Cardiovascular disease is a major cause of mortality affecting human beings. In recent years, gene therapy has emerged as a promising approach for treating and curing damaged cardiovascular tissues, capable of restoring the heart from a nonfunctional state to a normal state. In the treatment of cardiovascular diseases, AAV viral vectors have advantages over plasmids, lentiviral and adenoviral vectors. They can effectively, persistently, stably and safely express exogenous genes in the heart. Therefore, AAV viral vectors are mostly used for related therapies. Numerous animal experiments and clinical data have shown that carrying therapeutic genes via AAV has positive effects on cardiovascular diseases such as heart failure, myocardial ischemia, and cardiac remodeling after myocardial infarction. When using rAAV for cardiac research, the correct selection of AAV serotypes, promoters and injection methods are essential for the successful execution of experiments.

Selection of serotypes｜AAV

There are various AAV serotypes, and in cardiac studies, AAV1, AAV2, AAV6, AAV8, and AAV9 have good infection efficiency in the heart, with AAV8 and AAV9 being better than the other serotypes, and AAV9 is currently recognized as a highly efficient and specific targeting of the cardiac expression of the viral vector serotype. Although AAV8 was also effective in infection, it did not spread as well as AAV9. The AAV serotypes that target the myocardium in clinical practice are generally 1, 6, and 9, of which AAV1 and AAV6 are both effective in targeting cardiac and skeletal muscle cells, whereas AAV9 is more inclined to infect the myocardium.

At Creative Biogene, we offer a range of tissue-specific adeno-associated virus (AAV) particles that can be used in heart-related disease research. Our AAV have been carefully designed to contain cardiac-specific promoters (αMHC, cTNT) that efficiently and selectively infect cardiac cells, providing an invaluable resource for the study of cardiac-associated diseases at the molecular, cellular, and tissue levels.

View more of our tissue specific AAV particles.

Selection of promoters｜AAV

The common broad-spectrum promoters CMV and CAG can be well expressed in the heart, and the CMV promoter can be chosen if the specificity requirement is not high, and its transduction efficiency to the heart is high. To realize specific expression of target genes in the heart, it is recommended to choose heart-specific promoters, such as cTNT, αMHC, Des, MLC2v, etc., among which cTNT and αMHC promoters are more widely used.

Selection of injection method｜AAV

AAV can be injected into the heart in a variety of ways, myocardial origin injection, coronary artery injection, jugular vein injection and caudal vein injection can deliver the drug to the heart, and usually myocardial origin injection and caudal vein injection are more frequently reported in the literature.

• Myocardial injection

Myocardial injection is the direct injection of drugs into cardiac tissues by means of a syringe or similar device, mostly by in situ multipoint injection, with an injection volume of 20 μL/site and 3-5 points of injection, usually injecting a total viral load of more than 10E11VG. Compared with vascular injection, the direct myocardial injection method is more specific and can achieve efficient transduction of the region of interest, but the expression is not uniform and only in the area around the injection site.

• Intravenous injection

The drug is delivered to the heart by intravenous injection from the side of the tail after immobilization of the mouse. Tail vein injection is easy to perform, but is slightly less specific than myocardial injection. The volume of injection is around 100 μL -200 μL, and a higher viral titer is usually required because the vector is widely distributed throughout the body and the injected agent is diluted in the blood volume.

• Jugular vein injection

For jugular vein injection, an incision is made in the neck of anesthetized mice and the jugular vein is isolated and the carrier is injected into the jugular vein with a 31G needle. This method requires surgery, is more demanding to operate, and is injurious to the animal. For jugular vein injection, the total amount of virus injected is approximately 20 μL-100 uL.

• Coronary injection

Paracoronary injection involves the direct injection of vectors through a catheter, enabling minimally invasive delivery of cardioselective vectors. This method offers significant advantages and has been widely used in clinical and experimental gene therapy studies. However, the endothelium acts as a barrier for the vector to reach the cardiomyocytes, resulting in a relatively low transduction efficiency. This can be improved by continuously injecting small doses or using certain agents, such as adenosine, histamines, or nitroglycerin, to increase the permeability of the vascular bed. Retrograde coronary injection is a reverse infusion of pressure regulation by blocking the coronary sinus, which is more effective compared to a downstream injection. However, this method requires temporary interruption of coronary blood flow to eliminate the effect of collateral flow on pressure. Although transduction is efficient, it is more traumatic and dangerous.

Treatment of Heart Failure

In the medical field, heart failure is a challenging disease to treat due to its high morbidity and mortality rates. Gene therapy has brought hope to patients with advanced heart failure by strengthening myocardial contractility.

• Case 1: Therapies that incorporate the rAAV-mediated cardiac sarcoplasmic reticulum Ca2+-ATPase2a gene (SERCA2a) have been clinically effective by the mechanism shown below.

Fig. 1 Targeting the calmodulin SERCA2a in the treatment of heart failure.

• Case 2: Expression of the SERCA2a gene in a model of chronic heart failure reduces myocardial sarcoplasmic reticulum calcium leakage and decreases ventricular arrhythmias.

Fig. 2 Restoration of SERCA2a protein levels in the cardiac sarcoplasmic reticulum by transduction of SERCA2a gene expression by Ad or AAV9 was found to improve left ventricular function using pressure-volume analysis.

• Case 3: Inhibition of miR-25 improves myocardial contractility in hearts with heart failure.

Figure 3. miR-25 carried by the AAV9 vector directly targets SERCA2a and regulates the dynamics of contractile calcium ions.

Treatment of Myocardial Infarction

Myocardial infarction (MI) is a disease in which atherosclerosis of the coronary arteries causes thrombosis and blockage of the branches of the coronary arteries, which results in necrosis of a part of the myocardium without blood supply. Myocardial cell death is the pathologic basis of myocardial infarction, and acute myocardial infarction is characterized by both myocardial cell necrosis and apoptosis. In recent years, gene therapy for myocardial infarction through angiogenesis has become a popular research field both at home and abroad, and gene therapy on myocardial growth factor is one of the most successful cases of human gene therapy.

• Case 1: Co-expression of VEGF and angiopoietin in a porcine model of myocardial infarction promotes angiogenesis and cardiomyocyte proliferation while reducing apoptosis.

Figure 4. myocardial perfusion and cardiac function

• Case 2: AAV-mediated overexpression of the tissue hypoxia regulatory element CD151 has effects in mice with localized cardiac ischemia.CD151 plays an important role in integrin-promoted cell migration and angiogenesis signaling.

Figure 5. Overexpression of CD151 increases the density of small arteries and capillaries.