Human type H vessels are a sensitive biomarker of bone mass

Liang Wang, Fei Zhou, Peng Zhang, Hongzhen Wang, Zhipeng Qu, Peng Jia, Zhe Yao, Guangsi Shen, Guangfei Li, Guoyang Zhao, Jian Li, Yongtao Mao, Zonggang Xie, Wei Xu, Youjia Xu, Ying Xu, Liang Wang, Fei Zhou, Peng Zhang, Hongzhen Wang, Zhipeng Qu, Peng Jia, Zhe Yao, Guangsi Shen, Guangfei Li, Guoyang Zhao, Jian Li, Yongtao Mao, Zonggang Xie, Wei Xu, Youjia Xu, Ying Xu

Abstract

Vascularization is fundamental for bone formation and bone tissue homeostasis. However, in human subjects, a direct molecular relationship has not been identified between angiogenesis and agents that promote bone disease or factors related to age. Osteopenia is a condition in which bone mineral density is lower than normal, and it represents a sign of normal aging. Here we tested whether the type H vessel, which was recently identified as strongly positive for CD31 and Endomucin (CD31hiEmcnhi) in mice, is an important indicator of aging and osteopenia in human subjects. We found that age-dependent losses of type H vessels in human bone sections conform to the observations in aged mice. The abundance of human type H vessels and osteoprogenitors may be relevant to changes in the skeletal microarchitecture and advanced osteopenia. Furthermore, ovariectomized mice, a widely used model for postmenopausal osteoporosis, exhibited significantly reduced type H vessels accompanied by reduced osteoprogenitors, which is consistent with impaired bone microarchitecture and osteoporosis, suggesting that this feature is an indicator of bone mass independent of aging. More importantly, administration of desferrioxamine led to significantly increased bone mass via enhanced angiogenesis and increased type H vessels in ovariectomized mice. Altogether, these data represent a novel finding that type H vessels are regulated in aged and osteopenia subjects. The abundance of human type H vessels is an early marker of bone loss and represents a potential target for improving bone quality via the induction of type H vessels.

Trial registration: ClinicalTrials.gov NCT02750046.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Age series of mouse tibia and femur immunostaining. (a–c) Tibia immunostaining for CD31 (green), Endomucin (red), Osterix (gray), and DAPI (blue) for juvenile (3 weeks, a), adult (11 weeks, b), and aged (52 weeks, c) mice. (d–f), Femur (proximal region) immunostaining for CD31 (green), Endomucin (red), Osterix (gray), and DAPI (blue) for juvenile (5 weeks, d), adult (16 weeks, e), and aged (47 weeks, f) mice. Line in each panel indicates the boundary of metaphysis and growth plate. Scale bar: 100 μm
Figure 2
Figure 2
Decreased type H vessels in aged patients. (a), Schematic illustration of bone sample collection from patients during surgery. (b), Typical human bone sample collected from PFN surgery. (c,d), Pre- and post-surgical X-ray images of a typical patient. (e–g), Bone immunostaining for CD31 (green), Endomucin (red), Osterix (gray), and DAPI (blue) for young (e), adult (f) and aged (g) patients. (h) Quantification of type H vessel percentage (HV/TV%) in young (N=7), adult (N=8), and aged patients (N=24). Graphs represent mean±S.D.; *P<0.05; ***P<0.0001; Scale bar: 100 μm
Figure 3
Figure 3
Synergistic decrease of type H vessel and bone mineral density in human bone. (a–c) Micro-CT 3D reconstructed image of bone samples from normal (a), osteopenia (b), and osteoporosis (c) patients. (d–i) Quantification of the trabecular index of bone samples from normal (N=6), osteopenia (N=10), and osteoporosis (N=8) patients. (j–l) Bone immunostaining for CD31 (green), Endomucin (red), Osterix (gray), and DAPI (blue) for normal (j), osteopenia (k), and osteoporosis (l) patients. Graphs represent mean±S.D.; *P<0.05; **P<0.01; Scale bar: 100 μm
Figure 4
Figure 4
Change of type H vessels in osteoporotic mice. (a–f) Tibia immunostaining for CD31 (green), Endomucin (red), Osterix (gray), and DAPI (blue) of Sham (a–b), OVX (c–d), and OVX+DFO (e–f) group mice. Lines in b, d, and f indicate the boundary of the metaphysis and growth plate. (g–i) Micro-CT 3D reconstructed images of cancellous bone from the Sham (g), OVX (h), and OVX+DFO (i) mice. (j–n) Quantification of the BMD and trabecular index of bone from the Sham, OVX, and OVX+DFO mice. Graphs represent mean±S.D.; *P<0.05; **P<0.01; Scale bar: 200 μm (e), 100 μm (f)

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