Thrombin induced connective tissue growth factor expression in rat vascular smooth muscle cells via the PAR-1/JNK/AP-1 pathway

Abstract

Aim: To investigate the signaling pathways involved in thrombin-induced connective tissue growth factor (CTGF) expression in rat vascular smooth muscle cells (VSMCs). Methods: Experiments were preformed on primary rat aortic smooth muscle cells (RASMCs) and a rat VSMC line (A10). CTGF protein levels were measured using Western blotting. Luciferase reporter genes and dominant negative mutants (DNs) were used to investigate the signaling pathways mediating the induction of CTGF expression by thrombin. Results: Thrombin (0.3-3.0 U/mL) caused a concentration-and time-dependent increase in CTGF expression in both RASMCs and A10 cells. Pretreating A10 cells with the protease-activated receptor 1 (PAR-1) antagonist SCH79797 (0.1 µmol/L) significantly blocked thrombin-induced CTGF expression, while the PAR-4 antagonist tcY-NH 2 (30 µmol/L) had no effect. The PAR-1 agonist SFLLRN-NH 2 (300 µmol/L) induced CTGF expression, while the PAR-4 agonist GYPGQV-NH 2 (300 µmol/L) had no effect. Thrombin (1 U/mL) caused time-dependent phosphorylation of c-Jun N-terminal kinase (JNK). Pretreating with the JNK inhibitor SP600125 (3-30 µmol/L) or transfection with DNs of JNK1/2 significantly attenuated thrombin-induced CTGF expression. Thrombin (0.3-3.0 U/mL) increased activator protein-1 (AP-1)-luciferase activity, which was inhibited by the JNK inhibitor SP600125. The AP-1 inhibitor curcumin (1-10 µmol/L) concentration-dependently attenuated thrombin-induced CTGF expression. Conclusion: Thrombin acts on PAR-1 to activate the JNK signaling pathway, which in turn initiates AP-1 activation and ultimately induces CTGF expression in VSMCs.

Galectin-3 Interacts with Vascular Cell Adhesion Molecule-1 to Increase Cardiovascular Mortality in Hemodialysis Patients

“…Mechanistically, myocardial stretch induced upregulation of galectin-3 and downstream connective tissue growth factor [ 10 ], which acted as a profibrotic and proinflammatory factor in our previous studies [ 11 , 12 ]. Furthermore, galectin-3 regulates many biological functions and signaling pathways with regard to cell proliferation, migration, adhesion, and cell cell interactions through binding with galactose-containing glycoproteins on the cell surface [ 13 ].…”

Inhibition of Mitochondria- and Endoplasmic Reticulum Stress-Mediated Autophagy Augments Temozolomide-Induced Apoptosis in Glioma Cells

“…Minimum essential medium (MEM) and supplements were purchased from Hyclone (Logan, UT). The JNK1 dominant-negative mutant (DN) and JNK2 DN were provided by kindly Dr. CH Lin (Taipei Medical University, Taipei, Taiwan) [43] . CsA, SP600125, 4-PBA, and 3-(4,5-dimethyl-2-thiazolyl)-2,5-dimethyl-2H-tetrazolium bromide (MTT) were purchased from Merck (Darmstadt, Germany).…”

The Effects of NF-κB and c-Jun/AP-1 on the Expression of Prothrombotic and Proinflammatory Molecules Induced by Anti-β2GPI in Mouse

“…It is widely accepted that anti-β 2 GPI antibodies play a pathogenic role in APS and have been shown to induce activation of ECs, in vitro and in vivo , which may contribute to hypercoagulability in APS patients [ 12 , 25 – 26 ]. Thrombin is correlated with the overall functional coagulation status of plasma in APS patients, and induced expression of connective tissue growth factors in rat vascular smooth muscle cells via the AP-1 pathway [ 27 – 28 ]. NF-κB may be involved in aPL/anti-β 2 GPI-induced expression of TF and adhesion molecules in ECs and monocytes in vitro [ 16 , 29 – 30 ].…”