Effect of EGF on [3H]-thymidine incorporation and cell cycle regulatory proteins in primary cultured chicken hepatocytes: Involvement of Ca2+/PKC and MAPKs.
The reported studies on the metabolism in chicken hepatocytes in comparison with those of mammals are quite different. Therefore, this study examined the effect of EGF on DNA synthesis along with its related signal cascades in primary cultured chicken hepatocytes. EGF stimulated DNA synthesis in a dose (> or =10 ng/ml)-dependent manner, which correlated with the increase in CDK-2 and CDK-4 expression. The EGF-induced increase in [3H]-thymidine incorporation was blocked by AG 1478 (an EGF receptor tyrosine kinase antagonist), genistein, and herbimycin A (tyrosine kinase inhibitors), suggesting a role in the activation and tyrosine phosphorylation of the EGF receptor. In addition, the EGF-induced stimulation of [3H]-thymidine incorporation was prevented by staurosporine, H-7, or bisindolylmaleimide I (protein kinase C inhibitors), suggesting a role of PKC. In addition, PD 98059 (a MEK inhibitor), SB 203580 (a p38 MAPK inhibitor), and SP 600125 (a JNK inhibitor) blocked the EGF-induced stimulation of [3H]-thymidine incorporation and CDK-2/4 expression. Indeed, EGF increased the translocation of PKC from the cytosol to the membrane fraction, and increased the activation of p44/42 MAPK, p38 MAPK, and JNK. Moreover, EGF increased the CDK-2, CDK-4, cyclin D1, and cyclin E expression levels but decreased the p21 and p27 expression levels. These EGF-induced increases were blocked by an EGF receptor antagonist, tyrosine kinase inhibitors, PKC inhibitors, and MAPKs inhibitors. In conclusion, EGF stimulates DNA synthesis of primary cultured chicken hepatocytes via Ca2+/PKC and the MAPKs signaling pathways.
Lee
Min Young
MY
Department of Veterinary Physiology, Biotherapy Human Resources Center, College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, Korea.
Lee
Sang Hun
SH
Kim
Yun Hee
YH
Heo
Jung Sun
JS
Park
Soo Hyun
SH
Lee
Jang Hern
JH
Han
Ho Jae
HJ
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002645:Chickens; D004261:DNA Replication / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013936:Thymidine / Q000378:metabolism*; D014316:Tritium
D018797:Cell Cycle Proteins; D014316:Tritium; D004815:Epidermal Growth Factor; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium; D013936:Thymidine