Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1

Songtao LI; Jing SUN; Wugui CHEN; Min MA; Ying ZHANG; Xiaojian NIU; Chiyu ZHOU; Tongwei CHU

doi:10.16016/j.1000-5404.201811206

您当前的位置：

首页 >

文章列表页 >

Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1

Basic Medicine | 更新时间：2026-01-09

- Hypoxia inhibits osteoclast differentiation by HIF-1α-mediated upregulation of NRP-1
- Journal of Third Military Medical University Vol. 41, Issue 11, Pages: 1052-1058(2019)
- 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：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(81570800)
- DOI：10.16016/j.1000-5404.201811206
  CLC： R322.71;R329.21;R329.28
- Received：28 November 2018，
  
  Revised：2019-01-08，
  
  Published：15 June 2019
- 稿件说明：
移动端阅览
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：

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

UPADHYAY J, FARR O M, MANTZOROS C S. The role of leptin in regulating bone metabolism[J]. Metab Clin Exp, 2015, 64(1):105-113.DOI:10.1016/j.metabol.2014.10.021.

YAGI M, MIYAMOTO T, TOYAMA Y, et al . Role of DC-STAMP in cellular fusion of osteoclasts and macrophage giant cells[J ] . J Bone Miner Metab, 2006, 24(5): 355-358. DOI:10.1007/s00774-006-0697-9.

陈莉丽, 黄玫, 雷利红, 等. OPG/RANKL/RANK系统参与牙槽骨吸收及重建过程作用初探[J].第三军医大学学报, 2013, 35(4): 288-292.

CHEN L L, HUANG M, LEI L H, et al . Role of OPG/RANKL/RANK system during alveolar bone resorption and remodeling[J ] .J Third Mil Med Univ, 2013, 35(4): 288-292.

KOLAR P, GABER T, PERKA C, et al . Human early fracture hematoma is characterized by inflammation and hypoxia[J ] . Clin Orthop Relat Res, 2011, 469(11): 3118-3126. DOI:10.1007/s11999-011-1865-3.

NOMURA T, AOYAMA M, WAGURI-NAGAYA Y, et al . Tumor necrosis factor stimulates osteoclastogenesis from human bone marrow cells under hypoxic conditions[J ] . Exp Cell Res, 2014, 321(2):167-177. DOI:10.1016/j.yexcr.2013.11.020.

SCHÖNENBERGER M J, KOVACS W J. Hypoxia signaling pathways: modulators of oxygen-related organelles[J]. Front Cell Dev Biol, 2015, 3: 42.DOI:10.3389/fcell.2015.00042.

WENGER R H. Cellular adaptation to hypoxia: O 2 -sensing protein hydroxylases, hypoxia-inducible transcription factors, and O 2 -regulated gene expression[J ] . The FASEB J, 2002, 16(10): 1151-1162. DOI:10.1096/fj.01-0944rev..

王东江, 段莉, 谭包生.低氧条件下HIF调控破骨细胞分化的分子机制研究进展[J].口腔颌面修复学杂志, 2016, 17(5):313-315. DOI:10.3969/j.issn.1009-3761.2016.05.016.

WANG D J, DUAN L, TAN B S. Molecular mechanisms of regulation of osteoclast differentiation by HIF under hypoxic condition[J].Chin J Prosthodont, 2016, 17(5):313-315. DOI:10.3969/j.issn.1009-3761.2016.05.016.

WANG Y, WAN C, GILBERT S R, et al . Oxygen sensing and osteogenesis[J ] . Ann N Y Acad Sci, 2007, 1117: 1-11. DOI:10.1196/annals.1402.049.

MISRA R M, BAJAJ M S, KALE V P. Vasculogenic mimicry of HT1080 tumour cells in vivo: critical role of HIF-1α-neuropilin-1 axis[J]. PLoS ONE, 2012, 7(11): e50153.DOI:10.1371/journal.pone.0050153.

HAYASHI M, NAKASHIMA T, TANIGUCHI M, et al . Osteoprotection by semaphorin 3A[J ] . Nature, 2012, 485(7396): 69-74.DOI:10.1038/nature11000.

MORISHIMA S, MORITA I, TOKUSHIMA T, et al . Expression and role of mannose receptor/terminal high-mannose type oligosaccharide on osteoclast precursors during osteoclast formation[J ] . J Endocrinol, 2003, 176(2): 285-292. DOI:10.1677/joe.0.1760285.

QUARLES L D, YOHAY D A, LEVER L W, et al . Distinct proliferative and differentiated stages of murine MC3T3-E1 cells in culture: an in vitro model of osteoblast development[J ] . J Bone Miner Res, 1992, 7(6): 683-692. DOI:10.1002/jbmr.5650070613.

KARTSOGIANNIS V, NG K W. Cell lines and primary cell cultures in the study of bone cell biology[J]. Mol Cell Endocrinol, 2004, 228(1/2): 79-102. DOI:10.1016/j.mce.2003.06.002.

WANG Y, WAN C, DENG L, et al . The hypoxia-inducible factor alpha pathway couples angiogenesis to osteogenesis during skeletal development[J ] . J Clin Invest, 2007, 117(6): 1616-1626. DOI:10.1172/JCI31581.

MORTEN K J, BADDER L, KNOWLES H J. Differential regulation of HIF-mediated pathways increases mitochondrial metabolism and ATP production in hypoxic osteoclasts[J]. J Pathol, 2013, 229(5): 755-764. DOI:10.1002/path.4159.

KNOWLES H J, ATHANASOU N A. Acute hypoxia and osteoclast activity: a balance between enhanced resorption and increased apoptosis[J]. J Pathol, 2009, 218(2): 256-264. DOI:10.1002/path.2534.

Views

下载量

CSCD

Alert me when the article has been cited

提交

Tools

Publicity Resources

Mutual regulation of HIF-1α and circ-UBE2G1 under hypoxic microenvironment promotes thyroid cancer metastasis

RANKL通过ROS介导HIF-1α下调促进自噬和破骨细胞分化

Hypoxia drives spermatogenic impairment by triggering lysosome-dependent death of Leydig cells via M1 macrophage polarization

Celecoxib improves right heart function in mice after acute high-altitude hypoxia exposure by increasing 12,13-diHOME level

Hypoxia characteristics and prognostic significance of glioblastoma MES-like subpopulation through multi-transcriptomics sequencing

Related Author

DENG Xiaoling

ZHANG Nengying

LONG Kaiguo

ZENG Feng

LI Songtao

SUN Jing

CHU Tongwei

MENG Dan

Related Institution

Department of Thyroid Breast Surgery， the Second Affiliated Hospital of Zunyi Medical University

Institute of Immunology, College of Basic Medical Science，Army Medical University （Third Military Medical University）

Reproductive Medical Center， First Affiliated Hospital, College of Basic Medical Science，Army Medical University （Third Military Medical University）

Department of Pathophysiology， Faculty of High Altitude Military Medicine, Second Affiliated Hospital， Army Medical University （Third Military Medical University）

⁰