**2.2 Endocytosis**

*Gene Expression and Control*

Microbubbles can serve as vehicles for carrying genetic cargo and can be destructed by ultrasound, resulting in the local release of genetic payload. Meanwhile, sonoporation effect will occur upon which the bubbles are exposed to the appropriate ultrasonic energy, producing the transient small holes on the adjacent cell membrane and thus increasing the vascular and cellular permeability **Figure 1**. In this chapter, we will briefly introduce the mechanism and review the recent advances

*Schematic model of ultrasound-mediated gene delivery. Bioeffect produced by ultrasound and microbubble interaction could enhance the permeability of vascular and promote the accumulation of gene (green) in tissue.* 

When ultrasound is irradiated locally with certain energy, the cavitation nuclei, such as ultrasound contrast agents and bubbles, could alternately occur expansion, contraction, splitting, fusion, and even rupture. This physical process is called cavitation effect. Accompanied by the cavitation effect, acoustic microstreaming, micro-jet, high temperature, and shockwave will occur in the medium, resulting in the formation of some temporary, reversible pores on the cell membrane, which is sonoporation [6, 7]. Generally, it is an accepted notion that the sonoporation from

There are a large number of studies, which have confirmed that sonoporation can increase the efficiency of gene delivery through enhancing the permeability of the cell membrane [9–12]. The number of pores, having a high impact on the gene delivery efficiency, can be affected by a lot of factors, such as acoustic pressure, irradiation duration time, and pulse repetition frequency [13–15]. Sonoporation pores trend to be larger along with the increase of acoustic pressure and irradiation time, which also enhance gene transfection efficiency [16]. However, excessive acoustic pressure or ultrasonic duration may reduce cell viability and even cause cell death, vascular rupture, and other side effects [17–19]. Therefore, to achieve a

about local gene delivery by ultrasound.

*(Quoted from: Sirsi and Borden [5]).*

**2.1 Sonoporation**

**Figure 1.**

**2. Mechanism of ultrasound-mediated gene transfection**

cavitation effect allows genes and drugs to enter cells [8].

**100**

In addition to sonoporation, cavitation effect can change the cell membrane structure through microstreaming and shear force. The mechanical force may cause cytoskeleton rearrangement and regulate various downstream cellular signaling pathways, helping the endocytosis of genetic cargo [20, 21]. Generally, there are three forms of endocytosis, including macropinocytosis, clathrin-mediated endocytosis, and caveolae-mediated endocytosis [22]. After ultrasound irradiation, the reactive oxygen species are produced to stimulate the calcium influx and induce the occurrence of endocytosis [23]. In addition, cavitation effect and shear force induced by ultrasound can change cell structure and influence endocytosis through mechanosensors and signaling cascade [24]. Meijering et al. demonstrated that endocytosis was involved in the uptake of the macromolecular substances, while small molecules enter cells mainly through the pores of the membrane surface [25].
