Phospholipases A2 and C Content of Human Corneal Epithelial Cells

Abstract

Abstract: : Purpose : In order to achieve a better understanding of the inflammation process in human corneal epithelial cells, this study was undertaken to analyze the phospholipases A2 (PLA2) and phospholipases C (PLC) content of human corneal epithelium. Methods : Specific primers were designed to amplify the known cytosolic (cPLA2) and secreted (sPLA2) phospholipases A2 and phospholipases C mRNA’s by RT-PCR. Corresponding PCR products were cloned and amplified in order to be sequenced. Immunofluorescence of flatmounted corneal sections and Western blotting were used to detect the PLA2s and PLCs expressed by corneal epithelial cells. Results : The RNA of the following phospholipases was detected by RT-PCR in the corneal epithelial cells : group X sPLA2, cPLA2α, cPLA2ß and cPLA2γ, PLC-ß1, PLC-ß2, PLC-ß3, PLC-γ1, PLC-γ2, PLC-δ3 and PLC-ε. Immunofluorescence studies on corneal epithelium sections demonstrated the presence of sPLA2 X, PLC-ß2, PLC-ß3, PLC-γ1 and PLC-γ2. It. was not possible to verify the presence of the different cPLA2s and the PLC-δ3 by immunological techniques since the specific antibodies for these phospholipases are not yet available. The results obtained by immunofluorescence on tissue section were confirmed by Western blotting on freshly isolated corneal epithelial cells. Conclusions : Many phospholipase isoforms are present in corneal epithelial cells. These should play a major role in signal transduction (PLC) as well as in the release of precursors of potent mediators of inflammation such as leukotrienes and prostaglandines (PLA2). The individual role of each phospholipase will have to be determined in order to achieve a better understanding of their involvement in inflammation.

Keywords: cornea: epithelium • enzymes/enzyme inhibitors • gene/expression