3.1 Heart tissue structure
Cardiomyocytes, fibroblasts, endothelial cells, and conduction systems have different expression needs.
3.2 AAV serotype selection
Systemic drug delivery is often evaluated based on AAV9, and local myocardial injection can be compared with AAV6; cardiomyocyte specificity cannot be judged based on whole-heart fluorescence alone.
3.3 Cardiac-specific promoter
cTnT/TNNT2, MYL2 or MHCK7 are commonly used in cardiac muscle; fibroblast projects should use independent regulatory elements and cell markers.
3.4 Administration method and dosage reference
Systemic vein (IV): AAV9 has natural tropism for myocardium. 1×10^11-1×10^12 vg/mouse, 100-300 μL; Inagaki confirmed that the global cardiac gene transfer of AAV9 mice after systemic administration is better than that of AAV8[3-4].
Local injection into the myocardium: focal myocardial transduction. Mouse 5×10^10-1×10^11 vg/heart, a total of 30 μL divided into 3 points (10 μL each ventricular/dorsal/side of the left ventricular wall), 31 G Hamilton, the fourth intercostal small incision was made through thoracotomy and the heart was externally injected; rat 1×10^11-5×10^11 vg/heart, 30-50 μL multiple points; dog 7-Fr endocardial 27 The G injection needle catheter enters the left ventricle through the aortic valve, and is guided by fluoroscopy + MRI fusion; pig thoracotomy is used for multi-point injection [3-5].
Coverage and limitations: IV myocardial transduction is extensive but accompanied by liver off-target; local myocardial transduction is focal around the injection point, not covering the whole heart, and is suitable for locating the infarct boundary zone. The expression peaks in 3-4 weeks, and dystrophin can be restored in the DMD model [3-5,3-6]. It is recommended to simultaneously evaluate cardiac function, tissue expression, inflammation, and liver off-targets.
3.5 Key research evidence
Gao et al. established a simple and efficient method for cardiac-specific gene manipulation by intramyocardial injection in mice [3-5]; MRI roadmap-guided transendocardial rAAV delivery restored dystrophin expression in the canine DMD model [3-6]; Inagaki et al. systematically compared the cardiac transduction efficiency of systemic administration of AAV9 and AAV8 [3-4].
3.6 References for this chapter
[3-1] Litviňuková M, et al. Cells of the adult human heart. Nature. 2020;588:466-472. [H]
[3-2] Wang J, et al. AAV6 versus AAV9 after intramyocardial administration. Mol Ther Methods Clin Dev. 2025;33:101532. [N]
[3-3] Ravindran D, et al. High-throughput evaluation of cardiac-specific promoters for AAV gene therapy. Gene Ther. 2025. [N]
[3-4] Inagaki K, et al. Robust systemic transduction with AAV9 vectors in mice: efficient global cardiac gene transfer superior to that of AAV8. Mol Ther. 2006;14(1):45-53. DOI:10.1016/j.ymthe.2006.03.011. [H]
[3-5] Gao F, et al. A simple and efficient method for in vivo cardiac-specific gene manipulation by intramyocardial injection in mice. J Vis Exp. 2015;(102):e53022. PMC5933510. [F]
[3-6] MRI roadmap-guided transendocardial delivery of exon-skipping rAAV restores dystrophin in canine DMD model. PMC3424392. [H]