Hypoxia-inducible factor 1α enhances RANKL-induced osteoclast differentiation by upregulating the MAPK pathway

Dong Wang; Lin Liu; Zechao Qu; Bo Zhang; Xiangcheng Gao; Wangli Huang; Mingzhe Feng; Yining Gong; Lingbo Kong; Yanjun Wang; Liang Yan

doi:10.21037/atm-22-4603

Hypoxia-inducible factor 1α enhances RANKL-induced osteoclast differentiation by upregulating the MAPK pathway

Ann Transl Med. 2022 Nov;10(22):1227. doi: 10.21037/atm-22-4603.

Authors

Dong Wang^#^{1

2}, Lin Liu^#¹, Zechao Qu^{1

2}, Bo Zhang^{1

2}, Xiangcheng Gao^{1

2}, Wangli Huang^{1

2}, Mingzhe Feng^{1

2}, Yining Gong¹, Lingbo Kong¹, Yanjun Wang¹, Liang Yan¹

Affiliations

¹ Department of Orthopedic Surgery, Xi'an Honghui Hospital, Xi'an Jiaotong University, School of Medicine, Xi'an, China.
² School of Medicine, Yanan University, Yanan, China.

^# Contributed equally.

Abstract

Background: Hypoxia (low-oxygen tension) and excessive osteoclast activation are common conditions in many bone loss diseases, such as osteoporosis, rheumatoid arthritis (RA), and pathologic fractures. Hypoxia-inducible factor 1 alpha (HIF1α) regulates cellular responses to hypoxic conditions. However, it is not yet known how HIF1α directly affects osteoclast differentiation and activation. This study sought to. explore the effects of HIF1α on osteoclast differentiation and it's molecular mechanisms.

Methods: L-mimosine, a prolyl hydroxylase (PHDs) domain inhibitor, was used to stabilize HIF1α in normoxia. In the presence of receptor activator of nuclear factor-kB (NF-kB) ligand (RANKL), RAW264.7 cells were cultured and stimulated by treatment with L-mimosine at several doses to maintain various levels of intracellular HIF1α. The multi-nucleated cells were assessed by a tartrate-resistant acid phosphatase (TRAP) and F-actin ring staining assays. The osteoclast-specific genes, such as Cathepsin K, β3-Integrin, TRAP, c-Fos, nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), and matrix metallo-proteinase 9 (MMP9), were analyzed by real time-polymerase chain reaction (RT-PCR). The expression of relevant proteins was analyzed by Western blot.

Results: L-mimosine increased the content of intracellular HIF1α in a dose-dependent manner, which in turn promoted RANKL-induced osteoclast formation and relevant protein expression by upregulating the mitogen-activated protein kinase (MAPK) pathways.

Conclusions: Our findings suggest that HIF1α directly increases the osteoclast differentiation of RANKL-mediated RAW264.7 cells in vitro by upregulating the MAPK pathways.

Keywords: Hypoxia-inducible factor 1 alpha (HIF1α); L-mimosine; RAW264.7 cells; mitogen-activated protein kinase pathway (MAPK pathway); osteoclast.