Abstract
Purpose: :
To understand the effects of VEGF and IL-6 neutralization on the secretion of cytokines and angiogenic growth factors by ARPE-19 cells in vitro
Methods: :
ARPE-19 cells were exposed to different concentrations of anti-VEGF antibody (ranibizumab; RZB) (0.125 mg/ml and 0.25 mg/ml) or anti-IL6 antibody (0.05 µg/ml, 0.1 µg/ml and 0.2 µg/ml) for 24 hrs. The conditioned medium was analyszd for 12 cytokines (IL-1α,IL1-β,IL-2, IL-4,IL-6,IL-8,IL-10,IFN-γ,TNF-α,VEGF,MCP-1 and EGF) by Cytokine Biochip Array. mRNA expression was also studied using SYBR green real-time PCR analysis.
Results: :
Among the cytokines measured, significant levels of VEGF (687.2±9.3pg/ml), IL-6 (25.2±0.3pg/ml) IL-2 (5.1±1.2pg/ml), IFN-γ (22.4±4.7pg/ml), MCP-1(>840pg/ml) and IL-8(253.6±5.6pg/ml) were observed in 24 hrs culture supernatant. A significant reduction of VEGF (P<0.001) and also IL-6 and IL-2 (P<0.001) was observed on treatment with RZB. After 24hrs treatment, cells were cultured without RZB for 48 hrs and still VEGF was not detected in the conditioned medium. Moreover, neutralizing antibody to IL-6 reduced the secreted levels of VEGF (P<0.001) in the cell supernatant. However, subsequent culturing without anti-IL6 antibody did not affect the secretion of VEGF by ARPE 19 cells. These antibodies did not influence the secretion of other cytokines (IFN-γ and MCP-1). No remarkable changes in the mRNA expression of VEGF and IL-6 were found when treated with antibodies.
Conclusions: :
Anti-VEGF antibody reduced the secretion of IL-6 in addition to VEGF. Neutralizing IL-6 antibody also reduced VEGF expression by ARPE-19 cells. Further studies are required to understand the molecular mechanism between cytokines and angiogenic growth factor regulation in RPE.
Keywords: retinal pigment epithelium • age-related macular degeneration • cytokines/chemokines