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Effect of Connective Tissue Growth Factor on Protein Kinase Expression and Activity in Human Corneal Fibroblasts
Author(s) -
Radhakrishnan S.S.,
Blalock T.D.,
Schultz G.S.
Publication year - 2004
Publication title -
wound repair and regeneration
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.847
H-Index - 109
eISSN - 1524-475X
pISSN - 1067-1927
DOI - 10.1111/j.1067-1927.2004.0abstractcx.x
Subject(s) - ctgf , protein kinase a , focal adhesion , microbiology and biotechnology , kinase , map2k7 , ask1 , connective tissue , chemistry , cyclin dependent kinase 2 , signal transduction , biology , growth factor , biochemistry , receptor , genetics
Purpose : To evaluate expression patterns of protein kinases and protein kinase activities in human corneal fibroblasts treated with connective tissue growth factor (CTGF). Methods : Human corneal fibroblast cultures were grown to confluence and treated with CTGF for 0, 5, and 15 minutes. Cytoplasmic protein extracts were obtained; protein kinase expression and activity arrays were performed using Kinetworks™ analysis (screening for expression of 75 different protein kinases and 31 different phosphoproteins). Further studies using extended time courses of CTGF exposure in corneal fibroblasts (0, 1, 2, 3, 4, 5, 10, 15, 30, and 60 minutes) were performed using immunoblot analysis to detect expression of protein kinase A catalytic subunit (PKA‐cat), and focal adhesion kinase (FAK). All results and were normalized by comparison to beta‐actin. Results : After 5 minutes of exposure to CTGF, levels of active proteins increased for 21 of the 75 kinases analyzed. Some notable protein kinases that were induced include: death‐associated kinase 1, focal adhesion kinase (FAK), G‐protein coupled receptor kinase 2, protein kinase A catalytic subunit (PKA‐cat), protein kinase B alpha, and protein kinase C (ε, μ, and ζ, subunits). Extended time course analysis of PKA‐cat and FAK showed statistically significant increases in expression following CTGF stimulation within 15 minutes. Conclusion : CTGF increased the levels of active protein kinases in human corneal fibroblast cultures, including PKA‐cat and FAK after 5 minutes of exposure. These results further our understanding of the signal transduction mechanism activated by CTGF in corneal fibroblasts. This suggests that CTGF mediates the effects of transforming growth factor beta (TGF‐β) on protein kinase expression and phosphorylation of second messengers in processes such as cell proliferation and collagen synthesis. Acknowledgments: Funding source: NEI 05587