Effects of oxygen levels on angiogenesis. *P <0.05; ** P <0.01; a, P <0.01 compared to normoxia or hyperoxia; b, P <0.05 compared to hypoxia; c, P <0.01 compared to normoxia, hypoxia, or hyperoxia.
Sat., 10/10/09 Basic Sci./Injury Prevent./Spine, Paper #67, 10:33 am OTA-2009
Environmental Oxygen Levels Affect Angiogenesis during Fracture Healing
Chuanyong Lu, MD (n); Zhiqing Xing, MD (n); Diane Hu, MD (n);
Theodore Miclau, III, MD (n); Ralph Marcucio, PhD (n);
Department of Orthopaedic Surgery, University of California at San Francisco,
San Francisco, California, USA
Purpose: Local hypoxia is thought to be a strong stimulator of angiogenesis, the major process of blood vessel repair after tissue injury. In this study, we examined the effects of environmental oxygen concentration on fracture healing and angiogenesis.
Methods: Male 129J/B6 mice (3 months old) were used in this study. A closed fracture was created in the middiaphysis of the right tibia by 3-point bending. All fractures were unstabilized. Animals were housed in chambers with controlled oxygen levels at 13% (hypoxic), 21% (normoxic), or 50% (hyperoxic) immediately after injury until sacrifice. Fracture healing was analyzed by histomorphometry at 5, 10, and 21 days after injury (n = 5/time/group). To analyze angiogenesis, animals (n = 4-6/group) were sacrificed at 3 days after injury. Uninjured tibiae (n = 5) were used as controls. PECAM (platelet-cell adhesion molecule) immunohistochemistry was used to visualize blood vessels on frozen sections. The length density (the length of blood vessels per unit volume of the reference space) and surface density (the area of the outer surface of blood vessels per unit volume of the reference space) of blood vessels within the fracture limbs were estimated using stereology.
Results: Hypoxia and hyperoxia alter fracture healing. At 10 days after fracture, both hypoxia and hyperoxia significantly decreased the amount of new bone in fracture calluses compared to normoxia. Hyperoxic fractures also exhibited significantly smaller calluses and less cartilage compared to normoxic fractures. At day 21, all fractures healed by bony bridging and with minimal amount of cartilage. Hypoxia and hyperoxia affect angiogenesis. All 3 fracture groups exhibited significantly higher length density and surface density of vasculature compared to unfractured limbs, suggesting that bone injury induced angiogenesis. Angiogenesis appeared to be reduced by hypoxia and increased by hyperoxia (see figure below).
Effects of oxygen levels on angiogenesis. *P <0.05; ** P <0.01; a, P <0.01 compared to normoxia or hyperoxia; b, P <0.05 compared to hypoxia; c, P <0.01 compared to normoxia, hypoxia, or hyperoxia.
Conclusion and Significance: Data from the current study demonstrate that environmental oxygen levels affect angiogenesis, which may ultimately lead to altered fracture healing. In contrast to local hypoxia, environmental hypoxia inhibits angiogenesis, which could be due to reduced oxygen gradient at the injury site or impaired collagen synthesis in a hypoxic environment.
Acknowledgment: This work is supported by OTA (a research grant to C.L.), NIH (grant R01-AR053645 to T.M.), and Zimmer Inc.
• The FDA has not cleared this drug and/or medical device for the use described in this presentation (i.e., the drug or medical device is being discussed for an “off label” use). ◆FDA information not available at time of printing. Δ OTA Grant