VEGF165b Protects Primary Human Retinal Epithelial Cells from TNF-α Mediated Inhibition of Occludin Expression

Abstract

 

Purpose:

 

Retinal pigmented epithelial cells (RPE) are considered to be the first line of defense against infections and inflammation in the retina. In ocular inflammation, lymphocytes and macrophages migrate to the posterior compartment of the eye and secrete pro-inflammatory mediators such as tumor necrosis factor (TNF)-α that can target and impair RPE barrier functions. TNF-α causes epithelial barrier dysfunction in vitro and in vivo and is accompanied by redistribution of the tight junction protein occludin. VEGF₁₆₅b is generated by alternative splicing of VEGF-A in the terminal exon 8. VEGF₁₆₅b is cytoprotective and antiangiogenic in retina. Therefore, our aim was to investigate the protective effects of VEGF₁₆₅b on TNF-α induced barrier dysfunction in RPE.

 

Methods:

 

Primary human RPE were treated with either 100 ng/ml TNF-α alone or TNF-α and 2.5 nM VEGF₁₆₅b. Whole-cell extracts were assayed for occludin by Western blotting and cells were evaluated by immunocytochemistry for occludin after treatment.

 

Results:

 

TNF-α decreased the expression of occludin whereas VEGF₁₆₅b inhibited TNF-α effect in RPE cells as determined by immunofluroescence (figure 1A). Western blotting indicated that overall expression was reduced to 60% of control with TNF-α which increased to 87% of control when VEGF₁₆₅b was included with the TNF-α

 

Conclusions:

 

These findings indicate that VEGF₁₆₅b inhibits the TNF-α mediated downregulation of occludin expression RPE cells, suggesting a protective property of VEGF₁₆₅b in reducing TNF-α mediated inflammation in the eye.  

 

Keywords: retinal pigment epithelium • inflammation • vascular endothelial growth factor