Global Gene Expression Analysis of Keloid Fibroblasts in Response to Electron Beam Irradiation Reveals the Involvement of Interleukin-6 Pathway

Abstract

Keloid is a dermal fibroproliferative lesion of unknown etiology that commonly recurs after surgical excision. Post-operative adjuvant electron beam (EB) irradiation has been successfully used to reduce keloid recurrences. To provide new insights into the molecular mechanism behind the effect of EB irradiation, we used a cDNA microarray screening of more than 5000 genes to assess early changes in gene expression between EB-irradiated and non-irradiated keloid and non-lesional dermal fibroblasts. Primary fibroblast cultures from keloid and associated non-lesional dermis obtained from five patients were exposed to 15 Gy EB irradiation and analyzed after 15 min incubation. Early response to EB irradiation showed that 96 (1.8%) genes were modulated 2-fold or more in keloid fibroblasts. Upregulated genes accounted for 29.2% (28 genes), whereas downregulated genes comprised 70.8% (68 genes), indicating a silencing of many genes in keloid fibroblasts after EB irradiation. Many of the downregulated genes play roles in the enhancement of cell proliferation and extracellular matrix production, whereas several of the upregulated genes involves in the promotion of apoptosis and extracellular matrix (ECM) degradation. Using emerging bioinformatic tools and further corroboration, the interleukin 6 (IL-6) signaling pathway was found to be mainly involved in EB irradiation response. We also showed co-expression of IL-6 and its specific receptor (IL-6Ralpha) in keloid fibroblasts that points to the existence of an IL-6 autocrine loop in these cells. These results suggested that at the molecular level, EB irradiation might hinder keloid formation by regularizing disturbances in the homeostatic equilibrium between inducer and inhibitor activities in the matrix system most likely through the IL-6 pathway. Our study provides clues for the molecular mechanism(s) behind the beneficial effect of EB irradiation in reducing keloid recurrences and may help develop alternative strategies for the therapy and prophylaxis of this lesion.