Review
Progress in the development of ultrasound-mediated gene delivery systems utilizing nano- and microbubbles

https://doi.org/10.1016/j.jconrel.2010.05.009Get rights and content

Abstract

Recently, ultrasound-mediated gene delivery with nano- and microbubbles was developed as a novel non-viral vector system. In this gene delivery system, microstreams and microjets, which are induced by disruption of nano/microbubbles exposed to ultrasound, are used as the driving force to transfer genes into cells by opening transient pores in the cell membrane. This system can directly deliver plasmid DNA and siRNA into cytosol without endocytosis pathway. Therefore, these genes are able to escape from degradation in lysosome and result in enhancing the efficiency of gene expression. In addition, it is expected that ultrasound-mediated gene delivery using nano/microbubbles would be a system to establish non-invasive and tissue specific gene expression because ultrasound can transdermally expose to target tissues and organs. This review focuses on the current ultrasound-mediated gene delivery system using nano/microbubbles. We discuss about the feasibility of this gene delivery system as novel non-viral vector system.

Graphical abstract

Introduction

Gene therapy has a potential in the treatment of cancer and diseases that are due to genomic causes. Viral vectors are efficient carriers of genes for transduction, but some problems have become evident [1], [2], [3]. Delivery vectors that are highly potent in terms of gene transduction efficiency should also be safe and easy to apply. Non-viral vectors have recently received focus as gene carriers, but their transduction efficiency is very low. Efforts have recently been directed towards improving this aspect [4], [5], [6]. Towards this end, ultrasound has been investigated for improving the efficiency of transgene delivery, and holds promise as a non-invasive gene delivery system.

Ultrasound shows potential for improving the efficiency of gene delivery into tissues and cells, a technique known as sonophoresis/sonoporation [7]. It is believed that ultrasound perturbs cell membranes and causes transient pores to open in the membrane, thus facilitating gene entry into the cell [8]. In addition, it has been reported that microbubbles utilized as ultrasound contrast agents play an important role in enhancing the efficiency of gene delivery, without causing cell damage [9]. In general, cell damage is dependent on ultrasound intensity, concentration of microbubbles and cell type. Especially, ultrasound intensity and exposure time are key factors. Therefore, it is important to optimize the condition of ultrasound exposure in ultrasound-mediated gene delivery [10], [11], [12], [13]. Some researchers studied about the cell damage by the disruption of microbubbles with ultrasound exposure [14], [15], [16], [17], [18], [19]. These reports are useful as informative references for ultrasound-mediated gene delivery utilizing microbubbles.

Microbubbles which are destroyed by ultrasound exposure generate microstreams or microjets, resulting in shear stress to cells and the generation of transient holes in cell membranes [20]. Since this approach can be used to deliver extracellular molecules such as genes into cells, microbubbles could facilitate ultrasound-mediated gene delivery. In addition, submicron sized bubbles (nanobubbles), which are smaller than conventional microbubbles, were recently reported [21], [22], and we have also developed novel liposomal nanobubbles (Bubble liposomes) [11], [23], [24], [25], [26], [27], [28], [29], [30], [31], [32]. These nanobubbles can also be utilized as enhancing tool of gene delivery efficiency in ultrasound-mediated gene delivery. In this review, we introduced about ultrasound-mediated delivery systems combined with nano/microbubbles and discussed the feasibility as non-viral vector system.

Section snippets

Microbubbles as ultrasound contrast agents

Ultrasonography is a widely used diagnostic medical imaging technique that is non-invasive, relatively low-cost, easy to use, provides real-time imaging, and importantly, avoids the use of hazardous ionizing radiation. Ultrasound wave pulses generated by an ultrasound transducer are partially reflected or scattered by the interfaces between different tissues. The transducer detects the ultrasound waves returned by scattering, and these signals are converted to ultrasound images. Since blood

Properties of microbubbles combined with ultrasound

The behavior of microbubbles depends on the amplitude of ultrasound used. At very low acoustic pressure (mechanical index (MI) < 0.05–0.1), the microbubbles cause linear oscillation, and the reflected frequency is equal to the transmitted frequency (Fig. 1(a)). An increase in acoustic pressure (0.1 < MI < 0.3), referred to as low-power imaging, causes non-linear expansion and compression of the microbubbles (Fig. 1(b)). In fact, the bubble becomes somewhat more resistant to compression than to

Gene delivery using sonoporation as a non-viral vector system

The first studies investigating the utility of ultrasound for gene delivery used frequencies in the range 20–50 kHz [7], [44]. However, these frequencies, along with cavitation, are known to cause tissue damage if not properly controlled [45], [46]. To overcome this problem, many gene delivery studies have used therapeutic ultrasound, which operates at frequencies of 1–3 MHz, intensities of 0.5–2.5 W/cm2 or MI 0.3–2, and in pulse-mode [47]. However, as these conditions result in very inefficient

Efforts to tissue- or organ-selective gene delivery

To establish ideal gene therapy, it is important to deliver therapeutic gene into target tissue or organ. In the early study, gene and nano/microbubbles were directly injected into target tissue and organ [53], [66]. However, in this method, there are some limitations such as injection volume and injection technique. To improve these problems, some researchers recently developed ultrasound-mediated gene delivery by the supplying gene and nano/microbubbles via blood flow [11], [67]. In this

Conclusion

Ultrasound has long been utilized as a useful diagnostic tool. Therapeutic ultrasound was recently developed and is being utilized in clinical settings. The combination of therapeutic ultrasound and nano/microbubbles is an interesting and important system for establishing a novel and non-invasive gene delivery system. Gene expression efficiency with this system can effectively deliver gene compared with conventional non-viral vector system such as lipofection method due to deliver gene into

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