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Transforming growth factor beta 1 upregulates the expression of hyaluronan synthase 2 and hyaluronan synthesis in culture models of equine articular chondrocytes
Siriwan Ongchai1, Oraphan Somnoo1, Patiwat Kongdang1, Siriporn Peansukmanee2, Siriwan Tangyuenyong2,*
1Thailand Excellence Center for Tissue Engineering and Stem Cells, Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand
2Equine Clinic, Department of Companion Animal and Wildlife Clinic, Faculty of Veterinary Medicine, Chiang Mai University, Chiang Mai 50100, Thailand
Correspondence to: Tel: +66-5394-8015; Fax: +66-5394-8065; E-mail: makhaboocha@gmail.com, siriwan.tangy@cmu.ac.th
Received: June 12, 2018; Revised: July 3, 2018; Accepted: July 13, 2018; Published online: July 20, 2018.
Abstract
We investigated the effect of transforming growth factor beta 1 (TGF-β1) on equine hyaluronan synthase 2 (HAS2) gene expression and hyaluronan (HA) synthesis in culture models of articular chondrocytes. Equine chondrocytes were treated with TGF-β1 at different concentrations and time points in monolayer cultures. In three-dimensional cultures, chondrocyte-seeded gelatin scaffolds were cultured in chondrogenic media containing 10 ng/mL of TGF-β1. The amount of HA in conditioned media and in scaffolds was determined using ELISA. HAS2 mRNA expression was analyzed by semi-quantitative RT-PCR. The uronic acid content and DNA content of the scaffolds were measured by colorimetric and Hoechst 33258 assays, respectively. Cell proliferation was evaluated using the AlamarBlue assay. Scanning electron microscopy (SEM), histology, and immunohistochemistry provided microscopic analysis of the samples. The upregulation of HAS2 mRNA levels by TGF-β1 stimulation was dose and time dependent. TGF-β1 was shown to enhance HA and uronic acid content in the scaffolds. Cell proliferation and DNA content were significantly lower in TGF-β1 treatments. SEM and histological findings showed formation of a cartilaginous-like extracellular matrix in the TGF-β1-treated scaffolds. Together, our results suggest that TGF-β1 has a stimulatory effect in equine chondrocytes, enhancing HA synthesis and promoting cartilage matrix generation.
Keywords: chondrocyte; horse; hyaluronan; hyaluronan synthase; transforming growth factor


© 2018 The Korean Society of Veterinary Science.