המחקרים מציגים נתמכים על ידי ללב, ריאות הלאומי, ודם המכון גרנט R01-HL0921, R01-R01 DK083385 ו-HL098294 כדי HK Eltzschig, 1K08HL102267-01 ל ט Eckle, ואת הקרן לחינוך הרדמה מענקי מחקר ל ט Eckle ו HK Eltzschig, ו American Heart Association גרנט ט Eckle ו HK Eltzschig ו דויטשה Forschungsgemeinschaft (DFG) מלגת מחקר מ Koeppen. אנו מודים שלי Eltzschig לעבודת אמנות.

אין ניגודי אינטרסים הכריז.

המחקר הנוכחי מתאר טכניקה חדשנית של ביצוע IP במודל Murine שלם באמצעות מערכת תלוי במשקל ובכך להימנע מחסימה של העורקים הכליליים על ידי פקעת. למעשה, מחקר זה מדגים הגדלים האוטם לשעתקו מאוד והגנה לב על ידי ה-IP, ובכך לצמצם את השונות הקשורות בקשר מבוססי מודלים חסימה כלילית. חוקרים הרואים בלימוד cardioprotection על ידי ה-IP של עכברים עשויים להפיק תועלת ממודל זה.

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use of a hanging weight system for coronary artery occlusion in mice

מחקרים Murine של פגיעה חריפה על שטח של חקירה אינטנסיבית, כמו עכברים knockout עבור גנים שונים נעשים זמינים יותר ויותר 1-38. Cardioprotection ידי נפשית מראש איסכמי (IP) נותר שטח של חקירה אינטנסיבית. כדי להמשיך להבהיר הבסיס המולקולרי שלו, השימוש מחקרים עכבר נוקאאוט חשוב במיוחד 7, 14, 30, 39. למרות העובדה כי מחקרים קודמים כבר ביצעה בהצלחה reperfusion איסכמיה לב בעכברים, מודל זה הוא מאוד מאתגר מבחינה טכנית. במיוחד, זיהוי חזותי של העורק הכלילי, המיקום של תפר סביב כלי השיט ואת חסימה כלילית על ידי קשירת את הקשר עם כלי נתמך קשה מבחינה טכנית. בנוסף, פתיחה מחדש את הקשר עבור reperfusion לסירוגין של העורק הכלילי במהלך IP מבלי לגרום טראומה ניתוחית מוסיף אתגר נוסף. יתר על כן, אם הקשר אינו כבול reperfusion חזק, מספיק בשוגג עקב חסימה מושלמת של כלילית עשוי להשפיע על התוצאות. למעשה, זה יכול בקלות להתרחש עקב תנועה של הלב הפועם. בהתבסס על בעיות פוטנציאליות הקשורות באמצעות מערכת קשרים חסימה כלילית, אימצנו מודל שפורסמו בעבר של קרדיומיופתיה כרונית המבוסס על מערכת משקל תלוי על חסימה לסירוגין לב כלילית במהלך 39 IP. למעשה, חסימת עורקים כליליים יכולה אפוא להיות מושגת מבלי לחסום את העורקים הכליליים על ידי פקעת. יתר על כן, reperfusion של כלי השיט ניתן להשיג בקלות על ידי תמיכה משקולות תלוי אשר נמצאים לוקליזציה מרוחק מן הרקמות לב. בדקנו את המערכת באופן שיטתי, כולל וריאציה של פעמים איסכמיה ו reperfusion, גדודי נפשית מראש, טמפרטורת הגוף ועל רקע גנטי 39. בנוסף האוטם מכתים, בדקנו לב טרופונין I (cTnI) כסמן של אוטם שריר הלב במודל זה. למעשה, רמות הפלזמה של cTnI בקורלציה עם בגדלים האוטם (R2 = 0.8). לבסוף, אנחנו יכולים להראות במספר מחקרים כי טכניקה זו התשואות גדלים האוטם לשעתקו מאוד במהלך IP Murine לבין אוטם שריר הלב 6, 8, 30, 40, 41. לכן, טכניקה זו עשויה לסייע לחוקרים מי לרדוף המנגנונים המולקולריים המעורבים cardioprotection על ידי ה-IP באמצעות גישה גנטיות בעכברים עם מחיקת הגן ממוקדות. מחקרים נוספים על ה-IP לב באמצעות העכברים הטרנסגניים לשקול את הטכניקה הזו.

Watch this Scientific Journal Video about Use of a Hanging Weight System for Coronary Artery Occlusion in Mice at JoVE.com

Use of a Hanging Weight System for Coronary Artery Occlusion in Mice

מודל Murine עבור איסכמיה בשריר הלב נפשית מראש איסכמי הוא כלי מחקר חשוב מנגנוני הלב<em&gt; In vivo</em&gt;. כאן אנו מדווחים החלים קל<em&gt; באתרו</em&gt; מודל ה-IP לב באמצעות מערכת תלוי במשקל עבור חסימת העורקים הכליליים.

השימוש במערכת משקל תליה על חסימת העורקים הכליליים של עכברים

Murine studies of acute injury are an area of intense investigation, as knockout mice for different genes are becoming increasingly available 1-38. ...

use-of-a-hanging-weight-system-for-coronary-artery-occlusion-in-mice

Research

JoVE Journal

Medicine

21.5K Views.  University of Colorado Denver. מודל Murine עבור איסכמיה בשריר הלב נפשית מראש איסכמי הוא כלי מחקר חשוב מנגנוני הלב In vivo. כאן אנו מדווחים החלים קל באתרו מודל ה-IP לב באמצעות מערכת תלוי במשקל עבור חסימת העורקים הכליליים.

Video: Use of a Hanging Weight System for Coronary Artery Occlusion in Mice

Modeling Stroke in Mice - Middle Cerebral Artery Occlusion with the Filament Model

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are very limited. To meet the need for novel therapeutic approaches, experimental stroke research frequently employs rodent models of focal cerebral ischaemia. Most researchers use permanent or transient occlusion of the middle cerebral artery (MCA) in mice or rats.
Proximal occlusion of the middle cerebral artery (MCA) via the intraluminal suture technique (so called filament or suture model) is probably the most frequently used model in experimental stroke research. The intraluminal MCAO model offers the advantage of inducing reproducible transient or permanent ischaemia of the MCA territory in a relatively non-invasive manner. Intraluminal approaches interrupt the blood flow of the entire territory of this artery. Filament occlusion thus arrests flow proximal to the lenticulo-striate arteries, which supply the basal ganglia. Filament occlusion of the MCA results in reproducible lesions in the cortex and striatum and can be either permanent or transient. In contrast, models inducing distal (to the branching of the lenticulo-striate arteries) MCA occlusion typically spare the striatum and primarily involve the neocortex. In addition these models do require craniectomy. In the model demonstrated in this article, a silicon coated filament is introduced into the common carotid artery and advanced along the internal carotid artery into the Circle of Willis, where it blocks the origin of the middle cerebral artery. In patients, occlusions of the middle cerebral artery are among the most common causes of ischaemic stroke. Since varying ischemic intervals can be chosen freely in this model depending on the time point of reperfusion, ischaemic lesions with varying degrees of severity can be produced. Reperfusion by removal of the occluding filament at least partially models the restoration of blood flow after spontaneous or therapeutic (tPA) lysis of a thromboembolic clot in humans.
In this video we will present the basic technique as well as the major pitfalls and confounders which may limit the predictive value of this model.

Filamentous occlusion of the Middle cerebral artery is a common model for studying ischemic stroke in mice.

דוגמנות שבץ בעכברים - העורקים התיכון ספיגה מוחין עם דגם נימה

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are ...

Use of a Hanging-weight System for Isolated Renal Artery Occlusion

In hospitalized patients, over 50% of cases of acute kidney injury (AKI) are caused by renal ischemia 1-3. A recent study of hospitalized patients revealed that only a mild increase in serum creatinine levels (0.3 to 0.4 mg/dl) is associated with a 70% greater risk of death than in persons without any increase 1. Along these lines, surgical procedures requiring cross-clamping of the aorta and renal vessels are associated with a renal failure rates of up to 30% 4. Similarly, AKI after cardiac surgery occurs in over 10% of patients under normal circumstances and is associated with dramatic increases in mortality. AKI are also common complications after liver transplantation. At least 8-17% of patients end up requiring renal replacement therapy 5. Moreover, delayed graft function due to tubule cell injury during kidney transplantation is frequently related to ischemia-associated AKI 6. Moreover, AKI occurs in approximately 20% of patients suffering from sepsis 6.The occurrence of AKI is associated with dramatic increases of morbidity and mortality 1. Therapeutic approaches are very limited and the majority of interventional trials in AKI have failed in humans. Therefore, additional therapeutic modalities to prevent renal injury from ischemia are urgently needed 3, 7-9. 
To elucidate mechanisms of renal injury due to ischemia and possible therapeutic strategies murine models are intensively required 7-13. Mouse models provide the possibility of utilizing different genetic models including gene-targeted mice and tissue specific gene-targeted mice (cre-flox system). However, murine renal ischemia is technically challenging and experimental details significantly influence results. We performed a systematic evaluation of a novel model for isolated renal artery occlusion in mice, which specifically avoids the use of clamping or suturing the renal pedicle 14. This model requires a nephrectomy of the right kidney since ischemia can be only performed in one kidney due to the experimental setting. In fact, by using a hanging-weight system, the renal artery is only instrumented once throughout the surgical procedure. In addition, no venous or urethral obstruction occurs with this technique. We could demonstrate time-dose-dependent and highly reproducible renal injury with ischemia by measuring serum creatinine. Moreover, when comparing this new model with conventional clamping of the whole pedicle, renal protection by ischemic preconditioning is more profound and more reliable. Therefore his new technique might be useful for other researchers who are working in the field of acute kidney injury.

A precise murine model for acute kidney injury (AKI) due to ischemia is an important tool to investigate acute kidney injury and possibly find therapeutic tools to treat renal injury. The hanging weight system offers a tool for immediate and reliable renal artery occlusion and reperfusion without causing renal congestion.

השימוש במערכת תלוי במשקל עבור חסימת עורק הכליה מבודדת

In hospitalized patients, over 50% of cases of acute kidney injury (AKI) are caused by renal ischemia 1-3. A recent study of hospitalized patients ...

Use of a Hanging-weight System for Liver Ischemia in Mice

Acute liver injury due to ischemia can occur during several clinical procedures e.g. liver transplantation, hepatic tumor resection or trauma repair and can result in liver failure which has a high mortality rate1-2. Therefore murine studies of hepatic ischemia have become an important field of research by providing the opportunity to utilize pharmacological and genetic studies3-9. Specifically, conditional mice with tissue specific deletion of a gene (cre, flox system) provide insights into the role of proteins in particular tissues10-13 . Because of the technical difficulty associated with manually clamping the portal triad in mice, we performed a systematic evaluation using a hanging-weight system for portal triad occlusion which has been previously described3. By using a hanging-weight system we place a suture around the left branch of the portal triad without causing any damage to the hepatic lobes, since also the finest clamps available can cause hepatic tissue damage because of the close location of liver tissue to the vessels. Furthermore, the right branch of the hepatic triad is still perfused thus no intestinal congestion occurs with this technique as blood flow to the right hepatic lobes is preserved. Furthermore, the portal triad is only manipulated once throughout the entire surgical procedure. As a result, procedures like pre-conditioning, with short times of ischemia and reperfusion, can be easily performed. Systematic evaluation of this model by performing different ischemia and reperfusion times revealed a close correlation of hepatic ischemia time with liver damage as measured by alanine (ALT) and aspartate (AST) aminotransferase serum levels3,9. Taken together, these studies confirm highly reproducible liver injury when using the hanging-weight system for hepatic ischemia and intermittent reperfusion. Thus, this technique might be useful for other investigators interested in liver ischemia studies in mice. Therefore the video clip provides a detailed step-by-step description of this technique.

We established a novel murine model of a hanging weight system for portal triad occlusion. This technique may be useful for future investigations of ischemia in murine hepatic models.

השימוש במערכת תלייה משקל של איסכמיה כבד בעכברים

Acute liver injury due to ischemia can occur during several clinical procedures e.g. liver transplantation, hepatic tumor resection or trauma repair and ...

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 ...

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.

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 ...

Permanent Ligation of the Left Anterior Descending Coronary Artery in Mice: A Model of Post-myocardial Infarction Remodelling and Heart Failure

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It is characterized by fluid retention, shortness of breath, and fatigue, in particular on exertion. Heart failure is a growing public health problem, the leading cause of hospitalization, and a major cause of mortality. Ischemic heart disease is the main cause of heart failure.
Ventricular remodelling refers to changes in structure, size, and shape of the left ventricle. This architectural remodelling of the left ventricle is induced by injury (e.g., myocardial infarction), by pressure overload (e.g., systemic arterial hypertension or aortic stenosis), or by volume overload. Since ventricular remodelling affects wall stress, it has a profound impact on cardiac function and on the development of heart failure. A model of permanent ligation of the left anterior descending coronary artery in mice is used to investigate ventricular remodelling and cardiac function post-myocardial infarction. This model is fundamentally different in terms of objectives and pathophysiological relevance compared to the model of transient ligation of the left anterior descending coronary artery. In this latter model of ischemia/reperfusion injury, the initial extent of the infarct may be modulated by factors that affect myocardial salvage following reperfusion. In contrast, the infarct area at 24 hr after permanent ligation of the left anterior descending coronary artery is fixed. Cardiac function in this model will be affected by 1) the process of infarct expansion, infarct healing, and scar formation; and 2) the concomitant development of left ventricular dilatation, cardiac hypertrophy, and ventricular remodelling.
Besides the model of permanent ligation of the left anterior descending coronary artery, the technique of invasive hemodynamic measurements in mice is presented in detail.

Heart failure is the leading cause of hospitalization and a major cause of mortality. A model of permanent ligation of the left anterior descending coronary artery in mice is applied to investigate ventricular remodelling and cardiac dysfunction post-myocardial infarction. The technique of invasive hemodynamic measurements in mice is presented.

קשירת קבע של העורק השמאלי קדמי יורד כלילית בעכברים: דגם של Remodelling אוטם שריר לב ופוסט-אי ספיקת לב

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It ...

Ultrasound Based Assessment of Coronary Artery Flow and Coronary Flow Reserve Using the Pressure Overload Model in Mice

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow reserve (CFR) in humans. Reduced CFR is accompanied by marked intramyocardial and pericoronary fibrosis and is used as an indication of the severity of dysfunction. This study explores, step-by-step, the real-time changes measured in the coronary flow velocity, CFR and systolic to diastolic peak velocity (S/D) ratio in the setting of an aortic banding model in mice. By using a Doppler transthoracic imaging technique that yields reproducible and reliable data, the method assesses changes in flow in the septal coronary artery (SCA), for a period of over two weeks in mice, that previously either underwent aortic banding or thoracotomy. 
During imaging, hyperemia in all mice was induced by isoflurane, an anesthetic that increased coronary flow velocity when compared with resting flow. All images were acquired by a single imager. Two ratios, (1) CFR, the ratio between hyperemic and baseline flow velocities, and (2) systolic (S) to diastolic (D) flow were determined, using a proprietary software and by two independent observers. Importantly, the observed changes in coronary flow preceded LV dysfunction as evidenced by normal LV mass and fractional shortening (FS). 
The method was benchmarked against the current gold standard of coronary assessment, histopathology. The latter technique showed clear pathologic changes in the coronary artery in the form of peri-coronary fibrosis that correlated to the flow changes as assessed by echocardiography. 
The study underscores the value of using a non-invasive technique to monitor coronary circulation in mouse hearts. The method minimizes redundant use of research animals and demonstrates that advanced ultrasound-based indices, such as CFR and S/D ratios, can serve as viable diagnostic tools in a variety of investigational protocols including drug studies and the study of genetically modified strains.

Coronary flow reserve (CFR) is useful for assessment of myocardial oxygen demand and evaluation of cardiovascular risk. This study establishes a step-by-step transthoracic Doppler echocardiographic (TTDE) method for longitudinal monitoring of the changes in CFR, as measured from coronary artery in mice, under the experimental pressure overload of aortic banding.

הערכת אולטראסאונד מבוססת של עורקים הכליליים זרימה והזרימה כלילית ריזרב שימוש בדגם עומס יתר הלחץ בעכברים

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow ...

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

גישה ניתוחית עבור העורקים התיכון ספיגה מוחין ושבץ reperfusion מושרה של עכברים

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

Use of a Low-flow Digital Anesthesia System for Mice and Rats

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers utilize a syringe pump to directly administer volatile anesthetics into a gas stream. Unlike a traditional vaporizer, it can be used at very low flow rates, making it ideal for use on mice and rats. 
The equipment's capability to use low flow rates could result in a substantial cost savings due to the reduced need for anesthetic agents, compressed gas, and charcoal scavenging filters1. A lower flow rate means less waste of anesthetic gas and likely reduces the risk of anesthetic exposure to laboratory personnel. Thus, the high levels of precision and safety associated with direct injection vaporizers, along with a reduced need for anesthetic agents, compressed gas, and charcoal filters are beneficial for research requiring small animal anesthesia. 
The goal of this protocol is to demonstrate the use of a syringe-driven direct injection vaporizer as part of a digital, low-flow anesthesia system. The direct injection vaporizer is capable of accurately delivering anesthesia at very low flow rates compared to a traditional vaporizer, making it a promising alternative for controlled gas anesthetic delivery to rodents.

Here, we present a protocol to more safely and efficiently administer anesthetic gas to mice using a digital, low flow anesthesia system utilizing a syringe-driven direct injection vaporizer.

שימוש של מערכת הרדמה דיגיטלית זרימה נמוכה עבור עכברים וחולדות

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers ...

Identifying Coronary Artery Calcification on Non-gated Computed Tomography Scans

Coronary artery calcification (CAC) provides an objective measure of coronary artery disease and can readily be identified on non-gated computed tomography (CT) scans with a high correlation with gated cardiac CT scans. This standardized protocol takes a step-wise approach to not only optimizing an image for the identification of calcification but also to distinguishing CAC from other common causes of calcification in the cardiac silhouette. Recognition of CAC on non-gated CT scans helps to identify a very powerful prognostic factor that can influence therapeutic interventions or downstream diagnostic testing without requiring a gated cardiac scan. These non-gated CT scans are often acquired as part of the routine care of the patient, and this data is readily available without another dose of&#160;ionizing radiation. This protocol allows for the precise and accurate extraction of this data for the purposes of retrospective data analysis in clinical research studies, but also in the clinical evaluation and management of patients.

Here, we present a protocol to reliably and systematically identify coronary artery calcification (CAC) on non-gated computed tomography (CT) scans of the chest or abdomen. CAC provides an objective measure of coronary artery disease for both research and clinical purposes.