Transient receptor potential vanilloid 4-dependent calcium influx, ATP release and expression of inflammatory factors in enteric glial cells

Qianshan TAN; Jun CHEN; Huichao XIE; Liqi LI; Shuaishuai CHEN; Hui DONG; Weidong XIAO

doi:10.16016/j.2097-0927.202211062

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Transient receptor potential vanilloid 4-dependent calcium influx, ATP release and expression of inflammatory factors in enteric glial cells

Basic Medicine | 更新时间：2026-01-09

- Transient receptor potential vanilloid 4-dependent calcium influx, ATP release and expression of inflammatory factors in enteric glial cells
- Journal of Army Medical University Vol. 45, Issue 5, Pages: 417-425(2023)
- 作者机构：
  
  1.400037 重庆，陆军军医大学(第三军医大学)第二附属医院普通外科
  2.400037 重庆，陆军军医大学(第三军医大学)第二附属医院消化科
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(82270585);the General Program of National Natural Science Foundation of China(81770524);the Chongqing Talent Program(15014);the High-Tech Incubation Project of Military Medicine(CX2019JS212)
- DOI：10.16016/j.2097-0927.202211062
  CLC： R322.45;R329.27;R392.3
- Received：09 November 2022，
  
  Revised：2022-12-22，
  
  Published：15 March 2023
- 稿件说明：
移动端阅览
Qianshan TAN, Jun CHEN, Huichao XIE, et al. Transient receptor potential vanilloid 4-dependent calcium influx, ATP release and expression of inflammatory factors in enteric glial cells[J]. Journal of Army Medical University, 2023, 45(5): 417-425.
DOI：

Qianshan TAN, Jun CHEN, Huichao XIE, et al. Transient receptor potential vanilloid 4-dependent calcium influx, ATP release and expression of inflammatory factors in enteric glial cells[J]. Journal of Army Medical University, 2023, 45(5): 417-425. DOI： 10.16016/j.2097-0927.202211062.

摘要

目的

探讨瞬时感受器电位离子通道香草素受体4(transient receptor potential vanilloid 4，TRPV4)在调节肠胶质细胞外Ca

内流中的作用及其意义。

方法

收集大鼠肠胶质细胞系CRL-2690、大鼠肠上皮细胞系IEC6和人胚肾细胞系HEK293T样本，采用RT-qPCR、Western blot和免疫荧光方法检测TRPV4的mRNA、蛋白表达与定位；将CRL-2690细胞依次分为GSK组与GSK+HC组，低渗组与低渗+HC组，GdCl

组与GdCl

+HC组以及CaCl

组与CaCl

+HC组，采用Fura-2荧光探针和单细胞荧光Ca

检测仪评估CRL-2690细胞中TRPV4通道活性。将CRL-2690细胞分为对照组、GSK组与GSK+HC组，通过荧光素-荧光素酶实验测量细胞ATP的释放水平；采用RT-qPCR检测炎症相关基因GFAP、IL-1β、IL-6、IL-10的mRNA表达。

结果

在大鼠肠胶质细胞系CRL-2690中检测到TRPV4 mRNA和蛋白表达；TRPV4特异性激动剂GSK1016790A和低渗透压刺激明显增强了CRL-2690细胞内钙荧光强度，而TRPV4特异性抑制剂HC067047可抑制此作用(0.430 38±0.063 65

0.004 23±0.005 78，

0.000 1)；钙敏感受体(calcium-sensing receptor，CaSR)激活诱导的外Ca

内流也可被HC067047所抑制(

0.05)；功能上，激活TRPV4可促ATP释放、EGC反应性增生标志物GFAP和炎症因子IL-1β、IL-6的表达，并抑制抗炎因子IL-10的表达，该作用可被HC067047所抑制(

0.05)。

结论

激活TRPV4可引起肠胶质细胞外Ca

内流，并促进ATP释放与炎症因子表达，可能参与肠道炎症的发生。

Abstract

Objective

To investigate the role and physiological significance of transient receptor potential vanilloid receptor 4 (TRPV4) in regulation of calcium influx in rat intestinal glial cells.

Methods

The expression of TRPV4 at mRNA and protein levels and its localization were detected by RT-qPCR

Western blotting and immunofluorescence assay in rat enteric glial cell line CRL-2690

rat intestinal epithelial cell line IEC6 and human embryonic kidney cell line HEK293T. The CRL-2690 cells were divided into GSK1016790A (GSK

TRPV4-specific agonist) group and GSK+HC067047 (HC

TRPV4-specific inhibitor) group

hypotonic group and hypotonic+HC group

GdCl

group and GdCl

+HC group

and CaCl

group and

CaCl

+HC group

while the activity of TRPV4 channel was assessed by Fura-2 fluorescent probe and single-cell fluorescent calcium ion assay. CRL-2690 cells were also divided into control group

GSK group and GSK+HC group

and the level of cellular ATP release was measured by luciferin-luciferase assay

and the mRNA levels of inflammatory genes GFAP

IL-1β

IL-6 and IL-10 were detected by RT-qPCR.

Results

TRPV4 was expressed at mRNA and protein levels in rat intestinal glial cell line CRL-2690. GSK and low osmolarity stimulation significantly enhanced intracellular calcium fluorescence intensity in CRL-2690 cells

which was inhibited by HC (0.430 38±0.063 65

0.004 23±0.005 78

0.000 1). Calcium-sensing receptor (CaSR) activation-induced Ca

influx was also inhibited by HC (

0.05). Functionally

activation of TRPV4 promoted ATP release

expression of EGC reactive proliferation marker

GFAP

and inflammatory factors IL-1 β as well as IL-6

and inhibited the expression of anti-inflammatory factor IL-10 respectively. But all these effects were inhibited by HC treatment (

0.05).

Conclusion

Activation of TRPV4 induces extracellular Ca

influx and promotes ATP release and expression of inflammatory cytokines in enteric glial cell

which may be closely involved in the underlying mechanisms of bowel inflammation.

关键词

Keywords

references

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