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A. Takase, A. Kato, T. Yasukawa, F. Hirata, Y. Ogura; Suppressive Effect of Short Interfering RNA on Hyperglycemia–Induced Expression of Intercellular Adhesion Molecule–1 (ICAM–1) on Cultured Vascular Endothelial Cells . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1729.
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© ARVO (1962-2015); The Authors (2016-present)
,The pathology of diabetic retinopathy involves vascular endothelial dysfunction, in which leukocyte adhesion to vascular endothelium via intercellular adhesion molecule–1 (ICAM–1) may play a key role. Short interfering RNAs (siRNAs) are unique modulators of gene expression in mammalian cells. The purpose of this study is to evaluate the suppressive effect of siRNAs on hyperglycemia–induced expression of ICAM–1 on cultured vascular endothelial cells.
Human umbilical vein endothelial cells (HUVECs) were seeded onto 24–well culture plates. On the next day, transfection of ICAM–1 specific siRNAs was performed by use of Lipofectamine 2000. Then glucose (15, 30, or 45 mM) was added to the medium for stimulation of ICAM–1.IL–1ß was used as a positive control. After 48 hours, HUVECs were collected and ICAM–1 was measured by enzyme–linked immunosolvent assay. mRNA of ICAM–1 was measured by quantitative RT–PCR. Fluoresceinated siRNAs was transfected into HUVECs to confirm the transfection by fluorescein microscopy.
Glucose enhanced the expression of ICAM–1 and its mRNA in the dose–dependent manner. The expression of ICAM–1 increased by hyperglycemia was significantly reduced in HUVECs transfected with corresponding siRNAs. The expression of mRNA was also reduced by siRNAs. Effectiveness of siRNAs transfection was confirmed with enhanced fluorescence in HUVECs incubated with control siRNAs.
We conclude that hyperglycemia stimulated mRNA and protein expression of ICAM–1 and that siRNAs suppressed gene expression of ICAM–1 in HUVECs. These findings suggest that RNA–targeting approach using siRNAs might provide a novel therapeutic option for diabetic retinopathy.
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