Effects of T-2 toxin on inflammatory cytokines, CD44 and integrin in chondrocytes cultured in vitro / 中华地方病学杂志

ABSTRACT

Objective:To investigate the effects of T-2 toxin on expression of inflammatory cytokines, human leukocyte differentiation antigen (CD) 44 and integrin in chondrocytes cultured in vitro. Methods:Primary chondrocytes from SPF Wistar rats aged 1 to 2 days were isolated and cultured in vitro, and the chondrocytes were identified by toluidine blue staining. The effects of different dose of T-2 toxin (0, 2, 4, 6, 10, 12, 20 ng/ml) on proliferation of chondrocytes for 24, 48 and 72 h were detected via the cell counting kit-8 (CCK-8) method. According to the cell survival rate, T-2 toxin concentrations of 0 (control), 2, 4, 6 and 8 ng/ml were selected for subsequent experiments, and the exposure time was 48 h. The contents of interleukin (IL)-6, IL-1β, tumor necrosis factors (TNF)-α and CD44 in cell supernatant were detected via the enzyme linked immunosorbent assay (ELISA). The protein expression levels of integrin α5 and integrin β1 in chondrocytes were detected by Western blotting. Results:At the same exposure time, there were significant differences of the survival rate of chondrocytes between different dose groups ( F = 130.759, 258.250, 123.337, P < 0.01). At 48 h after exposure, there were statistically significant differences in IL-6, IL-1β, TNF-α and CD44 contents in the culture supernatant of chondrocytes between different dose of T-2 toxin groups ( F = 10.613, 4.805, 2.943, 12.395, P < 0.01 or < 0.05); among them, the IL-6 levels of 2, 4, 6 and 8 ng/ml groups were higher than that of control group ( P < 0.05); the IL-1β levels of 6, 8 ng/ml groups were higher than that of control and 2 ng/ml groups, and the 6 ng/ml group was higher than that of 4 ng/ml group ( P < 0.05); the TNF-α levels of 6, 8 ng/ml groups were lower than that of control group ( P < 0.05); the CD44 levels of 2, 4, 6 and 8 ng/ml groups were lower than that of control group ( P < 0.05). At 48 h after exposure, there were statistically significant differences in integrin α5 and integrin β1 protein expression levels between different dose of T-2 toxin groups ( F = 4.635, 4.376, P < 0.05). Among them, the protein expression levels of integrin α5 in 6, 8 ng/ml groups were significantly lower than that of control group ( P < 0.05); the protein expression levels of integrin β1 in 6, 8 ng/ml groups were significantly higher than that of control group ( P < 0.05). Conclusion:T-2 toxin may up-regulate the expressions of IL-6, IL-1β and integrin β1, while down-regulate the expressions of TNF-α, CD44 and integrin α5, then disrupt the balance between chondrocytes and extracellular matrix, and cause chondrocytes damage.