Optimization and apoptosis induction by RNAi with UTMD technology in vitro

Abstract

Abstract. Apoptosis induction by short hairpin RNA (shRNA) expression vectors may be an efficient and promising strategy for cancer gene therapy. Ultrasound-targeted microbubble destruction (UTMD) is an appealing technique; however, there few data are available to demonstrate the feasibility and to optimize the methodology for this technology. The aim of this study was to optimize this technique and to elucidate the effects on gene inhibition and apoptosis induction in vitro. Human cervical cancer cell lines were obtained and cultured. shRNA vectors were constructed, and the UTMD technique was examined to determine whether or not it was suitable for shRNA transfection into cells. Cells were then examined using flow cytometry. The results revealed that the optimal irradiation parameters obtained higher transfection efficiency and did not affect the integrity of plasmid DNA. We concluded that survivin downregulation with shRNA expression vectors, mediated by the optimal UTMD parameters, markedly induced cell apoptosis and caused cell cycle arrest, laying a foundation for further investigation of this cancer therapy.

<p>Ultrasound-Targeted Microbubble Destruction Mediated si-CyclinD1 Inhibits the Development of Hepatocellular Carcinoma via Suppression of PI3K/AKT Signaling Pathway</p>

“… 34 – 37 At present, a large number of studies have confirmed that UTMD can effectively mediated gene transfer, which has the advantages of safety, practicability and targeting, and is one of the main trends of gene therapy. 38 – 41 Consistent with our study, one report focused on UTMD-mediated suicide gene and hepatic cancer also demonstrated that the tumor inhibition rate in the treatment group was markedly higher compared with that in the control group. 42 Furthermore, the present study also noted that the in vitro findings were further confirmed via in vivo study.…”

The present and future role of ultrasound targeted microbubble destruction in preclinical studies of cardiac gene therapy

“…Recent studies point out the use of low frequency probe can produce a wider and more uniform sound field and that strong cavitation is the key to thrombolysis. However, the optimal parameters have not been formed yet (7). In addition, when MBs exist in the capillaries, ultrasonic irradiation can cause microvascular leakage, intracardiac hemolysis capillary rupture, bleeding, formation, inflammatory cell infiltration, myocardial cell damage and other adverse reactions (7,89,90).…”

“…However, the optimal parameters have not been formed yet (7). In addition, when MBs exist in the capillaries, ultrasonic irradiation can cause microvascular leakage, intracardiac hemolysis capillary rupture, bleeding, formation, inflammatory cell infiltration, myocardial cell damage and other adverse reactions (7,89,90). Therefore, more technological revolution should be taken to enhance the biological application of UTMD.…”

“…Ultrasound targeted microbubble destruction (UTMD) is a novel therapy to deliver the objective gene to organs of living animals. It has been proven to bind non-invasively microbubbles (MBs) with DNA or RNA carriers (assemble adenoviral, plasmid or nanoparticles) into the shell and destroy the located MBs with low frequency and high mechanical index ultrasound, releasing target agents into peculiar organs (5)(6)(7). Not only can UTMD improve the transfection efficiency by several orders of magnitude, but also achieve specific target markedly (8).…”

Gene therapy for cardiovascular disease mediated by ultrasound and microbubbles

“…In recent years, a large number of proof-of-principle studies have confirmed that ultrasound targeted microbubble destruction (UTMD) could enhance transfection efficiency of naked plasmid DNA by several orders of magnitude [5-7]. Therefore, it is considered as a promising strategy for gene therapy.…”