Ultrasonic microbubbles for glioma-targeted drug delivery / 药学学报

Ultrasonic microbubbles were used to open blood-brain barriers (BBB) with a reversed and limited behavior feature in the study, which could improve the brain-targeted delivery of anti-tumor drugs. The glioma rat model was prepared. Low-frequency ultrasound was combined with microbubbles to affect the permeability of BBB compared with the permeability of independently administered Evans blue (EB) crossing BBB. Time point and length of ultrasound were investigated whether they affect the permeability of BBB and the damage of brain tissue. The effect of the growth time of glioma on BBB permeability was explored. Only glioma had a very little impact on BBB permeability. However, ultrasonic microbubbles opened the BBB with the features of temporary, limited and reversed behavior and improved EB and magnetic resonance imaging contrast agent penetrating BBB. A length of 30 s ultrasound is appropriate for opening BBB and no damage of brain tissue. Drugs should be injected before ultrasound so that they enter into brain as BBB opening. Ultrasonic microbubbles can open BBB effectively and safely, which improve drugs penetrating BBB under proper time point and length.

Optimization of the preparation of sonogenic phospholipids-based microbubbles by using central composite experimental design and response surface methodology / 药学学报

Sonogenic microbubble agent is a newly developed drug targeting delivery system, which uses ultrasonic beam to enhance the delivery of drug and gene to targeted cells and tissues. In this paper, the preparation of sonogenic phospholipids-based microbubbles was optimized by using central composite experimental design (CCD) and response surface methodology (RSM). Hydrogenated egg phosphatidylcholine (EPC), Tween 80 and polyethylene glycol 1500 (PEG 1500) were important components affecting the concentration of 2 - 8 microm microbubbles in the preparation. The combined effects of these three factors were analyzed by CCD and optimized by RSM. Evaluation variable was the concentration of 2 - 8 microm microbubbles. Overall desirability was fitted to a second-order polynomial equation, through which three dimensional response surface graphs were produced. Optimal experimental conditions were selected from the stationary point of the response surfaces. The stability of the sonogenic phospholipids-based microbubbles by the optimal formulation was investigated by accelerated experiment. The contrast effect in vivo of the optimal formulation was investigated. Foreign market product SonoVue was used as the control. From the results, all the three factors had positive effects on the concentration of 2 - 8 microm microbubbles. The optimal condition in the preparation of phospholipids-based microbubbles was obtained as following: EPC 8.35 mg, Tween 80 21.68 mg and PEG 1500 201 mg. The mean value of the concentration of 2 - 8 microm microbubbles in rechecking experiment reached 8.60 x 10(9) x mL(-1). From the accelerated experiment, phospholipids-based microbubbles showed good physical stability. The intensity (relative unit) and duration of the contrast effect by the optimal formulation were 4.47 +/- 0.15 and (302 +/- 7) s respectively, which showed little difference with foreign market product SonoVue [4.28 +/- 0.13, (309 +/- 8) s]. The optimal formulation selected by CCD and RSM showed high microbubble concentration, good physical stability and effective sonogenic contrast effect.

Recent advances in the applications of ultrasonic microbubbles as gene delivery systems / 药学学报

Ultrasonic beam can be focused on a particular tissue. As ultrasound contrast agents for medical ultrasound imaging, gas-filled microbubbles can also be used as drugs or gene carriers. Therefore, ultrasonic microbubbles become a topic of intense interest in drug delivery because they can be used as gene targeting delivery systems. Based on the relevant materials, the applications of ultrasonic microbubbles as gene delivery systems in various tissues in vivo were reviewed, such as cardiac, vascular, skeletal muscle and tumor. Their potential problems in future use were also discussed.

Optimizing the operating variables that affect the transfection experiment of antisense oligodeoxyribonucleotide by gas-filled microbubbles / 药学学报

To optimize the operating variables that affect the transfection of antisense oligodeoxyribonucleotide (AS-ODNs) by insonated gas-filled lipid microbubbles, SF6-filled microbubbles were prepared by sonication-lyophilization method. An AS-ODNs sequence and a breast cancer cell line SK-BR-3 were used to define the various operating variables determining the transfection efficiency of SF6-filled microbubbles. Three levels of mixing speed, different durations of mixing and various delay time before ultrasound were examined, separately. Transfection efficiency was detected by fluorescence microscopy. Transfection results with and without incubation of AS-ODNs and microbubbles before mixing cells were compared. From the results, there is no significant difference between the transinfection efficiency with or without incubation of AS-ODNs and microbubbles before mixing cells. AS-ODNs transfection efficiency showed an increasing trend with mixing speed and mixing duration, but there is a negative relationship with delay time before ultrasound. The optimum parameters for AS-ODNs transfection by SF6-filled microbubbles were found at a mixing speed of 40-50 r x min(-1) for 30-60 s with less than 60 s delay before ultrasound. For a successful transfection, long time of incubation with gene is essential for normal nonviral vectors such as liposomes or cationic lipid-polymer hybrids, because these vectors depend on endocytosis and membrane fusion to realize transfection. Unlike liposomes and cationic lipid-polymer hybrids, gas-filled lipid microbubbles depend on sonorporation effect to realize transfection. Therefore, the incubation of gene and microbubbles before mixing cells may not be necessary. Ultrasound-mediated AS-ODNs transfection enhanced by gas-filled lipid microbubbles represents an effective avenue for gene transfer.

Comparison of sonoporation effect of liposomes and phospholipids-based microbubbles on cultured cell membrane / 药学学报

<p><b>AIM</b>To compare sonoporation effect of two phospholipids-based vectors-liposomes and microbubbles on cultured cell membrane.</p><p><b>METHODS</b>A breast cancer cell line SK-BR-3 was exposed to ultrasound alone, 2% or 5% liposome + ultrasound and 2% or 5% microbubble + ultrasound, separately. Immediately after the experiment and 24 h after ultrasound exposure, atomic-force microscopy (AFM) scanning was used to observe the membrane change of SK-BR-3 cells.</p><p><b>RESULTS</b>After ultrasound exposure, normal SK-BR-3 cells more or less lost their natural shape, showing elliptic outline with obtuse curved boundary. In groups added with phospholipids-based microbubbles, more obtuse curved boundary of cells was observed. The membrane pores of SK-BR-3 cells had apparent changes after ultrasound exposure. With AFM technique, membrane pores under ultrasound alone or ultrasound with liposomes conditions were enlarged, the diameter of some pores exceeding 1 microm. But all the membrane pores in these conditions returned to normal appearance after 24 hours. In ultrasound with 2% microbubble condition, most membrane pores were about 1 - 3 microm in size and returned to normal appearance after 24 h. In ultrasound with 5% microbubble condition, however, pores of most cell membrane porosity was about 2 - 4 pm and did not totally return to normal appearance after 24 h.</p><p><b>CONCLUSION</b>At 2% concentration, phospholipids-based microbubble could enhance ultrasonic sonoporation effect and produce reparable membrane pores on SK-BR-3 cells, which appeared to be a promising vehicle for drug and gene delivery.</p>

Echogenic phospholipids-based gas-filled microbubbles as delivery system of antisense oligodeoxynucleotides / 药学学报

<p><b>AIM</b>To investigate the feasibility of transfer antisense oligodeoxynucleotides (AS-ODNs) by the phospholipids-based gas-filled microbubbles (PGM) under ultrasound activation.</p><p><b>METHODS</b>An antisense oligodeoxynucleotides sequence ZL combined with luciferase reporter plasmid was used. A breast cancer cell line SK-BR-3 was exposed to different conditions to investigate the effects of such factors as ZL concentration, PGM concentration, mechanical index (MI) and ultrasound exposure duration on transfection efficiency and cell viability. The transfection efficiency and cell viability by other lipid vectors such as lipofectamine and liposome were also tested, whose results were comparied with that of PGM. Transfection efficiency was detected by fluorescence microscopy. Cell viability was verified by PI (propidium iodide) assay.</p><p><b>RESULTS</b>Among the factors tested, ultrasound exposure duration, MI and PGM concentration had obvious impacts on transfection efficiency and cell viability. The results showed that the optimal ultrasound condition was the exposure to ultrasound at MI 1.0 for 30 s with 2% PGM concentration, which gave an overall transfection efficiency of 78% +/- 10%, increased nearly 18 folds over the transfection by PGM (4.0%) or lipofectamine (4.3%) without ultrasound. Under same ultrasound conditions, different vectors showed significant difference in transfection efficiency while there are similar results in cell viability.</p><p><b>CONCLUSION</b>Under proper ultrasound conditions, PGM can markedly enhance AS-ODNs transfection efficiency.</p>