Combination Of Bubble Liposomes Research Articles

Enhancement of Blood-Brain Barrier Permeability and Delivery of Antisense Oligonucleotides or Plasmid DNA to the Brain by the Combination of Bubble Liposomes and High-Intensity Focused Ultrasound.

The blood–brain barrier (BBB) is a major obstacle that prevents therapeutic drugs or genes from being delivered to the central nervous system. Therefore, it is important to develop methods to enhance the permeability of the BBB. We have developed echo-contrast gas (C3F8) entrapping liposomes (Bubble liposomes, BLs) that can work as a gene delivery tool in combination with ultrasound (US) exposure. Here, we studied whether the permeability of the BBB can be enhanced by the combination of BLs and high-intensity focused ultrasound (HIFU). Mice were intravenously injected with Evans blue (EB). BLs were subsequently injected, and the right hemispheres were exposed to HIFU. As a result, the accumulation of EB in the HIFU-exposed brain hemispheres was increased over that observed in the non-HIFU-exposed hemispheres, depending on the intensity and the duration of the HIFU. Similarly, the combination of BLs and HIFU allowed fluorescent-labeled antisense oligonucleotides to be delivered into the HIFU-exposed left hemispheres of the treated mice. Furthermore, a firefly luciferase-expressing plasmid DNA was delivered to the brain by the combination method of BLs and HIFU, which resulted in the increased gene expression in the brain at the focused-US exposure site. These results suggest that the method of combining BLs and HIFU together serves as a useful means for accelerating the permeability of BBB and thereby enabling antisense oligonucleotides or genes to be delivered to the focused brain site.

Enhancement of blood-brain barrier permeability by the combination of bubble liposomes and high-intensity focused ultrasound

The blood-brain barrier is a major obstacle that prevents therapeutic drugs or genes being delivered into the central nervous systems. Therefore, it is important to develop enabling enhance the permeability of blood-brain barrier (BBB). So far, we have developed echo-contrast gas (C3F8) entrapping liposomes (Bubble liposomes, BLs) which can be work as a gene delivery tool in the combination of ultrasound (US) exposure. Here, we investigated whether the delivery efficiency of intravenously injected large molecular agents can be induced by the combination of BLs and high-intensity focused ultrasound (HIFU).

Nonviral gene delivery systems by the combination of bubble liposomes and ultrasound.

The combination of therapeutic ultrasound (US) and nano/microbubbles is an important system for establishing a novel and noninvasive gene delivery system. Genes are delivered more efficiently using this system compared with a conventional nonviral vector system such as the lipofection method, resulting in higher gene expression. This higher efficiency is due to the gene being delivered into the cytosol and bypassing the endocytosis pathway. Many in vivo studies have demonstrated US-mediated gene delivery with nano/microbubbles, and several gene therapy feasibility studies for various diseases have been reported. In addition, nano/microbubbles can deliver genes site specifically by the control of US exposure site. In the present review, we summarize the gene delivery systems by the combination of nano/microbubbles and US, describe their properties, and assess applications and challenges of US theranostics.

Bubble Liposomes and Ultrasound Exposure Improve Localized Morpholino Oligomer Delivery into the Skeletal Muscles of Dystrophic mdx Mice

Duchenne muscular dystrophy (DMD) is a genetic disorder that is caused by mutations in the DMD gene that lead to an absence of functional protein. The mdx dystrophic mouse contains a nonsense mutation in exon 23 of the dystrophin gene; a phosphorodiamidate morpholino oligomer (PMO) designed to skip this mutated exon in the mRNA induces dystrophin expression. However, an efficient PMO delivery method is needed to improve treatment strategies for DMD. We previously developed polyethylene glycol (PEG)-modified liposomes (Bubble liposomes) that entrap ultrasound contrast gas and demonstrated that the combination of Bubble liposomes with ultrasound exposure is an effective gene delivery tool in vitro and in vivo. In this study, to evaluate the ability of Bubble liposomes as a PMO delivery tool, we tested the potency of the Bubble liposomes combined with ultrasound exposure to boost the delivery of PMO and increase the skipping of the mutated exon in the mdx mouse. The results indicated that the combination of Bubble liposomes and ultrasound exposure increased the uptake of the PMO targeting a nonsense mutation in exon 23 of the dystrophin gene and consequently increased the PMO-mediated exon-skipping efficiency compared with PMO injection alone, leading to significantly enhanced dystrophin expression. This increased efficiency indicated the potential of the combination of Bubble liposomes with ultrasound exposure to enhance PMO delivery for treating DMD. Thus, this ultrasound-mediated Bubble liposome technique may provide an effective, noninvasive, nonviral method for PMO therapy for DMD muscle as well as for other muscular dystrophies.

Combination of Bubble Liposomes and High-Intensity Focused Ultrasound (HIFU) Enhanced Antitumor Effect by Tumor Ablation

Ultrasound (US) is used in the clinical setting not only for diagnosis but also for therapy. As a therapeutic US technique, high-intensity focused ultrasound (HIFU) can be applied to treat cancer in a clinical setting. Microbubbles increased temperature and improved the low therapeutic efficiency under HIFU; however, microbubbles have room for improvement in size, stability, and targeting ability. To solve these issues, we reported that "Bubble liposomes" (BLs) containing the US imaging gas (perfluoropropane gas) liposomes were suitable for ultrasound imaging and gene delivery. In this study, we examined whether BLs and HIFU could enhance the ablation area of the tumor and the antitumor effect. First, we histologically analyzed the tumor after BLs and HIFU. The ablation area of the treatment of BLs and HIFU was broader than that of HIFU alone. Next, we monitored the temperature of the tumor, and examined the antitumor effect. The temperature increase with BLs and HIFU treatment was faster and higher than that with HIFU alone. Moreover, treatment with BLs and HIFU enhanced the antitumor effect, which was better than with HIFU alone. Thus, the combination of BLs and HIFU could be efficacious for cancer therapy.

Novel Dianostics and Therapeutics with Ultrasound Technologies and Nanotechnologies

Ultrasound is a good tool for theranostics due to have multi-potency both of diagnostics with sonography and therapeutics with high intensity focused ultrasound (HIFU). In addition, microbubbles and nanobubbles are utilized as not only contrast imaging agent but also enhancer of drug and gene delivery by combination of ultrasound. Recently, we developed novel liposomal nanobubbles (Bubble liposomes) which were containing perfluoropropane. Bubble liposomes induced jet stream by low intensity ultrasound exposure and resulted in enhancing permeability of cell membrane. This phenomenon has been utilized as driving force for drug and gene delivery. On the other hand, the combination of Bubble liposomes and high intensity ultrasound induces strong jet stream and increase temperature. This condition can directly damage to tumor cells, we are applying this for cancer therapy. Therefore, their combination has potency for various cancer therapies such as gene therapy, immunotherapy and hyperthermia. In this review, we discuss about cancer therapy by the combination of Bubble liposomes and ultrasound.

Gene delivery to periodontal tissue using Bubble liposomes and ultrasound

Periodontitis is the most common inflammatory disease caused by oral biofilm infection. For efficient periodontal treatment, it is important to enhance the outcome of existing regenerative therapies. The physical action of an ultrasound may be able to deliver a therapeutic gene or drugs into the local area of the periodontium being treated for periodontal regeneration. Previously, we developed "Bubble liposomes" as a useful carrier for gene or drug delivery, and reported that delivery efficiency was increased with high-frequency ultrasound in vitro and in vivo. Hence, the aim of the present study was to examine the possibility of delivering genes into gingival tissues using Bubble liposomes and ultrasound. We attempted to deliver naked plasmid DNA encoding luciferase or enhanced green fluorescent protein (EGFP) into the lower labial gingiva of Wistar rats using Bubble liposomes, with or without ultrasound exposure. Ultrasound parameters were optimized for intensity (0-4.0 W/cm(2) ) and exposure time (0-120 s) to establish the most efficient conditions for exposure. The efficacy and duration of gene expression in the gingiva were investigated using a luciferase assay and fluorescence microscopy. The strongest relative luciferase activity was observed when rats were treated under the following ultrasound conditions: 2.0 W/cm(2) intensity and 30 s of exposure time. Relative luciferase activity, 1 d after gene delivery, was significantly higher in gingiva treated using Bubble liposomes and ultrasound than in gingiva of the other treatment groups. Histological analysis also showed that distinct EGFP-expressing cells were observed in transfected gingiva when rats were treated under optimized conditions. From these results, the combination of Bubble liposomes and ultrasound provides an efficient technique for delivering plasmid DNA into the gingiva. This technique can be applied for the delivery of a variety of therapeutic molecules into target tissue, and may serve as a useful treatment strategy for periodontitis.

Delivery of an Angiogenic Gene into Ischemic Muscle by Novel Bubble Liposomes Followed by Ultrasound Exposure

To develop a safe and efficient gene delivery system into skeletal muscle using the combination of Bubble liposomes (BL) and ultrasound (US) exposure, and to assess the feasibility and the effectiveness of BL for angiogenic gene delivery in clinical use. A solution of luciferase-expressing plasmid DNA (pDNA) and BL was injected into the tibialis (TA) muscle, and US was immediately applied to the injection site. The transfection efficiency was estimated by a luciferase assay. The ischemic hindlimb was also treated with BL and US-mediated intramuscular gene transfer of bFGF-expressing plasmid DNA. Capillary vessels were assessed using immunostaining. The blood flow was determined using a laser Doppler blood flow meter. Highly efficient gene transfer could be achieved in the muscle transfected with BLs, and US mediated the gene transfer. Capillary vessels were enhanced in the treatment groups with this gene transfer method. The blood flow in the treated groups with this gene transfer method quickly recovered compared to other treatment groups (non-treated, bFGF alone, or bFGF+US). The gene transfer system into skeletal muscle using the combination of BL and US exposure could be an effective means for angiogenic gene therapy in limb ischemia.

Cancer gene therapy by IL-12 gene delivery using liposomal bubbles and tumoral ultrasound exposure

Interleukin-12 (IL-12) gene therapy is expected to be effective against cancers because it primes the immune system for cancer cells. In this therapy, it is important to induce IL-12 gene expression in the tumor tissue. Sonoporation is an attractive technique for developing non-invasive and non-viral gene delivery systems, but simple sonoporation using only ultrasound is not an effective cancer gene therapy because of the low efficiency of gene delivery. We addressed this problem by combining ultrasound and novel ultrasound-sensitive liposomes (Bubble liposomes) which contain the ultrasound imaging gas perfluoropropane. Our previous work showed that this is an effective gene delivery system, and that Bubble liposome collapse (cavitation) is induced by ultrasound exposure. In this study, we assessed the utility of this system in cancer gene therapy using IL-12 corded plasmid DNA. The combination of Bubble liposomes and ultrasound dramatically suppressed tumor growth. This therapeutic effect was T-cell dependent, requiring mainly CD8 + T lymphocytes in the effector phase, as confirmed by a mouse in vivo depletion assay. In addition, migration of CD8 + T cells was observed in the mice, indicating that the combination of Bubble liposomes and ultrasound is a good non-viral vector system in IL-12 cancer gene therapy.

839. Control of the Gene Delivery Area by the Combination of Bubble Liposomes and Ultrasound

839. Control of the Gene Delivery Area by the Combination of Bubble Liposomes and Ultrasound

Effective gene delivery with novel liposomal bubbles and ultrasonic destruction technology

From the viewpoint of safety, non-viral vector systems represent an attractive gene delivery system for gene therapy. However, the transfection efficiency of non-viral vectors in vivo is generally very low. Previously, it was reported that microbubbles, utilized as imaging agents for diagnostic echocardiography, could promote gene delivery into cells when combined with ultrasound exposure. We therefore developed novel liposomal bubbles (Bubble liposomes) containing the lipid nanobubbles of perfluoropropane which is used as ultrasound imaging agent. These Bubble liposomes were smaller in diameter than conventional microbubbles and induced cavitation upon exposure to ultrasound. These results suggested that cavitation of these Bubble liposomes could be an efficient approach for delivering plasmid DNA into cells. In addition, in in vivo gene delivery, the combination of Bubble liposomes and ultrasound provided more effective gene delivery than conventional lipofection methods, further suggesting that Bubble liposomes could be effective as a non-viral vector system in in vivo gene delivery. In this review, we discuss the characteristics of Bubble liposomes and their potential utility as a gene delivery tool in vitro and in vivo.

Tumor specific ultrasound enhanced gene transfer in vivo with novel liposomal bubbles

Bubble liposomes (liposomes which entrap an ultrasound imaging gas) may constitute a unique system for delivering various molecules efficiently into mammalian cells in vitro. In this study, Bubble liposomes were compared with cationic lipid (CL)–DNA complexes as potential gene delivery carriers into tumor in vivo. The delivery of genes by Bubble liposomes depended on the intensity of the applied ultrasound. Transfection efficiency plateaued at 0.7 W/cm 2 ultrasound intensity. Bubble liposomes efficiently transferred genes into cultured cells even when the cells were exposed to ultrasound for only 1 s. In addition, Bubble liposomes could introduce the luciferase gene more effectively than CL-DNA complexes into mouse ascites tumor cells and solid tumor tissue. We conclude that the combination of Bubble liposomes and ultrasound is a minimally-invasive and tumor specific gene transfer method in vivo.