Thymosin Beta 4 Modulates Inflammatory Mediators in Human Corneal Epithelial Cells

Abstract

Abstract: : Purpose: Previous studies have shown that thymosin beta 4 (Tß₄) promotes corneal wound healing and decreases inflammation. This study investigated cytokine mRNA and protein expression levels in human corneal epithelial cells (HCEC) after treatment with and without Tß4 in the presence or absence of recombinant interleukin (IL)-1ß. Methods: HCEC were grown to 60% of confluence in serum-free medium under four different conditions: 1) medium alone (control), 2) Tß₄ (1000 ng/ml), 3) IL-1ß (1000 units/ml), 4) Tß₄ (1000 ng/ml) plus IL-1ß (1000 U/ml) for 2, 8, 16, and 24 hours. mRNA was isolated and analyzed for cytokine gene transcript levels. An average of ≥2-fold change in gene expression between the experimental groups and controls was considered significant. Additionally, cell lysates and culture supernatants were analyzed by Western blot to determine cytokine protein expression levels. Results: Gene transcript levels for IL-1ß, IL-18, IL-19, IL-22, IL-9/p40, and transforming growth factor (TGF)–α in HCEC were decreased in Tß₄ and Tß₄ plus IL-1ß-treated HCEC compared to their respective control groups at the 8 hour time point. Further, mRNA expression of IL-6 and IL-8, which were induced in HCEC by treatment with recombinant IL-1ß only, also decreased after 8 hours of Tß₄ plus IL-1ß treatment. Western blot analysis was consistent with the mRNA expression data and confirmed decreased protein levels of IL-1ß, IL-8, and IL-18 after Tß₄ treatment. Conclusion: These data suggest that Tß₄ inhibits the transcription of many key pro-inflammatory cytokines including IL-1ß induced cytokines in human corneal epithelial cells. These findings suggest that Tß₄ may promote corneal wound healing by modulating inflammatory mediators.

Keywords: cornea: epithelium • cytokines/chemokines • inflammation