אנו מודים לצוות וטרינרים במחלקת סקוט &amp; White בית החולים השוואתי רפואה, בית המקדש, טקסס, לקבלת הייעוץ יסולא בפז שלהם וסיוע בפיתוח של טכניקה זו. עבודה זו מומנה בחלקו על ידי המכון למשובי קרנות תכנית לרפואה, סקוט &amp; White RGP # מענק 90,172, NIH 2P40RR017447-07 וNIH R01AR066033-01 (NIAMS). אנו מודים לד&quot;ר סוזן זיתוני לבידוד את כתב היד.

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The authors have no competing financial interests.

בזאת, אנו מתארים שיטה פשוטה כדי ליצור פגם-התייצב סיכה קריטית בגודל של עצם הירך העכברית באמצעות מעבדה סטנדרטית וציוד וטרינרים. בעוד ההרכבה של הסיכות וההליך הכירורגי עצמו דורש תרגול, זה גם בתוך את היכולות של מדען מאומן היטב ביו-רפואי או מחקר וטרינר. <p class="jove_content" style=";t...

הערכה היא כי מחצית מאוכלוסיית ארה&quot;ב חווה שבר בגיל 65 1. עבור אותם מטופלים עם שברים שטופלו בניתוח, 500,000 הליכים כרוכים בשימוש בשתל עצם 2 ומספר זה צפוי לעלות עם הזדקנות אוכלוסייה יותר ויותר 3 . למרות שעצם הוא אחד מכמה איברים שיש את היכולת לרפא לחלוטין ללא צלקות, יש מקרים שבם התהליך נכשל 3,4. בהתאם לנסיבות ולאיכות טיפול, 2-30% משברים בעצמות ארוכות ייכשלו, וכתוצאה מכך 3,5 שאינו איגוד. אמנם יש עדיין ויכוח על ההגדרה, pseudoarthrosis, פציעות עצם הקריטיות בגודל או אי-איחוד בדרך כלל מתייחס לפגיעה שאינה נרפא במשך החיים הטבעיים של הנושא 6. למטרות ניסוי, משך זה מתקצר לזמן הממוצע שנדרש להחלמה של פציעה עצם בגודל ממוצעת מלאה. נגעי עצם ללא איחוד להתרחש numסיבות erous, אך גורמים עיקריים כוללות טראומה קיצונית וכתוצאה מכך פער קריטי בגודל, זיהום, אנגיוגנזה עניה, שימוש בטבק, או יכולת osteoregenerative עכבות עקב מחלה או גיל 7. גם אם מטופלים אינם איגודים בהצלחה, זה יכול לעלות בעודף של 60,000 $ להליך, בהתאם לסוג פציעה והגישות המועסקות 8. במקרים מתונים, השתלת עצם אוטולוגית מועסקת. אסטרטגיה זו כרוכה בהחלמה של עצם מאתר תורם והשתלה באתר של פציעה. בעוד גישה זו היא מאוד יעילה, הנפח של עצם תורם נגזרות זמין מוגבל וההליך כרוך ניתוח נוסף, שתוצאתו כאב מתמשך בחולים רבים 9,10. בנוסף, היעילות של שתל עצם אוטולוגית תלויה של בריאותו של המטופל. תחליפי עצם עשויים מחומרים סינטטיים או עצם מגופות מעובד זמינים 11-13 בשפע, אבל הם חהve מגבלות משמעותיות, כוללים נכסים עניים הידבקות מארח תאים, osteoconductivity המופחת, והפוטנציאל לדחייה חיסונית 14. אין אפוא צורך דחוף בטכנולוגיות התחדשות עצם שבטוחות, יעילים וזמינות לציבור רחב. היכולת שלנו לשפר את אסטרטגיות משובי עצם תלויה באופן קריטי על היכולת לחקות טראומת עצם חמורה בבעלי חיים בדיקה, אבל הדור והייצוב של נגעי עצם גדולים הוא מאתגר מבחינה טכנית. ברוב המקרים, טראומת עצם ארוך רצינית חיקתה בניסוי על ידי הקמת פגם שלא באופן טבעי לרפא. למרות שזה יכול להשתנות עם מינים 15, זו מושגת על ידי הסרת קטע עצם שהוא גדול יותר מ -1.5 פעמים הקוטר של עצם החתך 16 מלאה. העצם מכן התייצב עם שתל מתכת כדי לשמור על כיוון נכון של קצות שבר ולאפשר לניידות. בשל גודלם הקטן והשברירי שלהעצמות הארוכות שלהם, הקמת כזה נגעים בעכברים הן מעבר ליכולות של רוב קבוצות מחקר. ככזה, מודלים פגם עצם ארוך מוגבלים לחולדות ובעלי חיים גדולים יותר. עם זאת, עכברים יעניקו יתרונות משמעותיים במחקר שהם יכולים להיות מהונדסים גנטיים וגדלו כזנים שנפרץ חיסון שלא לדחות תאים ורקמות אנושיים. עבור יישומים מבוססי תאים אנושיים, עכברים שנפרץ חיסון הם אטרקטיביים לעבוד איתו כי הם מבחינה פיזיולוגית מאופיינים היטב, קל לבית, חסכוני, בקלות ובנתח radiologically והיסטולוגיה. חשיבות עליונה הוא שעכברים שנפרץ חיסון לא דוחים תאים ממינים שונים כולל בני אדם. גודלם הקטן מאפשר גם הבדיקה של מספרים קטנים מאוד של תאים או כרכים של פיגומים ניסיוניים ביישומים אורטופדיים. כמה מודלים עכבריים אורתופדים כבר דיווחו כי להרשות לעצמו דרגות שונות של יציבות עצם 17,18. תוכנה לאלהms המוביל לרמות גבוהות מאוד של יציבות, כגון תופסנים חיצוניים וצלחות נעילה בעיקר לרפא על ידי התאבנות intramembranous למרות ריפוי endochondral דווח 19. בניגוד לכך, אלה המאפשרים כמה מיקרו ו / או מקרו-תנועה, כגון אלה המעסיקים סיכות לשדיות מבולבלות או חלקי-קבועות, בדרך כלל לרפא עם דומיננטיות של התאבנות endochondral 20,21. איגוד מושהה או פגמים שאינם איחוד של עצם ארוך הם קשה במיוחד להשגה בעכברים עקב הרמה נוספת של ייצוב הנדרשת. עם זאת, מספר גישות כבר דיווחו, כולל סיכות לשדיות עם ציפורני לובים, צלחות נעילה ותופסנים חיצוניים 22. מערכות אלה בדרך כלל עובדים היטב, אבל בהתחשב בעיצוב המסובך שלהם הם יכולים להיות מאתגרים מבחינה טכנית להתקנה. לדוגמא, גרסיה et al. 23 המציא מערכת סיכה שלובים אלגנטית לשימוש בעכברים, אבל ההליך כרוך חתכים בשני אתר נפרדים ושינוי נרחב של עצם הירך כדי להכיל את הסיכות. נהלים אלה בוצעו תחת מיקרוסקופ לנתח. במסמך זה, אנו מתארים סיכה לשדית הירך פשוטה עם צווארון מרכזי שנועד למנוע סגירת גירעון עצם 3 מ&quot;מ וגם להתוות את הקצוות המקוריים של הפגם. בעוד הסיכה לא הייתה קבועה לעצם עצמו, לשינוי גודל מדויק של קוטר הסיכה וreaming של תוצאות החלל הלשדיות בהתערבות מספיקה כדי למזער תנועת torsional (איור 1). עם בחירה זהירה של גיל טהור, מגדר ועכברים בהתאמה מתח, התוצאה היא שאינו uniondefect hypertrophic מאוד לשחזור 22 ניתן יהיה להעריך בקלות radiologically. יתר על כן אזורים של עניין יכולים להיות מוגדרים reproducibly לאחר טומוגרפיה-מחושב מיקרו (μCT) למדידת דה נובו היווצרות עצם ופרמטרי histomorphological. הסיכות היו prototyped במעבדה שלנו תוך שימוש בכלים זמינים.<img alt="איור 1" src="/files/ftp_upload/52368/52368fig1highres.jpg" /> איור 1: עיקרון ניסויי סיכום תרשים של מודל הפגם המגזרי.. קטע 3 מ&quot;מ המרכזי של עצם ירך עכברית 9-10 מ&quot;מ הוא נכרת בניתוח (משמאל). צינור פלדה כירורגית ארוך 3 מ&quot;מ, 19 מד הוא חלף על פני מ&quot;מ ארוך 9, 22 צינור הנירוסטה G וקבוע עם דבק במרכז המדויק (מימין). סיכת התוצאה היא מצויד לתוך התעלות הלשדיות של החלקים הפרוקסימלית ודיסטלי הנותרים של עצם הירך עם 19 צווארון G החלפת קטע 3 מ&quot;מ של עצם (להלן, במרכז).

<table border="0" cellpadding="0" cellspacing="0" width="799">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:235px;">Name of Equipment/Material*</td>
			<td style="width:207px;">Company</td>
			<td style="width:180px;">Catalog or model</td>
			<td style="width:179px;">Notes</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Pin Assembly</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dremel rotary tool</td>
			<td>Dremel</td>
			<td>8220</td>
			<td>or equivalent</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Heavy duty cut off wheel</td>
			<td>Dremel</td>
			<td>420</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical tubing 19G</td>
			<td>Small Parts (Amazon)</td>
			<td>B000FMZ8LY</td>
			<td>OD 1.07mm, ID 0.889mm</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical tubing 21G</td>
			<td>Small Parts (Amazon)</td>
			<td>B000FMZ8YQ</td>
			<td>OD 0.82mm, ID 0.635mm</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical tubing 22G</td>
			<td>Small Parts (Amazon)</td>
			<td>B000FMYLZS</td>
			<td>OD 0.719mm, ID 0.502mm</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical tubing 23G</td>
			<td>Small Parts (Amazon)</td>
			<td>B000FN0SY0</td>
			<td>OD 0.643mm, ID 0.444mm</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Cyanoacrylate adhesive</td>
			<td>Loctite</td>
			<td>1365882</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Emery disc</td>
			<td>Dremel</td>
			<td>413</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rubber polishing point</td>
			<td>Dremel</td>
			<td>462</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Felt polishing disc</td>
			<td>Dremel</td>
			<td>414</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Gelatin sponge</td>
			<td>Surgifoam/Ethicon</td>
			<td>1974</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Punch biopsy cutter</td>
			<td>Miltex</td>
			<td>33-34</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgery/post-operative</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Warm pad and circulator pump</td>
			<td>Stryker/Thermocare</td>
			<td>TP700, TP700C, TPP722</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Coverage quaternary spray</td>
			<td>Steris</td>
			<td>1429-77</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Bead sterilizer</td>
			<td>Germinator/CellPoint Scentific</td>
			<td>Germinator 500</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Anesthesia system</td>
			<td>VetEquip Inc</td>
			<td>901806 or 901807/901809</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Isofluorane anesthetic</td>
			<td>VETone/MWI</td>
			<td>501017, 502017</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical disinfectant</td>
			<td>Chloraprep/CareFusion</td>
			<td>260449</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Surgical tools</td>
			<td>Fine Science Tools</td>
			<td>various</td>
			<td>recommend German made</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Face protection</td>
			<td>Splash Shield</td>
			<td>4505</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Rechargable high speed drill</td>
			<td>Fine Science Tools</td>
			<td>18000-17</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Diamond cutting wheel</td>
			<td>Strauss Diaiond</td>
			<td>361.514.080HP</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Absorbable sutures&#160;</td>
			<td>Covidien</td>
			<td>UM-213</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Outer sutures</td>
			<td>Ethicon</td>
			<td>668G</td>
			<td>or equivalent</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Vetbond</td>
			<td>3M</td>
			<td>1469SB</td>
			<td>or equivalent</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Hydration gel</td>
			<td>Clear H2O</td>
			<td>70-01-1082</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Diet gel</td>
			<td>Clear H2O</td>
			<td>72-01-1062</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Buprenorphine</td>
			<td>Reckitt and Benckser</td>
			<td>12496-0757-01</td>
			<td>controlled substance</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Mouse igloos</td>
			<td>Bio Serv</td>
			<td>K3328, 3570,3327</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Euthanasia cocktail</td>
			<td>Euthasol/Virbac</td>
			<td>710101</td>
			<td>controlled substance</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Analysis</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Live animal imager&#160;</td>
			<td>Orthoscan</td>
			<td>FD Pulse</td>
			<td>or equivalent</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Micro-CT unit and software</td>
			<td>Bruker</td>
			<td>Skyscan1174</td>
			<td>or equivalent</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sealing film/Parafilm M</td>
			<td>VWR or Fisher</td>
			<td>100501-338, S37441</td>
			<td></td>
		</tr>
		<tr height="20">
			<td colspan="4" height="20" style="height:20px;">*Generic sources are suitable for all other items such as gause, drapes, protective clothing, animal care equipment.</td>
		</tr>
	</tbody>
</table>

a simple critical-sized femoral defect model in mice

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all limb bone fractures fail to heal completely due to the extent of the trauma, disease, or age of the patient. Our ability to improve bone regenerative strategies is critically dependent on the ability to mimic serious bone trauma in test animals, but the generation and stabilization of large bone lesions is technically challenging. In most cases, serious long bone trauma is mimicked experimentally by establishing a defect that will not naturally heal. This is achieved by complete removal of a bone segment that is larger than 1.5 times the diameter of the bone cross-section. The bone is then stabilized with a metal implant to maintain proper orientation of the fracture edges and allow for mobility. 
Due to their small size and the fragility of their long bones, establishment of such lesions in mice are beyond the capabilities of most research groups. As such, long bone defect models are confined to rats and larger animals. Nevertheless, mice afford significant research advantages in that they can be genetically modified and bred as immune-compromised strains that do not reject human cells and tissue.
Herein, we demonstrate a technique that facilitates the generation of a segmental defect in mouse femora using standard laboratory and veterinary equipment. With practice, fabrication of the fixation device and surgical implantation is feasible for the majority of trained veterinarians and animal research personnel. Using example data, we also provide methodologies for the quantitative analysis of bone healing for the model.

עכברים בדרך כלל להתאושש תודעה ואחורית-איבר ניידות 5-10 דקות לאחר הנסיגה של הרדמה. במהלך 5 הימים הראשונים, רצוי לבית עכברים בנפרד ולהציג את ההעשרה סביבתית כדי למנוע שימוש יתר בגפיים. לצורך כך, קיני איגלו הסוג להפחית את הצורך בבניית קן ולעודד מנוחה. יש לנו גם ציינו כי אספק?...

Watch this Scientific Journal Video about A Simple Critical-sized Femoral Defect Model in Mice at JoVE.com

A Simple Critical-sized Femoral Defect Model in Mice

Animal models are frequently employed to mimic serious bone injury in biomedical research. Due to their small size, establishment of stabilized bone lesions in mice are beyond the capabilities of most research groups. Herein, we describe a simple method for establishing and analyzing experimental femoral defects in mice.

דגם פגם עצם הירך קריטית בגודל פשוט בעכברים

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all ...

a-simple-critical-sized-femoral-defect-model-in-mice

Research

JoVE Journal

Medicine

15.0K Views.  Texas A&M Health Science Center. Animal models are frequently employed to mimic serious bone injury in biomedical research. Due to their small size, establishment of stabilized bone lesions in mice are beyond the capabilities of most research groups. Herein, we describe a simple method for establishing and analyzing experimental femoral defects in mice. 

Video: A Simple Critical-sized Femoral Defect Model in Mice

A Simple Guide Screw Method for Intracranial Xenograft Studies in Mice

The grafting of human tumor cells into the brain of immunosuppressed mice is an established method for the study of brain cancers including glioblastoma (glioma) and medulloblastoma. The widely used stereotactic approach only allows for the injection of a single animal at a time, is labor intensive and requires highly specialized equipment. The guide screw method, initially developed by Lal et al.,1 was developed to eliminate cumbersome stereotactic procedures. We now describe a modified guide screw approach that is rapid and exceptionally safe; both of which are critical ethical considerations. Notably, our procedure now incorporates an infusion pump that allows up to 10 animals to be simultaneously injected with tumor cells.
To demonstrate the utility of this procedure, we established human U87MG glioma cells as intracranial xenografts in mice, which were then treated with AMG102; a fully human antibody directed to HGF/scatter factor currently undergoing clinical evaluation2-5. Systemic injection of AMG102 significantly prolonged the survival of all mice with intracranial U87MG xenografts and resulted in a number of complete cures. 
This study demonstrates that the guide screw method is an inexpensive, highly reproducible approach for establishing intracranial xenografts. Furthermore, it provides a relevant physiological model for validating novel therapeutic strategies for the treatment of brain cancers.

In order to evaluate novel therapeutic paradigms for the treatment of glioma, physiological relevant models are essential. We utilize an implantable guide screw procedure for establishment of intracranial xenograft models that is more rapid and safer than stereotactic approaches.

מדריך פשוט בורג שיטה ללימודי xenograft תוך גולגולתי של עכברים

The grafting of human tumor cells into the brain of immunosuppressed mice is an established method for the study of brain cancers including glioblastoma ...

Creating Rigidly Stabilized Fractures for Assessing Intramembranous Ossification, Distraction Osteogenesis, or Healing of Critical Sized Defects

Assessing modes of skeletal repair is essential for developing therapies to be used clinically to treat fractures. Mechanical stability plays a large role in healing of bone injuries. In the worst-case scenario mechanical instability can lead to delayed or non-union in humans. However, motion can also stimulate the healing process. In fractures that have motion cartilage forms to stabilize the fracture bone ends, and this cartilage is gradually replaced by bone through recapitulation of the developmental process of endochondral ossification. In contrast, if a bone fracture is rigidly stabilized bone forms directly via intramembranous ossification. Clinically, both endochondral and intramembranous ossification occur simultaneously. To effectively replicate this process investigators insert a pin into the medullary canal of the fractured bone as described by Bonnarens4. This experimental method provides excellent lateral stability while allowing rotational instability to persist. However, our understanding of the mechanisms that regulate these two distinct processes can also be enhanced by experimentally isolating each of these processes. We have developed a stabilization protocol that provides rotational and lateral stabilization. In this model, intramembranous ossification is the only mode of healing that is observed, and healing parameters can be compared among different strains of genetically modified mice 5-7, after application of bioactive molecules 8,9, after altering physiological parameters of healing 10, after modifying the amount or time of stabilization 11, after distraction osteogenesis 12, after creation of a non-union 13, or after creation of a critical sized defect. Here, we illustrate how to apply the modified Ilizarov fixators for studying tibial fracture healing and distraction osteogenesis in mice.

This article describes a method for stabilizing long bone fractures that is based on the application of modified Ilizarov external fixators 1-3. After application of the fixators and creation of the bone injury, healing can be assessed, distraction osteogenesis can be performed, or non-union or critical sized defect can be created and used to study therapeutic interventions.

יצירת שברים התייצב נוקשה להערכת התאבנות Intramembranous, הסחת הדעת osteogenesis, או ריפוי של פגמים בגודל קריטי

Assessing modes of skeletal repair is essential for developing therapies to be used clinically to treat fractures. Mechanical stability plays a large role ...

Repair of a Critical-sized Calvarial Defect Model Using Adipose-derived Stromal Cells Harvested from Lipoaspirate

Craniofacial skeletal repair and regeneration offers the promise of de novo tissue formation through a cell-based approach utilizing stem cells. Adipose-derived stromal cells (ASCs) have proven to be an abundant source of multipotent stem cells capable of undergoing osteogenic, chondrogenic, adipogenic, and myogenic differentiation. Many studies have explored the osteogenic potential of these cells in vivo with the use of various scaffolding biomaterials for cellular delivery. It has been demonstrated that by utilizing an osteoconductive, hydroxyapatite-coated poly(lactic-co-glycolic acid) (HA-PLGA) scaffold seeded with ASCs, a critical-sized calvarial defect, a defect that is defined by its inability to undergo spontaneous healing over the lifetime of the animal, can be effectively show robust osseous regeneration. This in vivo model demonstrates the basis of translational approaches aimed to regenerate the bone tissue - the cellular component and biological matrix. This method serves as a model for the ultimate clinical application of a progenitor cell towards the repair of a specific tissue defect.

This protocol describes the isolation of adipose-derived stromal cells from lipoaspirate and the creation of a 4 mm critical-sized calvarial defect to evaluate skeletal regeneration.

תיקון של דגם פגם Calvarial הקריטי בגודל תאי שומן באמצעות נגזרת סטרומה נקטפו מן Lipoaspirate

Craniofacial skeletal repair and regeneration offers the promise of de novo tissue formation through a cell-based approach utilizing stem cells. ...

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.

שיטה פשוטה של ​​אינטובציה ריאות המאוסות

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

Femoral Bone Marrow Aspiration in Live Mice

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed step-by-step technical procedure for an analogous procedure in live mice which allows for serial characterization of cells present in the BM. This procedure facilitates studies aimed to detect the presence of exogenously administered cells within the BM of mice as would be done in xenograft studies for instance. Moreover, this procedure allows for the retrieval and characterization of cells enriched in the BM such as hematopoietic stem and progenitor cells (HSPCs) without sacrifice of mice. Given that the cellular composition of peripheral blood is not necessarily reflective of proportions and types of stem and progenitor cells present in the marrow, procedures which provide access to this compartment without requiring termination of the mice are very helpful. The use of femoral bone marrow aspiration is illustrated here for cytological analysis of marrow cells, flow cytometric characterization of the hematopoietic stem/progenitor compartment, and culture of sorted HSPCs obtained by femoral BM aspiration compared with conventional marrow harvest.

This protocol describes a procedure for serial sampling of femoral bone marrow (BM) without requiring the sacrifice of mice. This procedure facilitates longitudinal studies of the BM composition of mice over time and provides serial access to cells within the BM for ex vivo and transplantation studies.

הירך מח עצם שאיפה בעכברים חיים

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed ...

Murine Model of Femoral Artery Wire Injury with Implantation of a Perivascular Drug Delivery Patch

Percutaneous interventions including balloon angioplasty and stenting have been used to restore blood flow in vessels with occlusive vascular disease. While these therapies lead to the rapid restoration of blood flow, these technologies remain limited by restenosis in the case of bare metal stents and angioplasty, or reduced healing and possibly enhanced risk of thrombosis in the case of drug eluting stents. A key pathophysiological mechanism in the formation of restenosis is intimal hyperplasia caused by the activation of vascular smooth muscle cells and inflammation due to arterial stretch and injury. Surgeries that induce arterial injury in genetically modified mice are useful for the mechanistic study of the vascular response to injury but are often technically challenging to perform in mouse models due to the their small size and lack of appropriate sized devices. We describe two approaches for a surgical technique that induces endothelial denudation and arterial stretch in the femoral artery of mice to produce robust neointimal hyperplasia. The first approach creates an arteriotomy in the muscular branch of the femoral artery to obtain vascular access. Following wire injury this arterial branch is ligated to close the arteriotomy. A second approach creates an arteriotomy in the main femoral artery that is later closed through localized cautery. This method allows for vascular access through a larger vessel and, consequently, provides a less technically demanding procedure that can be used in smaller mice. Following either method of arterial injury, a degradable drug delivery patch can be placed over or around the injured artery to deliver therapeutic agents.

We describe a surgical technique that produces wire injury in the femoral artery of mice to induce neointimal hyperplasia to serve as a model testing system for the perivascular delivery of therapeutic compounds for the inhibition of restenosis.

דגם Murine של עורק הירך חוט פגיעה בהשרשה של perivascular החדרת תרופות תיקון

Percutaneous interventions including balloon angioplasty and stenting have been used to restore blood flow in vessels with occlusive vascular disease. ...

A Murine Model of Arterial Restenosis: Technical Aspects of Femoral Wire Injury

Cardiovascular disease caused by atherosclerosis is the leading cause of death in the developed world. Narrowing of the vessel lumen, due to atherosclerotic plaque development or the rupturing of established plaques, interrupts normal blood flow leading to various morbidities such as myocardial infarction and stroke. In the clinic endovascular procedures such as angioplasty are commonly performed to reopen the lumen. However, these treatments inevitably damage the vessel wall as well as the vascular endothelium, triggering an excessive healing response and the development of a neointimal plaque that extends into the lumen causing vessel restenosis (re-narrowing). Restenosis remains a major cause of failure of endovascular treatments for atherosclerosis. Thus, preclinical animal models of restenosis are vitally important for investigating the pathophysiological mechanisms as well as translational approaches to vascular interventions. Among several murine experimental models, femoral artery wire injury is widely accepted as the most suitable for studies of post-angioplasty restenosis because it closely resembles the angioplasty procedure that injures both endothelium and vessel wall. However, many researchers have difficulty utilizing this model due to its high degree of technical difficulty. This is primarily because a metal wire needs to be inserted into the femoral artery, which is approximately three times thinner than the wire, to generate sufficient injury to induce prominent neointima. Here, we describe the essential surgical details to effectively overcome the major technical difficulties of this model. By following the presented procedures, performing the mouse femoral artery wire injury becomes easier. Once familiarized, the whole procedure can be completed within 20 min.

The mouse femoral artery wire injury model of restenosis is technically challenging. In this protocol we show the key technical details essential for successfully performing wire injury to induce consistent&#160;neointima for studies of restenosis.

דגם Murine של עורקי היצרות: היבטים טכניים של עצם הירך חוט פגיעה

Cardiovascular disease caused by atherosclerosis is the leading cause of death in the developed world. Narrowing of the vessel lumen, due to ...

Establishment of a Segmental Femoral Critical-size Defect Model in Mice Stabilized by Plate Osteosynthesis

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a model organism, the mouse has already been widely used in bone-related research. Large diaphyseal bone defect models in mice, however, are sparse and often use bone fixation which fills the bone marrow cavity and does not provide optimal mechanical stability. The objectives of the current study were to develop a critical-size, segmental, femoral defect in nude mice. A 3.5-mm mid-diaphyseal femoral ostectomy (approximately 25% of the femur length) was performed using a dedicated jig, and was stabilized with an anterior located locking plate and 4 locking screws. The bone defect was subsequently either left empty or filled with a bone substitute (syngenic bone graft or coralline scaffold). Bone healing was monitored noninvasively using radiography and in vivo micro-computed-tomography and was subsequently assessed by ex vivo micro-computed-tomography and undecalcified histology after animal sacrifice, 10 weeks postoperatively. The recovery of all mice was excellent, a full-weight-bearing was observed within one day following the surgical procedure. Furthermore, stable bone fixation and consistent fixation of the implanted materials were achieved in all animals tested throughout the study. When the bone defects were left empty, non-union was consistently obtained. In contrast, when the bone defects were filled with syngenic bone grafts, bone union was always observed. When the bone defects were filled with coralline scaffolds, newly-formed bone was observed in the interface between bone resection edges and the scaffold, as well as within a short distance within the scaffold. 
The present model describes a reproducible critical-size femoral defect stabilized by plate osteosynthesis with low morbidity in mice. The new load-bearing segmental bone defect model could be useful for studying the underlying mechanisms in bone regeneration pertinent to orthopaedic applications.

Although mouse models are invaluable tools for bone tissue engineering, models of long bone defects are sparse. This need motivated development of the present protocol which uses a locking plate with four screws and a dedicated jig to perform and stabilize a reproducible, femoral, critical-size defect with low morbidity.

הקמת דגם פגם קריטי בגודל הירך מגזרית עכברים מיוצב על ידי פלייט Osteosynthesis

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a ...

A Repetitive Concussive Head Injury Model in Mice

Despite the concussion/ mild traumatic brain injury (mTBI) being the most frequent occurrence of traumatic brain injury, there is still a lack of knowledge on the injury and its effects. To develop a better understanding of concussions, animals are often used because they provide a controlled, rigorous, and efficient model. Studies have adapted traditional animal models to perform mTBI to stimulate mild injury severity by changing the injury parameters. These models have been used because they can produce morphologically similar brain injuries to the clinical condition and provide a spectrum of injury severities. However, they are limited in their ability to present the identical features of injuries in patients. Using a traditional impact system, a repetitive concussive injury (rCHI) model can induce mild to moderate human-like concussion. The injury degree can be determined by measuring the period of loss of consciousness (LOC) with a sign of a transient termination of breathing. The rCHI model is beneficial to use for its accuracy and simplicity in determining mTBI effects and potential treatments.

Concussion presents the most common type of traumatic brain injury. Therefore, a repetitive concussive animal model, which replicates the important features of an injury in patients, may provide a means to study concussion in a rigorous, controlled, and efficient manner.

מודל פגיע ראש החוזרים מזעזע של עכברים

Despite the concussion/ mild traumatic brain injury (mTBI) being the most frequent occurrence of traumatic brain injury, there is still a lack of ...

The Generation of Closed Femoral Fractures in Mice: A Model to Study Bone Healing

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies that promote efficient bone healing are non-existent and in high demand. Effective and reproducible animal models of fractures healing are needed to understand the complex biological processes associated with bone regeneration. Many animal models of fracture healing have been generated over the years; however, murine fracture models have recently emerged as powerful tools to study bone healing. A variety of open and closed models have been developed, but the closed femoral fracture model stands out as a simple method for generating rapid and reproducible results in a physiologically relevant manner. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice and facilitate a post-fracture stabilization of the femur by inserting an intramedullary steel rod. Although devices such as a nail or a screw offer greater axial and rotational stability, the use of an intramedullary rod provides a sufficient stabilization for consistent healing outcomes without producing new defects in the bone tissue or damaging nearby soft tissue. Radiographic imaging is used to monitor the progression of callus formation, bony union, and subsequent remodeling of the bony callus. Bone healing outcomes are typically associated with the strength of the healed bone and measured with torsional testing. Still, understanding the early cellular and molecular events associated with fracture repair is critical in the study of bone tissue regeneration. The closed femoral fracture model in mice with intramedullary fixation serves as an attractive platform to study bone fracture healing and evaluate therapeutic strategies to accelerate healing.

The murine closed femoral fracture model is a powerful platform to study fracture healing and novel therapeutic strategies to accelerate bone regeneration. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice using an intramedullary steel rod to stabilize the femur.

הדור של שברי הירך סגור בעכברים: מודל ללמוד ריפוי עצם

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies ...