Ultrasound-targeted transfection of tissue-type plasminogen activator gene carried by albumin nanoparticles to dog myocardium to prevent thrombosis after heart mechanical valve replacement

Abstract

Background

There are more than 300,000 prosthetic heart valve replacements each year worldwide. These patients are faced with a higher risk of thromboembolic events after heart valve surgery and long-term or even life-long anticoagulative and antiplatelet therapies are necessary. Some severe complications such as hemorrhaging or rebound thrombosis can occur when the therapy ceases. Tissue-type plasminogen activator (t-PA) is a thrombolytic agent. One of the best strategies is gene therapy, which offers a local high expression of t-PA over a prolonged time period to avoid both systemic hemorrhaging and local rebound thrombosis. There are some issues with t-PA that need to be addressed: currently, there is no up-to-date report on how the t-PA gene targets the heart in vivo and the gene vector for t-PA needs to be determined.

Aims

To fabricate an albumin nano-t-PA gene ultrasound-targeted agent and investigate its targeting effect on prevention of thrombosis after heart mechanic valve replacement under therapeutic ultrasound.

Methods

A dog model of mechanical tricuspid valve replacement was constructed. A highly expressive t-PA gene plasmid was constructed and packaged by nanoparticles prepared with bovine serum albumin. This nanopackaged t-PA gene plasmid was further cross-linked to ultrasonic microbubbles prepared with sucrose and bovine serum albumin to form the ultrasonic-targeted agent for t-PA gene transfection. The agent was given intravenously followed by a therapeutic ultrasound treatment (1 MHz, 1.5 w/cm², 10 minutes) of the heart soon after valve replacement had been performed. The expression of t-PA in myocardium was detected with multiclonal antibodies to t-PA by the indirect immunohistochemical method. Venous blood t-PA and D-dimer contents were tested before and 1, 2, 4, and 8 weeks after the operation.

Results

The high expression of t-PA could be seen in myocardium with increases in blood t-PA and D-dimer contents and thrombosis was prevented 8 weeks after operation.

Conclusion

We successfully fabricated an albumin nano-t-PA gene ultrasound-targeted agent that could prevent dog thrombosis after mechanical heart valve replacement. Our study provides an experimental basis for prevention of human thrombosis-related diseases.

Keywords:

• albumin nanoparticles
• ultrasonic microbubbles
• valve replacement

Acknowledgments

This project was supported by the National 863 plan (No 2007AA021809, No 2007AA021803, No 2007AA021904) and Shenzhen Science and Technology Bureau in China (No 200622). We give special thanks for the technical support from the National Nanolaboratory at ZhongNan University.

Disclosure

The authors report no conflicts of interest in this work.