Effects of TGF‐β1 on plasminogen activation in human dental pulp cells: Role of ALK5/Smad2, TAK1 and MEK/ERK signalling

Abstract

Transforming growth factor-β1 (TGF-β1) plays an important role in the pulpal repair and dentinogenesis. Plasminogen activation (PA) system regulates extracellular matrix turnover. In this study, we investigated the effects of TGF-β1 on PA system of dental pulp cells and its signalling pathways. Dental pulp cells were treated with different concentrations of TGF-β1. MTT assay, reverse transcription-polymerase chain reaction, Western blotting and enzyme-linked immunosorbant assay (ELISA) were used to detect the effect of TGF-β1 on cell viability, mRNA and protein expression of urokinase-type plasminogen activator (uPA), uPA receptor (uPAR), plasminogen activator inhibitor-1 (PAI-1) as well as their secretion. The phosphorylation of Smad2 and TAK1 was analysed by Pathscan ELISA or Western blotting. Cells were pretreated with SB431542 (ALK5/Smad2/3 inhibitor), 5z-7-oxozeaenol (TAK1 inhibitor) and U0126 (MEK/ERK inhibitor) for examining the related signalling. TGF-β1 slightly inhibited cell growth that was reversed by SB431542. TGF-β1 upregulated both RNA and protein expression of PAI-1 and uPAR, whereas it downregulated uPA expression. Accordingly, TGF-β1 stimulated PAI-1 and soluble uPAR (suPAR) secretion of pulp cells, whereas uPA secretion was inhibited. TGF-β1 induced the phosphorylation of Smad2 and TAK1. In addition, SB431542, 5z-7-oxozeaenol and U0126 attenuated the TGF-β1-induced secretion of PAI-1 and suPAR. These results indicate that TGF-β1 is possibly involved in the repair/regeneration and inflammatory processes of dental pulp via regulation of PAI-1, uPA and uPAR. These effects of TGF-β1 are related to activation of ALK5/Smad2, TAK1 and MEK/ERK signalling pathways. Clarifying the signal transduction for the effects of TGF-β1 is helpful for pulpo-dentin regeneration and tissue engineering. Copyright © 2016 John Wiley & Sons, Ltd.

Effects of transforming growth factor‐β1 on plasminogen activation in stem cells from the apical papilla: role of activating receptor‐like kinase 5/Smad2 and mitogen‐activated protein kinase kinase (MEK)/extracellular signal‐regulated kinase (ERK) signalling

“…Morphological changes of SCAP were examined and photographed by a phase‐contrast microscope. Culture medium was collected for measurement of PAI‐1, uPA and suPAR concentrations by ELISA kits (R & D Systems, Minneapolis, MN, USA) according to the manufacture’s instruction (Chang et al ). Cell number was estimated by 3‐(4,5‐dimethyldiazol‐2‐yl)‐2,5 diphenyl tetrazolium bromide (MTT, Sigma‐Aldrich) assay as described before (Chang et al , , ).…”

Bone Metastasis Phenotype and Growth Undergo Regulation by Micro-Environment Stimuli: Efficacy of Early Therapy with HGF or TGFβ1-Type I Receptor Blockade

“…Cultured cells under starvation were treated with HGF (100 ng/mL) or with TGFβ1 (5 ng/mL), which was used alone or in combination with 5 µM SB431542. The addition of SB431542 to the cultured cells was performed 1 h before TGFβ1 [5,52,53]. We authenticated 1833 cells with the method of short-tandem repeat profiling (STR) of nine highly polymorphic STR loci plus amelogenin on September 2014 (Cell Service from IRCCS-Azienda Ospedaliera Universitaria San Martino-IST-Istituto Nazionale per la Ricerca sul Cancro, Genova, Italy).…”

Lysophosphatidylcholine induces cytotoxicity/apoptosis and IL-8 production of human endothelial cells: Related mechanisms

“…After 24 h, culture medium was changed with fresh medium containing various amounts of LPC (final concentrations of 10, 20, 30, 40, 50 mg/ml) for 3 days. Culture medium was collected for analysis of IL-8 by ELISA following the instruction of assay kits [ 35 , 37 , 39 , 40 ].…”