Platelet-activating factor-induced intracellular signaling and release of cytokines and prostaglandin E2 in immortalized human corneal epithelial cells.
OBJECTIVE
Immortalized human corneal epithelial (CEPI-17-CL4) cells were exposed to different concentrations of platelet-activating factor (PAF) and the mobilization of intracellular calcium ([Ca(2+)](i)) was studied using fluorometrics. Additionally, the production of the cytokines [interleukin-6 (IL-6), interleukin-8 (IL-8), granulocyte macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor-alpha (TNF-alpha)], and matrix metalloproteinase-1 (MMP-1) and prostaglandin E(2) (PGE(2)) from PAF-stimulated cells was also determined using ELISA assays.


RESULTS
While PAF, histamine, and bradykinin stimulated the mobilization of [Ca(2+)](i) in these cells, PAF was the least efficacious. [Ca(2+)](i) mobilization induced by PAF was inhibited by 2 PAF receptor antagonists, PCA-42481 and CV-6209 (both 10 microM), and by a phospholipase C inhibitor, U73122 (60% at 4 microM). PAF increased the production of PGE(2) (with maximum effect at 30 and 100 nM) and GM-CSF (maximum effect at 1 microM) in CEPI-17-CL4 cells. However, PAF did not stimulate the generation of IL-6, IL-8, TNF-alpha, and MMP-1 to any significant level and in a consistent manner. PAF increased the incorporation of [(3)H]-thymidine into CEPI-17-CL4 cells with a maximal effect at 30 nM.


CONCLUSIONS
These data indicate that functional PAF receptors are present on CEPI-17-CL4 cells that can activate mobilization of [Ca(2+)](i). Other consequences of PAF receptor activation in CEPI-17-CL4 cells are the generation of PGE(2) and certain proinflammatory cytokines such as GM-CSF, and increasing cell proliferation.
Sharif
Najam A
NA
Pharmaceutical Products Research, Alcon Research, Ltd, Fort Worth, Texas 76134, USA. naj.sharif@alconlabs.com
Wiernas
Terry K
TK
D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000378:metabolism; D002461:Cell Line, Transformed; D049109:Cell Proliferation; D016207:Cytokines / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D019573:Epithelium, Corneal / Q000166:cytology / Q000378:metabolism*; D005470:Fluorometry; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism*; D020781:Matrix Metalloproteinase 1 / Q000096:biosynthesis; D009994:Osmolar Concentration; D010972:Platelet Activating Factor / Q000008:administration & dosage*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013936:Thymidine / Q000378:metabolism
D016207:Cytokines; D018836:Inflammation Mediators; D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D020781:Matrix Metalloproteinase 1; D015232:Dinoprostone; D002118:Calcium; D013936:Thymidine