Synthesis of ENA-78 and GCP-2 Are Differentially Regulated in Human Corneal Keratocyte and Epithelial Cells

Abstract

Abstract: : Purpose: To determine if interleukin-1α- (IL-1α) and tumor necrosis factor-α- (TNF-α) stimulated human corneal epithelial cells (HCEC) and human corneal keratocytes (HCK) produce the neutrophil chemotactic chemokines epithelial cell-derived neutrophil attractant-78 (ENA-78) and granulocyte chemotactic protein (GCP-2). Methods: Cultures of HCEC and HCK were stimulated with human recombinant IL-1α or TNF-α. At selected times post-stimulation, culture supernatants were harvested and assayed for ENA-78 and GCP-2 by enzyme-linked immunosorbent assay. RNA was extracted from cell cultures to measure steady-state levels of intracellular ENA-78 and GCP-2 pre-mRNA and mRNA by the reverse transcriptase-polymerase chain reaction. Results: Exposure of HCEC with either IL-1α or TNF-α stimulated a >4.5-fold increase in ENA-78 RNA and protein synthesis without stimulating a significant increase in either GCP-2 RNA synthesis or protein production. Exposure of HCK to IL-1α stimulated a 10-fold increase in ENA-78 and GCP-2 RNA synthesis and >300-fold increase in ENA-78 and GCP-2 protein production. In contrast, exposure of keratocytes to TNF-α significantly enhanced ENA-78 RNA synthesis resulting in a >68-fold increase in ENA-78 protein synthesis without significantly enhancing either GCP-2 RNA synthesis or protein secretion. Conclusions: The results suggest that ENA-78 synthesis is stimulated in corneal epithelial cells and keratocytes whenever IL-1α or TNF-α is released into corneal tissue as a result of disease or injury. In contrast, due to fact that its synthesis can only be induced in keratocytes exposed to IL-1α, GCP-2 gene expression is more tightly regulated than that for ENA-78.

Keywords: inflammation • cytokines/chemokines • cornea: basic science