WTAP promotes microglial M1 polarization by regulating SIRT1

Hongxiu ZHOU; Xu GAO; Shengping HOU

doi:10.16016/j.2097-0927.202109187

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WTAP promotes microglial M1 polarization by regulating SIRT1

Neurosciences | 更新时间：2026-01-09

- WTAP promotes microglial M1 polarization by regulating SIRT1
- Journal of Army Medical University Vol. 44, Issue 7, Pages: 620-629(2022)
- 作者机构：
  
  1.400016 重庆，重庆医科大学附属第一院眼科，眼科学重庆市重点实验室，重庆市眼科研究所
  2.402760 重庆，重庆市璧山区人民医院眼科
- 作者简介：
- 基金信息：
  
  the Project of Innovative Research Group of Chongqing Education Commission(CXQT19015);the General Program of Natural Science Foundation of Chongqing(cstc2019jcyjmsxmX0120);the Innovation Supporting Plan of Overseas Returnees of Chongqing(cx2018010);the Project of Chongqing Education Commission(KJQN202000406)
- DOI：10.16016/j.2097-0927.202109187
  CLC： R322.91;R329.25;R341
- Received：24 September 2021，
  
  Revised：2021-11-17，
  
  Published：15 April 2022
- 稿件说明：
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Hongxiu ZHOU, Xu GAO, Shengping HOU. WTAP promotes microglial M1 polarization by regulating SIRT1[J]. Journal of Army Medical University, 2022, 44(7): 620-629.
DOI：

Hongxiu ZHOU, Xu GAO, Shengping HOU. WTAP promotes microglial M1 polarization by regulating SIRT1[J]. Journal of Army Medical University, 2022, 44(7): 620-629. DOI： 10.16016/j.2097-0927.202109187.

摘要

目的

探讨Wilms肿瘤抑制因子1相关蛋白(Wilms' tumor 1-associating protein，WTAP)在小胶质细胞活化调控中的作用及其可能的分子机制。

方法

用100 ng/mL IFN

联合1 μg/mL LPS诱导HMC3小胶质细胞M1极化作为实验组，以未处理的细胞作为空白对照组，采用实时荧光定量PCR和Western blot检测细胞中促炎细胞因子IL-6、IL-1β、TNF-α和N6-甲基腺苷(m6A)相关基因的表达，在HMC3小胶质细胞中转染RNAi慢病毒敲低WTAP，使用实时荧光定量PCR和Western blot检测敲低效率，Western blot检测敲低HMC3小胶质细胞WTAP后促炎细胞因子IL-6、IL-1β和TNF-α蛋白表达，Western blot检测敲低WTAP后HMC3小胶质细胞沉默信息调节因子2相关酶1 (silent mating-type information regulation 2 homolog 1

SIRT1)、COP1、USP18、EP4、促炎转录因子和部分炎症相关通路受体的表达，使用SIRT1激动剂SRT1720和SIRT1抑制剂EX527分别处理空载病毒组和敲低WTAP组HMC3细胞后，Western blot检测c/EBPβ的表达。

结果

与空白对照组比较，100 ng/mL IFN

联合1 μg/mL LPS诱导的HMC3细胞IL-6、IL-1β、TNF-α表达上调(

0.05)。M1极化的HMC3小胶质细胞中WTAP高表达(

0.05)。敲低WTAP后，HMC3细胞促炎细胞因子IL-6、IL-1β和TNF-α表达降低(

0.05)，SIRT1表达升高(

0.05)，且促炎转录因子c/EBPβ表达降低(

0.05)。SIRT1激动剂SRT1720使空载病毒组相对高表达的c/EBPβ降低(

0.01)。SIRT1抑制剂EX527使敲低WTAP的HMC3细胞相对低表达的c/EBPβ升高(

0.05)。

结论

WTAP促进HMC3小胶质细胞M1极化，其作用机制可能与通过调控SIRT1表达进而影响c/EBPβ信号通路有关。

Abstract

Objective

To investigate the role and the possible molecular mechanism of RNA methylase

Wilms' tumor 1-associating protein (WTAP)

in microglial activation.

Methods

HMC3 cells was induced to M1 polarization by treatment of 100 ng/mL

IFN

combined with 1 μg/mL LPS and then served as experimental group

and the untreated cells served as control group. The expression of pro-inflammatory cytokines IL-6

IL-1β

TNF-α and N6-methyladenosine (m6A) at mRNA and protein levels was detected by real-time quantitative PCR and Western blotting

respectively. After HMC3 cells were transfected with RNAi lentivirus to knock down WTAP

the knockdown efficiency was detected by real-time quantitative PCR and Western blotting. Western blot assay was employed to measure the expression levels of pro-inflammatory cytokines IL-6 IL-1β and TNF-α in WTAP knockdown HMC3 cells. Western blotting was also adopted to detect the protein levels of silent mating-type information regulation 2 homolog 1 (SIRT1)

COP1

USP18

EP4

pro-inflammatory transcription factors and some inflammatory pathway receptors in the knockdown cells. SIRT1 agonist SRT1720 and SIRT1 inhibitor EX527 were employed to treat HMC3 cells in the blank-virus group and the knockdown WTAP group

respectively

and the expression level of C/EBPβ was detected by Western blotting.

Results

Treatment of 100 ng/mL IFN

combined with 1 μg/mL LPS induced significant up-regulation of IL-6

IL-1β and TNF-α expression in HMC3 cells as compared with the untreated cells (

0.05). WTAP was highly expressed in M1 polarized HMC3 microglia (

0.05). WTAP knockdown resulted in obviously decreased expression levels of pro-inflammatory cytokines IL-6

IL-1β and TNF-α (

0.05)

increased level of SIRT1 (

0.05)

and reduced pro-inflammatory transcription factor C/EBPβ (

0.05). SIRT1 agonist SRT1720 reduced the relatively high expression of C/EBPβ in the blank-virus group (

0.01). While

the inhibitor EX527 increased the relative low expression of C/EBPβ in HMC3 cells with low WTAP knockdown (

0.05).

Conclusion

WTAP promotes M1 polarization in HMC3 cells

possibly through regulating the expression of SIRT1 and thus affecting the C/EBPβ signaling pathway.

关键词

Keywords

references

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