Ultrasonic microbubble contrast agent might produce biological effects on cells, because such agents are able to induce cellular and cell membrane injuries [
1,
6]. ROS induced by ultrasound irradiation are thought be responsible for the biological effects of ultrasound irradiation [
16,
17]. Ultrasonic microbubble contrast agents have the ability to produce oxyradicals, which has been confirmed by spin resonancespectroscopy [
18]. These radicals might induce apoptosis and necrosis in tumor cells via several targets, such as cell membranes, intracellular bioactive molecules, and DNA [
19]. In this study, we found that low-frequency ultrasonic irradiation with microbubble agent promoted apoptosis and inhibited cellular proliferation in tumor cells and vascular endothelial cell. This effect was associated with the attenuation of the SOD activity in the culture medium. Consumption of active oxygen scavengers indicates an increase in the level of ROS produced during cellular damage caused by treatment with ultrasound and microbubble agent. This phenomenon has been assessed by other researchers who have suggested a sonochemical mechanism underlying this effect [
20].
Apoptosis can be induced by treating cells with ultrasound and contrast agent. ROS-mediated injury may be one of the important factors responsible for the biological effects of ultrasound [
17,
18]. Increase in the expression of NF-kappa B and I kappa B-alfa has been often associated with the induction of apoptosis [
14]. These nuclear factors might be related with the sequential induction of cell death caused by ultrasound.
ROS can activate the expression of intracellular NF-kappa B. This ubiquitous cytokine is involved in the regulation of various cellular effectors molecules and is present in an inactive form as a dimer with I kappa B-alfa, in the absence of any stimulation. The activity of NF-kappa B proteins is regulated by I kappa B-alfa proteins [
21]. When cells are stimulated, NF-kappa B is activated and is translocated to the nucleus, where it binds to the target gene promoter and activates its transcription [
15]. A variety of extracellular signals can stimulate the activation of NF-kappa B, such as ROS, ultraviolet irradiation, double-stranded RNA, cytokine interleukin-1, tumor necrosis factor-α, lipopolysaccharide, and viruses [
11‐
13]. Because the regulation by NF-kappa B activation, de novo I kappa B-alfa can reintegrate with NF-kappa B to form a dimeric compound to inhibit the activation of NF-kappa B. Although I kappa B-alfa is synthesized de novo, NF-kappa B activity remains sustainable for several hours [
22]. We found that the protein level of NF-kappa B was markedly augmented at 1 hour after treatment with ultrasound irradiation and microbubble agent, suggesting that the cellular damage was associated with the oxidation induced by ultrasound and microbubbles. Hence, the biological effects of ultrasound were speculated to be associated with the ultrasound chemical reaction. At 24 hours after ultrasound treatment, the enhanced expression of I kappa B suggested that emerge of the activities of intracellular biological mediation produced through ultrasound chemical reactions, corresponding with the general regulation of modulating cellular proliferation in this type of cytokines [
21,
22]. We found that I kappa B-alfa expression promoted apoptosis in the experimental groups. Ultrasonic irradiation increased the level of ROS and induced the expression of NF-kappa B and I kappa B in succession. This finding suggests that the biological effect of sonochemistry is one of related factors in ultrasound killing cells with microbubble agent. Due to the dynamic characteristics of ultrasound contrast agents [
5], ultrasound microbubble agents undergo reactions involved in ultrasound physics and ultrasound chemistry, damaging cell membranes and intracellular structures, and ultimately resulting in cellular injury or even cell death [
23].