Evidence for ERK1/2 phosphorylation controlling contact inhibition of proliferation in Madin-Darby canine kidney epithelial cells

• Published in: American Journal of Physiology: Cell Physiology 2004-08, Vol.287 (2), p.C432-C439
• Main Authors: Li, Shixiong, Gerrard, Edward R., Jr, Balkovetz, Daniel F
• Format: Article
• Language: English
• Subjects: Animals; Butadienes - pharmacology; Cell Division - drug effects; Cell Division - physiology; Cell Line; Contact Inhibition - physiology; Cyclin D1 - metabolism; Dogs; Enzyme Inhibitors - pharmacology; Epithelial Cells - cytology; Epithelial Cells - enzymology; Flavonoids - pharmacology; Hepatocyte Growth Factor - pharmacology; Kidney - cytology; MAP Kinase Signaling System - drug effects; MAP Kinase Signaling System - physiology; Mitogen-Activated Protein Kinase 1 - metabolism; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases - metabolism; Nitriles - pharmacology; Phosphorylation; Trypsin
• ISSN: 0363-6143; 1522-1563
• DOI: 10.1152/ajpcell.00020.2004

Departments of 1 Medicine, 2 Cell Biology, and 3 Surgery, University of Alabama at Birmingham, and 4 Birmingham Veterans Affairs Medical Center, Birmingham, Alabama 35294 Submitted 14 January 2004 ; accepted in final form 2 April 2004 Increasing cell density arrests epithelial cell proliferation by a process termed contact inhibition. We investigated mechanisms of contact inhibition using a model of contact-inhibited epithelial cells. Hepatocyte growth factor (HGF) treatment of contact-inhibited Madin-Darby canine kidney (MDCK) cells stimulated cell proliferation and increased levels of phosphorylated ERK1/2 (phospho-ERK1/2) and cyclin D1. MEK inhibitors PD-98059 and U0126 inhibited these HGF-dependent changes, indicating the dependence on phosphorylation of ERK1/2 during HGF-induced loss of contact inhibition. In relation to contact-inhibited high-density cells, low-density MDCK cells proliferated and had higher levels of phospho-ERK1/2 and cyclin D1. PD-98059 and U0126 inhibited low-density MDCK cell proliferation. Trypsinization of high-density MDCK cells immediately increased phospho-ERK1/2 and was followed by a transient increase in cyclin D1 levels. Reformation of cell junctions after trypsinization led to decreases in phospho-ERK1/2 and cyclin D1 levels. High-density MDCK cells express low levels of both cyclin D1 and phospho-ERK1/2, and treatment of these cells with fresh medium containing HGF but not fresh medium alone for 6 h increased phospho-ERK1/2 and cyclin D1 levels compared with cells without medium change. These data provide evidence that HGF abrogates MDCK cell contact inhibition by increasing ERK1/2 phosphorylation and levels of cyclin D1. These results suggest that in MDCK cells, contact inhibition of cell proliferation in the presence of serum occurs by cell density-dependent regulation of ERK1/2 phosphorylation. cell density; cyclin D1; hepatocyte growth factor; cell cycle; extracellular signal-regulated kinases Address for reprint requests and other correspondence: D. F. Balkovetz, Dept. of Medicine, Univ. of Alabama at Birmingham, 1530 Third Ave. South, LHRB 642, Birmingham, AL 35294-0007 (E-mail: balkovet{at}uab.edu ).