薯蓣皂苷元通过调控miR-148b-3p/DNMT1/FOXO1轴改善糖尿病肾病中的脂质代谢

Diosgenin Improves Lipid Metabolism in Diabetic Nephropathy via Regulation of miR-148b-3p/DNMT1/FOXO1 Axis.

作者信息

Luo Min, Hu Zongren, Yang Jichang, Yang Jinhan, Sheng Wen, Lin Chengxiong, Li Dian, He Qinghu

机构信息

Hunan Engineering Research Center of Internet-Chinese and Western Medicine Collaboration-Health Service, Hunan University of Medicine, Huaihua, China.

Department of Traditional Chinese Medicine, The First Affiliated Hospital of Hunan University of Medicine, Huaihua, China.

出版信息

Nephron. 2025;149(4):226-239. doi: 10.1159/000541690. Epub 2024 Nov 27.

DOI:10.1159/000541690

PMID:39602888

Abstract

BACKGROUND

The progression of diabetic nephropathy (DN) is closely associated with lipid accumulation. Diosgenin (Dio) plays a beneficial role in the lipid metabolism associated with multiple diseases. Thus, the mechanism underlying Dio's function in DN associated with aberrant lipid accumulation warrants further investigation.

METHODS

To model DN in vitro, HK-2 cells were treated with high glucose (HG) and palmitic acid. Cell viability was evaluated using MTT assay. The triglyceride (TG) content in HK-2 cells was measured using a commercial assay kit. The formation of lipid droplets in HK-2 cells was observed using Oil Red O staining. The expression levels of mRNA and protein were detected using RT-qPCR and Western blot, respectively. The DNA methylation of FOXO1 was assessed using MSP. The interaction between DNMT1 and the FOXO1 promoter was confirmed by ChIP assay.

RESULTS

Dio treatment reduced TG levels and lipid droplet formation in HK-2 cells co-treated with HG and palmitic acid. Simultaneously, the levels of miR-148b-3p and FOXO1 were increased by Dio, while Dio decreased the expression levels of DNMT1 and SREBP-2. Meanwhile, miR-148b-3p can bind to DNMT1, which in turn inhibits the expression of FOXO1 by mediating the DNA methylation of FOXO1. In addition, FOXO1 negatively regulates the expression of SREBP-2 by interacting with the SREBP-2 promoter. MiR-148b-3p inhibition or silencing of FOXO1 abolished the inhibitory effect of Dio on TG production and lipid droplet formation. This effect was further exacerbated by the downregulation of DNMT1. FOXO1 overexpression may counteract the promotive effects of miR-148b-3p inhibitor on lipid accumulation.

CONCLUSIONS

Dio treatment reduced TG production and lipid droplet formation in HK-2 cells during the progression of DN by modulating the miR-148b-3p/DNMT1/FOXO1/SREBP-2 axis. This finding provides new evidence supporting the therapeutic potential of Dio for DN.

摘要

背景

糖尿病肾病（DN）的进展与脂质蓄积密切相关。薯蓣皂苷元（Dio）在与多种疾病相关的脂质代谢中发挥有益作用。因此，Dio在与异常脂质蓄积相关的DN中发挥作用的机制值得进一步研究。

方法

为在体外构建DN模型，用高糖（HG）和棕榈酸处理HK-2细胞。使用MTT法评估细胞活力。使用商业检测试剂盒测量HK-2细胞中的甘油三酯（TG）含量。使用油红O染色观察HK-2细胞中脂滴的形成。分别使用RT-qPCR和蛋白质印迹法检测mRNA和蛋白质的表达水平。使用甲基化特异性PCR（MSP）评估FOXO1的DNA甲基化。通过染色质免疫沉淀（ChIP）试验确认DNA甲基转移酶1（DNMT1）与FOXO1启动子之间的相互作用。

结果

Dio处理降低了与HG和棕榈酸共同处理的HK-2细胞中的TG水平和脂滴形成。同时，Dio增加了miR-148b-3p和FOXO1的水平，而Dio降低了DNMT1和固醇调节元件结合蛋白2（SREBP-2）的表达水平。同时，miR-148b-3p可以与DNMT1结合，进而通过介导FOXO1的DNA甲基化来抑制FOXO1的表达。此外，FOXO1通过与SREBP-2启动子相互作用负向调节SREBP-2的表达。抑制miR-148b-3p或沉默FOXO1消除了Dio对TG产生和脂滴形成的抑制作用。DNMT1的下调进一步加剧了这种作用。FOXO1过表达可能抵消miR-148b-3p抑制剂对脂质蓄积的促进作用。