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Experimental endostatin-GFP gene transfection into human retinal vascular endothelial cells using ultrasound-targeted cationic microbubble destruction.

Xu Y, Xie Z, Zhou Y, Zhou X, Li P, Wang Z, Zhang Q - Mol. Vis. (2015)

Bottom Line: Cationic microbubbles (CMBs) were prepared and then compared with neutral microbubbles (NMBs) and liposomes.Moreover, the inhibition of HREC growth was enhanced following treatment with CMBs compared with NMBs or liposomes in vitro (p<0.01).These results suggest that the combination of UTMD and ES-GFP compounds might be a useful tool for gene therapy targeting retinal neovascularization.

View Article: PubMed Central - PubMed

Affiliation: Institute of Ultrasound Imaging and Department of Ultrasound, Second Affiliated Hospital of Chongqing Medical University; Chongqing Key Laboratory of Ultrasound Molecular Imaging, Chongqing, P.R. China.

ABSTRACT

Purpose: The purpose of this study was to investigate whether ultrasound-targeted cationic microbubble destruction could effectively deliver endostatin-green fluorescent protein (ES-GFP) plasmids to human retinal vascular endothelial cells (HRECs).

Methods: Cationic microbubbles (CMBs) were prepared and then compared with neutral microbubbles (NMBs) and liposomes. First, the two types of microbubbles were characterized in terms of size and zeta potential. The cell viability of the HRECs was measured using the 3-(4,5-dimthylthiazol-2-yl)-2,5 diphenyl-tetrazolium bromide (MTT) assay. The transcription and expression of endostatin, VEGF, Bcl-2, and Bcl-xl were measured via quantitative real-time PCR (qPCR) and western blotting, respectively.

Results: CMBs differed significantly from NMBs in terms of the zeta potential, but no differences in size were detected. Following ultrasound-targeted microbubble destruction (UTMD)-mediated gene therapy, the transcription and expression of endostatin were highest in the CMB group (p<0.05), while the transcription and expression of VEGF, Bcl-2, and Bcl-xl were lowest compared with the other groups. Moreover, the inhibition of HREC growth was enhanced following treatment with CMBs compared with NMBs or liposomes in vitro (p<0.01).

Conclusions: This study demonstrated that ultrasound-mediated cationic microbubbles could enhance the transfection efficiency of ES-GFP, which had obvious impacts on the inhibition of the growth process of HRECs in vitro. These results suggest that the combination of UTMD and ES-GFP compounds might be a useful tool for gene therapy targeting retinal neovascularization.

No MeSH data available.


Related in: MedlinePlus

ES-GFP gene transfer suppresses HREC growth. Mean cell numbers were assessed for 5 consecutive days. Transfection with embryonic stem-green fluorescent protein (ES-GFP) using ultrasound-targeted cationic microbubble destruction caused a significant decrease in the human retinal vascular endothelial cell (HREC) cell viability rate compared with the other groups. (* p<0.05 relative to the control and cationic microbubble (CMB) groups, #p<0.01 relative to the control in every time point; n=6 per group).
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f3: ES-GFP gene transfer suppresses HREC growth. Mean cell numbers were assessed for 5 consecutive days. Transfection with embryonic stem-green fluorescent protein (ES-GFP) using ultrasound-targeted cationic microbubble destruction caused a significant decrease in the human retinal vascular endothelial cell (HREC) cell viability rate compared with the other groups. (* p<0.05 relative to the control and cationic microbubble (CMB) groups, #p<0.01 relative to the control in every time point; n=6 per group).

Mentions: The influence of all treatment factors on HREC cell viability was evaluated using the MTT assay. The cells treated with the ES-GFP plasmid together with NMBs and ultrasound, or with the ES-GFP plasmid with liposomes, or the ES-GFP plasmid with CMBs and ultrasound all underwent apoptotic-like cytological changes characterized by a loss of anchorage dependence and the assumption of a small spherical shape. The cells were collected and counted from day 1 through day 5, and the cell viability rate of the control group was 100%; the viability rate of the cells in each group was as shown in Figure 3. In multiple trials, the cell viability rate was observed to decrease significantly in the CMBs compared with the other groups.


Experimental endostatin-GFP gene transfection into human retinal vascular endothelial cells using ultrasound-targeted cationic microbubble destruction.

Xu Y, Xie Z, Zhou Y, Zhou X, Li P, Wang Z, Zhang Q - Mol. Vis. (2015)

ES-GFP gene transfer suppresses HREC growth. Mean cell numbers were assessed for 5 consecutive days. Transfection with embryonic stem-green fluorescent protein (ES-GFP) using ultrasound-targeted cationic microbubble destruction caused a significant decrease in the human retinal vascular endothelial cell (HREC) cell viability rate compared with the other groups. (* p<0.05 relative to the control and cationic microbubble (CMB) groups, #p<0.01 relative to the control in every time point; n=6 per group).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4548789&req=5

f3: ES-GFP gene transfer suppresses HREC growth. Mean cell numbers were assessed for 5 consecutive days. Transfection with embryonic stem-green fluorescent protein (ES-GFP) using ultrasound-targeted cationic microbubble destruction caused a significant decrease in the human retinal vascular endothelial cell (HREC) cell viability rate compared with the other groups. (* p<0.05 relative to the control and cationic microbubble (CMB) groups, #p<0.01 relative to the control in every time point; n=6 per group).
Mentions: The influence of all treatment factors on HREC cell viability was evaluated using the MTT assay. The cells treated with the ES-GFP plasmid together with NMBs and ultrasound, or with the ES-GFP plasmid with liposomes, or the ES-GFP plasmid with CMBs and ultrasound all underwent apoptotic-like cytological changes characterized by a loss of anchorage dependence and the assumption of a small spherical shape. The cells were collected and counted from day 1 through day 5, and the cell viability rate of the control group was 100%; the viability rate of the cells in each group was as shown in Figure 3. In multiple trials, the cell viability rate was observed to decrease significantly in the CMBs compared with the other groups.

Bottom Line: Cationic microbubbles (CMBs) were prepared and then compared with neutral microbubbles (NMBs) and liposomes.Moreover, the inhibition of HREC growth was enhanced following treatment with CMBs compared with NMBs or liposomes in vitro (p<0.01).These results suggest that the combination of UTMD and ES-GFP compounds might be a useful tool for gene therapy targeting retinal neovascularization.

View Article: PubMed Central - PubMed

Affiliation: Institute of Ultrasound Imaging and Department of Ultrasound, Second Affiliated Hospital of Chongqing Medical University; Chongqing Key Laboratory of Ultrasound Molecular Imaging, Chongqing, P.R. China.

ABSTRACT

Purpose: The purpose of this study was to investigate whether ultrasound-targeted cationic microbubble destruction could effectively deliver endostatin-green fluorescent protein (ES-GFP) plasmids to human retinal vascular endothelial cells (HRECs).

Methods: Cationic microbubbles (CMBs) were prepared and then compared with neutral microbubbles (NMBs) and liposomes. First, the two types of microbubbles were characterized in terms of size and zeta potential. The cell viability of the HRECs was measured using the 3-(4,5-dimthylthiazol-2-yl)-2,5 diphenyl-tetrazolium bromide (MTT) assay. The transcription and expression of endostatin, VEGF, Bcl-2, and Bcl-xl were measured via quantitative real-time PCR (qPCR) and western blotting, respectively.

Results: CMBs differed significantly from NMBs in terms of the zeta potential, but no differences in size were detected. Following ultrasound-targeted microbubble destruction (UTMD)-mediated gene therapy, the transcription and expression of endostatin were highest in the CMB group (p<0.05), while the transcription and expression of VEGF, Bcl-2, and Bcl-xl were lowest compared with the other groups. Moreover, the inhibition of HREC growth was enhanced following treatment with CMBs compared with NMBs or liposomes in vitro (p<0.01).

Conclusions: This study demonstrated that ultrasound-mediated cationic microbubbles could enhance the transfection efficiency of ES-GFP, which had obvious impacts on the inhibition of the growth process of HRECs in vitro. These results suggest that the combination of UTMD and ES-GFP compounds might be a useful tool for gene therapy targeting retinal neovascularization.

No MeSH data available.


Related in: MedlinePlus