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J Vet Sci 2018; 19(6): 735-743  https://doi.org/10.4142/jvs.2018.19.6.735
TGF-β1 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
Received: June 12, 2018; Revised: July 3, 2018; Accepted: July 13, 2018; Published online: November 30, 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 times in monolayer cultures. In three-dimensional cultures, chondrocyte-seeded gelatin scaffolds were cultured in chondrogenic media containing 10 ng/mL of TGF-β1. The amounts of HA in conditioned media and in scaffolds were determined by enzyme-linked immunosorbent assays. HAS2 mRNA expression was analyzed by semi-quantitative reverse transcription polymerase chain reaction. The uronic acid content and DNA content of the scaffolds were measured by using colorimetric and Hoechst 33258 assays, respectively. Cell proliferation was evaluated by using the alamarBlue assay. Scanning electron microscopy (SEM), histology, and immunohistochemistry were used for 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 results revealed the formation of a cartilaginous-like extracellular matrix in the TGF-β1-treated scaffolds. Together, our results suggest that TGF-β1 has a stimulatory effect on equine chondrocytes, enhancing HA synthesis and promoting cartilage matrix generation.
Keywords: chondrocytes, horses, hyaluronan, hyaluronan synthases, transforming growth factors


© 2018 The Korean Society of Veterinary Science.

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