Regulation of expression level of fms-like tyrosine kinase-4 is related to osteoclast differentiation.
BACKGROUND
The aim of this study is to determine whether regulation of the expression level of fms-like tyrosine kinase-4 (Flt-4) is related to osteoclast differentiation.


METHODS
Osteoclast formation and differentiation of mouse bone marrow cells and RAW264.7 cells were performed. To induce osteoclast differentiation, RANKL (50 ng/ml) with or without vascular endothelial growth factor-C (VEGF-C) and vascular endothelial growth factor-D (VEGF-D) was added to mouse bone marrow cells and RAW264.7 cells. Then cells were examined under a microscope. TRAP-positive cells with 3 nuclei or more were considered as osteoclasts and counted. The Flt-4 gene was knocked down by transfection of siRNAs against Flt-4. Immunoblot analyses were performed.


RESULTS
The osteoclast formation assay indicated that VEGF-C resulted in 500 or 450 vs. 100 (p < 0.05) of osteoclasts in mouse bone marrow cells and RAW264.7 cells, respectively. Vascular endothelial growth factor-D resulted in about 600 or 630 vs. 100 (p < 0.05) of osteoclasts for both mouse bone marrow cells and RAW264.7 cells. The knock-down of Flt-4 expression abolished the induction by VEGF-C or VEGF-D, resulting in induction similar to that of the negative control PBS.


CONCLUSIONS
Both VEGF-C and VEGF-D can induce osteoclast differentiation in the presence of the receptor activator of nuclear factor κB ligand. Down-regulation of expression level of Flt-4 protein abolishes osteoclast differentiation induced by VEGF-C or VEGF-D.
Department of Orthopaedics, the Second Hospital Affiliated to the University of South China, Hengyang, China.
Yi
Y
Zhu
Department of Laboratory, the Second Hospital Affiliated to the University of South China, Hengyang, China.
Yuan
Y
Wu
Institute of Pathogen Biology, Medical College, the University of South China, Hengyang, China.
Yu
Y
Liang
Department of Orthopaedics, the Second Hospital Affiliated to the University of South China, Hengyang, China.
Wenfu
W
Tan
Department of Laboratory, the Second Hospital Affiliated to the University of South China, Hengyang, China.
Zhuoran
Z
Liu
Institute of Pathogen Biology, Medical College, the University of South China, Hengyang, China.
Jianhua
J
Xiao
fms-like tyrosine kinase-4; osteoclast differentiation; vascular endothelial growth factor-C; vascular endothelial growth factor-D