ونحن نشكر الموظفين والأطباء البيطريين في وزارة سكوت ومستشفى الأبيض الطب المقارن، معبد بولاية تكساس، للحصول على المشورة التي لا تقدر بثمن والمساعدة خلال تطوير هذه التقنية. وقد تم تمويل هذا العمل في جزء من معهد الطب التجديدي أموال البرنامج، سكوت اند وايت RGP منحة # 90172، NIH 2P40RR017447-07 وNIH R01AR066033-01 (NIAMS). نشكر الدكتورة سوزان زيتوني لعزل المخطوطة.

<ol>
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The authors have no competing financial interests.

هنا، نحن تصف طريقة بسيطة لتوليد عيب الحرجة الحجم استقرت دبوس من عظم الفخذ الفئران باستخدام المختبر القياسية والمعدات البيطرية. في حين أن التجمع من المسامير وإجراء العمليات الجراحية في حد ذاته يتطلب الممارسة، فإنه على ما يرام في حدود قدرات عالم أبحاث الطب الحيوي الم?...

ويقدر أن نصف سكان الولايات المتحدة يعاني من كسر في سن 65 1. بالنسبة لأولئك المرضى الذين يعانون من كسور المعالجة جراحيا، تشمل 500،000 إجراءات استخدام الكسب غير المشروع العظام 2 و من المتوقع أن يرتفع مع شيخوخة السكان على نحو متزايد 3 هذا العدد . على الرغم من أن العظام هي واحدة من عدد قليل من الأجهزة التي لديها القدرة على شفاء تماما بدون تندب، هناك حالات حيث فشل عملية 3،4. تبعا للظروف ونوعية العلاج، 2-30٪ من كسور العظام الطويلة تفشل، مما أدى إلى عدم النقابات 3،5. في حين لا يزال هناك بعض الجدل حول تعريف، فصال كاذب، وإصابات حرجة الحجم أو غير نقابية العظام تشير بصفة عامة إلى إصابة لا يلتئم على مدى عمر الطبيعي لهذا الموضوع (6). لأغراض تجريبية، هو تقصير هذه المدة إلى متوسط ​​الوقت اللازم للشفاء التام من متوسط ​​الحجم إصابة العظام. تحدث الآفات العظام غير نقابية لالأسطواناتأسباب erous، ولكن العوامل الرئيسية وتشمل الصدمة الشديدة مما أدى إلى وجود فجوة خطيرة الحجم، والعدوى، وضعف الأوعية الدموية، واستخدام التبغ، أو قدرة osteoregenerative تحول دون ذلك بسبب المرض أو العمر 7. حتى لو يتم التعامل غير النقابات بنجاح، فإنه يمكن أن تكلف ما يزيد على 60،000 دولار في الإجراء، وهذا يتوقف على نوع الإصابة والمناهج المستخدمة 8. وفي الحالات المعتدلة، وتوظيف ذاتي ترقيع العظام. وتتضمن هذه الاستراتيجية استرداد العظام من موقع المانحة وغرس في موقع الإصابة. في حين أن هذا النهج هو فعالة للغاية، وحجم متاح العظام المستمدة المانحة محدود وينطوي على إجراء عملية جراحية إضافية، مما يؤدي إلى ألم مستمر في العديد من المرضى 9،10. وبالإضافة إلى ذلك، فإن فعالية من الكسب غير المشروع العظام ذاتي يعتمد على صحة المريض. هي بدائل العظام مصنوعة من مواد تركيبية أو معالجتها العظام جثي متوافرة بكثرة 11-13، لكنها هكتارلقد قيود كبيرة، بما في ذلك سوء خصائص التصاق الخلية المضيفة، وانخفاض osteoconductivity، وإمكانية الرفض المناعي 14. ولذلك هناك حاجة ملحة لتقنيات تجديد العظام التي هي آمنة وفعالة ومتاحة على نطاق واسع. لدينا القدرة على تحسين استراتيجيات التجدد العظام تعتمد بشكل كبير على القدرة على تقليد الصدمة العظام خطيرة في حيوانات التجارب، ولكن الجيل واستقرار الآفات العظام كبيرة يمثل تحديا تقنيا. في معظم الحالات، يتم تحاكي صدمة خطيرة العظام الطويلة تجريبيا من خلال إنشاء الخلل التي لن تلتئم بشكل طبيعي. على الرغم من أنه يمكن أن تختلف مع الأنواع 15، ويتحقق ذلك عن طريق الإزالة الكاملة للشريحة العظام أكبر من 1.5 مرة من قطر العظام المقطع العرضي 16. ثم يستقر العظام مع زرع المعادن للحفاظ على التوجه السليم من حواف كسر والسماح للتنقل. نظرا لصغر حجمها وهشاشةالعظام الطويلة، وإنشاء مثل هذه الآفات في الفئران هم أبعد من قدرات معظم المجموعات البحثية. على هذا النحو، تقتصر نماذج عيب العظام الطويلة للفئران والحيوانات الكبيرة. ومع ذلك، والفئران تحمل مزايا البحثية الهامة في أنها يمكن المعدلة وراثيا وتربيتها كما سلالات خطر المناعية التي لا نرفض الخلايا البشرية والأنسجة. لالتطبيقات المستندة إلى خلية الإنسان والفئران خطر المناعية جذابة للعمل مع لأنها من الناحية الفسيولوجية تتميز بشكل جيد، وسهلة المنزل، وفعالة من حيث التكلفة، وسهولة تحليلها شعاعيا وتشريحيا. من الأهمية بمكان أن الفئران خطر المناعية لا يرفض الخلايا من مختلف الأنواع بما في ذلك البشر. يسمح صغر حجمها أيضا اختبار أعداد صغيرة جدا من الخلايا أو مجلدات من السقالات التجريبية في تطبيقات العظام. تم الإبلاغ عن عدة نماذج تقويم العظام الفئران التي تحمل درجات مختلفة من الاستقرار العظام 17،18. تلك التنفسيمللي التي تؤدي إلى مستويات عالية جدا من الاستقرار، مثل المثبتات الخارجية وتأمين لوحات تلتئم في الغالب من قبل التحجر داخل الغشاء على الرغم من أن الشفاء غضروفي وقد تم الإبلاغ عن 19. في المقابل، تلك التي تسمح بعض المتناهية الصغر و / أو الحركة الماكرو، مثل تلك التي تستخدم المسامير النخاعية غير المثبتة أو جزئيا ثابتة، وشفاء بشكل عام مع غلبة التحجر غضروفي 20،21. اتحاد تأخر أو عيوب غير نقابية من العظام الطويلة من الصعب بشكل خاص لتحقيق في الفئران بسبب مستوى إضافي من الاستقرار المطلوب. ومع ذلك، فقد تم الإبلاغ عن عدد من النهج، بما في ذلك دبابيس النخاعية مع المسامير المتشابكة، وتأمين لوحات والمثبتات الخارجية 22. هذه الأنظمة تعمل بشكل جيد عموما، ولكن نظرا تصميم معقد من أنها يمكن أن يكون تحديا تقنيا لتثبيت. على سبيل المثال، جارسيا وآخرون. 23 وضعت على المتشابكة نظام دبوس أنيق للاستخدام في الفئران، ولكن ينطوي على إجراء شقوق في اثنين الموقع مستقلالصورة وتعديل واسع من عظم الفخذ لاستيعاب الدبابيس. وأجريت هذه الإجراءات تحت المجهر تشريح. هنا، نحن تصف دبوس النخاع الفخذ بسيط مع طوق المركزي تهدف إلى منع إغلاق عجز العظم 3 مم وكذلك ترسيم حواف الأصلية للعيب. في حين دبوس لم يحدد حتى العظم نفسه، وتحجيم دقيق لقطر دبوس والتوسيع من نتائج تجويف النخاع في التدخل كافية للحد من الحركة الالتوائية (الشكل 1). مع اختيار دقيق من العمر الفطرية ونوع الجنس والفئران المتطابقة سلالة، والنتيجة هي تكرار للغاية التصنع غير uniondefect-22 التي يمكن تقييمها بسهولة شعاعيا. وعلاوة على ذلك المناطق ذات الاهتمام يمكن تعريف بتكاثر بعد التصوير المقطعي-حسابها الصغرى (μCT) لقياس دي نوفو تكوين العظام والمعلمات histomorphological. تم نموذج أولي من المسامير في مختبرنا باستخدام أدوات متوفرة بسهولة.<img alt="الشكل (1)" src="/files/ftp_upload/52368/52368fig1highres.jpg" /> الشكل 1: مبدأ التجريبي ملخص بياني للنموذج الخلل القطعي. يتم استئصاله وسط شريحة 3 ملم من 9-10 ملم عظم الفخذ الفئران جراحيا (يسار). يتم تمرير 3 مم طويل، 19 مقياس أنابيب الصلب الجراحية أكثر من 9 ملم طويلة، 22 G أنبوب الفولاذ المقاوم للصدأ وثابتة بمادة لاصقة في وسط الدقيق (يمين). تم تجهيز دبوس الناتجة في القنوات النخاعية ما تبقى من أجزاء القريبة والبعيدة للعظم الفخذ مع G طوق 19 استبدال جزء 3 ملم من العظم (أدناه، وسط).

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			<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.

الفئران عادة استرداد وعيه والقائمة الخلفية التنقل 5-10 دقائق بعد انسحاب التخدير. خلال 5 أيام الأولى، فمن المستحسن لإيواء الفئران بشكل فردي وإدخال تخصيب البيئي لمنع استخدام الفائض من أطرافهم. لهذا الغرض، القباني من نوع أعشاش تقلل من الحاجة لبناء العش وتشجع يستريح. وقد ?...

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.

بسيطة الحرجة الحجم فخذي عيب النموذجي في الفئران

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.

برغي دليل بسيط أسلوب للدراسات طعم أجنبي داخل الجمجمة في الفئران

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.

خلق كسور استقرت بشكل صارم لتقييم التعظم الغشائي، التعظم الهاء، أو الشفاء من العيوب الحجم الحرج

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.

إصلاح نموذج الحرجة الحجم عيب قبي طريق الدهنية المشتقة من الخلايا اللحمية المقطوع 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.

طريقة بسيطة لتركيب الانبوبة الرئة ماوس

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.

الفخذ نخاع العظم الطموح في الفئران لايف

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

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study model. As the use of mouse models are often a vital step in preclinical drug discovery and drug development1-8, it is necessary to design a system to introduce drugs into mice in a uniform, reproducible manner. Ideally, the system should permit the collection of blood samples at regular intervals over a set time course. The ability to measure drug concentrations by mass-spectrometry, has allowed investigators to follow the changes in plasma drug levels over time in individual mice1, 9, 10. In this study, paclitaxel was introduced into transgenic mice as a continuous arterial infusion over three hours, while blood samples were simultaneously taken by retro-orbital bleeds at set time points. Carotid artery infusions are a potential alternative to jugular vein infusions, when factors such as mammary tumors or other obstructions make jugular infusions impractical. Using this technique, paclitaxel concentrations in plasma and tissue achieved similar levels as compared to jugular infusion. In this tutorial, we will demonstrate how to successfully catheterize the carotid artery by preparing an optimized catheter for the individual mouse model, then show how to insert and secure the catheter into the mouse carotid artery, thread the end of the catheter out through the back of the mouse&#8217;s neck, and hook the mouse to a pump to deliver a controlled rate of drug influx. Multiple low volume retro-orbital bleeds allow for analysis of plasma drug concentrations over time.

This method was developed with the goal of delivering a steady drug solution via the carotid artery, to assess the pharmacokinetics of novel drugs in mouse models.

الحقن الشريان السباتي لوأضافا الدوائية وتحليل Taxanes في الفئران

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

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.

والفئران نموذج من عودة التضيق الشرياني: الجوانب الفنية من الفخذ إصابة سلك

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.

إنشاء القطاعي فخذي الحرجة حجم عيب النموذجي في الفئران استقرت بواسطة لوحة تثبيت طرفي العظم

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.

والمتكرر الارتجاج إصابات الرأس النموذجي في الفئران

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

Induction of Accelerated Atherosclerosis in Mice: The "Wire-Injury" Model

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. Moreover, by rupture of the defective vascular wall, atherosclerosis induces occlusive thrombus formation, which represents the main cause of myocardial infarction or stroke and the most frequent cause of death. Despite the advances in the cardiovascular field, many questions remain unanswered, and additional basic research is essential to improve our understanding of the molecular mechanisms during atherosclerosis and its effects. Due to limited clinical studies, there is a need for representative animal models recreating atherosclerotic conditions such as neointima formation after stent implantation, balloon angioplasty, or endarterectomy. Since the mouse presents many advantages and is the most frequently used model for studying molecular processes, the current study proposes an invasive procedure of endothelial denudation, also known as the wire-injury model, which is representative of the human condition of neointima formation in arteries after revascularization procedures.

This study describes an invasive procedure for the induction of accelerated atherosclerosis in mice. In comparison to other methods using electric- or cryo-induced injury, mechanical-induced injury mimics the human condition of restenosis after revascularization therapies and is ideal for the study of the molecular mechanisms involved.

تحريض تصلب الشرايين المتسارع في الفئران: نموذج "الأسلاك- إصابة"

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. ...