缺氧通过HIF-1α介导NRP-1上调抑制破骨细胞分化

李松涛; 孙靖; 陈武桂; 马敏; 张莹; 牛晓健; 周驰雨; 初同伟

doi:10.16016/j.1000-5404.201811206

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缺氧通过HIF-1α介导NRP-1上调抑制破骨细胞分化

基础医学 | 更新时间：2026-01-09

- 缺氧通过HIF-1α介导NRP-1上调抑制破骨细胞分化
- Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1
- 第三军医大学学报 2019年41卷第11期页码：1052-1058
- 作者机构：
  
  400037 重庆，陆军军医大学(第三军医大学)第二附属医院骨科
- 作者简介：
  
  [初同伟，E-mail：chtw@sina.comCHU Tongwei, E-mail: chtw@sina.com
- 基金信息：
  
  国家自然科学基金面上项目(81570800)
- DOI：10.16016/j.1000-5404.201811206
  中图分类号： R322.71;R329.21;R329.28
- 收稿：2018-11-28，
  
  修回：2019-01-08，
  
  纸质出版：2019-06-15
- 稿件说明：
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李松涛, 孙靖, 陈武桂, 等. 缺氧通过HIF-1α介导NRP-1上调抑制破骨细胞分化[J]. 第三军医大学学报, 2019,41(11):1052-1058.

Songtao LI, Jing SUN, Wugui CHEN, et al. Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1[J]. Journal of Third Military Medical University, 2019, 41(11): 1052-1058.
李松涛, 孙靖, 陈武桂, 等. 缺氧通过HIF-1α介导NRP-1上调抑制破骨细胞分化[J]. 第三军医大学学报, 2019,41(11):1052-1058. DOI： 10.16016/j.1000-5404.201811206.

Songtao LI, Jing SUN, Wugui CHEN, et al. Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1[J]. Journal of Third Military Medical University, 2019, 41(11): 1052-1058. DOI： 10.16016/j.1000-5404.201811206.

摘要

目的

研究低氧/HIF-1α通路对RAW264.7细胞向成熟破骨细胞分化的调控作用，并初步探讨其可能的分子机制。

方法

采用短发夹RNA(short hairpin RNA，shRNA)慢病毒感染法，分别建立HIF-1α和NRP-1稳定低表达的RAW264.7细胞系。将RAW264.7细胞分组培养：对照组，分化组(RANKL 100 ng/mL+50%成骨细胞CM)，缺氧组(RANKL 100 ng/mL+50%成骨细胞CM+100 μmol/L CoCl

氯化钴)，缺氧+HIF-1α-NC组，缺氧+HIF-1α-shRNA干扰组，缺氧+NRP-1-NC组及缺氧+NRP-1-shRNA干扰组。TRAP染色观察破骨细胞的分化情况；RT-PCR检测Cath K、MMP-9、TRAP、HIF-1α和NRP-1 mRNA的表达；Western blot检测HIF-1α和NRP-1蛋白的表达。

结果

① 缺氧组破骨细胞分化相关基因Cath K、MMP-9、TRAP mRNA的表达量及TRAP染色阳性细胞数均明显低于分化组(

0.05)。②缺氧组细胞中HIF-1α和NRP-1的表达均明显高于分化组(

0.05)。③与缺氧组相比，缺氧+HIF-1α-shRNA干扰组TRAP染色阳性细胞数及破骨细胞分化相关基因的表达均明显增加(

0.05)；且HIF-1α和NRP-1表达均明显减少(

0.05)。④与缺氧组相比，缺氧+NRP-1-shRNA干扰组TRAP染色阳性细胞数及破骨细胞分化相关基因的表达均明显增加(

0.05)；且NRP-1表达明显减少(

0.05)。

结论

在RAW264.7细胞中，CoCl

诱导的缺氧能够通过上调HIF-1α增强NRP-1的表达，从而抑制其向成熟破骨细胞分化。

Abstract

Objective

To investigate the regulation effect of hypoxic/HIF-1α pathway on the differentiation of RAW264.7 cells into mature osteoclasts

and to explore the possible molecular mechanism.

Methods

RAW264.7 cell lines with stable low expression of HIF-1α and NRP-1 were respectively established by short hairpin RNA (shRNA) lentivirus infection. Then the RAW264.7 cells were cultured in groups

that is

control group

differentiation group [100 ng/mL RANKL+50% osteoblast conditioned medium (CM)

]

hypoxia group (100 ng/mL RANKL+50% osteoblast CM+100 μmol/L CoCl

)

hypoxia+HIF-1α-NC group

hypoxia+HIF-1α shRNA group

hypoxia+NRP-1-NC group

and hypoxia+NRP-1-shRNA group. TRAP staining was used to observe osteoclast differentiation. The mRNA expression of Cath K

MMP-9

TRAP

HIF-1α and NRP-1 were detected by RT-PCR. Western blotting was used to detect the expression of HIF-1α and NRP-1

proteins.

Results

① The mRNA expression levels of Cath K

MMP-9 and TRAP and the number of TRAP positive cells in the hypoxia groups were significantly lower than those in the differentiation groups (

0.05). ② The expression levels of HIF-1α and NRP-1 were significantly higher in the hypoxic groups than the differentiation groups (

0.05). ③ Compared with the hypoxia groups

the number of TRAP positive cells and the expression levels of osteoclast differentiation related genes were significantly increased

while those of HIF-1α and NRP-1 were significantly reduced in the hypoxic+HIF-1α shRNA group (

0.05). ④ Compared with the hypoxia groups

the number of TRAP positive cells and the expression levels of osteoclast differentiation related genes were significantly increased

while the level of NRP-1 was significantly decreased in the hypoxia+NRP-1-shRNA group (

0.05).

Conclusion

The hypoxia-mediated HIF-1α-dependent up-regulation of NRP-1 inhibits the differentiation of RAW264.7 cells into mature osteoclasts.

关键词

Keywords

references

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