乌头汤通过lncRNA NEAT1调控IRE1α/XBP1途径减轻软骨细胞内质网应激的作用
Effects of Wutou Decoction on regulation of lncRNA NEAT1 in regulating the IRE1α/XBP1 pathway to relieve endoplasmic reticulum stress in chondrocyte
金灵璐 1涂海水 1谢新宇 1马德尊 1叶锦霞 1郑春松 1仲卫红 2付长龙1
作者信息
- 1. 福建中医药大学中西医结合研究院,福州 350122;福建省中西医结合老年性疾病重点实验室,福州 350122
- 2. 福建省康复技术重点实验室,福州 350003;福建中医药大学附属康复医院,福州 350003
- 折叠
摘要
目的:探讨乌头汤通过lncRNA NEAT1调控IRE1α/XBP1途径减轻毒胡萝卜素(TG)诱导的软骨细胞内质网应激的作用与机制.方法:采用荧光原位杂交(FISH)检测25 μmol/L TG诱导的软骨细胞中lneRNA NEAT1分布情况;构建lncRNA NEAT1沉默软骨细胞,Real-time PCR检测其IRE1α、XBP1 mRNA水平变化;Western Blot检测其IRE1α、XBP1蛋白表达.使用第一代软骨细胞进行后续实验,设置空白组(10％DMEM)、TG组(25 μmol/L TG+10％DMEM)、乌头汤含药血清组(25 μmol/L TG+不同浓度含药血清).Real-time PCR检测TG诱导的软骨细胞中lncRNA NEAT1 mRNA水平变化;Western Blot检测TG诱导的软骨细胞中IRE1α、XBP1、EDEM1、CHOP蛋白表达.免疫荧光染色观察各组软骨细胞中IRE1α、Caspase-12的荧光表达量.流式细胞术检测各组软骨细胞凋亡率情况.结果:FISH结果显示,lncRNA NEAT1主要分布于软骨细胞的胞核中.与TG+Lenti-control组比较,TG+sh-NEAT1组IRE1α、XBP1 mRNA与蛋白表达显著降低(P＜0.05,P＜0.01).与TG组比较,乌头汤含药血清组lncRNA NEAT1水平及IRE1α、XBP1、EDEM1、CHOP蛋白表达显著降低(P＜0.01),IRE1α、Caspase-12的荧光表达量显著降低(P＜0.05),软骨细胞凋亡率显著下降(P＜0.05).结论:乌头汤可通过lneRNA NEAT1调控IRE1α/XBP1途径减轻TG诱导的软骨细胞内质网应激,延缓软骨细胞退变.
Abstract
Objective:To investigate the effect and mechanism of Wutou Decoction in alleviating endoplasmic reticulum stress in thapsigargin(TG)induced chondrocytes through the lncRNA NEAT 1-controlled IRE1α/XBP1 pathway.Methods:Fluorescent in situ hybridization(FISH)was used to detect the distribution and expression of lncRNA NEAT1 in 25 μmol/L TG-induced chondrosinocytes;lncRNA NEAT1 silencing chondrocytes were constructed,and Real-time PCR was used to detect IRE1α and XBP1 mRNA levels.Western Blot was used to detect its IRE1α and XBP1 protein expression.The first generation of chondrocytes was used for follow-up experiments,blank group(10％DMEM),TG group(25 μmol/L TG+10％DMEM),Wutou Decoction with medicinal serum group(25 μmol/L TG+different concentrations containing serum)were set up.Real-time PCR was used to detect the level of lncRNA NEAT1 mRNA in TG-induced chondrocytes;Western Blot was used to detect the expression of IRE1α,XBP1,EDEM1,and CHOP proteins in TG-induced chondrocytes.Immunofluorescence staining observed the fluorescence expression of IRE1α and Caspase-12 in each group of chondrocytes.Flow cytometry measures the apoptosis rate of chondrocytes in each group.Results:The FISH results showed that lncRNA NEAT1 was mainly distributed in the nucleus of chondrocytes.Compared with the TG+Lenti-control group,the mRNA levels and protein expression of IRE1α,XBP1 in the TG+sh-NEAT1 group were significantly reduced(P＜0.05,P＜0.01).Compared with the TG group,the levels of lncRNA NEAT1 and the expression of IRE1α,XBP1,EDEM1 and CHOP proteins in Wutou Decoction with medicinal serum group were significantly reduced(P＜0.01),the fluorescence expression of IRE1α and Caspase-12 was significantly reduced(P＜0.05),and the apoptosis rate of chondrocytes was significantly reduced(P＜0.05).Conclusion:Through lncRNA NEAT1 regulating IRE1α/XBP1 pathway,Wutou Decoction can relieve endoplasmic reticulum stress induced by TG and delay the degeneration of chondrocytes.
关键词
骨关节炎/乌头汤/内质网应激/软骨细胞/lncRNA/NEAT1Key words
Osteoarthritis(OA)/Wutou Decoction/Endoplasmic reticulum stress/Chondrocytes/lncRNA NEAT1引用本文复制引用
基金项目
国家自然科学基金(82104888)
福建省自然科学基金(2022J01370)
福建省自然科学基金(2022J01374)
福建中医药大学高层次人才科研启动资金项目(X2019011-人才)
出版年
2024
NETL
NSTL
万方数据