title: Mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3 kinase (PI3K) pathways differently regulate retinal pigment epithelial cell-mediated collagen gel contraction.
abstract: Retinal pigment epithelial (RPE) cell-mediated extracellular matrix contraction is believed to contribute to developing proliferative vitreoretinopathy. It has been shown that platelet-derived growth factor (PDGF) and its intracellular signaling pathway, including mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3 kinase (PI3K), are mainly involved in this process. The aim of this study is to investigate how these downstream signaling pathways are related to RPE-mediated collagen gel contraction. We performed the gel contraction assay to evaluate the effect of PDGF in cultured ARPE-19 cells under the presence or absence of PD98059, MAPK inhibitor or wortmannin, PI3K inhibitor. Experiments treated with neutralizing antibody for various subtypes of integrin were also performed and the effect on PDGF-induced gel contraction was investigated. Expression changes of integrin alpha1, alpha2 and beta1 after PDGF stimulation was evaluated using quantitative real-time PCR and flow cytometry. The results showed that PDGF up-regulated ARPE-19 cell-mediated gel contractile activity. PDGF-induced collagen gel contraction was attenuated under presence of PD98059, wortmannin, or neutralizing antibody for integrin alpha1, alpha2, or beta1, all of which are critical subset for binding with type I collagen. The expression of integrin alpha1 and alpha2 was increased after PDGF stimulation in both real-time PCR and flow cytometry, however beta1 expression was not increased. PD98059 significantly attenuated integrin alpha1 and alpha2 expressions. However, wortmannin did not have the same effect. In conclusion, PDGF promotes ARPE-19 cell-mediated gel contraction via both MAPK and PI3K. This was probably due to an increased expression of integrin alpha1 and alpha2, which is mediated by MAPK, but not by PI3K. PI3K may regulate collagen gel contraction by another mechanism other than the up-regulation of integrin expression.
authors.affiliation: Department of Ophthalmology, Osaka University Medical School, Osaka, Japan. bando@ophthal.med.osaka-u.ac.jp
authors.forename: Hajime
authors.initials: H
authors.lastname: Bando
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Ikuno
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Hori
authors.forename: Kaori
authors.initials: K
authors.lastname: Sayanagi
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Tano
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D002460:Cell Line; D003094:Collagen; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005109:Extracellular Matrix / Q000201:enzymology / Q000473:pathology; D005419:Flavonoids / Q000494:pharmacology; D005434:Flow Cytometry; D005782:Gels; D006801:Humans; D039423:Integrin alpha1 / Q000378:metabolism; D039421:Integrin alpha2 / Q000378:metabolism; D019012:Integrin beta1 / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010857:Pigment Epithelium of Eye / Q000201:enzymology* / Q000473:pathology; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D013268:Stimulation, Chemical; D018630:Vitreoretinopathy, Proliferative / Q000201:enzymology* / Q000473:pathology; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D003094:Collagen; D004791:Enzyme Inhibitors; D005419:Flavonoids; D005782:Gels; D039423:Integrin alpha1; D039421:Integrin alpha2; D019012:Integrin beta1; D020928:Mitogen-Activated Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010982:Platelet-Derived Growth Factor; D000077191:Wortmannin; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one