Nuclear-Factor Kappa B: A Potential Role in Pterygium Formation

Abstract

Purpose: : Pterygium is a common ocular surface disease manifesting as a fibrovascular conjunctival outgrowth that invades the cornea. We seek to elucidate the role of Nuclear Factor kappa B (NF-ΚB) pathway in the pathogenesis of pterygium.

Methods: : Surgically excised pterygium tissues, obtained with written informed consent in accordance with declaration of Helsinki, were studied compared to normal uninvolved conjunctiva. Immunohistochemistry, immunoblotting, and nuclear DNA binding assays against p65, Rel B, p50, p52 and c-Rel were performed to evaluate activation of various NF-ΚB members. Levels of phosphorylated inhibitor IΚB were evaluated by ELISA. Expression of NF-ΚB target genes were studied using U133A genechip microarray. Primary fibroblasts culture from pterygium explants were stimulated with increasing doses of TNF- (20ng/ml to 100ng/ml) and UVB irradiation (5mJ/cm² to 30mJ/cm²) and immunocytochemistry, immunoblotting against above-mentioned proteins and real-time PCR against NF-ΚB target genes were studied.

Results: : p65 and p105/p50 were expressed in pterygium and conjunctival epithelium on immunohistochemistry. Increased nuclear p65 DNA binding activity indicated increased NF-ΚB activation in pterygium tissues compared to normal uninvolved conjunctiva. Levels of phosphorylated IΚB were increased in pterygium tissues using ELISA. Cultured pterygium fibroblasts showed p65 nuclear translocation after exposure to 20ng/ml TNF- and 10mJ/cm² of UVB irradiation on immunocytochemistry. Real-time PCR showed increased transcription of IΚB, IL-6, IL-8 within 30min after UVB and TNF- stimulation, and delayed increase in MCP-1 transcription beyond 2 hours, and Rel B, MMP2, MMP3 and MMP9 beyond 12 hours after TNF- stimulation, and MMP 1 beyond 12 hours after both UVB and TNF- stimulation.

Conclusions: : The NF-ΚB pathway is activated in pterygium. Stimulation of pterygium fibroblasts by TNF- and UVB can activate NF-ΚB pathway. The control of transcription of interleukins and MMP genes may be controlled by different members of the NF-ΚB family.

Keywords: pterygium • wound healing • transcription factors