ApoE gene deficiency enhances the reduction of bone formation induced by a high-fat diet through the stimulation of p53-mediated apoptosis in osteoblastic cells.
Osteoblast apoptosis increased in the tibias of apoE(-/-) mice fed with a high-fat diet, decreasing bone formation. The expression of p53 mRNA in marrow adherent cells increased. LDL or oxidized LDL increased apoptosis in the calvarial cells of apoE(-/-) mice. The increase in p53-mediated apoptosis is apparently related to a high-fat diet-induced osteopenia in apoE(-/-) mice.


BACKGROUND
The effects of high-fat loading and the apolipoprotein E (apoE) gene on bones have not been elucidated. We hypothesized that apoE gene deficiency (apoE(-/-)) modulates the effects of high-fat loading on bones.


METHODS
We assessed this hypothesis using wildtype (WT) and apoE(-/-) mice fed a standard (WTS and ApoES groups) or a high-fat diet (WTHf and ApoEHf groups). The concentration of serum lipid levels and bone chemical markers were measured. Histomorphometry of the femurs was performed using microCT and a microscope. Bone marrow adherent cells from the femurs were used for colony-forming unit (CFU)-fibroblastic (CFU-f) assay and mRNA expressions analysis. The apoptotic cells in the tibias were counted. TUNEL fluorescein assay and Western analysis were performed in cultures of calvarial cells by the addition of low-density lipoprotein (LDL) or oxidized LDL.


RESULTS
In the ApoEHf group, the values of cortical bone volume and trabecular and endocortical bone formation of the femurs decreased, and urinary deoxypyridinoline increased. Subsequent analysis revealed that the number of apoptotic cells in the tibias of the ApoES group increased, and more so in the ApoEHf group. The ratio of alkaline phosphatase-positive CFU-f to total CFU-f was decreased in the ApoEHf group. p53 mRNA expression in adherent cells of the apoE(-/-) mice increased and had a significantly strong positive correlation with serum LDL. TUNEL fluorescein assay of osteoblastic cells revealed an increase of apoptotic cells in the apoE(-/-) mice. The number of apoptotic cells in the apoE(-/-) mice increased with the addition of 100 microg/ml LDL or oxidized LDL. The p53 protein expression in apoE(-/-) cells exposed to 100 microg/ml LDL or oxidized LDL increased.


CONCLUSIONS
We concluded that apoE gene deficiency enhances the reduction of bone formation induced by a high-fat diet through the stimulation of p53-mediated apoptosis in osteoblastic cells.
Hirasawa
Hideyuki
H
Department of Orthopaedic Surgery, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
Tanaka
Shinya
S
Sakai
Akinori
A
Tsutsui
Masato
M
Shimokawa
Hiroaki
H
Miyata
Hironori
H
Moriwaki
Sawako
S
Niida
Shumpei
S
Ito
Masako
M
Nakamura
Toshitaka
T
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001057:Apolipoproteins E / Q000172:deficiency* / Q000235:genetics*; D017209:Apoptosis / Q000187:drug effects*; D015415:Biomarkers / Q000378:metabolism; D001835:Body Weight / Q000187:drug effects; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D003114:Colony-Forming Units Assay; D004041:Dietary Fats / Q000494:pharmacology*; D005269:Femur / Q000187:drug effects / Q000254:growth & development; D005333:Fetus / Q000166:cytology / Q000187:drug effects; D020287:In Situ Nick-End Labeling; D008055:Lipids / Q000097:blood; D008077:Lipoproteins, LDL / Q000494:pharmacology; D008297:Male; D051379:Mice; D009929:Organ Size / Q000187:drug effects; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012886:Skull / Q000166:cytology / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
D001057:Apolipoproteins E; D015415:Biomarkers; D024042:Collagen Type I; D004041:Dietary Fats; D008055:Lipids; D008077:Lipoproteins, LDL; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; C099252:oxidized low density lipoprotein; D015675:Osteocalcin; D000469:Alkaline Phosphatase