Thrombin regulates interleukin-6 synthesis through phosphatidylcholine hydrolysis by phospholipase D in osteoblasts.
We previously reported that thrombin stimulates Ca2+ influx and activates phosphatidylcholine-hydrolyzing phospholipase D in osteoblast-like MC3T3-E1 cells. In this study, we investigated the effect of thrombin on interleukin-6 (IL-6) synthesis in these cells. Thrombin stimulated IL-6 synthesis dose-dependently in the range between 0.01 and 1 U/ml. The depletion of extracellular Ca2+ by EGTA suppressed the thrombin-induced IL-6 synthesis. TMB-8, an inhibitor of intracellular Ca2+ mobilization, also inhibited the IL-6 synthesis by thrombin. Propranolol, a phosphatidic acid phosphohydrolase inhibitor, enhanced the IL-6 synthesis by thrombin. Calphostin C, a highly potent and specific inhibitor for protein kinase C, significantly amplified the IL-6 synthesis by thrombin. The thrombin-induced IL-6 synthesis was enhanced in PKC down-regulated MC3T3-E1 cells. These results strongly suggest that thrombin stimulates IL-6 synthesis, which depends on intracellular Ca2+ mobilization mainly from extracellular space in osteoblasts, and that the IL-6 synthesis by thrombin is regulated due to thrombin-activated protein kinase C through phosphatidylcholine-hydrolyzing phospholipase D.
Kozawa
O
O
Department of Pharmacology, Gifu University School of Medicine, Gifu, 500, Japan.
Tokuda
H
H
Kaida
T
T
Matsuno
H
H
Uematsu
T
T
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002460:Cell Line; D004533:Egtazic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology; D005707:Gallic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D015850:Interleukin-6 / Q000096:biosynthesis*; D051379:Mice; D009281:Naphthalenes / Q000494:pharmacology; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D010711:Phosphatidate Phosphatase / Q000037:antagonists & inhibitors; D010713:Phosphatidylcholines / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D011433:Propranolol / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D013917:Thrombin / Q000008:administration & dosage / Q000494:pharmacology*
D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D015850:Interleukin-6; D009281:Naphthalenes; D010713:Phosphatidylcholines; D000431:Ethanol; D004533:Egtazic Acid; C006014:8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate; D005707:Gallic Acid; D011433:Propranolol; D011493:Protein Kinase C; D010711:Phosphatidate Phosphatase; D010739:Phospholipase D; D013917:Thrombin; C058819:calphostin C; D002118:Calcium