Abstract
Purpose: :
Nucleolin is a one of the most abundant non-ribosomal proteins of the nucleolus. This protein has different functions depending on its cell localization. It has been well documented that nuclear expression is associated with cell proliferation due to its role in transcription and RNA maturation. In the same way, it has been recently reported that vascular endothelial growth factor (VEGF) induces transmembranal expression of nucleolin in tumor endothelial cells and this expression is associated with cell proliferation. Likewise, it has been demonstrated that a selective inhibition of cell surface nucleolin inhibits tumor vascularization. The aim of this study was to determine whether VEGF induced limbal fibroblasts proliferation and its association with nucleolin expression.
Methods: :
Human limbal fibroblasts were obtained from cadaveric donor sclera-corneal rims. Limbal fibroblasts were cultured in the presence of different concentrations of rhVEGF165 for 24 h. The expression of nucleolin was identified by immunocytochemistry using a rabbit polyclonal mouse anti-human nucleolin. Cell proliferation was determined by MTT technique.
Results: :
Vascular endothelial growth factor was able to induce nucleolin expression in limbal fibroblasts in a dose response manner with a peak in 10 ng/ml. The expression was identified mainly in the nucleus of dividing cells. When the cells were stimulated with 100 ng/ml, perinuclear and cytoplasmic nucleolin expression was observed. Meanwhile, low concentrations of rhVEGF165 such as 1 ng/ml induced fibroblasts proliferation.
Conclusions: :
These results indicate that rhVEGF165 is able to induce proliferation on a primary cell culture of limbal fibroblasts. Nucleolin protein expression was induced by VEGF. Moreover, nucleolin cell localization was dependent in the concentration of VEGF. These results indicate that nucleolin expression and cell localization depend in different VEGF concentrations and as reported by others, inhibiting nucleolin protein expression reduces angiogenesis, this molecule could be a target to treat corneal neovascularization.
Keywords: vascular endothelial growth factor • protein structure/function • cell survival