Upregulation of Connexin43 Expression in Corneal Fibroblasts by Corneal Epithelial Cells

• Published in: Investigative ophthalmology & visual science 2009-05, Vol.50 (5), p.2054-2060
• Main Authors: Ko, Ji-Ae, Yanai, Ryoji, Morishige, Naoyuki, Takezawa, Toshiaki, Nishida, Teruo
• Format: Article
• Language: English
• Subjects: Adolescent; Adult; Aged; Biological and medical sciences; Cells, Cultured; Child; Child, Preschool; Coculture Techniques; Connexin 43 - genetics; Connexin 43 - metabolism; Cornea - metabolism; Enzyme Inhibitors - pharmacology; Epithelial Cells - physiology; Eye and associated structures. Visual pathways and centers. Vision; Female; Fibroblasts - metabolism; Fluorescent Antibody Technique, Indirect; Fundamental and applied biological sciences. Psychology; Gap Junctions; Gene Expression Regulation - physiology; Humans; Immunoblotting; Insulin-Like Growth Factor I - genetics; Male; Medical sciences; Middle Aged; Mitogen-Activated Protein Kinases - antagonists & inhibitors; Mitogen-Activated Protein Kinases - physiology; Ophthalmology; p38 Mitogen-Activated Protein Kinases - antagonists & inhibitors; p38 Mitogen-Activated Protein Kinases - physiology; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; RNA, Small Interfering - physiology; Tissue Donors; Transfection; Up-Regulation; Vertebrates: nervous system and sense organs
• ISSN: 0146-0404; 1552-5783; 1552-5783
• DOI: 10.1167/iovs.08-2418

The authors have previously shown that the expression of tight-junction proteins in corneal epithelial cells is regulated by corneal fibroblasts in a coculture system. They have now examined the effect of corneal epithelial cells on the expression of the gap-junction protein connexin43 in corneal fibroblasts. Human corneal fibroblasts and simian virus 40-transformed human corneal epithelial (HCE) cells were cultured on opposite sides of a collagen vitrigel membrane. Expression of junctional proteins in corneal fibroblasts was examined by reverse transcription-polymerase chain reaction, immunoblot, and immunofluorescence analyses. The effect of a small interfering RNA specific for insulin-like growth factor-1 (IGF-1) mRNA on connexin43 expression was determined by transfection. The amounts of connexin43 mRNA and protein in corneal fibroblasts were increased by the presence of HCE cells. HCE cells had no effect on the expression of the tight-junction proteins ZO-1, occludin, and claudin in corneal fibroblasts. The effect of HCE cells on connexin43 expression was mimicked by exposure of corneal fibroblasts to IGF-1. Furthermore, depletion of IGF-1 in HCE cells by RNA interference largely abolished the effect of these cells on connexin43 expression in corneal fibroblasts. Finally, the upregulation of connexin43 expression in corneal fibroblasts by HCE cells was blocked by inhibitors of signaling by the mitogen-activated protein kinases ERK (PD98059) and p38 MAPK (SB203580). The presence of corneal epithelial cells upregulated the expression of connexin43 in corneal fibroblasts, suggesting that corneal epithelial cells are important for the maintenance of gap junction-mediated communication among corneal fibroblasts.