Cardiovascular Research Center, Mount Sinai School of Medicine
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Ishikawa, K., Ladage, D., Tilemann, L., Fish, K., Kawase, Y., Hajjar, R. J. Gene Transfer for Ischemic Heart Failure in a Preclinical Model. J. Vis. Exp. (51), e2778, doi:10.3791/2778 (2011).
Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine their efficacy and safety.
Gene therapy using viral vectors is one of the most promising approaches for treating cardiac diseases. Viral delivery of various different genes by changing the carrier gene has immeasurable therapeutic potential.
In this video, the full process of an animal model of heart failure creation followed by gene transfer is presented using a swine model. First, myocardial infarction is created by occluding the proximal left anterior descending coronary artery. Heart remodeling results in chronic heart failure. Unique to our model is a fairly large scar which truly reflects patients with severe heart failure who require aggressive therapy for positive outcomes. After myocardial infarct creation and development of scar tissue, an intracoronary injection of virus is demonstrated with simultaneous nitroglycerine infusion. Our injection method provides simple and efficient gene transfer with enhanced gene expression. This combination of a myocardial infarct swine model with intracoronary virus delivery has proven to be a consistent and reproducible methodology, which helps not only to test the effect of individual gene, but also compare the efficacy of many genes as therapeutic candidates.
1. Myocardial infarct creation
Common complications during and after the myocardial infarct creation procedure are coronary dissection, puncture site hematoma, arrhythmia, and heart failure. A refine technique and thorough monitoring is required to prevent and manage these complications.
2. Virus Injection
3. Representative Case
Figure 1 shows the left ventriculography at 1 month after the myocardial infarct creation. The left ventricular wall motion is significantly impaired with an ejection fraction (EF) of 31%. Figure 2 shows the cross sections of the left ventricle at 1 month. A large scar from the base to the apex of the anterior wall is seen. The average EF at 1 month after myocardial infarction in this model is 36% and the average end diastolic volume is 83 ml, whereas the average of those in the same sized animal without a myocardial infarction is 68% and 42 ml, respectively. Figure 3 shows the expression of beta-gal in the left ventricular border zone area 1 month after the injection of adeno-associated virus (2 x 1012 DNAse resitant particles of rAAV1 CMV Lac-Z). An enhanced expression was found in the virus injected area.
Figure 1. Ejection fraction measured from left ventriculography 1 month after myocardial infarct creation. Left ventricle is dilated to 92.5 ml and anterior wall motion is significantly impaired.
Figure 2. The cross sections of the left ventricle at 1 month after myocardial infarct creation, stained by triphenyl tetrazolium chloride to delineate the scar area. A large scar from the base to the apex of the anterior wall is seen.
Figure 3. Beta-gal expression induced by adeno-associated virus (2 x 1012 DNAse resistant particles of rAAV1 CMV Lac-Z). An enhanced expression is found at the border zone area between the infarction and the normal tissue, 1 month after the intracoronary virus injection. Right lower picture shows the negative control, where only normal saline was injected in the coronary arteries using the same protocol.
Among the various causes of heart failure, ischemia is by far the most frequent in etiology. To overcome this serious disease, various therapeutic approaches are being developed. Careful evaluations of therapies are required before these promising therapies are can be applied to the clinics. Therefore, establishment of an ischemic heart failure model that represents the clinical conditions of patients is critical to development of heart failure therapies. Our swine myocardial infarct model meets these translational requirements.
To induce heart failure by creating a myocardial infarct, the infarct area needs to be large. Our model has a larger scar compared to the previous studies which also used an endovascular myocardial infarct model1. We have found that administering Amiodarone and maintaining blood pressure over 80 mmHg during the coronary occlusion is essential to increase both survival rate and the infarct area. While Isoflurane is an effective anesthetic for this procedure1-2, we found that with Propofol, higher blood pressure can be maintained. In addition, Isoflurane is reported to have more cardio-protective effects compared to Propofol3, and this could lead to a decreased size of the infarction.
Cardiac gene therapy is now one of the most promising approaches to heart failures, and there are variety of strategies to transfer the gene. Above all, intracoronary injection of viral vectors carrying a gene, is a simple and less invasive technique. Various attempts are made to increase gene expression. Our lab has found that infusing nitroglycerin during the virus injection increases the gene expression (I. Karakikes, unpublished).
Gene transfer of SERCA2a using adeno-associated viral vector has resulted in positive outcomes in humans4. Further improvements to cardiac gene therapy are expected with additional carrier genes as well as vector refinements. Our delivery method provides simple and efficient gene transfer with enhanced gene expression, and can be readily translated the clinic.
All authors have no commercial associations that might pose as a conflict of interest in connection with the manuscript.
This work is supported by Leducq Foundation through the Caerus network (RJH), by NIH R01 HL093183, HL088434, HL071763, HL080498, HL083156, and P20HL100396 (RJH). DL was supported by the German Research Foundation. We would like to thank Lauren Leonardson for her technical assistance in this project.
|Telazol||Fort Dodge Animal Health||0856-9050-49|
|Potassium acetate||American Regent Inc.||2053-25|
|Nitroglycerine||American Regent Inc.||4810-25|
|Salix (Furosemide)||Intervet Inc.||710461|
|5 Fr Hockey stick catheter||Cordis||556-278-0L|
|7 Fr Hockey stick catheter||Cordis||778-278-00|
|Coronary wire||Abbott Laboratories||1044588|
|Coronary balloon||Abbott Laboratories||1009447-08|
|8Fr sheath||Terumo Medical Corp.||RSS801|