Thrombin-induced Ca2+ mobilization in human gingival fibroblasts is mediated by protease-activated receptor-1 (PAR-1).
By reverse transcription (RT)-PCR analyses, human gingival fibroblasts (HGFs) were demonstrated to express mRNAs for protease-activated receptor-1 (PAR-1) and PAR-3, although the expression of PAR-3 was much weaker than that of PAR-1. The mRNAs for PAR-2 and PAR-4 were not found by RT-PCR. Furthermore, PAR activation was studied by monitoring cytosolic Ca(2+) concentration ([Ca(2+)]i) in cultured HGFs loaded with fura-2. At concentrations > 0.1 nM, alpha-thrombin caused a transient increase in [Ca(2+)]i in a concentration-dependent manner, and the maximum response was obtained with 10 nM alpha-thrombin. After the [Ca(2+)]i response, the HGFs were completely desensitized to the second stimulation with alpha-thrombin. The PAR-1 agonist peptide SFLLRN produced approximately the same transient [Ca(2+)]i response as alpha-thrombin. After stimulation with SFLLRN, the HGFs did not respond to alpha-thrombin, indicating that treatment with SFLLRN results in complete desensitization to alpha-thrombin. The PAR-2 and PAR-4 agonist peptides had no effect on [Ca(2+)]i in HGFs. These results suggest that alpha-thrombin-induced Ca(2+) mobilization in HGFs is solely mediated by PAR-1.
Tanaka
Nobuhisa
N
Department of Orthodontics, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan.
Morita
Takao
T
Nezu
Akihiro
A
Tanimura
Akihiko
A
Mizoguchi
Itaru
I
Tojyo
Yosuke
Y
D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005881:Gingiva / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D010446:Peptide Fragments / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; D018179:Receptors, Thrombin / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013917:Thrombin / Q000494:pharmacology*
D010446:Peptide Fragments; D012333:RNA, Messenger; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; D018179:Receptors, Thrombin; C105295:protease-activated receptor 3; C082835:thrombin receptor peptide (42-47); D013917:Thrombin; C112908:protease-activated receptor 4; D002118:Calcium