Thrombin stimulates RPE cell motility by PKC-zeta- and NF-kappaB-dependent gene expression of MCP-1 and CINC-1/GRO chemokines.
Retinal pigment epithelial cells (RPE) are the major cell type involved in the pathogenesis of proliferative vitreoretinopathy (PVR), which involves the epithelial-mesenchymal transition, proliferation, and directional migration of transformed RPE cells to the vitreous upon RPE exposure to serum components, thrombin among them. Although the aqueous humor and vitreous of PVR patients contain high levels of chemokines, their possible involvement in PVR development has not been explored. We here analyzed the effect of thrombin on chemokine gene expression and its correlation with RPE cell migration using rat RPE cells in culture as a model system. We demonstrated that thrombin induces RPE cell migration through the dose-dependent stimulation of MCP1 and GRO expression/release, and the autocrine activation of CXCR-2 and CCR-2 chemokine receptors. Whereas inhibition of CXCR2 by Sb-225002 and of CCR2 by Rs-504393 partially prevented hirudin-sensitive cell migration, the joint inhibition of these receptors abolished thrombin effect, suggesting the contribution of distinct but coincident mechanisms. Thrombin effects were not modified by Ro-32-0432 inhibition of conventional/novel PKC isoenzymes or by the MAPkinase pathway inhibitor U0126. MCP1 and GRO expression/secretion, and cell migration were completely prevented by the inhibitory PKC-zeta pseudosubstrate and by the nuclear factor-kappa B (NF-kappaB) inhibitor BAY11-7082, but not by wortmannin inhibition of PI3K. Results show that signaling pathways leading to RPE cell migration differ from the MEK-ERK-PI3K-mediated promotion RPE of cell proliferation, both of which concur at the activation of PKC-zeta.
Palma-Nicolás
José Prisco
JP
División de Neurociencias, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México (UNAM), Coyoacan, México, DF.
López
Edith
E
López-Colomé
Ana María
AM
D000818:Animals; D001483:Base Sequence; D002465:Cell Movement / Q000187:drug effects*; D018932:Chemokine CCL2 / Q000235:genetics*; D054360:Chemokine CXCL1 / Q000235:genetics*; D017931:DNA Primers; D015870:Gene Expression; D016328:NF-kappa B / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D020318:Rats, Long-Evans; D055213:Retinal Pigment Epithelium / Q000166:cytology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013917:Thrombin / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects; D014945:Wound Healing; D000099221:Protein Kinase C zeta
D018932:Chemokine CCL2; D054360:Chemokine CXCL1; D017931:DNA Primers; D016328:NF-kappa B; D011493:Protein Kinase C; D013917:Thrombin; D000099221:Protein Kinase C zeta; C513297:Cxcl1 protein, rat