Effects of HT-2 toxin on expressions of SIRT1, autophagy and apoptosis pathway related proteins in chondrocytes
10.3760/cma.j.cn231583-20201221-00338
- VernacularTitle:HT-2毒素对软骨细胞SIRT1及自噬与凋亡通路相关蛋白表达的影响
- Author:
Chunhui LI
1
;
Yue WANG
;
Yujiao LIU
;
Jiaxin LI
;
Xin ZHANG
;
Jun YU
Author Information
1. 哈尔滨医科大学中国疾病预防控制中心地方病控制中心大骨节病防治研究所 150081
- Keywords:
HT-2 toxin;
Silent information regulator of transcription 1;
Autophagy;
Apoptosis;
Chondrocytes
- From:
Chinese Journal of Endemiology
2021;40(9):705-711
- CountryChina
- Language:Chinese
-
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.
