Differential transcriptional responses of keloid and normal keratinocytes to serum stimulation

Background. Keloids are benign tumors that occur only in response to injury, for which there is no effective treatment. We demonstrated previously that keloid keratinocytes (KKs) promote fibroblast proliferation more than normal keratinocytes (NKs) and that transforming growth factor (TGF)-beta is a component of that signal. We used the transcriptional response to serum stimulation to examine how TGF-beta expression is stimulated in KKs. Materials and methods. Quiescent KKs and NKs were stimulated using serum; harvested using RNA at 0, 1, 6, 12, and 24 h; and analyzed usingquantitative real-time polymerase chain reaction. TGF-beta activity in the conditioned medium was measured with an MLEC/PAI-luciferase assay. Inhibition of ERK1/2, p38 kinase, and JNK pathways was performed with PD98059, SB203580, and SP600125, respectively. Results. Increased transcription of TGF-beta 2 occurs within 1 h of serum stimulation in KKs but not in NKs. In contrast, TGF-beta 3 transcription was suppressed in KKs compared with NKs. No significant differences were observed in the transcriptional response of TGF-beta 1. Increased TGF-beta 2 mRNA correlated with increased TGF-beta biological activity in the conditioned medium. Inhibition of the ERK, p38 kinase or JNK signal transduction pathways blocked the transcriptional up-regulation of TGF-beta 2, T beta R1, and T beta R2 in KKs. Conclusions. KKs produce more TGF-beta 2 mRNA than NKs in response to serum stimulation, resulting in increased TGF-beta activity in conditioned medium. Combining these results with our previous data lead us to propose a model of keloid formation characterized by an exaggerated response to cellular stress and abnormal epithelial-mesenchymal signaling promoting keloid formation. (c) 2006 Elsevier Inc. All rights reserved.