| 武晓慧,马子怡,张钊华,张琳贻,党怡雄,廉 婷.双硫仑治疗小鼠肥胖的安全性和有效性评价[J].,2024,(2):219-227 |
| 双硫仑治疗小鼠肥胖的安全性和有效性评价 |
| Study on the Safety and Efficacy of Disulfiram in the Treatment of Obesity in Mice |
| 投稿时间：2023-08-15 修订日期：2023-09-09 |
| DOI：10.13241/j.cnki.pmb.2024.02.004 |
| 中文关键词: 双硫仑 肥胖 棕色脂肪细胞 肝细胞水肿 |
| 英文关键词: Disulfiram Obesity Brown adipocytes Hepatocellular edema |
| 基金项目:陕西省卫生健康科研基金项目（2022D018）；陕西省大学生创新创业训练计划项目（S202211840088）；西安医学院校级大学生创新创业训练计划项目（121522088）；陕西省自然科学基础研究计划—— 一般项目（面上项目）（2023-JC-YB-699）；西安医学院博士科研启动基金（2020DOC04）；陕西省科技厅-科技资源开放共享平台项目（2022PT-44） |
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| 中文摘要: |
| 摘要 目的：观察双硫仑治疗小鼠肥胖的安全性和有效性。方法：取6周龄C57BL/6J雄性小鼠10只，高脂饲料诱导肥胖后，随机分为双硫仑组（双硫仑玉米油溶液，300 mg/(kg?d）和对照组（等量玉米油），每组5只小鼠。每日灌胃给药1次，连续2周，期间仍给与高脂饲料。监测小鼠食物消耗量和体重。给药结束后取小鼠血清、附睾白色脂肪垫、肩胛间区棕色脂肪和肝脏。白色、棕色脂肪和肝脏进行HE染色，观察细胞形态。电镜下观察棕色脂肪细胞内的脂滴和线粒体。Realtime-qPCR法检测棕色脂肪组织中Ucp1、Fabp4、Prdml6和Cidea的mRNA相对表达量，Western blot法检测Ucp1的蛋白表达量。检测血清中转氨酶ALT和AST含量。取8周龄C57BL/6J雄性小鼠10只，随机分为双硫仑组（双硫仑300 mg/(kg?d）和对照组（等量玉米油），每日灌胃1次，连续2周。给药结束后进行棕色脂肪和肝脏HE染色并检测血清中ALT和AST含量。取8周龄C57BL/6J雄性小鼠10只，随机分为双硫仑组（双硫仑300 mg/(kg?d）和对照组（等量玉米油），每日灌胃1次，连续4周，进行肝脏HE染色并检测血清中ALT和AST含量。取孕13.5天的C57BL/6J胚胎小鼠，进行成纤维细胞原代培养，分为双硫仑组（双硫仑5 mg/L）和对照组（等量DMSO）并诱导分化为棕色脂肪细胞。分化8天后进行油红O染色，观察脂滴形成情况，检测Ucp1、Fabp4、Prdml6和Cidea的mRNA相对表达量和Ucp1的蛋白表达量。结果：肥胖小鼠给药过程中，双硫仑组和对照组的进食量及体重变化并无明显差别（P＞0.05）。给药结束后，两组白色脂肪细胞大小无明显差别。双硫仑组小鼠棕色脂肪细胞直径和细胞内脂滴明显增大（P＜0.05），脂滴数量、线粒体形态及数量无明显差别（P＞0.05）。双硫仑组小鼠棕色脂肪中Cidea和Prdm16的mRNA表达减少（P＜0.05）。正常体重小鼠双硫仑给药2周后棕色脂肪细胞脂滴也增大。细胞实验结果显示，双硫仑组脂滴形成明显减少，Ucp1、Cidea、Prdm16的mRNA表达明显减少（P<0.05）；Ucp1的蛋白表达明显减少（P<0.05）。肥胖与正常小鼠双硫仑给药2周后均出现明显的肝细胞水肿，血清中ALT和AST升高（P<0.05），正常小鼠给药4周后仍有明显肝细胞水肿，ALT和AST升高（P<0.05）。结论：短期使用双硫仑对饮食诱导的肥胖小鼠无明显减肥作用；双硫仑在体内、外均可抑制小鼠棕色脂肪细胞的分化。短期使用双硫仑可引起肝损害。双硫仑用于减肥治疗的安全性及有效性尚不够理想。 |
| 英文摘要: |
| ABSTRACT Objective: To observe the safety and efficacy of disulfiram in the treatment of obesity in mice. Methods: Ten 6-week-old C57BL/6J male mice were randomly divided into disulfiram group (disulfiram dissolved in corn oil, 300 mg/(kg?d) and control group (equivalent amount of corn oil), with 5 mice in each group. The mice were administered once a day by intragastric administration for 2 weeks, during which high-fat diet was still given. Food consumption and body weight of the mice were monitored during the disulfiram administration. Serum, white adipose pad of epididymis, brown adipose tissue in the scapular area, and liver were taken after administration. White and brown adipose tissue, and liver were stained with HE, and the cell morphology was observed. Lipid droplets and mitochondria in brown adipocytes were observed under electron microscopy. Realtime-qPCR was used to detect the relative mRNA expression levels of Ucp1, Fabp4, Prdml6, and Cidea in brown adipose tissue, while Western blot was used to detect the protein expression level of Ucp1. Levels of aminotransferase ALT and AST in serum was also detected. Ten C57BL/6J male mice aged 8 weeks were randomly divided into disulfiram group (disulfiram 300 mg/(kg?d) and control group (equivalent amount of corn oil). They were intragastrically administered once a day for two weeks. After administration, HE staining of the brown adipose tissue and liver were performed, and ALT and AST contents in serum were detected. Ten C57BL/6J male mice aged 8 weeks were randomly divided into disulfiram group (disulfiram 300 mg/(kg?d) and control group (equivalent amount of corn oil). They were intragastically administered once a day for four weeks, and HE staining of the liver was performed. Serum levels of ALT and AST were measured. C57BL/6J embryonic mice with a gestational age of 13.5 days were selected for primary fibroblast culture. They were divided into disulfiram group (5 mg/L disulfiram) and control group (equivalent amount of DMSO) and induced to differentiate into brown adipocytes. After 8 days of differentiation, oil red O staining was performed to observe the formation of lipid droplets, and the relative mRNA and protein expression levels of Ucp1, Fabp4, Prdml6, and Cidea were detected. Results: During the administration, there was no significant difference in food intake and weight changes between the disulfiram group and the control group (P>0.05). After administration, there was no significant difference in the size of white adipocytes between the two groups. The diameter of brown adipocytes and intracellular lipid droplets in the disulfiram group of mice were significantly increased (P<0.05), but there was no significant difference in the number of lipid droplets, mitochondrial morphology and mitochondrial number(P>0.05). The mRNA expression of Cidea and Prdm16 in brown adipose tissue of mice in the disulfiram group were decreased (P<0.05). For mice with normal weight, after 2 weeks of administration of disulfiram, the lipid droplets of brown adipocytes were also enlarged. The results of cell experiments showed that the formation of lipid droplets was significantly reduced in the disulfiram group, and the mRNA expression of Ucp1, Cidea, and Prdm16 was significantly reduced (P<0.05); The protein expression of Ucp1 was significantly reduced (P<0.05). Both obese and normal weight mice showed significant liver cell edema after 2 weeks of administration of disulfiram, with elevated levels of ALT and AST in serum (P<0.05). Normal weight mice also showed significant liver cell edema after 4 weeks of disulfiram administration, with elevated levels of ALT and AST (P<0.05). Conclusion: Short term use of disulfiram has no significant effect on weight loss of diet-induced obese mice; disulfiram can inhibit the differentiation of mice's brown adipocytes both in vivo and in vitro. Short-term use of disulfiram can cause liver damage. The safety and effectiveness of disulfiram in weight loss treatment are not ideal enough. |
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