Protection of human conjunctival fibroblasts from NO-induced apoptosis by interleukin-4 or interleukin-13.
OBJECTIVE
To examine the possible roles of T helper type 2 (Th2) cell-derived cytokines in formation of the giant papillae characteristic in individuals with vernal keratoconjunctivitis, the effects of these cytokines on the proliferation and apoptosis of cultured human conjunctival fibroblasts were investigated.


METHODS
Apoptosis was induced by the NO donor sodium nitroprusside. Cell viability was determined by measurement of mitochondrial metabolic activity, and apoptotic cells were identified on the basis either of nuclear morphology after staining with 4',6-diamidino-2-phenylindole or of TUNEL staining. The activation of antiapoptotic signaling mediated by the protein kinase Akt was assessed by immunoblot analysis and by an in vitro kinase assay. Expression of interleukin (IL)-4 and IL-13 receptor subunits was examined by reverse transcription and polymerase chain reaction analysis and by flow cytometry.


RESULTS
IL-4 and IL-13, but not IL-5, IL-9, or IL-10, induced the proliferation of conjunctival fibroblasts as well as protecting these cells from NO-induced apoptosis. Both IL-4 and IL-13 induced the phosphorylation of Akt and increased the kinase activity of this enzyme in a manner that was sensitive to the phosphatidylinositol 3-kinase inhibitors LY294002 or wortmannin. These inhibitors also blocked the antiapoptotic effects of IL-4 and IL-13. Transcripts encoding IL-4 and IL-13 receptor components were detected in conjunctival fibroblasts, and the proteins were expressed at the cell surface.


CONCLUSIONS
Among the various Th2 cytokines tested, only IL-4 and IL-13 induced the proliferation of human conjunctival fibroblasts and protected these cells from apoptosis. These effects may contribute to the formation of giant papillae in individuals with vernal keratoconjunctivitis.
Fujitsu
Youichiro
Y
Department of Biomolecular Recognition, Yamaguchi University School of Medicine, Ube City, Yamaguchi 755-8505, Japan.
Fukuda
Ken
K
Kimura
Kazuhiro
K
Seki
Keisuke
K
Kumagai
Naoki
N
Nishida
Teruo
T
D000730:Androstadienes / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D003228:Conjunctiva / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D019610:Cytoprotection; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D018793:Interleukin-13 / Q000494:pharmacology*; D053661:Interleukin-13 Receptor alpha1 Subunit; D015847:Interleukin-4 / Q000494:pharmacology*; D009025:Morpholines / Q000494:pharmacology; D020030:Nitric Oxide Donors / Q000633:toxicity*; D009599:Nitroprusside / Q000633:toxicity; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D018123:Receptors, Interleukin / Q000235:genetics / Q000378:metabolism; D053660:Receptors, Interleukin-13; D019948:Receptors, Interleukin-4 / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; C508181:IL13RA1 protein, human; D018793:Interleukin-13; D053661:Interleukin-13 Receptor alpha1 Subunit; D009025:Morpholines; D020030:Nitric Oxide Donors; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D018123:Receptors, Interleukin; D053660:Receptors, Interleukin-13; D019948:Receptors, Interleukin-4; D009599:Nitroprusside; D015847:Interleukin-4; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D000077191:Wortmannin