Communication between osteoblasts and endothelial cells (ECs) is essential for bone turnover, but the molecular mechanisms of such communication are not well defined. Here we identify Cxcl9 as an angiostatic factor secreted by osteoblasts in the bone marrow microenvironment. We show that Cxcl9 produced by osteoblasts interacts with vascular endothelial growth factor and prevents its binding to ECs and osteoblasts, thus abrogating angiogenesis and osteogenesis both in mouse bone and in vitro. The mechanistic target of rapamycin complex 1 activates Cxcl9 expression by transcriptional upregulation of STAT1 and increases binding of STAT1 to the Cxcl9 promoter in osteoblasts. These findings reveal the essential role of osteoblast-produced Cxcl9 in angiogenesis and osteogenesis in bone, and Cxcl9 can be targeted to elevate bone angiogenesis and prevent bone loss-related diseases.

Natural bone apatite crystals, which mediate the development and regulate the load-bearing function of bone, have recently been associated with strongly bound citrate molecules. However, such understanding has not been translated into bone biomaterial design and osteoblast cell culture. In this work, we have developed a new class of biodegradable, mechanically strong, and biocompatible citrate-based polymer blends (CBPBs), which offer enhanced hydroxyapatite binding to produce more biomimetic composites (CBPBHAs) for orthopedic applications. CBPBHAs consist of the newly developed osteoconductive citrate-presenting biodegradable polymers, crosslinked urethane-doped polyester (CUPE) and poly (octanediol citrate) (POC), which can be composited with up to 65 wt.-% hydroxyapatite (HA). CBPBHA networks produced materials with a compressive strength of 116.23 ± 5.37 MPa comparable to human cortical bone (100 – 230 MPa), and increased C2C12 osterix (OSX) gene and alkaline phosphatase (ALP) gene expression in vitro. The promising results above prompted an investigation on the role of citrate supplementation in culture medium for osteoblast culture, which showed that exogenous citrate supplemented into media accelerated the in vitro phenotype progression of MG-63 osteoblasts. After 6-weeks of implantation in a rabbit lateral femoral condyle defect model, CBPBHA composites elicited minimal fibrous tissue encapsulation and were well integrated with the surrounding bone tissues. The development of citrate-presenting CBPBHA biomaterials and preliminary studies revealing the effects of free exogenous citrate on osteoblast culture shows the potential of citrate biomaterials to bridge the gap in orthopedic biomaterial design and osteoblast cell culture in that the role of citrate molecules has previously been overlooked.

Background Intervertebral disc degeneration is a major cause of chronic low back pain, and excessive loading contributes to intervertebral disc degeneration. However, the lack of an effective bipedal in vivo animal model limits research about this condition. Questions/purposes To evaluate the utility of a new type of bipedal standing mouse model for intervertebral disc degeneration, we asked: (1) Are there spinal degeneration changes in bipedal mice as determined by lumbar disc height, histologic features, and immunohistochemistry measures compared with control mice? (2) Are the bipedal mice comparable to aged mice for simulating the spinal degeneration caused by increased stress? Methods Thirty-two 8-week-old male C57BL/6 mice were divided into experimental and control groups. Based on their hydrophobia, mice in the experimental group were placed in a limited water-containing space (5 mm deep) and were thereby induced to actively take a bipedal standing posture. This was conducted twice a day for a total of 6 hours a day, 7 days a week. Control mice were similarly placed in a limited but water-free space. Video surveillance was used to calculate the percentage of time spent in the bipedal stance for the two groups of mice. Compared with the control group, the percentage of time standing on both feet in the experimental group was higher (48% 6 5%, 95% confidence interval [CI], 42%-54% versus 95% 6 1%, 95% CI, 92%-97%; p < 0.001). Eight mice from both groups were then randomly euthanized at either 6 or 10 weeks and lumbar spine specimens (L3-L6) were collected. The lumbar disc height index (DHI%) of the two groups was compared using micro-CT measurements, and the extent of disc degeneration was assessed based on histologic staining (cartilage endplate height, disc degeneration score) and by immunohistochemistry (Col2a1,CollagenX, matrix metalloprotease-13 [MMP-13], osteocalcin [OCN]). In addition, the histopathologic features of spinal degeneration were compared with 12-This work was funded by National Natural Sciences Foundation of China (81672228 [ZZ], 81874013[ZZ], 31801012[MH]) and Natural Science Foundation of Guangdong Province (2016A030313608 [MH]). Clinical Orthopaedics and Related Research® neither advocates nor endorses the use of any treatment, drug, or device. Readers are encouraged to always seek additional information, including FDAapproval status, of any drug or device prior to clinical use. Each author certifies that his or her institution approved the animal protocol for this investigation and that all investigations were conducted in conformity with ethical principles of research.