Mechanism of DNA methylation-mediated downregulation of O6-methylguanine-DNA methyltransferase in cartilage injury of Kashin–Beck disease
| Autor: | Dandan Zhang, Di Zhang, Chen Wang, Rongqiang Zhang, Qiang Li, Yongmin Xiong |
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| Rok vydání: | 2021 |
| Předmět: | |
| Zdroj: | Rheumatology. 61:3471-3480 |
| ISSN: | 1462-0332 1462-0324 |
| DOI: | 10.1093/rheumatology/keab913 |
| Popis: | Objective Kashin–Beck disease (KBD) is an endemic osteoarthropathy, in which excessive apoptosis of chondrocytes occurs. O6-methylguanine-DNA methyltransferase (MGMT), a DNA damage repair gene, plays an important role in apoptosis, but the mechanism is unclear in KBD cartilage injury. This study was to investigate the expression and promoter methylation of MGMT in KBD patients and its role in DNA damage and apoptosis of chondrocytes. Methods MGMT mRNA and protein level were detected by quantitative real-time PCR and immunohistochemistry. Demethylation of MGMT was carried out using 5-Aza-2′-deoxycytidine, and the methylation level of MGMT promoter was measured by quantitative methylation specific PCR. Next, small hairpin RNA was used to knockdown the expression of MGMT. Cell viability, apoptosis and DNA damage were determined by MTT assay, flow cytometry, Hoechst 33342 staining and alkaline comet assay following T-2 toxin and selenium treatment. Results MGMT protein expression and mRNA levels were decreased (P = 0.02, P = 0.007) and promoter methylation was increased (P = 0.008) in KBD patients. Meanwhile, MGMT level was upregulated by 5-Aza-2′-deoxycytidine in chondrocytes (P = 0.0002). DNA damage and apoptosis rates were increased in MGMT-silenced chondrocytes (all P Conclusions MGMT might play a crucial part in the pathogenesis of KBD cartilage injury, which could provide a therapeutic target for KBD. |
| Databáze: | OpenAIRE |
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