ABSTRACT
Objective:To explore the effects of HT-2 toxin on expressions of silent information regulator of transcription 1 (SIRT1) and autophagy and apoptosis pathway related proteins in cultured chondrocytes in vitro. Methods:The third-generation chondrocytes of SD neonatal rats aged 1 to 2 days were cultured in vitro and identified by toluidine blue staining and type Ⅱ collagen immunofluorescence staining. CCK-8 method was used to detect the proliferation of chondrocytes. According to the cell survival rate, 2, 4 and 8 ng/ml HT-2 toxin were selected for subsequent experiments, and the exposure time was 48 h. At the same time, a negative control group and a solvent (absolute ethanol) control group were set up. Western blotting was used to detect the expressions of SIRT1 and autophagy and apoptosis pathway related proteins [microtubule-associated protein 1 light chain 3 (LC3)-Ⅱ, LC3-Ⅰ, p62, Beclin1, Caspase-3, B-cell lymphoma-2 (Bcl-2) and Bcl-2-Associated X protein (Bax)] in each group. Results:After staining, the cells were identified as chondrocytes with high purity. The expression levels of SIRT1 protein in 2, 4, 8 ng/ml HT-2 toxin groups (0.69 ± 0.18, 0.46 ± 0.13, 0.35 ± 0.19) were significantly lower than that in negative control group (1.00 ± 0.39, P < 0.05). In 2, 4 and 8 ng/ml HT-2 toxin groups, the ratios of autophagy pathway related proteins LC3-Ⅱ and LC3-Ⅰ expressions (LC3-Ⅱ/LC3-Ⅰ, 1.47 ± 0.15, 1.37 ± 0.13, 1.81 ± 0.34) were higher than that in negative control group (1.00 ± 0.21, P < 0.05), and the expression levels of p62 protein in 4, 8 ng/ml HT-2 toxin groups (0.70 ± 0.04, 0.57 ± 0.01) were lower than that in negative control group (1.00 ± 0.15, P < 0.05). In 2, 4, 8 ng/ml HT-2 toxin groups, the expression levels of apoptosis pathway related protein Bcl-2 (0.61 ± 0.06, 0.54 ± 0.16, 0.47 ± 0.06) were significantly lower than that in negative control group (1.00 ± 0.14, P < 0.05), and the ratio of Bax to Bcl-2 protein expressions in 8 ng/ml HT-2 toxin group (Bax/Bcl-2, 3.27 ± 0.18) was higher than that in negative control group (1.00 ± 0.27, P < 0.05). The expression level of SIRT1 protein was significantly negatively correlated with the expression level of autophagy pathway related protein LC3-Ⅱ ( r = - 0.819, P < 0.01), and was significantly positively correlated with the expression level of p62 protein( r = 0.772, P < 0.01), but not with the expression level of Beclin1 protein ( r = 0.399 , P > 0.05); there was no correlation between SIRT1 protein expression and apoptosis pathway related protein Caspase-3 and Bax expressions ( r = - 0.297、- 0.284, P > 0.05), but there was a significant positive correlation with Bcl-2 protein expression ( r = 0.755, P < 0.01). Conclusion:HT-2 toxin may increase the expression of autophagy pathway related protein LC3-Ⅱ/LC3-Ⅰ, decrease the expression of p62 protein, and increase the apoptosis pathway related protein Bax/Bcl-2 by inhibiting the expression of SIRT1 protein in chondrocytes, resulting in abnormal autophagy and apoptosis, and finally leads to the injury of chondrocytes.
ABSTRACT
As the main pollutant of food crops, T-2 toxin has toxic effects on human and animal digestive system, nervous system, reproductive development and so on. In Kashin-Beck disease related areas in China, the etiological substance of Kashin-Beck disease is the abnormal accumulation of T-2 toxin in the grain produced in the endemic area. The prevention and treatment of Kashin-Beck disease has achieved remarkable results through the comprehensive prevention and control measures, such as changing the production and life style, grain exchange, etc., but there are still pathogenic factors in the external environment of the disease area. In this paper, the metabolic kinetics of T-2 toxin is reviewed, and the physicochemical properties, distribution in the body, metabolic kinetics, biotransformation and damage of T-2 toxin and its metabolites to various organs are described, so as to provide new thinking for the study on the effects of T-2 toxin on various organs of the body.