Effect of siRNA-mediated α-TAT1 gene silencing on migration behavior of endothelial cells in rats with hepatopulmonary syndrome

Chang LIU; Jiaxi ZHU; Yanan LIU; Mu XU; Jingli CHEN

doi:10.16016/j.2097-0927.202305047

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Effect of siRNA-mediated α-TAT1 gene silencing on migration behavior of endothelial cells in rats with hepatopulmonary syndrome

Basic Medicine | 更新时间：2026-01-09

- Effect of siRNA-mediated α-TAT1 gene silencing on migration behavior of endothelial cells in rats with hepatopulmonary syndrome
- Journal of Army Medical University Vol. 46, Issue 3, Pages: 215-224(2024)
- 作者机构：
  
  1.431600 武汉，华中科技大学同济医学院附属武汉市中心医院麻醉科
  2.431600 武汉，湖北省分子诊断重点实验室
- 作者简介：
- 基金信息：
  
  the National Natural Science Foundation for Young Scholars of China(82000584)
- DOI：10.16016/j.2097-0927.202305047
  CLC： R322.12;R394.3;R575
- Received：15 May 2023，
  
  Revised：2023-07-07，
  
  Published：15 February 2024
- 稿件说明：
移动端阅览
Chang LIU, Jiaxi ZHU, Yanan LIU, et al. Effect of siRNA-mediated α-TAT1 gene silencing on migration behavior of endothelial cells in rats with hepatopulmonary syndrome[J]. Journal of Army Medical University, 2024, 46(3): 215-224.
DOI：

Chang LIU, Jiaxi ZHU, Yanan LIU, et al. Effect of siRNA-mediated α-TAT1 gene silencing on migration behavior of endothelial cells in rats with hepatopulmonary syndrome[J]. Journal of Army Medical University, 2024, 46(3): 215-224. DOI： 10.16016/j.2097-0927.202305047.

摘要

目的

探究siRNA沉默微管乙酰化转移酶1(alpha tubulin acetyltransferase 1，α-TAT1)对肝肺综合征(hepatopulmonary syndrome

HPS)中内皮细胞迁移行为的影响。

方法

通过在线数据库Tabula Muris分析肺组织各细胞亚群α-TAT1的表达。24只雄性SD大鼠按照随机数字表法分为对照组(Sham组，

=6)、胆管结扎组(HPS组，

＝18)。造模后在第2、4周分别取材HPS组大鼠，通过免疫荧光共定位检测2周和4周HPS大鼠肺血管内皮细胞中α-TAT1表达水平变化。收集Sham和HPS大鼠血清处理人脐静脉血管内皮细胞(human umbilical vein endothelial cells，HUVECs) 12 h和24 h，之后进行小分子RNA(small interfering RNA

siRNA)干扰实验，分为Sham血清+siRNA NC组、Sham血清+siRNA α-TAT1组、HPS血清+siRNA NC组、HPS血清+siRNA α-TAT1组。Western blot检测各组HUVECs中α-TAT1、Ace-α-tubulin表达水平变化。免疫荧光观察各组诺考达唑(10 μmol/L)预处理后HUVECs中α-Tubulin微管聚合水平，评估微管的结构稳定性；细胞划痕实验联合细胞免疫荧光观察HUVECs的细胞迁移能力和高尔基体的定位。

体外成血管实验检测HUVECs成血管能力。

结果

体内及体外实验表明HPS中肺血管内皮细胞α-TAT1表达显著增加(

0.01)。siRNA α-TAT1干扰组中α-TAT1和Ace-α-tubulin表达量均显著下调

且微管稳定性减弱(

0.01)。另外，siRNA α-TAT1干扰组中GM130标记的高尔基体在核周细胞突出段方向的分布比例降低，且迁移能力下降(

0.01)。同时，成血管长度及网络水平显著减弱(

0.01)。

结论

沉默α-TAT1可减弱HPS中内皮细胞迁移和成血管能力，其机制与减弱微管稳定性有关。

Abstract

Objective

To investigate the effect of silencing alpha tubulin acetyltransferase 1 (α-TAT1) on migration behavior of endothelial cells induced by hepatopulmonary syndrome (HPS).

Methods

Online database Tabula Muris was used to analyze the expression of α-TAT1 in various cell subsets in the lungs. Twenty-four male SD rats were randomly divided into control group (Sham group

=6) and common bile duct ligation group (HPS group

=18). The rats in HPS group were euthanasized at 2 and 4 weeks after modelling

and then the expression of α-TAT1 in pulmonary vascular endothelial cells was detected by immunofluorescence colocalization. The sera from the Sham and HPS rats were used to stimulate human umbilical vein endothelial cells (HUVECs) for 12 and 24 h

respectively. Then the obtained HUVECs were divided into 4 groups: Sham serum + siRNA NC group

Sham serum + siRNA α-TAT1 group

HPS serum+ siRNA NC group

HPS serum + siRNA α-TAT1 group. The expression levels of α-TAT1 and Ace-α-tubulin in HUVECs were detected by Western blotting. Immunofluorescence assay was applied to observe the levels of polymerized microtubules of α-Tubulin in HUVECs after nocodazole (10 μmol/L) pretreatment to evaluate the stability of microtubule structure. Cell scratch assay combined with cell immunofluorescence assay was employed to observe the nuclear localization of Golgi apparatus and cell migration ability of HUVECs. The angiogenesis ability of HUVECs wa

s tested by

in vitro

angiogenesis test.

Results

In vivo

and

in vitro

experiments showed that the expression of α-TAT1 in endothelial cells was significantly increased after HPS inducement. The expression levels of α-TAT1 and Ace-α-tubulin were significantly down-regulated

and the stability of microtubules was weakened in the siRNA α-TAT1 interference group (

0.01). In addition

the distribution of GM130 labeled Golgi apparatus in the protrusion of HUVECs was down-regulated in the siRNA α-TAT1 interference group

as well as the migration ability (

0.01). And the length of angiogenesis and network level were also significantly declined (

0.01).

Conclusion

Silencing α-TAT1 reduces the migration and angiogenesis of endothelial cells in HPS

which was associated with weakened stabilization of microtubule.

关键词

Keywords

references

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