血管软化丸调控lncRNA-TUG1防治动脉粥样硬化的分子机制

秦合伟; 李彦杰; 孙孟艳; 王梦楠

doi:10.3969/j.issn.1006-2157.2023.03.013

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血管软化丸调控lncRNA-TUG1防治动脉粥样硬化的分子机制

实验研究 | 更新时间：2023-04-07

- 血管软化丸调控lncRNA-TUG1防治动脉粥样硬化的分子机制
- Molecular mechanism of vascular softening pill regulating lncRNA-TUG1 to prevent atherosclerosis
- 北京中医药大学学报 2023年46卷第3期页码：366-376
- 作者机构：
  
  河南中医药大学第二附属医院　郑州　450002
- 作者简介：
  
  秦合伟，男，博士，副主任医师
  #李彦杰，女，教授，主任医师，硕士生导师，主要研究方向：中医药防治心脑血管疾病，E-mail：yanjieli2008@126.com
- 基金信息：
  
  国家自然科学基金项目(81704030);河南省科技攻关计划项目(212102310359);中原英才计划青年拔尖人才项目(豫组通〔2021〕44号)
- DOI：10.3969/j.issn.1006-2157.2023.03.013
  中图分类号： R285.5
- 收稿日期：2022-08-07，
  
  纸质出版日期：2023-03-30
- 稿件说明：
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秦合伟, 李彦杰, 孙孟艳, 等. 血管软化丸调控lncRNA-TUG1防治动脉粥样硬化的分子机制[J]. 北京中医药大学学报, 2023,46(3):366-376.

QIN Hewei, LI Yanjie, SUN Mengyan, et al. Molecular mechanism of vascular softening pill regulating lncRNA-TUG1 to prevent atherosclerosis[J]. Journal of beijing university of traditional chinese medicine, 2023, 46(3): 366-376.
秦合伟, 李彦杰, 孙孟艳, 等. 血管软化丸调控lncRNA-TUG1防治动脉粥样硬化的分子机制[J]. 北京中医药大学学报, 2023,46(3):366-376. DOI： 10.3969/j.issn.1006-2157.2023.03.013.

QIN Hewei, LI Yanjie, SUN Mengyan, et al. Molecular mechanism of vascular softening pill regulating lncRNA-TUG1 to prevent atherosclerosis[J]. Journal of beijing university of traditional chinese medicine, 2023, 46(3): 366-376. DOI： 10.3969/j.issn.1006-2157.2023.03.013.

摘要

目的

观察血管软化丸调控长链非编码RNA牛磺酸上调基因1(lncRNA-TUG1)抑制p38丝裂原活化蛋白激酶(p38 MAPK)信号通路活化及血管炎症反应的分子机制。

方法

体内实验：采用高脂饮食法建立动脉粥样硬化小鼠模型。按照随机数字表法将75只ApoE

－/－

小鼠分为模型组、牛磺酸上调基因1(TUG1)抑制剂组(10 μL TUG1干扰慢病毒)、阴性对照组(10 μL空载慢病毒)、血管软化丸组(43.2 g/kg)、联合组(10 μL TUG1干扰慢病毒＋43.2 g/kg血管软化丸)，每组各15只，连续干预8周。采用免疫组化法检测各组小鼠主动脉TUG1的蛋白表达；酶联免疫吸附测定(ELISA)法检测各组小鼠外周血清中细胞间黏附分子－1 (ICAM-1)、血管细胞黏附分子－1 (VCAM-1)、白细胞介素－8(IL-8)、单核细胞趋化蛋白－1(MCP-1)的含量；实时荧光PCR法检测各组小鼠主动脉TUG1、p38总蛋白(T-p38)的mRNA表达水平。体外实验：建立血管内皮细胞(VEC)功能紊乱模型，采用血管软化丸含药血清进行干预。干预后采用MTT法检测细胞存活率；透射电镜观察细胞超微结构；ELISA法检测细胞液中ICAM-1、VCAM-1、IL-8和MCP-1的含量；实时荧光PCR法检测细胞TUG1、T-p38的mRNA表达水平；蛋白质印迹法检测细胞T-p38、磷酸化p38 (p-p38)蛋白表达水平。

结果

体内实验发现，免疫组化结果显示，TUG1蛋白表达阳性信号为棕黄色或棕褐色，模型组和阴性对照组的动脉粥样硬化斑块形成明显，且TUG1蛋白表达阳性信号强；与模型组相比，血管软化丸组主动脉TUG1平均光密度降低(

0.05)，外周血清中ICAM-1、VCAM-1、IL-8和MCP-1含量降低(

0.05)，主动脉TUG1和T-p38 mRNA表达水平降低(

0.05)。体外实验发现，与模型组相比，血管软化丸血清组的VEC细胞存活率升高(

0.05)；透射电镜显示，血管软化丸血清组细胞内线粒体间距离较模型组小，细胞内纤维排列较模型组整齐；血管软化丸血清组细胞液中ICAM-1、VCAM-1、IL-8和MCP-1含量降低(

0.05)，细胞TUG1和T-p38 mRNA表达水平降低(

0.05)，细胞T-p38、p-p38蛋白表达水平降低(

0.05)。

结论

血管软化丸防治动脉粥样硬化的分子机制可能与通过lncRNA-TUG1调控p38 MAPK信号通路、抑制血管炎症反应、抑制VEC凋亡，保护血管内皮有关。

Abstract

Objective

We aimed to investigate the molecular mechanism by which vascular softening pill regulate the activation of the p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway and the vascular inflammatory response to prevent atherosclerosis by long-stranded non-coding RNA taurine upregulated gene 1 (lncRNA-TUG1).

Methods

In vitro

a high-fat diet was used to establish an atherosclerosis mouse model. Seventy-five ApoE

-/-

mice were randomly divided into the model group

the TUG1 inhibitor group (10 μL TUG1-interfering lentivirus)

the negative control group (10 μL empty lentivirus)

the vascular softening pill group (43.2 g/kg) and the combination group (10 μL TUG1-interfering lentivirus + 43.2 g/kg vascular softening pill)

15 mice in each group. The intervention was continued for 8 weeks. Protein expression of TUG1 were measured by immunohistochemistry; the levels of intercellular adhesion molecule-1 (ICAM-1)

vascular cell adhesion molecule-1 (VCAM-1)

interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) in peripheral serum were measured by ELISA; the mRNA expression levels of TUG1 and total p38 protein (T-p38) in mouse aorta were detected by real-time PCR. In vivo

a vascular endothelial cell (VEC) dysfunction model was established and the vascular softening pill drug serum was used for intervention. After the intervention

survival rate of cell was measured by MTT assay

the cell ultrastructure was observed by transmission electron microscopy

the levels of ICAM-1

VCAM-1

IL-8 and MCP-1 in the serum were measured by ELISA

the mRNA expression levels of TUG1 and T-p38 were measured by real-time PCR

and the protein levels of T-p38 and phosphorylated p38 (p-p38) were measured by Western blotting.

Results

After the intervention

Immunohistochemical detection showed that the positive signal of TUG1 mean optical density was brown yellow or brown. The most obvious atherosclerotic plaque formation and the most obvious positive signal of TUG1 protein expression were found in the model and negative control groups. Compared with the model group

the aortic TUG1 mean optical density was reduced in the vascular softening pill group (

0.05)

the peripheral serum levels of ICAM-1

VCAM-1

IL-8 and MCP-1 were reduced (

0.05)

and aortic TUG1 and T-p38 mRNA expression levels were reduced (

0.05). Cell culture experiments revealed that the survival rate of VEC cells was increased in the vascular softening pill serum group compared with the model group (

0.05). Transmission electron microscopy showed that the distance between mitochondria in the cells in the vascular softening pill serum group was smaller than that in the model group

and the arrangement of intracellular fibers was more orderly than that in the model group. Compared with the model group

the levels of ICAM-1

VCAM-1

IL-8 and MCP-1 in the cell sap of the vascular softening pill serum group were reduced (

0.05)

the cellular TUG1 and T-p38 mRNA expression levels were reduced (

0.05) and the cellular T-p38 and p-p38 protein expression levels were reduced (

0.05).

Conclusion

The molecular mechanism by which vascular softening pills protect against atherosclerosis may be related to the regulation of the p38 MAPK signaling pathway through lncRNA-TUG1

inhibition of the vascular inflammatory response

inhibition of vascular endothelial cells apoptosis and protection of vascular endothelium.

关键词

Keywords

references

DENG B , FANG F , YANG TL , et al . Ghrelin inhibRs AngII-induced expression of TNF-alpha, IL-8, MCP-1 in human umbilical vein endotheaal cells [J]. Int J Exp Med , 2018 , 8 ( 1 ): 579 － 588 .

李孟婷，李宏帆，杨彬，等 . LncRNA TUGl在血管内皮细胞功能紊乱中的作用研究 [J]. 中国分子心脏病学杂志， 2016 , 16 ( 2 ): 1649 － 1653 .

JERSMANN HP , HII CS , FERRANTE JV , et al . Bacterial Lipopolysaccharide and Tumor Necrosis Factor Alpha Synergistically Increase Expression of Human Endothelial Adhesion Molecules through Activation of NF-κB and p38 Mitogen-Activated Protein Kinase Signaling Pathways [J]. Infect Immun , 2001 , 69 ( 3 ): 1273 － 1279 .

秦合伟，李彦杰，李斯锦，等 . 化痰祛瘀中药抗动脉粥样硬化的作用机制研究 [J]. 北京中医药大学学报， 2019 , 42 ( 7 ): 566 － 576 .

孔艳芳，秦合伟，王媛，等 . 血管软化丸调控miR－17－5p与PCSK9/VLDLR信号通路抗动脉粥样硬化研究 [J]. 安徽中医药大学学报， 2021 , 40 ( 6 ): 69 － 75 .

秦合伟，李彦杰，李斯锦 . 血管软化丸调控CD40－CD40L系统及抗动脉粥样硬化的作用机制研究 [J]. 中华中医药学刊， 2019 , 37 ( 6 ): 1306 － 1309 .

刘建东，秦合伟 . 基于血脂代谢紊乱理论研究血管软化丸抗动脉粥样硬化的作用机制 [J]. 中西医结合心脑血管病杂志， 2018 ， 16 ( 20 )： 2943 － 2947 .

张永晨，秦合伟，赵平丽 . 血管软化丸调控miRNA－467b抗动脉粥样硬化的作用与初步机制研究 [J]. 安徽中医药大学学报， 2020 , 39 ( 5 )： 66 － 71 .

MOGHADASIAN MH , MCMANUS BM , NGUYEN LB , etal . Pathophysiology of apolipoprotein E deficiency in mice: relevance to apo E-related disorders in humans [J]. FASEB J , 2001 , 15 ( 14 ): 2623 － 2630 .

唐泽民 . 干扰长链非编码RNA Z38表达对鼻咽癌细胞增殖的影响 [D]. 长沙：湖南大学， 2017 .

MAN HSJ , SUKUMAR AN , LAM GC , et al . Angiogenic patterning by STEEL, an endothelial-enriched long noncoding RNA [J]. Proc Natl Acad Sci USA , 2018 , 115 ( 10 ): 2401 － 2406 .

ZHANG XL , LIU J , GU YF , et al . Down-regulation of lncRNA OGFRP1 induces autophagy and growth inhibition by AKT/mTOR signaling pathway in HCAECs [J]. Cell Biol Int , 2019 , 43 ( 2 ): 158 － 166 .

ZHAO X , SU L , HE XY , et al . Long noncoding RNA CA7－4 promotes autophagy and apoptosis via sponging MIR877－3P and MIR5680 in high glucose-induced vascular endothelial cells [J]. Autophagy , 2020 , 16 ( 1 ): 70 － 85 .

DORAN AC . Inflammation Resolution: Implications for Atherosclerosis [J]. Circ Res , 2022 , 130 ( 1 )： 130 － 148 .

LIBBY P . The changing landscape of atherosclerosis [J]. Nature , 2021 , 592 ( 7855 ): 524 － 533 .

LAHIRI V , HAWKINS WD , KLIONSKY DJ . Watch What You (Self－) Eat: Autophagic Mechanisms that Modulate Metabolism [J]. Cell Metab , 2019 , 29 ( 4 )： 803 － 826 .

MANDATORI S , PACELLA I , MARZOLLA V , et al . Altered Tregs Differentiation and Impaired Autophagy Correlate to Atherosclerotic Disease [J/OL]. Front Immunol , 2020 , 11 : 350 [ 2022－08－01 ]. https：//www.frontiersin.org/articles/10.3389/fimmu.2020.00350/full https：//www.frontiersin.org/articles/10.3389/fimmu.2020.00350/full .

WANG XH ， YANG C ， XU XL ， et a1 . Long noncoding RNA expression profiles of hypoxic pulmonary hypertension rat model [J]. Gene , 2017 , 579 ( 1 ): 23 － 28 .

GUO LW , WANG LY , LI HF , et al . Down regulation of GALNT3 contributes to endothelial cell injury via activation of p38 MAPK signaling pathway [J]. Atherosclerosis ， 2016 , 245 : 94 － 100 .

ZHAO Z , SUN W , GUO ZY , et al . Long Noncoding RNAs in Myocardial Ischemia-Reperfusion Injury [J/OL]. Oxid Med Cell Longev , 2021 ， 2021 : 8889123 [ 2022－08－01 ]. https：//www.hindawi.com/journals/omcl/2021/8889123/ https：//www.hindawi.com/journals/omcl/2021/8889123/ .

ZHANG H , LIU B , SHI XJ , et al . Long noncoding RNAs: Potential therapeutic targets in cardiocerebrovascular diseases [J/OL]. Pharmacol Ther , 2021 ， 221 ： 107744 [ 2022－08－01 ]. https：//www.sciencedirect.com/science/article/pii/S0163725820302758？via%3Dihub https：//www.sciencedirect.com/science/article/pii/S0163725820302758？via%3Dihub .

HOFMANN P , SOMMER J , THEODOROU K , et al . Long non-coding RNA H19 regulates endothelial cell aging via inhibition of STAT3 signalling [J]. Cardiovasc Res , 2019 , 115 ( 1 ): 230 － 242 .

ZHONG XM , MA X , ZHANG L , et al . MIAT promotes proliferation and hinders apoptosis by modulating miR－181b/STAT3 axis in ox-LDL-induced atherosclerosis cell models [J]. Biomed Pharmacother , 2018 , 97 : 1078 － 1085 .

WANG S , HAN X , MAO ZC , et al . MALAT1 lncRNA Induces Autophagy and Protects Brain Microvascular Endothelial Cells Against Oxygen-Glucose Deprivation by Binding to miR－200c－3p and Upregulating SIRT1 Expression [J]. Neuroscience , 2019 , 397 : 116 － 126 .

AFLAKI E , MOAVEN N , BORGER D K , et al . Lysosomal storage and impaired autophagy lead to inflammasome activation in Gaucher macrophages [J]. Aging Cell , 2016 , 15 ( 1 ): 77 － 88 .

DEROISSART J , PORSCH F , KOLLER T , et al . Anti-inflammatory and Immunomodulatory Therapies in Atherosclerosis [J]. Handb Exp Pharmacol , 2022 , 270 ： 359 － 404 .

赵晶，秦合伟，李彦杰，等 . 血管软化丸调控PI3K/Akt/mTOR通路影响细胞自噬及抗动脉粥样硬化的作用机制研究 [J]. 中华中医药学刊， 2020 , 38 ( 1 ): 65 － 69 .

秦合伟，李彦杰，任锟，等 . 基于TLR3/TLR9介导巨噬细胞自噬/极化效应探讨血管软化丸抗AS的作用机制 [J]. 辽宁中医杂志， 2019 , 46 ( 1 ): 156 － 160, 225 .

聂勇，秦合伟，吕哲，等 . 血管软化丸调控miRNA－155防治动脉粥样硬化的机制 [J]. 安徽中医药大学学报， 2021 , 40 ( 2 ): 72 － 76 .

秦合伟，李彦杰，张志鑫，等 . 化痰祛瘀法通过调控miR－181影响输入蛋白α3/NF－κB通路的抗动脉粥样硬化的研究 [J]. 北京中医药大学学报， 2020 , 43 ( 8 ): 653 － 660 .

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