Name: Inducing Myointimal Hyperplasia Versus Atherosclerosis in Mice: An Introduction of Two Valid Models (Video) | JoVE Journal
Uploaded: 2014-05-14T13:00:00.000+00:00
Duration: 8 min 34 s
Description: Learn two validated mouse models for studying aortic narrowing through distinct mechanisms: balloon catheterization inducing myointimal hyperplasia and dietary manipulation of APOE null mice creating atherosclerotic plaques.

ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

The authors thank Christiane Pahrmann for technical assistance. S.S. received funding from the Deutsche Forschungsgemeinschaft (DFG) (SCHR992/3-2 and SCHR992/4-2).

All animal work should be carried out according to relevant animal care guidelines. Obtain institutional approval for animal work prior to beginning protocol.
1. Myointimal Hyperplasia Model (A)
For the intimal hyperplasia model, purchase C57BL/6J (Stock number 000664, C57BL/6J) mice at the age of 8 weeks weighing approximately 25 g. House mice for these experiments under conventional conditions; feed mice standard mouse chow and provide water ad libitum.
<ol>
	<li>Mouse preparation: Introduce the mouse to an anesthetic state with isoflurane (2%) utilizing an induction chamber.
	<ol>
		<li>Remove the abdominal hair using a hair trimmer, lay the mouse on its back and cover its nose and mouth with the facemask to assure the anesthesia.</li>
		<li>With Provo-Iodine, disinfect the abdominal area universally, followed by 80% ethanol, in two cycles.</li>
		<li>Assure the absence of reflexes by pinching the hind limb to monitor the sufficient depth of anesthesia.</li>
		<li>Separate the skin and muscle along the linea alba in two steps to expose&#160;the abdominal organs.</li>
		<li>Lay the intestines in a saline moisturized glove. Wrap the glove around the intestines to keep them moist.</li>
		<li>Attentively clear away the fatty tissue sheeting the abdominal aorta. Expose the infrarenal aorta down to its bifurcation, taking care not to injure any branch vessels.</li>
		<li>First, place a microsurgical clamp on the proximal, infrarenal aorta to shut down the blood flow. Now, apply a second distal clamp next to the aortic bifurcation.</li>
		<li>When the blood flow is interrupted, make a small incision into the aorta close to the proximal clamp.</li>
		<li>Insert the balloon-tipped catheter, which has been moisturized with 0.9% saline, and advance towards the distal clamp of the aorta.</li>
		<li>For endothelial injury, denude the aorta by inflating the balloon carefully and slowly pulling the catheter into the direction of the proximal clamp.</li>
		<li>Repeat the denudation maneuver twice.</li>
		<li>Remove the catheter and close the aortic incision using 10-0 prolene running sutures.</li>
		<li>Carefully open the distal clamp. In case of bleeding at the aortotomy site, close the clamp again, locate the bleeding and place additional interrupted stitches, as necessary.</li>
		<li>If no bleeding can be observed, slowly open the proximal clamp.</li>
		<li>An aortic pulse should be visible distally from the incision. Now the intestines can be arranged back in situ.</li>
		<li>Rinse the abdominal cavity utilizing pre-warmed sterile saline.</li>
		<li>Close the abdominal wall beginning with the muscle layer using 6-0 prolene running sutures.</li>
		<li>Next adapt the skin under the use of 5-0 prolene performing running sutures.</li>
		<li>During the time the mouse is still anesthetized, apply 4-5 mg/kg Carprofen via a subcutaneous injection.</li>
		<li>Add Metamizol to the drinking water (50 mg Metamizol per 100 ml) for pain control and continue for 3 days post surgery. Monitor animal throughout recovery. 
		Note: The observation period for this model is 28 days.</li>
	</ol>
	</li>
</ol>
2. Atherosclerosis Model (B)
For the atherosclerosis model, purchase apolipoprotein E-deficient (ApoE -/-) mice (Stock number 002052, B6.129P2-Apoetm1Unc/J) at an age of&#160;4 weeks, and then fed for 4-6 months. House mice for these experiments under conventional conditions; feed mice high lipid western diet upon arrival (Harlan Laboratories TD.88137) and provide water ad libitum for a duration of 4 to 6 months.
<ol>
	<li>Mouse preparation:
	<ol>
		<li>Feed the ApoE -/- mice with western diet beginning at 4 weeks&#160;of age. Dietary manipulation should begin directly upon arrival.</li>
		<li>Maintain western diet throughout duration of experiment.</li>
	</ol>
	</li>
</ol>
3. Analysis
<ol>
	<li>Myointimal hyperplasia model (A) 
	The region of interest in this model is the &#8220;lesion area&#8221;. Options for analysis include luminal obliteration, I/M ratio (intima/media ratio), maximal myointima thickness, myointimal area, Trichrome staining for fibrosis, and H&#38;E staining for mononuclear cells (as illustrated in Figures 1A-1C). According to the scientific question numerous other stainings like Picrosirius red for collagen, immunohistochemistry, and confocal methods can be applied.</li>
	<li>Atherosclerosis model (B) The regions of interest in this model are the aortic valve, aortic arch as well as the descending aorta. Options for analysis: Sudan red staining for lipids and trichrome staining of aortic valve, aortic arch and descending aorta (as illustrated in Figure 1D). 
	This model also yields a variety of other staining options like for example Oil Red O for neutral lipids, Picrosirius red, immunohistochemistry, and confocal methods.
	<ol>
		<li>Sudan red staining: Perform the Sudan red staining on freshly harvested aortic tissue.
		<ol>
			<li>Remove the diet 6h before harvesting.</li>
			<li>Introduce the mouse to an anesthetic state with isoflurane (2%) utilizing an induction chamber. Assure the absence of reflexes by pinching the hind limb to monitor the sufficient depth of anesthesia.</li>
			<li>Open the chest; cut off the apex of the heart and perfuse via the left ventricle, first with saline (3 ml), followed by 4% PFA (3 ml) through the aortic root to fix the aorta.</li>
			<li>Free the aorta from the root to its bifurcation. Try to remove fatty tissue as much as possible.</li>
			<li>Cut off the heart and the aortic root, and store them in 4% PFA for histology.</li>
			<li>For histology the aortic root has to carefully be placed in the paraffin block in an upright position to allow the cutting of the aortic root generating slides that display all 3 valvular leaflets.</li>
			<li>Cut one end of the aortic tree and fix it onto the silicone layer with fine pins. Open the aorta longitudinally and fix the aortic tissue.</li>
			<li>Rinse the tissue with 50% ethanol. Immerse the tissue with Sudan III staining solution for about 15 min.</li>
			<li>Rinse lightly with 50% ethanol to remove the excess Sudan III. (Note: 50% ethanol may wash the dye away.) Rinse with dH2O, and take photos for further analysis.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>

<ol><li>Manuel, D. G., Leung, M., Nguyen, K., Tanuseputro, P., Johansen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Burden+of+cardiovascular+disease+in+Canada%5BTitle%5D)+AND+%22Can+J+Cardiol%22%5BJournal%5D)">Burden of cardiovascular disease in Canada.</a> Can J Cardiol. 19, 997-1004 (2003).</li>
<li>ER, O. B., Ma, X., Simard, T., Pourdjabbar, A., Hibbert, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Pathogenesis+of+neointima+formation+following+vascular+injury%5BTitle%5D)+AND+%22Cardiovasc+Hematol+Disord+Drug+Targets%22%5BJournal%5D)">Pathogenesis of neointima formation following vascular injury.</a> Cardiovasc Hematol Disord Drug Targets. 11, 30-39 (2011).</li>
<li>Ross, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+pathogenesis+of+atherosclerosis%3A+a+perspective+for+the+1990s%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">The pathogenesis of atherosclerosis: a perspective for the 1990s.</a> Nature. 362, 801-809 (1993).</li>
<li>Meng, X., Zhang, F., Valji, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ultrasound+guidance+for+the+creation+of+a+small+animal+model+of+aortic+injury%5BTitle%5D)+AND+%22J+Vasc+Interv+Radiol%22%5BJournal%5D)">Ultrasound guidance for the creation of a small animal model of aortic injury.</a> J Vasc Interv Radiol. 22, 1193-1197 (2011).</li>
<li>Deuse, T., Koyanagi, T., Erben, R. G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Sustained+inhibition+of+epsilon+protein+kinase+C+inhibits+vascular+restenosis+after+balloon+injury+and+stenting%5BTitle%5D)+AND+%22Circulation%22%5BJournal%5D)">Sustained inhibition of epsilon protein kinase C inhibits vascular restenosis after balloon injury and stenting.</a> Circulation. 122, 170-178 (2010).</li>
<li>Schrepfer, S., Deuse, T., Sultan, K. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Inhibition+of+restenosis+development+after+mechanical+injury%3A+a+new+field+of+application+for+malononitrilamides%5BTitle%5D)+AND+%22Cardiology%22%5BJournal%5D)">Inhibition of restenosis development after mechanical injury: a new field of application for malononitrilamides.</a> Cardiology. 108, 128-137 (2007).</li>
<li>Tao, M., Mauro, C. R., Yu, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+simplified+murine+intimal+hyperplasia+model+founded+on+a+focal+carotid+stenosis%5BTitle%5D)+AND+%22Am+J+Pathol%22%5BJournal%5D)">A simplified murine intimal hyperplasia model founded on a focal carotid stenosis.</a> Am J Pathol. 182, 277-287 (2013).</li>
<li>Kumar, A., Lindner, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Remodeling+with+neointima+formation+in+the+mouse+carotid+artery+after+cessation+of+blood+flow%5BTitle%5D)+AND+%22Arterioscler+Thromb+Vasc+Biol%22%5BJournal%5D)">Remodeling with neointima formation in the mouse carotid artery after cessation of blood flow.</a> Arterioscler Thromb Vasc Biol. 17, 2238-2244 (1997).</li>
<li>Hua, X., Deuse, T., Michelakis, E. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Human+internal+mammary+artery+%28IMA%29+transplantation+and+stenting%3A+a+human+model+to+study+the+development+of+in-stent+restenosis%5BTitle%5D)+AND+%22J+Vis+Exp%22%5BJournal%5D)">Human internal mammary artery (IMA) transplantation and stenting: a human model to study the development of in-stent restenosis.</a> J Vis Exp. , 3663(2012).</li>
<li>Daugherty, A., Whitman, S. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Quantification+of+atherosclerosis+in+mice%5BTitle%5D)+AND+%22Methods+Mol+Biol%22%5BJournal%5D)">Quantification of atherosclerosis in mice.</a> Methods Mol Biol. , (2003).</li>
<li>Nakashima, Y., Plump, A. S., Raines, E. W., Breslow, J. L., Ross, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((ApoE-deficient+mice+develop+lesions+of+all+phases+of+atherosclerosis+throughout+the+arterial+tree%5BTitle%5D)+AND+%22Arterioscler+Thromb%22%5BJournal%5D)">ApoE-deficient mice develop lesions of all phases of atherosclerosis throughout the arterial tree.</a> Arterioscler Thromb. 14, 133-1340 (1994).</li>
<li>Zhang, Y., Li, L., You, J., Cao, J., Fu, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+7-difluoromethyl-5%2C+4%27-dimethoxygenistein+on+aorta+atherosclerosis+in+hyperlipidemia+ApoE%28-%2F-%29+mice+induced+by+a+cholesterol-rich+diet%5BTitle%5D)+AND+%22Drug+Des+Devel+Ther%22%5BJournal%5D)">Effect of 7-difluoromethyl-5, 4'-dimethoxygenistein on aorta atherosclerosis in hyperlipidemia ApoE(-/-) mice induced by a cholesterol-rich diet.</a> Drug Des Devel Ther. 7, 233-242 (2013).</li>
<li>Rudolph, T. K., Rudolph, V., Edreira, M. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Nitro-fatty+acids+reduce+atherosclerosis+in+apolipoprotein+E-deficient+mice%5BTitle%5D)+AND+%22Arterioscler+Thromb+Vasc+Biol%22%5BJournal%5D)">Nitro-fatty acids reduce atherosclerosis in apolipoprotein E-deficient mice.</a> Arterioscler Thromb Vasc Biol. 30, 938-9345 (2010).</li>
<li>Rondelet, B., Kerbaul, F., Motte, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Bosentan+for+the+prevention+of+overcirculation-induced+experimental+pulmonary+arterial+hypertension%5BTitle%5D)+AND+%22Circulation%22%5BJournal%5D)">Bosentan for the prevention of overcirculation-induced experimental pulmonary arterial hypertension.</a> Circulation. 107, 1329-1335 (2003).</li>
<li>Zeibig, S., Li, Z., Wagner, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+the+oxLDL+binding+protein+Fc-CD68+on+plaque+extension+and+vulnerability+in+atherosclerosis%5BTitle%5D)+AND+%22Circ+Res%22%5BJournal%5D)">Effect of the oxLDL binding protein Fc-CD68 on plaque extension and vulnerability in atherosclerosis.</a> Circ Res. 108, 695-703 (2011).</li>
<li>Painter, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Myointimal+hyperplasia%3A+pathogenesis+and+implications.+2.+Animal+injury+models+and+mechanical+factors%5BTitle%5D)+AND+%22Artificial+organs%22%5BJournal%5D)">Myointimal hyperplasia: pathogenesis and implications. 2. Animal injury models and mechanical factors.</a> Artificial organs. 15, 103-118 (1991).</li>
<li>Nitschke, Y., Weissen-Plenz, G., Terkeltaub, R., Rutsch, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Npp1+promotes+atherosclerosis+in+ApoE+knockout+mice%5BTitle%5D)+AND+%22Journal+of+cellular+and+molecular%22%5BJournal%5D)">Npp1 promotes atherosclerosis in ApoE knockout mice.</a> Journal of cellular and molecular. 15, 2273-2283 (2011).</li>
</ol>

The authors have nothing to disclose.

Although both diseases result in similar symptoms in the patient, the underlying mechanisms of plaque development and therefore the treatment approaches are very different2. In different forms of arterial stenosis the clinical findings in the patients as well as the timeframe of plaque development depend on the underlying mechanism.
Depending on the clinical findings in the patient, different methods of sample analysis must be performed5,15. It is necessary to adapt the a...

In industrialized nations ischemic heart disease remains a leading cause of mortality1. The causes for coronary artery stenosis are manifold and include myointimal hyperplasia as well as atherosclerosis, with revascularization remaining the most common treatment strategy2. Research models that clearly distinguish between the mechanistic pathways of intimal hyperplasia and atherosclerosis are essential to investigate the pathobiological and pathophysiological processes in arterial plaque development. In this video two different mouse models used to study either the development of myointimal hyperplasia or atherosclerosis are introduced.
Myointimal hyperplasia is postulated to form via the &#8220;response-to-injury&#8221; paradigm originally described for atherosclerosis3. The mechanical disruption of the endothelial layer leads to an intense remodeling enhanced by paracrine effects. There are several different animal models commonly used to study myointimal hyperplasia. Some groups use metal devices in the ascending aorta of rats4. The aortic denudation model performed with a balloon catheter is also commonly used in laboratory rats5,6. The intimal hyperplasia model performed in laboratory mice7 implements ligation of the carotid artery and induces myointimal lesions8. In our lab, the abdominal aortic denudation model &#8211; to study the development of myointimal hyperplasia &#8211; as well as a humanized stent restenosis model9 are commonly used. This video emphasizes that choosing the proper experimental animal model is crucial for mechanistic or pathophysiologic studies of arterial stenosis.
Myointimal hyperplasia models must be distinguished from atherosclerosis models. For the latter, apolipoprotein E-deficient (ApoE -/-) mice in combination with western diet are commonly used to induce atherosclerotic lesions10,11,12,13. Examples for the induction of both of these types of myointimal disease in mice are shown here, along with different model-adapted options to analyze vessel stenosis14.

<table><tbody><tr><td>Thyoglycollate Broth 3%</td><td>Fluka</td><td>70157</td><td>powder</td></tr><tr><td>PFA 4%</td><td>Electron Microscopy Sciences</td><td>#157135S</td><td>20%</td></tr><tr><td>Sudan III staining solution</td><td>Sigma Aldrich</td><td>S4131</td><td>powder</td></tr><tr><td>mouse C57BL/6J</td><td>Jackson Laboratories</td><td>Stock # 0006664</td><td></td></tr><tr><td>mouse ApoE -/-</td><td>Jackson Laboratories</td><td>Stock #002052</td><td></td></tr><tr><td>Western Diet</td><td>Harlan Laboratories</td><td>TD.88137</td><td></td></tr><tr><td>hair clipper</td><td>WAHL</td><td>8786-451A ARCO SE</td><td></td></tr><tr><td>Forene</td><td>Abbott</td><td></td><td>Isoflurane</td></tr><tr><td>microsurgical clamp</td><td>Fine Science Tools</td><td>18055-04</td><td>Micro-Serrefine - 4 mm</td></tr><tr><td>clamp applicator</td><td>Fine Science Tools</td><td>18056-14</td><td></td></tr><tr><td>catheter</td><td></td><td></td><td></td></tr><tr><td>10-0 prolene suture</td><td>Ethicon</td><td>788G</td><td></td></tr><tr><td>6-0 prolene suture</td><td>Ethicon</td><td>8709H</td><td></td></tr><tr><td>5-0 prolene suture</td><td>Ethicon</td><td>EH7229H</td><td></td></tr><tr><td>Rimadyl</td><td>Pfizer</td><td></td><td>Carprofen</td></tr><tr><td>Metacam 1.5 mg/ml</td><td>Boehringer Ingelheim</td><td></td><td>Metamizol</td></tr></tbody></table>

inducing myointimal hyperplasia versus atherosclerosis in mice: an introduction of two valid models

Inducing Myointimal Hyperplasia Versus Atherosclerosis in Mice: An Introduction of Two Valid Models

Various in vivo laboratory rodent models for the induction of artery stenosis have been established to mimic diseases that include arterial plaque formation and stenosis, as observed for example in ischemic heart disease. Two highly reproducible mouse models&#160;&#8211;&#160;both resulting in artery stenosis but each underlying a different pathway of development&#160;&#8211; are introduced here. The models represent the two most common causes of artery stenosis; namely one mouse model for each myointimal hyperplasia, and atherosclerosis are shown. To induce myointimal hyperplasia, a balloon catheter injury of the abdominal aorta is performed. For the development of atherosclerotic plaque, the ApoE -/- mouse model in combination with western fatty diet is used. Different model-adapted options for the measurement and evaluation of the results are named and described in this manuscript. The introduction and comparison of these two models provides information for scientists to choose the appropriate artery stenosis model in accordance to the scientific question asked.

Figure 1 shows different analysis options and the induced disease state for both models. For the intimal hyperplasia model, the analysis of representative cross sections in trichrome staining is shown. From these cross sections, results like I/M ratios, maximum plaque thickness, maximum intimal thickness in the lumen, luminal obliteration in percent, as well as plaque area can be measured and calculated.
To evaluate the outcome of the atherosclerosis model, all methods named a...

This video shows two models of intimal plaque development in murine arteries and emphasizes the differences in myointimal hyperplasia and atherosclerosis.

Various in vivo laboratory rodent models for the induction of artery stenosis have been established to mimic diseases that include arterial plaque ...

The overall goal of the following experiment is to introduce two valid animal models leading to narrowing of the aorta via different mechanisms. The first model is balloon catheterization of the aorta to create the development of myo intimal hyperplasia. First, the infrarenal abdominal aorta is exposed and clamped.Then the aorta is opened with a transverse incision and the balloon catheter is introduced. The endothelium of the aorta is then denuded with the inflated catheter and the surgery is completed. The second model is a dietary manipulation of transgenic APOE null mice to create atherosclerotic plaques.These mice are housed under standard conditions and fed a high lipid western diet for four to six months. Ultimately, histological analysis of the surgically induced myo intimal hyperplasia and the dietary restriction model shows that they result in different luminal narrowing. Today we'll show you two different models to induce luminal narrowing the mouse model using a balloon catheter to induce vessel injury and the arterial sclerosis model.Using transgenic mice in the Western diet, The balloon catheter model can help answer key questions in the stenosis field. The Transgenic homozygous upper E mutant mouse model is used to study the pathophysiology of atherosclerosis. For the intimal hyperplasia model, obtain C 57 black six J mice that weigh about 25 grams.Maintain them under normal conditions with a normal diet. After anesthetizing the mouse with 2%isof fluorine, trim off the abdominal hair, provide carprofen by subcutaneous injection for analgesia. Next, alternatively, disinfect the skin with Provo iodine and 80%ethanol in two cycles.Now check for an absence of reflexes using a toe pinch. Start the surgery by separating the skin and muscle along the linear alba. Next, wrap the intestines in a glove moistened with saline, then attentively, clear away the fatty tissue, sheeting the abdominal aorta.Now expose the infrarenal aorta down to its bifurcation. Taking care not to injure any branch vessels. Now clamp the aorta.First place a microsurgical clamp on the proximal infrarenal aorta to shut down the blood flow. Second, apply a second distal clamp next to the aortic bifurcation. When the blood flow is interrupted, make a small incision into the aorta close to the proximal clamp.Insert the catheter towards the distal clamp of the aorta to make an endothelial injury. Denude the aorta by inflating the catheter balloon carefully and slowly pulling the catheter into the direction of the proximal clamp. Repeat this, maneuver twice for a total of three aorta inflations.Now remove the catheter and close the aortic incision using 10 au proline running sutures. With sutures in place, carefully open the distal clamp if there is bleeding at the AOR autotomy site. Close the clamp again, locate the bleeding and stop it with additional interrupted stitches.When there is no bleeding, slowly open the proximal clamp. An aortic pulse should be visible distal to the incision. Now arrange the intestines in C two.Next, close the abdominal wall beginning with the muscle layer using six OTT proline running sutures. Then close the skin with five OTT sutures and transfer the mouse back to its cage. Over the first three postoperative days, provide metrazole to the drinking water for pain control.Monitor the animal throughout its recovery, which takes about 28 days. For the atherosclerosis model, obtain four week old A POE deficient mice. House these mice under standard conditions with a high lipid western diet for four to six months.Maintaining this diet throughout the experiment. Six hours before harvesting the aorta, take away access to food. Then prepare the mouse for surgery as described in the previous section.Start the surgery by opening the chest. Next, cut off the apex of the heart and perfuse it under gentle pressure with three milliliters of 4%Paraform aldehyde through the aortic root to fix the aorta. Then free the aorta from the root to its bifurcation.Try to remove as much of the fatty tissue as possible. Next, cut off the aortic arch and store it in 4%para formaldehyde. Now transfer the heart and aortic tree to a silicone line dish with PBS in the dish.Cut one end of the aortic tree and fix it onto the silicone layer with fine pins. Open the aorta longitudinally and allow the aortic tissue to lie in PBS at room temperature for a few minutes. Then rinse the tissue once with 50%ethanol and immerse it into a bath of Sudan.Three, staining solution for about 15 minutes. After staining, quickly rinse the tissue with 50%ethanol for a few seconds to remove just the excess Sudan. Three over rinsing will remove too much stain, immediately rinse the tissue with distilled water and proceed with imaging For the myo intimal hyperplasia model, denuded mouse aortas were harvested, paraffin embedded, and stained with chrome Im ratios were calculated from the tri chrome stain sections by measuring intima and media thickness.Using computer analysis followed by calculation of the ratio, maximum plaque thickness was measured as the maximum intimal thickness in the lumen by computer morpho tree. The percentage of luminal obliteration was calculated as the ratio of the new inner lumen from the former lumen. The plaque area was calculated by subtracting the new inner lumen from the former lumen.The same analyses were performed on tri chrome stained cross sections of the atherosclerosis model at the aortic valve, the aortic arch, and the descending aorta for just the atherosclerosis model. The descending aorta was opened longitudinally and stained with Sudan red. The plaque area and the total aortic area were measured with the aid of a computer For analysis of both models.Other methods such as histopathology and the confocal immunofluorescence microscopy can be performed in order to answer questions like luminal oblation, morphology, and the cellularity. Thank you for watching and good luck with your experiments.

inducing-myointimal-hyperplasia-versus-atherosclerosis-mice-an

Research

JoVE Journal

Medicine

We describe protocols for our mouse graft arteriosclerois (GA) models which involve interposition of a mouse vessel segment into a recipient of the same inbred strain. By backcrossing additional genetic changes into the vessel donor, the model can assess the effect of specific genes on GA.

Mouse Models for Graft Arteriosclerosis

We describe procedures to quantify and characterize atherosclerotic lesions in mouse models using precision sectioning of the aortic sinus and ascending aorta combined with histochemical and immunohistochemical analysis.

Quantitative Analysis and Characterization of Atherosclerotic Lesions in the Murine Aortic Sinus

Animal models have proven to be invaluable tools in defining host and pathogen specific mechanisms that contribute to the development of chronic inflammation. Here we describe a mouse model of oral infection with the human pathogen Porphyromonas gingivalis and detail methodologies to assess the progression of inflammation at local and systemic sites.

A Mouse Model for Pathogen-induced Chronic Inflammation at Local and Systemic Sites

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 neointima for studies of restenosis.

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

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.

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

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

Murine models of atherosclerosis are useful tools to investigate pathogenic pathways on a molecular level, but require standardized quantification of lesion development. This protocol describes an optimized method to determine lesion size in the major arterial vessels including the aortic root, aortic arch, and brachiocephalic artery.

Quantification of Atherosclerosis in Mice

This article demonstrates an efficient surgical approach to establish acute ischemia in mice with a small incision. This approach can be applied by most research groups without any laboratory upgrades.

A Modified Surgical Model of Hind Limb Ischemia in ApoE-/- Mice using a Miniature Incision

Here we present a murine model for femoral artery anastomosis, offering researchers a valuable animal model to study and simulate vascular anastomotic stenosis. This development is crucial for advancing our understanding of the pathophysiology underlying this condition and facilitating more accurate and effective research into vascular diseases.

Development of a Murine Model for Femoral Artery Anastomotic Stenosis

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.

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

Learn two validated mouse models for studying aortic narrowing through distinct mechanisms: balloon catheterization inducing myointimal hyperplasia and dietary manipulation of APOE null mice creating atherosclerotic plaques.

Inducing Myointimal Hyperplasia Versus Atherosclerosis in Mice: An Introduction of Two Valid Models

Explore More Videos