ברצוננו להודות למתקן FCCC בעלי החיים במעבדה על תמיכתם בפרויקט זה. אנו מודים וולף מעבדות, Inc על עזרת בניתוח רמות פקליטקסל בפלזמה וברקמות. עבודה זו נתמכה על ידי המכון הלאומי לבריאות מעניקה K01CA120091 לEHB, וCA06927 פוקס צ&#39;ייס במרכז הסרטן.

<ol>
	<li>Gallo, J. M., Li, S., Guo, P., Ma Reed, K., J, <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Effect+of+P-Glycoprotein+on+Paclitaxel+Brain+and+Brain+Tumor+Distribution+in+Mice.">The Effect of P-Glycoprotein on Paclitaxel Brain and Brain Tumor Distribution in Mice.</a> Cancer Research. 63 (16), 5114-5117 (2003).</li><li>Sonnichen, D. S., Relling, M. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+Pharmacokinetics+of+Paclitaxel.">Clinical Pharmacokinetics of Paclitaxel.</a> Clinical Pharmacokinetics. 27 (4), 256-269 (1994).</li><li>Gianni, L., 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=Nonlinear+Pharmacokinetics+and+Metabolism+of+Paclitaxel+and+Its+Pharmacokinetic+/+Pharmacodynamic+Relationship+in+Humans.">Nonlinear Pharmacokinetics and Metabolism of Paclitaxel and Its Pharmacokinetic / Pharmacodynamic Relationship in Humans.</a> Journal Clinical Oncology. 13 (1), 180-190 (1995).</li><li>Sparreboom, A., Van Tellingen, O., Nooijen, W. J., Beijnen, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonlinear+Pharmacokinetics+of+Paclitaxel+in+Mice+Results+from+the+Pharmaceutical+Vehicle+Cremophor+EL.">Nonlinear Pharmacokinetics of Paclitaxel in Mice Results from the Pharmaceutical Vehicle Cremophor EL.</a> Cancer Research. 56 (9), 2112-2115 (1996).</li><li>Sparreboom, A., Van Tellingen, O., Nooijen, W. J., Beijnen, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+paclitaxel+and+metabolites+in+mouse+plasma,+tissues,+urine+and+faeces+by+semi-automated+reversed-phase+high-performance+liquid+chromatography.">Determination of paclitaxel and metabolites in mouse plasma, tissues, urine and faeces by semi-automated reversed-phase high-performance liquid chromatography.</a> Journal of Chromatography B: Biomedical Sciences and Applications. 664, 383-391 (1995).</li><li>Bradshaw-Peirce, E. L., Eckhardt, S. G., Gustafson, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Physiologically+Based+Pharmacokinetic+Model+of+Docetaxel+Disposition:+from+Mouse+to+Man.">A Physiologically Based Pharmacokinetic Model of Docetaxel Disposition: from Mouse to Man.</a> Clinical Cancer Research. 13, 2768-2778 (2007).</li><li>Eiseman, J. L., 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=Plasma+Pharmacokinetics+and+Tissue+Distribution+of+Paclitaxel+in+CD2F1+Mice.">Plasma Pharmacokinetics and Tissue Distribution of Paclitaxel in CD2F1 Mice.</a> Cancer Chemotherapy and Pharmacology. 34 (6), 465-471 (1994).</li><li>Schinkel, A. H., 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=Normal+Viability+and+Altered+Pharmacokinetics+in+Mice+Lacking+mdr1-type+(drug-transporting)+P-glycoproteins.">Normal Viability and Altered Pharmacokinetics in Mice Lacking mdr1-type (drug-transporting) P-glycoproteins.</a> Proc. Nat. Acad. Sci. USA. 94, 4028-4033 (1997).</li><li>Fraser, I. J., Dear, G. J., Plumb, R., L&#8217;Affineur, M., Fraser, D., Skippen, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Use+of+Capillary+High+Performance+Liquid+Chromatography+with+Electrospray+Mass+Spectrometry+for+the+Analysis+of+Small+Volume+Blood+Samples+from+Serially+Bled+Mice+to+Determine+the+Pharmacokinetics+of+Early+Discovery+Compounds.">The Use of Capillary High Performance Liquid Chromatography with Electrospray Mass Spectrometry for the Analysis of Small Volume Blood Samples from Serially Bled Mice to Determine the Pharmacokinetics of Early Discovery Compounds.</a> Rapid Communications in Mass Spectrometry. 13 (23), 2366-2375 (1999).</li><li>Bateman, K. P., 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=Reduction+of+Animal+Usage+by+Serial+Bleeding+of+Mice+for+Pharmacokinetic+Studies:+Application+of+Robotic+Sample+Preparation+and+Fast+Liquid+Chromatography+&#8211;+Mass+Spectrometry.">Reduction of Animal Usage by Serial Bleeding of Mice for Pharmacokinetic Studies: Application of Robotic Sample Preparation and Fast Liquid Chromatography &#8211; Mass Spectrometry.</a> Journal of Chromatography B: Biomedical Sciences and Applications. 754 (1), 245-251 (2001).</li><li>Berndt, K., Vogel, J., Buehler, C., Vogt, P., Born, W., Fuchs, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+method+for+repeated+drug+infusion+into+the+femoral+artery+of+mice.">A new method for repeated drug infusion into the femoral artery of mice.</a> J. Am. Assoc. Lab Animal Sci. 51 (6), 825-831 (2012).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Squibb+&amp;+Sons,+L.L.C..">Squibb &amp; Sons, L.L.C..</a> Drug Information for TAXOL (Paclitaxel) Injection. , (2011).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abraxis+BioScience+L.L.C.">Abraxis BioScience L.L.C.</a> Full Prescribing Information ABRAXANE for Injectable suspension (paclitaxel protein-bound particles for injectable suspension) (albumin-bound). , (2013).</li><li>Fergusson, G., Ethier, M., Zarrouki, B., Font&#233;s, G., Poitout, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Model+of+Chronic+Nutrient+Infusion+in+the+Rat.">A Model of Chronic Nutrient Infusion in the Rat.</a> J. Vis. Exp. (78), (2013).</li></ol>

יש לי המחברים אין לחשוף.

עירוי בעורק התרדמה הוא טכניקה משמעותית במחקר זה של הפרמקוקינטיקה פקליטקסל. עירוי בעורק התרדמה הוא שיטה להפצה במהירות בסמים לאורך מערכת הדם 14. עירוי שעה 3 הוא קרוב יותר חקיין של ממשל הקליני של תרופות כגון taxanes מאשר זריקות בולוס. הניתוח יכול להתבצע באופן אמין על י?...

עירוי תרופות דרך עורק התרדמה ניתן לבצע באופן אמין וreproducibly על ידי אופטימיזציה של ציוד וטכניקה. ההליך הוא לא מסובך, למרות שזה דורש בקרה טובה ותשומת לב לפרטים. טיפול ומיומנות מעולים יש צורך לבודד את העורק הראשי ולהכניס את הצנתר, אשר בדרך כלל ניתן לרכוש באמצעות תרגול. ניתוח על ידי טכנאי מנוסה לא יעלה על שעה אחת. לאחר ניתוח מוצלח, העכבר צריך להופיע נורמלי ובריא (אם כי העכבר עשוי להגיב לעירוי התרופה בפועל), ותרופה (ים) עשויה להיות מנוהלת במינון רציף, אחיד בשליטה. דגימות דם חייבות להילקח מאתר אחר מאשר העורק הראשי; דימומי רטרו מסלולית הוכיחו קל לאסוף ומניח את דעת לניתוח של ריכוזי תרופה. צנתרים של גודל אופטימלי וצורה הם נכס לא יסולא בפז בביצוע עירוי מוצלח 11. מצאנו commerciall זמין הצנתריםy לעתים קרובות להיות גדול ו / או גמיש מדי כדי לאפשר גישה נוחה לעורק הראשי עכבר מדי. זה הוכיח עדיף על צנתרים אופנה מצינורות פוליאתילן המשמשים לחיבור העכבר למזרק העירוי. כך, כל צינורות, המחברים והמחטים היו בממדים עולים בקנה אחד, שפשוט הרכבה עירוי. השימוש בטכניקה זו, אין צורך לדחוף את קצה הצנתר לתוך העורק מעבר לנקודה שבה עדיין גלוי, וזרימת דם לעורק הראשי לא שוחזרה עד לאחר קטטר תחילה מאובטח. הפעולה זו מפחיתה את הסכנות של ניקוב העורק או שיש קטטר נדחפו החוצה על ידי הלחץ הגבוה של זרימת דם. עיצוב קטטר במסמך זה אינו כולל &quot;להקפיץ&quot; להחזיק אותו במקום, ולכן הבטחת קטטר גם עם תפרים ואספלנית היא בראש סדר עדיפויות. חליטות עשויות להיות עדיפות על זריקות בולוס iv המשותפים, כטובות יותר חקיין של ממשל הקליני שלתרופות כגון taxanes 3, 12, 13. הטכניקה המתוארת כאן פותחו במקור כדי לאפשר לעירוי לתוך מודלים של עכברים שבי גישה לוריד הצוואר או הירך נמנעה בגלל צמיחת גידול החלב ו / או כלי דם יתר של אזור ההחדרה. שיטה זו עשויה לעתים להיות מתאימה גם בעכברים ללא גידול, למרות שבידוד וcatheterizing התרדמה הוא מעט יותר פולשנית, שמצאנו אותו עדיף על הצוואר, כי הנטייה של קיר הצוואר לקרוע הביאה הוספות וכישלונות כדי להשלים נכשלו יותר כמובן זמן 3 שעות. בעוד שהתוצאות מופיעות כאן הן מC57BL / 6-י (ב- הוליד בית) עכברים, השתמשנו בטכניקה זו כדי להחדיר בהצלחה פקליטקסל לכמה זנים של עכברים, כוללים FVB ומעורבים-זנים, כדי לעקוב הפרמקוקינטיקה במודלים של עכברים שהופעלו transgenically ללמטה לווסת פונקציות טרנספורטר סלולריות. דגימות הדם ורקמה שנאספו הראו רמות צפויות של paclitaxאל, בטווח של הרמות שנראו לאחר חליטות הצוואר 1. טכניקה זו ניתן לצפות כי עבודה לא פחות טובה במודלים של עכברים אחרים ועם פתרונות עירוי אחרים.

<table border="0" cellpadding="0" cellspacing="0" width="846">
	<tbody>
		<tr height="25">
			<td height="25" style="height:25px;width:293px;">Table of specific reagents and equipment:</td>
			<td style="width:185px;">Source:</td>
			<td style="width:111px;">Catalog Number</td>
			<td style="width:259px;">Comments</td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Polyethylene tubing 0.024&#8221;OD X 0.011&#8221;ID&#160;</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">PE10</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Stainless steel port plug (30 gauge)</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">NB-30</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">2 Stainless steel connector plugs (28 gauge)</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">PP-28</td>
			<td style="width:259px;">Slightly larger than PE tubing ID, to fit snugly and keep a tight seal.</td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">3 Blunted needles (30 gauge)</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">C-30</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">3 1cc syringes</td>
			<td style="width:185px;">Becton, Dickinson and Co.</td>
			<td style="width:111px;">309659</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Sterile 0.9% Saline solution</td>
			<td style="width:185px;">Hospira</td>
			<td style="width:111px;">0409-7984-37</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Cath-Loc HGS Heparin/Glycerol Solution&#160;</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">HGS</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Silk suture</td>
			<td style="width:185px;">Braintree Scientific, Inc.</td>
			<td style="width:111px;">SUT-S&#160;113</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Vanna Scissors (micro-scissors)</td>
			<td style="width:185px;">World Precision Instruments</td>
			<td style="width:111px;">14122</td>
			<td style="width:259px;">This model has a curved tip, but straight-tip scissors work as well.</td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Hartman Mesquito Hemostatic Forceps</td>
			<td style="width:185px;">World Precision Instruments</td>
			<td style="width:111px;">501705</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Betadine Swabsticks</td>
			<td style="width:185px;">Perdue Products L.P.</td>
			<td style="width:111px;">BSWS1S</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Bupivacaine</td>
			<td style="width:185px;">Hospira</td>
			<td style="width:111px;">0409-1160-01</td>
			<td style="width:259px;">May be replaced with Lidocaine, or similar local anesthesia.</td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Paclitaxel</td>
			<td style="width:185px;">LC Laboratories</td>
			<td style="width:111px;">P-9600</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Methanol</td>
			<td style="width:185px;">Sigma-Aldrich</td>
			<td style="width:111px;">32213</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">Micro-Hematocrit Capillary Tubes, Heparinized</td>
			<td style="width:185px;">Fisher Scientific</td>
			<td style="width:111px;">22-362-566</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">&#160;Capillary Tube Sealant&#160;</td>
			<td style="width:185px;">Fisher Scientific</td>
			<td style="width:111px;">02-678</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="70">
			<td height="70" style="height:70px;width:293px;">C57BL/6J mice</td>
			<td style="width:185px;">Fox Chase Cancer Center, Laboratory Animal Facility in-house-bred</td>
			<td style="width:111px;"></td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:293px;">API 4000 Q-Trap mass spetrometer</td>
			<td style="width:185px;">Applied Biosystems</td>
			<td style="width:111px;"></td>
			<td style="width:259px;"></td>
		</tr>
	</tbody>
</table>

carotid artery infusions for pharmacokinetic and pharmacodynamic analysis of taxanes in mice

כאשר מציעים את השימוש של תרופה, שילוב תרופות, או אספקת סמים למערכת רומן, יש להעריך את הפרמקוקינטיקה של התרופה במודל המחקר. ככל שהשימוש במודלים של עכברים הוא לעתים קרובות צעד חיוני בגילוי תרופות פרה-קליני ופיתוח תרופות 1-8, יש צורך לתכנן מערכת להציג תרופות לעכברים במדים, אופן לשחזור. באופן אידיאלי, המערכת צריכה לאפשר איסוף דגימות דם במרווחים זמן קבוע על מסלול זמן מוגדר. היכולת למדוד ריכוזי תרופה על ידי ספקטרומטר מסה, אפשרה לחוקרים לעקוב אחר השינויים ברמות תרופה בפלזמה לאורך זמן בעכברים בודדים 1, 9, 10. במחקר זה, פקליטקסל הוכנס עכברים הטרנסגניים כעירוי דם מתמשך על פני שלוש שעות, ואילו דגימות דם נלקחו בו זמנית על ידי דימומי רטרו מסלולית בנקודות זמן מוגדר. חליטות עורק התרדמה הן חלופה פוטנציאלית לחליטות וריד הצוואר, כאשר גורמים כמוגידולי החלב או מכשולים אחרים לעשות חליטות הצוואר מעשיות. שימוש בטכניקה זו, ריכוזי פקליטקסל בפלזמה וברקמות השיגו רמות דומות בהשוואה לעירוי הצוואר. במדריך זה, אנו נדגים כיצד לצנתר את העורק הראשי בהצלחה על ידי הכנת קטטר מותאם למודל העכבר הבודד, ולאחר מכן להציג כיצד להוסיף ולהבטיח את הצנתר לתוך עורק הצוואר עכבר, חוט סוף הצנתר דרך חזרה צווארו של העכבר, ולחבר את העכבר למשאבה כדי לספק קצב מבוקר של זרם סמים. דימומי רטרו מסלולית נפח נמוך מרובים מאפשרים ניתוח של ריכוזי תרופה בפלזמה לאורך זמן.

<html> הפצת פקליטקסל כדלקמן דפוסים צפויים במשטר מינון 3 שעות של עירוי במהירות גבוהה 15 דקות, ואחרי עירוי במהירות נמוכה 165 דקות. איור 1 מציג השוואה של ריכוזי וריד הצוואר-עירוי פקליטקסל הפלזמה ועורק-חליטות התרדמה. ריכוזי פקליטק?...

Watch this Scientific Journal Video about Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice at JoVE.com

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

שיטה זו פותחה במטרה לספק פתרון תרופתי קבוע באמצעות העורק הראשי, כדי להעריך את הפרמקוקינטיקה של תרופות חדשניות במודלים של עכברים.

חליטות עורק הראשי לpharmacokinetic וpharmacodynamic ניתוח של taxanes בעכברים

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 ...

carotid-artery-infusions-for-pharmacokinetic-pharmacodynamic-analysis

Research

Education

JoVE Core

JoVE Journal

Pharmacokinetics and Pharmacodynamics

Medicine

Unit 1

Pharmacokinetic Models

SCIENTISTS IN ACTION

22.7K Views.  Fox Chase Cancer Center.  שיטה זו פותחה במטרה לספק פתרון תרופתי קבוע באמצעות העורק הראשי, כדי להעריך את הפרמקוקינטיקה של תרופות חדשניות במודלים של עכברים. 

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

A Model of Disturbed Flow-Induced Atherosclerosis in Mouse Carotid Artery by Partial Ligation and a Simple Method of RNA Isolation from Carotid Endothelium

Despite the well-known close association, direct evidence linking disturbed flow to atherogenesis has been lacking. We have recently used a modified version of carotid partial ligation methods [1,2] to show that it acutely induces low and oscillatory flow conditions, two key characteristics of disturbed flow, in the mouse common carotid artery. Using this model, we have provided direct evidence that disturbed flow indeed leads to rapid and robust atherosclerosis development in Apolipoprotein E knockout mouse [3]. We also developed a method of endothelial RNA preparation with high purity from the mouse carotid intima [3]. Using this mouse model and method, we found that partial ligation causes endothelial dysfunction in a week, followed by robust and rapid atheroma formation in two weeks in a hyperlipidemic mouse model along with features of complex lesion formation such as intraplaque neovascularization by four weeks. This rapid in vivo model and the endothelial RNA preparation method could be used to determine molecular mechanisms underlying flow-dependent regulation of vascular biology and diseases. Also, it could be used to test various therapeutic interventions targeting endothelial dysfunction and atherosclerosis in considerably reduced study duration.

This describes a partial carotid ligation surgery, which causes disturbed flow conditions and subsequent atherosclerosis development (in two weeks) with intraplaque neo-vascularization (in four weeks) in the mouse common carotid artery. We also describe a novel method of RNA isolation from the carotid intima, providing high purity endothelial RNA.

המודל של טרשת עורקים Flow-induced מופרע על העורק הראשי עכבר ידי קשירת חלקי שיטה פשוטה של ​​בידוד RNA מן התרדמה האנדותל

Despite the well-known close association, direct evidence linking disturbed flow to atherogenesis has been lacking. We have recently used a modified ...

Modified Technique for Coronary Artery Ligation in Mice

Myocardial infarction (MI) is one of the most important causes of mortality in humans1-3. In order to improve morbidity and mortality in patients with MI we need better knowledge about pathophysiology of myocardial ischemia. This knowledge may be valuable to define new therapeutic targets for innovative cardiovascular therapies4. Experimental MI model in mice is an increasingly popular small-animal model in preclinical research in which MI is induced by means of permanent or temporary ligation of left coronary artery (LCA)5. In this video, we describe the step-by-step method of how to induce experimental MI in mice.
The animal is first anesthetized with 2% isoflurane. The unconscious mouse is then intubated and connected to a ventilator for artificial ventilation. The left chest is shaved and 1.5 cm incision along mid-axillary line is made in the skin. The left pectoralis major muscle is bluntly dissociated until the ribs are exposed. The muscle layers are pulled aside and fixed with an eyelid-retractor. After these preparations, left thoracotomy is performed between the third and fourth ribs in order to visualize the anterior surface of the heart and left lung. The proximal segment of LCA artery is then ligated with a 7-0 ethilon suture which typically induces an infarct size ~40% of left ventricle. At the end, the chest is closed and the animals receive postoperative analgesia (Temgesic, 0.3 mg/50 ml, ip). The animals are kept in a warm cage until spontaneous recovery.

The surgical procedure used to induce experimental myocardial infarction in mice begins with left thoracotomy between the third and the fourth ribs in order to visualize the anterior surface of the heart and left lung. The left coronary artery is ligated, the chest is closed and the mouse is allowed to recover spontaneously.

טכניקה שונה לקשירת עורקים הכליליים בעכברים

Myocardial infarction (MI) is one of the most important causes of mortality in humans1-3. In order to improve morbidity and mortality in patients with MI ...

Implantation of a Carotid Cuff for Triggering Shear-stress Induced Atherosclerosis in Mice

It is widely accepted that alterations in vascular shear stress trigger the expression of inflammatory genes in endothelial cells and thereby induce atherosclerosis (reviewed in 1 and 2). The role of shear stress has been extensively studied in vitro investigating the influence of flow dynamics on cultured endothelial cells 1,3,4 and in vivo in larger animals and humans 1,5,6,7,8. However, highly reproducible small animal models allowing systematic investigation of the influence of shear stress on plaque development are rare. Recently, Nam et al. 9 introduced a mouse model in which the ligation of branches of the carotid artery creates a region of low and oscillatory flow. Although this model causes endothelial dysfunction and rapid formation of atherosclerotic lesions in hyperlipidemic mice, it cannot be excluded that the observed inflammatory response is, at least in part, a consequence of endothelial and/or vessel damage due to ligation.
In order to avoid such limitations, a shear stress modifying cuff has been developed based upon calculated fluid dynamics, whose cone shaped inner lumen was selected to create defined regions of low, high and oscillatory shear stress within the common carotid artery 10. By applying this model in Apolipoprotein E (ApoE) knockout mice fed a high cholesterol western type diet, vascular lesions develop upstream and downstream from the cuff. Their phenotype is correlated with the regional flow dynamics 11 as confirmed by in vivo Magnetic Resonance Imaging (MRI) 12: Low and laminar shear stress upstream of the cuff causes the formation of extensive plaques of a more vulnerable phenotype, whereas oscillatory shear stress downstream of the cuff induces stable atherosclerotic lesions 11. In those regions of high shear stress and high laminar flow within the cuff, typically no atherosclerotic plaques are observed.
 
In conclusion, the shear stress-modifying cuff procedure is a reliable surgical approach to produce phenotypically different atherosclerotic lesions in ApoE-deficient mice.

The constricting cuff presented in this article is designed to induce atherosclerosis in the murine common carotid artery. Due to the conical shape of its inner lumen the implanted cuff generates well-defined regions of low, high and oscillatory shear stress triggering the development of atherosclerotic lesions of different inflammatory phenotypes.

השרשה של קאף הראש מפעילה שאר מתח טרשת עורקים המושרה עכברים

It is widely accepted that alterations in vascular shear stress trigger the expression of inflammatory genes in endothelial cells and thereby induce ...

A Murine Model of Stent Implantation in the Carotid Artery for the Study of Restenosis

Despite the considerable progress made in the stent development in the last decades, cardiovascular diseases remain the main cause of death in western countries. Beside the benefits offered by the development of different drug-eluting stents, the coronary revascularization bears also the life-threatening risks of in-stent thrombosis and restenosis. Research on new therapeutic strategies is impaired by the lack of appropriate methods to study stent implantation and restenosis processes. Here, we describe a rapid and accessible procedure of stent implantation in mouse carotid artery, which offers the possibility to study in a convenient way the molecular mechanisms of vessel remodeling and the effects of different drug coatings.

A model of stent implantation in mouse carotid artery is described. Compared to other similar methods, this procedure is very rapid, simple and accessible, offering the possibility to study in a convenient way the vascular wall reaction to different drug-eluting stents and the molecular mechanisms of restenosis.

דגם Murine של השתלת תומכן בעורק הראשי לחקר restenosis

Despite  the considerable progress made in the stent development in the last decades,  cardiovascular diseases remain the main cause of death in western ...

Adult Mouse Venous Hypertension Model: Common Carotid Artery to External Jugular Vein Anastomosis.

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model creating an artificial fistula between the common carotid artery (CCA) and the external jugular vein (EJV) has been widely described and proved technically feasible. This construct provokes a consistent cerebral venous hypertension (CVH), and therefore has helped studying the contribution of venous hypertension to formation, clinical symptoms, and prognosis of brain AVMs and dural AVFs. Equivalent mice models have been only scarcely described and have shown trouble with stenosis of the fistula. An established murine model would allow the study of not only pathophysiology but also potential genetic therapies for these cerebrovascular diseases.
We present a model of arteriovenous fistula that produces a durable intracranial venous hypertension in the mouse. Microsurgical anastomosis of the murine CCA and EJV can be difficult due to diminutive anatomy and frequently result in a non-patent fistula. In this step-by-step protocol we address all the important challenges encountered during this procedure. Avoiding excessive retraction of the vein during the exposure, using 11-0 sutures instead of 10-0, and making a carefully planned end-to-side anastomosis are some of the critical steps. Although this method requires advanced microsurgical skills and a longer learning curve that the equivalent in the rat, it can be consistently developed.
This novel model has been designed to integrate transgenic mouse techniques with a previously well-established experimental system that has proved useful to study brain AVMs and dural AVFs. By opening the possibility of using transgenic mice, a broader spectrum of valid models can be achieved and genetic treatments can also be tested. The experimental construct could also be further adapted to the study of other cerebrovascular diseases related with venous hypertension such as migraine, transient global amnesia, transient monocular blindness, etc.

We describe a method for creating a reliable model of cerebral venous hypertension in the adult mouse. This model has been widely described and tested in the rat. This new counterpart in the mice opens the possibility of using genetic modified animals and thereby broadens the applications of the model.

למבוגרים עכבר ורידי יתר לחץ דם דגם: עורק תרדמה משותף לחיצוני הצוואר וריד השקה.

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model ...

Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical options are available for a small proportion of stroke patients in the acute phase. Current research is intensively searching for novel therapeutic strategies and is increasingly focusing on the sub-acute and chronic phase after stroke because more patients might be eligible for therapeutic interventions in a prolonged time window. These delayed mechanisms include important pathophysiological pathways such as post-stroke inflammation, angiogenesis, neuronal plasticity and regeneration. In order to analyze these mechanisms and to subsequently evaluate novel drug targets, experimental stroke models with clinical relevance, low mortality and high reproducibility are sought after. Moreover, mice are the smallest mammals in which a focal stroke lesion can be induced and for which a broad spectrum of transgenic models are available. Therefore, we describe here the mouse model of transcranial, permanent coagulation of the middle cerebral artery via electrocoagulation distal of the lenticulostriatal arteries, the so-called &#8220;coagulation model&#8221;. The resulting infarct in this model is located mainly in the cortex; the relative infarct volume in relation to brain size corresponds to the majority of human strokes. Moreover, the model fulfills the above-mentioned criteria of reproducibility and low mortality. In this video we demonstrate the surgical methods of stroke induction in the &#8220;coagulation model&#8221; and report histological and functional analysis tools.

Various murine models of middle cerebral artery occlusion (MCAO) are widely used in experimental brain research. Here, we demonstrate the model of transcranial permanent distal MCAO which produces consistent cortical infarction of a size corresponding to damage imposed by the majority of human ischemic strokes.

דוגמנות שבץ בעכברים: קרישה קבועה של עורקי דיסטלי התיכון מוחין

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical ...

Catheterization of the Carotid Artery and Jugular Vein to Perform Hemodynamic Measures, Infusions and Blood Sampling in a Conscious Rat Model

The success of a small animal model to study critical illness is, in part, dependent on the ability of the model to simulate the human condition. Intra-tracheal inoculation of a known amount of bacteria has been successfully used to reproduce the pathogenesis of pneumonia which then develops into sepsis. Monitoring hemodynamic parameters and providing standard clinical treatment including infusion of antibiotics, fluids and drugs to maintain blood pressure is critical to simulate routine supportive care in this model but to do so requires both arterial and venous vascular access. The video details the surgical technique for implanting carotid artery and common jugular vein catheters in an anesthetized rat. Following a 72 hr recovery period, the animals will be re-anesthetized and connected to a tether and swivel setup attached to the rodent housing which connects the implanted catheters to the hemodynamic monitoring system. This setup allows free movement of the rat during the study while continuously monitoring pressures, infusing fluids and drugs (antibiotics, vasopressors) and performing blood sampling.

Vascular accesses to measure hemodynamics, provide fluids and perform blood sampling are important to any small animal model study. We present a technique for implanting catheters into the carotid artery and the common jugular vein in an anesthetized rat for connecting to a system to perform monitoring, infusions and sampling.

צנתור של עורק התרדמה וצוואר הווריד לבצע צעדים המודינמית, חליטות ודגימת דם במודל עכברוש מודע

The success of a small animal model to study critical illness is, in part, dependent on the ability of the model to simulate the human condition. ...

Vascular Balloon Injury and Intraluminal Administration in Rat Carotid Artery

The carotid artery balloon injury model in rats has been well established for over two decades. It remains an important method to study the molecular and cellular mechanisms involved in vascular smooth muscle dedifferentiation, neointima formation and vascular remodeling. Male Sprague-Dawley rats are the most frequently employed animals for this model. Female rats are not preferred as female hormones are protective against vascular diseases and thus introduce a variation into this procedure. The left carotid is typically injured with the right carotid serving as a negative control. Left carotid injury is caused by the inflated balloon that denudes the endothelium and distends the vessel wall. Following injury, potential therapeutic strategies such as the use of pharmacological compounds and either gene or shRNA transfer can be evaluated. Typically for gene or shRNA transfer, the injured section of the vessel lumen is locally transduced for 30 min with viral particles encoding either a protein or shRNA for delivery and expression in the injured vessel wall. Neointimal thickening representing proliferative vascular smooth muscle cells usually peaks at 2 weeks after injury. Vessels are mostly harvested at this time point for cellular and molecular analysis of cell signaling pathways as well as gene and protein expression. Vessels can also be harvested at earlier time points to determine the onset of expression and/or activation of a specific protein or pathway, depending on the experimental aims intended. Vessels can be characterized and evaluated using histological staining, immunohistochemistry, protein/mRNA assays, and activity assays. The intact right carotid artery from the same animal is an ideal internal control. Injury-induced changes in molecular and cellular parameters can be evaluated by comparing the injured artery to the internal right control artery. Likewise, therapeutic modalities can be evaluated by comparing the injured and treated artery to the control injured only artery.

This protocol uses a balloon catheter to cause an intraluminal injury on the rat carotid artery and henceforth elicit neointimal hyperplasia. This is a well-established model for studying the mechanisms of vascular remodeling in response to injury. It is also widely used to determine the validity of potential therapeutic approaches.

כלי דם בלון פגיעה וIntraluminal מנהל בעורקי תרדמת העכברוש

The carotid artery balloon injury model in rats has been well established for over two decades. It remains an important method to study the molecular and ...

Ferric Chloride-induced Thrombosis Mouse Model on Carotid Artery and Mesentery Vessel

Severe thrombosis and its ischemic consequences such as myocardial infarction, pulmonary embolism and stroke are major worldwide health issues. The ferric chloride injury is now a well-established technique to rapidly and accurately induce the formation of thrombi in exposed veins or artery of small and large diameter. This model has played a key role in the study of the pathophysiology of thrombosis, in the discovery and validation of novel antithrombotic drugs and in the understanding of the mechanism of action of these new agents. Here, the implementation of this technique on a mesenteric vessel and carotid artery in mice is presented. The method describes how to label circulating leukocytes and platelets with a fluorescent dye and to observe, by intravital microscopy on the exposed mesentery, their accumulation at the injured vessel wall which leads to the formation of a thrombus. On the carotid artery, the occlusion caused by the clot formation is measured by monitoring the blood flow with a Doppler probe.

The FeCl3 induced thrombosis model in mice is described herein. A method to monitor thrombus growth by intravital microscopy observation on a mesenteric vessel and by blood flow measurement in the carotid artery is presented.

פקקת עכבר דגם על עורק ראשי וכלי לפדר מושרה כלוריד Ferric

Severe thrombosis and its ischemic consequences such as myocardial infarction, pulmonary embolism and stroke are major worldwide health issues. The ferric ...

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves focal endothelial-selective transgenesis, thereby allowing an investigator to determine the biological roles of endothelial-expressed transgenes and to quantify the in vivo transcriptional activity of DNA sequences in large artery endothelial cells. The method uses surgical isolation of rabbit common carotid arteries and an arteriotomy to deliver a transgene-expressing viral vector into the arterial lumen. A short incubation period of the vector in the lumen, with subsequent aspiration of the lumen contents, is sufficient to achieve efficient and durable expression of the transgene in the endothelium, with no detectable transduction or expression outside of the isolated arterial segment. The method allows assessment of the biological activities of transgene products both in normal arteries and in models of human vascular disease, while avoiding systemic effects that could be caused either by targeting gene delivery to other sites (e.g. the liver) or by the alternative approach of delivering genetic constructs to the endothelium by germ line transgenesis. Application of the method is limited by the need for a skilled surgeon and anesthetist, a well-equipped operating room, the costs of purchasing and housing rabbits, and the need for expertise in gene-transfer vector construction and use. Results obtained with this method include: transgene-related alterations in arterial structure, cellularity, extracellular matrix, or vasomotor function; increases or reductions in arterial inflammation; alterations in vascular cell apoptosis; and progression, retardation, or regression of diseases such as intimal hyperplasia or atherosclerosis. The method also allows measurement of the ability of native and synthetic DNA regulatory sequences to alter transgene expression in endothelial cells, providing results that include: levels of transgene mRNA, levels of transgene protein, and levels of transgene enzymatic activity.

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

This method is to introduce a transgene into the endothelium of rabbit carotid arteries. Introduction of the transgene allows the assessment of the biological role of the transgene product either in normal arteries or disease models. The method is also useful for measuring activity of DNA regulatory sequences.

אין ויוו העברה גנטית כדי אנדותל העורק נפוצות הארנב

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves ...