל של כלי שיט על האזיקים רכובים בעבר על הכלי המסתיים של גרףt. השתמש תפר 10-0 ניילון ולבצע עניבה היקפית כדי לתקן את ספינת הנמען על השרוול. </li><li style=";text-align:right;direction:rtl"> הבא לשחרר את המלחציים. בשלב זה לוודא ויזואלית סיבוב שרוול ומיצוב אופטימלי כדי למנוע אי-סיבוב מסתלסל של הכלים. </li><li style=";text-align:right;direction:rtl"> בצע המוסטאסיס מוקפד באמצעות כוויית אלקטרו עם דגש מיוחד על הממשק התורם למקבל השריר ואת קצות העצם. </li><li style=";text-align:right;direction:rtl"> סגור את העור באמצעות תפרים ניילון שאינם נספגים (6-0 Ethilon). </li><li style=";text-align:right;direction:rtl"> לקבוע תנאי normothermic בכך שהוא מאפשר את החיה להתאושש בכלוב שלה תחת מנורת חימום. המשך מעקב שוטף לפחות 4 שעות לפני חזרת אותו למוקד הדיור. </li><li style=";text-align:right;direction:rtl"> לספק שיכוך כאבים שלאחר הניתוח עם עצירות במינון של 0.1mg / kg SC כל 6-8 שעות במשך 3 ימים. </li></ol></li></ol>

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D., Liu, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+hindlimb+transplantation+in+the+mouse.">Orthotopic hindlimb transplantation in the mouse.</a> J. Reconstr. Microsurg. 19, 49 (2002).</li><li>Tung, T. H., Mohanakumar, T., Mackinnon, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+a+Mouse+Model+for+Heterotopic+Limb+and+Composite-Tissue+Transplantation.">Development of a Mouse Model for Heterotopic Limb and Composite-Tissue Transplantation.</a> J. Reconstr. Microsurg. 17, 267-274 (2001).</li><li>Zhang, F., Shi, D. Y., Kryger, Z., Moon, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+a+mouse+limb+transplantation+model.">Development of a mouse limb transplantation model.</a> Microsurgery. 19 (5), 209-213 (1999).</li><li>Schneeberger, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atypical+acute+rejection+after+hand+transplantation.">Atypical acute rejection after hand transplantation.</a> Am. J. Transplant. 8, 688-696 (2008).</li><li>Mathes, D. W., 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=Stable+mixed+hematopoietic+chimerism+permits+tolerance+of+vascularized+composite+allografts+across+a+full+major+histocompatibility+mismatch+in+swine.">Stable mixed hematopoietic chimerism permits tolerance of vascularized composite allografts across a full major histocompatibility mismatch in swine.</a> Transpl. Int. 27, 1086-1096 (2014).</li><li>Yamada, Y., 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=Use+of+CTLA4Ig+for+induction+of+mixed+chimerism+and+renal+allograft+tolerance+in+nonhuman+primates.">Use of CTLA4Ig for induction of mixed chimerism and renal allograft tolerance in nonhuman primates.</a> Am. J. Transplant. 14, 2704-2712 (2014).</li><li>Sachs, D. H., Kawai, T., Sykes, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+tolerance+through+mixed+chimerism.">Induction of tolerance through mixed chimerism.</a> Cold Spring Harb. Perspect. Med. 4, a015529 (2014).</li><li>Kawai, T., 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=HLA-mismatched+renal+transplantation+without+maintenance+immunosuppression.">HLA-mismatched renal transplantation without maintenance immunosuppression.</a> N. Engl. J. Med. 358, 353-361 (2008).</li><li>Kawai, T., Sachs, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tolerance+induction:+hematopoietic+chimerism.">Tolerance induction: hematopoietic chimerism.</a> Curr Opin Organ Transplant. 18, 402-407 (2013).</li><li>Kawai, T., Sachs, D. H., Sykes, M., Cosimi, A. .. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=HLA-mismatched+renal+transplantation+without+maintenance+immunosuppression.">HLA-mismatched renal transplantation without maintenance immunosuppression.</a> N. Engl. J. Med. 368, 1850-1852 (2013).</li><li>Leventhal, 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=Chimerism+and+tolerance+without+GVHD+or+engraftment+syndrome+in+HLA-mismatched+combined+kidney+and+hematopoietic+stem+cell+transplantation.">Chimerism and tolerance without GVHD or engraftment syndrome in HLA-mismatched combined kidney and hematopoietic stem cell transplantation.</a> Sci. Transl. Med. 4, 124-128 (2012).</li><li>Cendales, L. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Banff+2007+working+classification+of+skin-containing+composite+tissue+allograft+pathology.">The Banff 2007 working classification of skin-containing composite tissue allograft pathology.</a> Am. J. Transplant. 8, 1396-1400 (2008).</li></ol>

The authors declare that they have no competing financial interest.

Vascularized Composite אלוגרפט, כגון גפיים עליונים והשתלות פן לשיקום פגמי רקמות הרסניות, התפתח כאפשרות טיפול תקפה בחולים לא amendable להליכים שיקומיים קונבנציונליים. התקדמות טכנית בתחום המיקרו-כירורגיה לשחזור וכן ניסיון רב עם אימונוסופרסיבי החזק וטיפולי modulatory חיסוניים בהשתלות א...

The late nineties heralded the pioneering days of reconstructive transplantation with the first successful hand transplant performed in France in 1998. Since then, the use of VCAs for reconstruction of devastating tissue defects has been successfully employed in a wide spectrum of patients. To date, the world counts 76 recipients of 112 upper extremities as well as 31 faces 1-3. In addition, several other types of VCAs such as abdominal wall 4, larynx 5, trachea 6, vascularized joints 7, and even penis 8 have been performed. Furthermore, the live birth of a baby was recently reported after uterus transplantation 9. This growing world experience is indicative for how reconstructive transplantation has become a valid therapeutic option for patients suffering of significant functional tissue defects not amendable to conventional reconstructive and restorative surgery and treatment. 
While the idea of replacing "like with like" sparked clinical enthusiasm, initial skepticism still prevails with regards to side effects of conventional high-dose immunosuppression required to maintain allografts and their function 10,11. However, as shown by seminal work of Lee et al., these composite grafts are less likely to reject than its individual components, and furthermore, some of the tissue components such as the vascularized bone compartment have fueled optimism as they might exert unique immunological effects onto the balance of immune acceptance and rejection 12. 
Our group pioneered several microsurgical animal models for solid organ transplantation, as well as vascularized composite allotransplantation 13-19. Here we describe a novel surgical procedure using a non-suture cuff technique to perform super micro-vascular anastomosis in an orthotopic mouse hind limb transplantation model. This transplant model provides a useful tool for investigating immune acceptance and rejection mechanisms, as well as the role of individual tissue components, such as the vascularized bone marrow compartment, towards tolerance induction in the immunologically versatile setting of the mouse species. Additionally, the orthotopic placement of the limb opens the possibilities for nerve regeneration and functional outcome studies, which are critically important to the setting of VCA.

<table><tbody><tr><td>Suture, 6-0 Nylon</td><td>MWI</td><td>31849</td><td></td></tr><tr><td>Suture, 6-0 Polysorb</td><td>MWI</td><td>72667</td><td></td></tr><tr><td>Suture, 10-0 Nylon</td><td>Aero Surgical</td><td>TK-107038</td><td></td></tr><tr><td>Polyimide Tubing, Size 25</td><td>Vention Medical</td><td>141-0023</td><td></td></tr><tr><td>Polyimide Tubing, Size 27</td><td>Vention Medical</td><td>141-0015</td><td></td></tr><tr><td>Microvascular Clamps (Single)</td><td>Synovis</td><td>00396</td><td></td></tr><tr><td>Microvascular Clamps (Double)</td><td>Synovis</td><td>00414</td><td></td></tr><tr><td>Micro-Scissors</td><td>Synovis</td><td>SAS-18</td><td></td></tr><tr><td>Micro-Forceps</td><td>Synovis</td><td>FRS-15 RM-8</td><td></td></tr><tr><td>Micro-Dilators</td><td>Synovis</td><td>FRS-15 RM-8d.1</td><td></td></tr><tr><td>Micro-Needledriver</td><td>Synovis</td><td>C-14</td><td></td></tr><tr><td>Micro-Clamp Applicator</td><td>Synovis</td><td>CAF-4</td><td></td></tr><tr><td>Micro-Flushing Needle</td><td>Hamilton</td><td></td><td>10MM, 30&amp;deg;, 33G</td></tr><tr><td>Lactated Ringers Solution</td><td>Fisher Scientific</td><td>NC9968051</td><td></td></tr><tr><td>Buprenorphine</td><td></td><td></td><td>DEA Number required; Obtained from hosptial pharmacy.</td></tr><tr><td>Enrofloxacin; Baytril</td><td>Bayer Health Care</td><td>186599</td><td></td></tr><tr><td>Heparin</td><td></td><td></td><td>Obtained from hosptial pharmacy</td></tr></tbody></table>

orthotopic hind limb transplantation in the mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research. 
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace "like with like" in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.

Performing אלוגרפט מרוכבים vascularized במודל של עכברים באמצעות טכניקה השרוול הלא תפר מאפשר להשיג הישרדות השתל ובעלי חיים טווח מצוין וארוך כפי שמוצג באיור 1. יתר על כן, הוא מייצג שיטה אמינה לקבל תוצאות לשחזור של דחייה של השתל הדרגתית מרוכבים ...

Watch this Scientific Journal Video about Orthotopic Hind Limb Transplantation in the Mouse at JoVE.com

Orthotopic Hind Limb Transplantation in the Mouse

מודל הרומן הזה להשתלה הגפיים האחוריות orthotopic בתוך העכבר, החלת טכניקה השרוול הלא תפר עבור השקה סופר-כלי הדם, מספק כלי רב עוצמה עבור מחקר אימונולוגי מכניסטית in vivo הקשורים אלוגרפט מרוכבים vascularized (VCA).

Orthotopic הינד לימב השתלה בבית העכבר

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and ...

orthotopic-hind-limb-transplantation-in-the-mouse

Research

JoVE Journal

Medicine

11.6K Views.  Johns Hopkins University School of Medicine.  מודל הרומן הזה להשתלה הגפיים האחוריות orthotopic בתוך העכבר, החלת טכניקה השרוול הלא תפר עבור השקה סופר-כלי הדם, מספק כלי רב עוצמה עבור מחקר אימונולוגי מכניסטית in vivo הקשורים אלוגרפט מרוכבים vascularized (VCA). 

Video: Orthotopic Hind Limb Transplantation in the Mouse

Orthotopic Hind-Limb Transplantation in Rats

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel therapeutic entity in reconstructive surgery. However, long term high-dose multi-drug immunosuppressive therapy is required to ensure graft survival, bearing the risk of serious side effects which halters broader application. Further progression in this field may depend on better understanding of basic immunology and ischemia reperfusion injury in composite tissue grafts.
To date, orthotopic hind limb transplantation in rats has been the preferred rodent model for reconstructive transplantation (RT), however, it is an extremely demanding procedure that requires extraordinary microsurgical skills for reattachment of vasculature, bones, muscles and nerves.
We have introduced the vascular cuff anastomosis technique to this model, providing a rapid and reliable approach to rat hind limb transplantation. This technique simplifies and shortens the surgical procedure and enables surgeons with basic microsurgical experience to successfully perform the operation with high survival and low complication rates. The technique seems to be well suited for immunological as well as ischemia reperfusion injury (IRI) studies.

Here we describe the orthotopic rat hind-limb transplantation procedure, which seems to be the gold standard in vivo model for composite tissue allotransplantation research.

השתלת orthotopic לימב הינד-בחולדות

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel ...

Immunology and Infection

Heterotopic Heart Transplantation in Mice

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable for studying rejection and immune response now that newer transgenic and gene knockout mice are available, and a large number of immunologic reagents have been developed. The heart transplant model is less stringent than the skin transplant models, although technically more challenging. We have developed a modified technique and have completed over 1000 successful cases of heterotopic heart transplantation in mice. When making anastomosis of the ascending aorta and abdominal aorta, two stay sutures are placed at the proximal and distal apexes of recipient abdominal aorta with the donor s ascending aorta, then using 11-0 suture for anastomosis on both side of aorta with continuing sutures. The stay sutures make the anastomosis easier and 11-0 is an ideal suture size to avoid bleeding and thrombosis.
When making anastomosis of pulmonary artery and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s pulmonary artery. The left wall of the inferior vena cava and donor s pulmonary artery is closed with continuing sutures in the inside of the inferior vena cava after, one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s pulmonary artery are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis. In this article, we provide details of the technique to supplement the video.

The mouse heterotopic heart transplantation model has been proven by many investigators to be an important method for studying mechanisms of rejection and immune response. However, the techniques involved are still challenging. By modifying standard techniques we have had success with more than 1000 transplants.

השתלת הלב Heterotopic ב עכברים

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable ...

Biology

Orthotopic Aortic Transplantation in Mice for the Study of Vascular Disease

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. This is largely true, but there are a number of important principles that can reduce the difficulty of these procedures and enhance reproducibility. Orthotopic aortic transplantation is an excellent procedure in which to learn these principles because it involves only two end-to-end anastomoses, but requires good suturing technique and handling of the vessels for consistent success. This procedure begins with the procurement of a length of abdominal aorta from a donor animal, followed by division of the native aorta in the recipient. The procured aorta is then placed between the divided ends of the recipient aorta and sutured into place using end-to-end anastomoses. To accomplish this objective successfully requires a high degree of concentration, good tools, a steady hand, and an appreciation of how easily the vasculature of a mouse can be damaged, resulting in thrombosis. Learning these important principles is what occupies most of the beginner's time when learning microsurgery in small rodents. Throughout this protocol, we refer to these important points. This model can be used to study vascular disease in a variety of different experimental systems1-8. In the context shown here, it is most often used for the study of post-transplant vascular disease, a common long-term complication of solid organ transplantation in which intimal hyperplasia occurs within the allograft. The primary advantage of the model is that it facilitates quantitative morphometric analyses and the transplanted vessel lies contiguous to the endogenous vessel, which can serve as an additional control9. The technique shown here is most often used for mice weighing 18-25 grams. We have accumulated most of our experience using the C57BL/6J, BALB/cJ, and C3H/HeJ strains.

We describe a technique in which a section of the abdominal aorta from a mouse is transplanted orthotopically to just below the renal arteries in an allogeneic or syngeneic recipient. This technique can be useful in studies in which transplantation of large arteries of uniform size is deemed advantageous.

השתלת אב עורקים בעכברי orthotopic לחקר מחלת כלי דם

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. ...

A Modified Method for Heterotopic Mouse Heart Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents available. A difficulty is presented due to the small size of the animal and the considerable technical challenges of the microsurgery involved in heart transplantation. In particular, a high rate of technical failure early after transplantation may result from recipient death and post-operative complications such as hind limb paralysis or a non-beating heart. Here, the complete technique for heterotopic mouse heart transplantation is demonstrated, involving harvesting the donor heart and its subsequent implantation into a recipient mouse. The donor heart is harvested immediately following in situ perfusion with cold heparinized saline and transection of the ascending aorta and pulmonary artery. The recipient operation involves preparation of the abdominal aorta and inferior vena cava (IVC), followed by end-to-side anastomosis of the donor aorta with the recipient aorta using a single running 10-0 microsuture and a similar anastomosis of the donor pulmonary artery with the recipient IVC. Following the operation the animal is injected with 0.6 ml normal saline subcutaneously and allowed to recover on a 37&#160;&#176;C heating pad. The results from 227 mouse heart transplants are summarized with a success rate at 48 hr of 86.8%. Of the 13.2% failures within 48 hr, 5 (2.2%) experienced hind limb paralysis, 10 (4.4%) had a non-beating heart due to graft ischemic injury and/or thrombosis, while 15 (6.6%) died within 48 hr.

A technique is demonstrated for the microsurgical procedure for heterotopic transplantation of hearts in mice, including simplified methods for donor harvesting and recipient vessel anastomosis.

שיטה שונה לHeterotopic מאוס לב Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents ...

A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression must be pursued. Employing vascularized bone marrow transplantation and co-transplantation of the thymus have shown promise in this regard in various animal models.1-11 Vascularized bone marrow transplantation allows for the uninterrupted transfer of donor bone marrow cells within the preserved donor microenvironment, and the incorporation of thymus tissue with vascularized bone marrow transplantation has shown to increase T-cell chimerism ultimately playing a supportive role in the induction of immune regulation. The combination of solid organ and vascularized composite allotransplantation can uniquely combine these strategies in the form of a novel transplant model. Murine models serve as an excellent paradigm to explore the mechanisms of acute and chronic rejection, chimerism, and tolerance induction, thus providing the foundation to propagate superior allograft survival strategies for larger animal models and future clinical application. Herein, we developed a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique. The experience in syngeneic and allogeneic transplant settings is described for future broader immunological investigations via an instructional manuscript and video supplement.

To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

Microsurgical דגם חדשני לHeterotopic, כמקשה אחת בחזה קיר, קורנית, ולב השתלה בעכברים

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression ...

Partial Heterotopic Hindlimb Transplantation Model in Rats

Vascularized composite allotransplantations (VCA) represent the most advanced reconstruction option for patients without autologous surgical possibilities after a complex tissue defect. Face and hand transplantations have changed disfigured patients' lives, giving them a new aesthetic and functional social organ. Despite promising outcomes, VCA is still underperformed due to life-long immunosuppression comorbidities and infectious complications. The rat is an ideal animal model for in vivo studies investigating immunological pathways and graft rejection mechanisms. Rats are also widely used in novel composite tissue graft preservation techniques, including perfusion and cryopreservation studies. Models used for VCA in rats must be reproducible, reliable, and efficient with low postoperative morbidity and mortality. Heterotopic limb transplantation procedures fulfill these criteria and are easier to perform than orthotopic limb transplants. Mastering rodent microsurgical models requires solid experience in microsurgery and animal care. Herein is reported a reliable and reproducible model of partial heterotopic osteomyocutaneous flap transplantation in rats, the postoperative outcomes, and the means of prevention of potential complications.

This paper presents a partial heterotopic osteomyocutaneous flap transplantation protocol in rats and its potential outcomes in the mid-term follow-up.

מודל השתלת הינדימבה הטרוטופית חלקית בחולדות

Vascularized composite allotransplantations (VCA) represent the most advanced reconstruction option for patients without autologous surgical possibilities ...

A Modified Murine Heterotopic Heart Transplant Protocol Matching Contemporary Standards of Aseptic Technique, Anesthesia, and Analgesia

The development of experimental models of cardiac transplantation in animals has contributed to many advances in the fields of immunology and solid organ transplantation. While the heterotopic vascularized murine cardiac transplantation model was initially utilized in studies of graft rejection using combinations of mismatched inbred mouse strains, access to genetically modified strains and therapeutic modalities can provide powerful new preclinical insights. Fundamentally, the surgical methodology for this technique has not changed since its development, especially with respect to important factors such as aseptic technique, anesthesia, and analgesia, which make material impacts on postsurgical morbidity and mortality. Additionally, improvements in perioperative management are expected to provide improvements in both animal welfare and experimental outcomes. This paper reports upon a protocol developed in collaboration with a subject matter expert in veterinary anesthesia and describes the surgical technique with an emphasis on perioperative management. Additionally, we discuss the implications of these refinements and provide details on troubleshooting critical surgical steps for this procedure.

The present paper describes a modified technique for heterotopic vascularized cardiac transplantation with updated aseptic technique, analgesia, and anesthesia.

פרוטוקול השתלת לב הטרוטופי מורין מותאם לסטנדרטים עכשוויים של טכניקה אספטית, הרדמה ושיכוך כאבים

The development of experimental models of cardiac transplantation in animals has contributed to many advances in the fields of immunology and solid organ ...

Taking the Next Step: a Neural Coaptation Orthotopic Hind Limb Transplant Model to Maximize Functional Recovery in Rat

Limb transplant in particular and vascularized composite allotransplant (VCA) in general have wide therapeutic promise that have been stymied by current limitations in immunosuppression and functional neuromotor recovery. Many animal models have been developed for studying unique features of VCA, but here we present a robust reproducible model of orthotopic hind limb transplant in rats designed to simultaneously investigate both aspects of current VCA limitation: immunosuppression strategies and functional neuromotor recovery. At the core of the model rests a commitment to meticulous, time-tested microsurgical techniques such as hand sewn vascular anastomoses and hand sewn neural coaptation of the femoral nerve and the sciatic nerve. This approach yields durable limb reconstructions that allow for longer lived animals capable of rehabilitation, resumption of daily activities, and functional testing. With short-term treatment of conventional immunosuppressive agents, allotransplanted animals survived up to 70 days post-transplant, and isotransplanted animals provide long lived controls beyond 200 days post-operatively. Evidence of neurologic functional recovery is present by 30 days post operatively. This model not only provides a useful platform for interrogating immunological questions unique to VCA and nerve regeneration, but also allows for in vivo testing of new therapeutic strategies specifically tailored for VCA.

This protocol presents a robust, reproducible model of vascularized composite allotransplant (VCA) geared toward simultaneous study of immunology and functional recovery. The time invested in meticulous technique in a right mid-thigh hind limb orthotopic transplant with hand sewn vascular anastomoses and neural coaptation yields the ability to study functional recovery.

לוקח את הצעד הבא: מודל השתלת גפיים אחוריות קופטציה עצבית כדי למקסם את ההתאוששות תפקודית בחולדה

Limb transplant in particular and vascularized composite allotransplant (VCA) in general have wide therapeutic promise that have been stymied by current ...

Blood Circuit Reconstruction in an Abdominal Mouse Heart Transplantation Model

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the established technique for a modified blood circuit reconstruction in a heterotopic abdominal heart transplantation model is presented. This method uses the intrathoracic inferior vena cava (IIVC) instead of the pulmonary artery of the donor heart for the anastomosis to the inferior vena cava of the recipient. It is facilitating and improving success rates for abdominal heart transplantation in mice.

A novel technique for blood circuit reconstruction in a heterotopic abdominal mouse heart transplantation model is demonstrated.

שחזור מעגל הדם במודל השתלת לב של עכבר בטן

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the ...

A Heterotopic Mouse Model for Studying Laryngeal Transplantation

Laryngeal heterotopic transplantation, although a technically challenging procedure, offers more scientific analysis and cost benefits compared to other animal models. Although first described by Shipchandler et al. in 2009, this technique is not widely used, possibly due to the difficulties in learning the microsurgical technique and time required to master it. This paper describes the surgical steps in detail, as well as potential pitfalls to avoid, in order to encourage effective use of this technique.
In this model, the bilateral carotid arteries of the donor larynx are anastomosed to the recipient carotid artery and external jugular vein, allowing for blood flow through the graft. Blood flow can be confirmed intraoperatively by the visualization of blood filling in the graft bilateral carotid arteries, reddening of the thyroid glands of the graft, and bleeding from micro vessels in the graft. The crucial elements for success include delicate preservation of the graft vessels, making the correct size arteriotomy and venotomy, and using the appropriate number of sutures on the arterial-arterial and arterial-venous anastomoses to secure vessels without leakage and prevent occlusion. 
Anyone can become proficient in this model with sufficient training and perform the procedure in approximately 3 h. If performed successfully, this model allows for immunologic studies to be performed with ease and at low cost.

The aim of this manuscript is to describe the microsurgical steps required to perform a heterotopic laryngeal transplant in mice. The advantages of this mouse model compared to other animal models of laryngeal transplantation are its cost-effectiveness and the availability of immunologic assays and data.

מודל עכבר הטרוטופי לחקר השתלת גרון

Laryngeal heterotopic transplantation, although a technically challenging procedure, offers more scientific analysis and cost benefits compared to other ...