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Silencing of Rac1 expression via RNA interference inhibits retinal neovascularization in rats.

Li J, Li Y, Zhang M, Hu Z - Mol. Vis. (2012)

Bottom Line: Two weeks after transfection, the neonatal vessels were tested using fluorescein isothiocyanate-dextran retinal angiography.The number of endothelial cells beyond the internal limiting membrane was counted with hematoxylin and eosin staining.In the shRNA interference group, the mean number of endothelial cells beyond the internal limiting membrane was significantly higher than that in the positive control group or the interference negative control group (p<0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Second People’s Hospital of Yunnan Province, Kunming, China. juanjuanlicn@yeah.net

ABSTRACT

Purpose: To investigate the inhibitory effect of Ras-related C3 botulinum toxin substrate 1-small interfering RNA (Rac1-siRNA) on retinal neovascularization in a rat model.

Methods: Rac1-short hairpin RNA (shRNA) was synthesized, constructed, and transfected into HeLa cells. Reverse transcription polymerase chain reaction was then conducted to test for Rac1 gene expression. Retinal vein obstruction was performed in 25 Sprague-Dawley rats using the retinal photodynamic method. The vitrea bulbus of the eye in the shRNA rats group was transfected with the Rac1-shRNA vector, the other eye in the blank control group was transfected with the blank vector, and the interference control group was prepared by transfecting the Rac1-shRNA vector. Two weeks after transfection, the neonatal vessels were tested using fluorescein isothiocyanate-dextran retinal angiography. The number of endothelial cells beyond the internal limiting membrane was counted with hematoxylin and eosin staining.

Results: A large area of neovascularization and fluorescein isothiocyanate leakage was found in the positive control group. However, a small area of neovascularization and a little fluorescence leakage were found in the shRNA group, whereas the retinal vessels were normal in the negative control interference group. In the shRNA interference group, the mean number of endothelial cells beyond the internal limiting membrane was significantly higher than that in the positive control group or the interference negative control group (p<0.05).

Conclusions: Silencing Rac1 expression with RNA interference inhibits retinal neovascularization in rats.

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Related in: MedlinePlus

Retina preparation by fluorescein isothiocyanate-dextran heart perfusion. A: The image of rat retinal angiography in the positive control group under a fluorescent microscope. Tortuous and thick abnormal new vessels were seen. B: The image of rat retinal angiography in the positive control group under fluorescent microscope. Fluorescent leakage of new vessel mass was seen. C: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Vessels were distributed in a disorganized manner. D: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Tortuous and thick new vessels were found. E: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Two layers of retinal vessels in normal distribution. F: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Deep vessels were normally arranged as a net.
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f2: Retina preparation by fluorescein isothiocyanate-dextran heart perfusion. A: The image of rat retinal angiography in the positive control group under a fluorescent microscope. Tortuous and thick abnormal new vessels were seen. B: The image of rat retinal angiography in the positive control group under fluorescent microscope. Fluorescent leakage of new vessel mass was seen. C: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Vessels were distributed in a disorganized manner. D: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Tortuous and thick new vessels were found. E: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Two layers of retinal vessels in normal distribution. F: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Deep vessels were normally arranged as a net.

Mentions: The retinal angiography of the positive control group showed that the retinal vessels had abnormal morphologies and distribution. Many abnormal neonatal vessel sprouts and neovascularization were observed, and fluorescent leakage exhibited strong fluorescence (Figure 2A,B). The shRNA group had abnormal retinal vessel morphology and distribution. New vessels and fluorescent leakage were observed but without a large area of neovascularization (Figure 2 C,D). The retinal vessel morphology in the negative control group was normal, and aberrant vessels or fluorescent leakage was not observed (Figure 2 E,F).


Silencing of Rac1 expression via RNA interference inhibits retinal neovascularization in rats.

Li J, Li Y, Zhang M, Hu Z - Mol. Vis. (2012)

Retina preparation by fluorescein isothiocyanate-dextran heart perfusion. A: The image of rat retinal angiography in the positive control group under a fluorescent microscope. Tortuous and thick abnormal new vessels were seen. B: The image of rat retinal angiography in the positive control group under fluorescent microscope. Fluorescent leakage of new vessel mass was seen. C: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Vessels were distributed in a disorganized manner. D: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Tortuous and thick new vessels were found. E: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Two layers of retinal vessels in normal distribution. F: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Deep vessels were normally arranged as a net.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369893&req=5

f2: Retina preparation by fluorescein isothiocyanate-dextran heart perfusion. A: The image of rat retinal angiography in the positive control group under a fluorescent microscope. Tortuous and thick abnormal new vessels were seen. B: The image of rat retinal angiography in the positive control group under fluorescent microscope. Fluorescent leakage of new vessel mass was seen. C: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Vessels were distributed in a disorganized manner. D: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Tortuous and thick new vessels were found. E: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Two layers of retinal vessels in normal distribution. F: The image of rat retinal angiography in the gene interference group under a fluorescent microscope. Deep vessels were normally arranged as a net.
Mentions: The retinal angiography of the positive control group showed that the retinal vessels had abnormal morphologies and distribution. Many abnormal neonatal vessel sprouts and neovascularization were observed, and fluorescent leakage exhibited strong fluorescence (Figure 2A,B). The shRNA group had abnormal retinal vessel morphology and distribution. New vessels and fluorescent leakage were observed but without a large area of neovascularization (Figure 2 C,D). The retinal vessel morphology in the negative control group was normal, and aberrant vessels or fluorescent leakage was not observed (Figure 2 E,F).

Bottom Line: Two weeks after transfection, the neonatal vessels were tested using fluorescein isothiocyanate-dextran retinal angiography.The number of endothelial cells beyond the internal limiting membrane was counted with hematoxylin and eosin staining.In the shRNA interference group, the mean number of endothelial cells beyond the internal limiting membrane was significantly higher than that in the positive control group or the interference negative control group (p<0.05).

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Second People’s Hospital of Yunnan Province, Kunming, China. juanjuanlicn@yeah.net

ABSTRACT

Purpose: To investigate the inhibitory effect of Ras-related C3 botulinum toxin substrate 1-small interfering RNA (Rac1-siRNA) on retinal neovascularization in a rat model.

Methods: Rac1-short hairpin RNA (shRNA) was synthesized, constructed, and transfected into HeLa cells. Reverse transcription polymerase chain reaction was then conducted to test for Rac1 gene expression. Retinal vein obstruction was performed in 25 Sprague-Dawley rats using the retinal photodynamic method. The vitrea bulbus of the eye in the shRNA rats group was transfected with the Rac1-shRNA vector, the other eye in the blank control group was transfected with the blank vector, and the interference control group was prepared by transfecting the Rac1-shRNA vector. Two weeks after transfection, the neonatal vessels were tested using fluorescein isothiocyanate-dextran retinal angiography. The number of endothelial cells beyond the internal limiting membrane was counted with hematoxylin and eosin staining.

Results: A large area of neovascularization and fluorescein isothiocyanate leakage was found in the positive control group. However, a small area of neovascularization and a little fluorescence leakage were found in the shRNA group, whereas the retinal vessels were normal in the negative control interference group. In the shRNA interference group, the mean number of endothelial cells beyond the internal limiting membrane was significantly higher than that in the positive control group or the interference negative control group (p<0.05).

Conclusions: Silencing Rac1 expression with RNA interference inhibits retinal neovascularization in rats.

Show MeSH
Related in: MedlinePlus