葛根素通过靶向 miR-342-3p/TGF-β/SMAD 轴缓解慢性肾衰竭诱导的肾小管上皮细胞细胞焦亡。
Puerarin alleviates chronic renal failure-induced pyroptosis in renal tubular epithelial cells by targeting miR-342-3p/TGF-β/SMAD axis.
机构信息
Department of Nephrology, The First Affiliated Hospital of Kunming Medical University, 295 Xichang Road, Wuhua District, Kunming, 650032, China.
出版信息
Genes Genomics. 2023 Dec;45(12):1563-1573. doi: 10.1007/s13258-023-01448-9. Epub 2023 Sep 25.
BACKGROUND
Chronic renal failure (CRF) is the result of kidney damage. Puerarin is a flavonoid with specific nephroprotective effect, but its effect on CRF needs further research. This study explored the effect of puerarin on CRF and the potential molecular mechanism.
METHODS
Adenine was used to establish an in vivo CRF model in rats, and rats were intragastrically administered with puerarin at a dose of 400 mg/kg body weight once a day from day 1 to day 28. Hematoxylin and eosin (HE) and Masson staining were used to observe the morphology and fibrosis of kidney tissue. Lipopolysaccharide (LPS) (400 ng/mL)/HO (200 µM) was applied to human kidney 2 (HK-2) cells to construct an in vitro CRF model. Enzyme-linked immunosorbent assay (ELISA) was performed to validate interleukin (IL)-1β and IL-18 levels. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) was performed to detect microRNA (miR)-342-3p levels. Transforming growth factor beta (TGF-β)1, SMAD2, SMAD3, and pyroptosis marker proteins were detected by Western blot. The interaction between miR-342-3p and TGF-β/SMAD was determined by a dual-luciferase reporter gene assay. Cell Counting Kit-8 (CCK-8) assay was utilized to determine cell viability.
RESULTS
In the CRF model, puerarin alleviated renal injury and fibrosis and reduced creatinine (Cr) and blood urea nitrogen (BUN) levels. At the same time, miR-342-3p was downregulated, while the TGF-β/SMAD axis was activated and levels of IL-1β and IL-18 were increased. After treatment of CRF rats with puerarin, the expression level of miR-342-3p was increased, the TGF-β/SMAD axis was inhibited, and the secretion of IL-1β and IL-18 was decreased. MiR-342-3p directly bound to and negatively regulated the expression of TGF-β1, SMAD2, and SMAD3. In the in vitro CRF model, miR-342-3p inhibited HK-2 cell pyroptosis by inhibiting the TGF-β/SMAD axis.
CONCLUSION
Puerarin reduced renal injury and pyroptosis in CRF rats by targeting the miR-342-3p/TGF-β/SMAD axis.
背景
慢性肾衰竭（CRF）是肾脏损伤的结果。葛根素是一种具有特定肾保护作用的黄酮类化合物，但它对 CRF 的作用仍需要进一步研究。本研究旨在探讨葛根素对 CRF 的作用及其潜在的分子机制。
方法
采用腺嘌呤建立大鼠体内 CRF 模型，从第 1 天到第 28 天每天给予大鼠 400mg/kg 体重的葛根素灌胃。苏木精-伊红（HE）和 Masson 染色观察肾组织形态和纤维化。用脂多糖（LPS）（400ng/mL）/HO（200µM）处理人肾 2 细胞（HK-2）构建体外 CRF 模型。酶联免疫吸附测定（ELISA）验证白细胞介素（IL）-1β和 IL-18 水平。实时定量逆转录聚合酶链反应（RT-qPCR）检测微小 RNA（miR）-342-3p 水平。Western blot 检测转化生长因子-β（TGF-β）1、SMAD2、SMAD3 和细胞焦亡标志物蛋白。双荧光素酶报告基因检测 miR-342-3p 与 TGF-β/SMAD 的相互作用。细胞计数试剂盒-8（CCK-8）检测细胞活力。
结果
在 CRF 模型中，葛根素减轻了肾脏损伤和纤维化，降低了肌酐（Cr）和血尿素氮（BUN）水平。同时，miR-342-3p 下调，而 TGF-β/SMAD 轴被激活，IL-1β 和 IL-18 水平升高。用葛根素处理 CRF 大鼠后，miR-342-3p 表达水平升高，TGF-β/SMAD 轴受到抑制，IL-1β 和 IL-18 的分泌减少。miR-342-3p 直接结合并负调控 TGF-β1、SMAD2 和 SMAD3 的表达。在体外 CRF 模型中，miR-342-3p 通过抑制 TGF-β/SMAD 轴抑制 HK-2 细胞焦亡。
结论
葛根素通过靶向 miR-342-3p/TGF-β/SMAD 轴减轻 CRF 大鼠的肾损伤和细胞焦亡。
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