TGF-beta 1 upregulates the expression of hyaluronan synthase 2 and hyaluronan synthesis in culture models of equine articular chondrocytes
2018
Siriwan Ongchai, Chiang Mai University, Chiang Mai, Thailand | Oraphan Somnoo, Chiang Mai University, Chiang Mai, Thailand | Patiwat Kongdang, Chiang Mai University, Chiang Mai, Thailand | Siriporn Peansukmanee, Chiang Mai University, Chiang Mai, Thailand | Siriwan Tangyuenyong, Chiang Mai University, Chiang Mai, Thailand
We investigated the effect of transforming growth factor beta 1 (TGF-beta1) on equine hyaluronan synthase 2 (HAS2) gene expression and hyaluronan (HA) synthesis in culture models of articular chondrocytes. Equine chondrocytes were treated with TGF-꺬1 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-beta1. 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-꺬1 stimulation was dose and time dependent. TGF-beta1 was shown to enhance HA and uronic acid content in the scaffolds. Cell proliferation and DNA content were significantly lower in TGF-beta1 treatments. SEM and histological results revealed the formation of a cartilaginous-like extracellular matrix in the TGF-beta1-treated scaffolds. Together, our results suggest that TGF-beta1 has a stimulatory effect on equine chondrocytes, enhancing HA synthesis and promoting cartilage matrix generation.
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