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| 亚慢性镉暴露对小鼠骨结构及m6A水平的影响 |
| Effects of subchronic cadmium exposure on bone structure and m6A levels in mice |
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| DOI：10.3969/j.issn.1006-7108.2025.06.006 |
| 中文关键词: 骨质疏松症 镉 N6-甲基腺苷修饰 甲基转移酶3 |
| 英文关键词:osteoporosis cadmium N6-methyladenosine modification METTL3 |
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| 中文摘要: |
| 目的 通过构建亚慢性镉暴露致小鼠骨质疏松模型，探讨N6-甲基腺嘌呤（N6-methyladenosine，m6A）在镉致骨质疏松过程中的变化及其机制。方法 40只雌性C57BL/6j小鼠随机分为4组：对照组、低剂量[5 mg/(kg·bw)]、中剂量[10 mg/(kg·bw)]和高剂量[20 mg/(kg·bw)]氯化镉暴露组，每组10只，连续灌胃90 d染毒，使用苏木精-伊红（HE）染色检测小鼠股骨骨质情况。采用斑点印迹法(Dot blot)检测股骨m6A RNA的甲基化水平。采用实时荧光定量PCR法（qPCR）和蛋白免疫印迹法 (Western blot) 分别检测各组小鼠股骨中成骨标志基因碱性磷酸酶 (ALP)、Runt相关转录因子2 (RUNX2) 、Osterix蛋白(SP7) 和甲基转移酶3 ( METTL3)表达量。 结果 HE染色结果显示，与对照组相比，镉处理组小鼠骨小梁数量减少，结构紊乱，骨髓腔扩大，骨髓脂肪细胞浸润，表现出明显的骨质疏松症病理变化。Dot blot结果显示，与对照组相比，在镉处理的小鼠股骨m6A的水平随着镉浓度的增加呈现剂量依赖性下降（P<0.05）。qPCR和Western blot结果显示，小鼠股骨METTL3的表达量随着镉浓度的增加呈现剂量依赖性下降（P<0.05），同时成骨标志基因 ALP、RUNX2、SP7的mRNA和蛋白表达水平也随着镉浓度的增加呈现剂量依赖性下降（P<0.05）。 结论 镉暴露可诱发小鼠骨质疏松的发生，其机制可能与抑制METTL3介导的m6A甲基化修饰，进而抑制成骨分化相关。 |
| 英文摘要: |
| Objective To investigate the changes of N6-methyladenosine (m6A) and its mechanism in cadmium-induced osteoporosis by constructing a subchronic cadmium exposure-induced osteoporosis model in mice. Methods Forty female C57BL/6j mice were randomly divided into four groups: control group, low-dose (5 mg/kg·bw), medium-dose (10 mg/kg·bw) and high-dose (20 mg/kg·bw) CdCl2-exposed group, which were poisoned by gavage for 90 days, and the femur bone quality of the mice was detected by hematoxylin-eosin (HE) staining. The methylation level of m6A RNA in the femur was detected by dot blot. The expression of alkaline phosphatase (ALP), Runt-associated transcription factor 2 (RUNX2), Osterix protein (SP7) and METTL3 in the femur of each group was detected by real-time fluorescence quantitative PCR (qPCR) and protein immunoblotting (Western blot), respectively. Results HE staining results showed that compared with the control group, the cadmium-treated mice showed a decrease in the number of bone trabeculae, structural disorganization, enlargement of the bone marrow cavity, and infiltration of bone marrow adipocytes, which demonstrated obvious pathological changes of osteoporosis. Dot blot results showed that compared with the control group, the level of femur m6A in cadmium-treated mice showed a dose-dependent decrease with increasing cadmium concentration (P<0.05). The results of qPCR and Western blot showed that the expression of METTL3 in the femur of mice decreased dose-dependently with the increase of cadmium concentration (P<0.05), and the mRNA and protein expression levels of the osteogenic marker genes, ALP, RUNX2, and SP7, also decreased dose-dependently with the increase of cadmium concentration (P<0.05). Conclusion Cadmium exposure induces the development of osteoporosis in mice by a mechanism that may be related to the inhibition of METTL3-mediated modification of m6A methylation and consequently osteogenic differentiation. |
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