RESUMO
Objective:To study the effects of a simulated plateau environment on fracture healing in rats.Methods:A rat model of mid-femoral fracture was established by hacksaw truncation and intramedullary fixation with Kirschner wires in 60 male Wistar rats which were divide into 2 groups ( n=30) by the random number table method. The rats in the control group were raised in the animal experiment center of The 940 Hospital of Joint Logistic Support Force of Chinese PLA at an altitude of 1,400 m, while the rats in the plateau group were placed in an animal experimental cabin in a simulated plateau environment at a simulated altitude of 5,000 m. The body weight was weighed once a week and X-ray films were taken every 2 weeks. Blood samples were collected after 4 weeks for detection of biochemical indicators of bone metabolism. After 8 weeks, the femurs of the surgical side were taken for bone biomechanical detection and the bone mineral density of the healthy side was detected. After 4 and 8 weeks, the femurs of the surgical side were taken for in vitro Micro-CT scanning and angiography detection. After 1, 2, 4 and 8 weeks, the femurs of the surgical side were taken for bone histopathologic detection. Results:During the entire experiment, no rats in the control group died while the mortality rate of the rats in the plateau group was as high as 26.7% (8/30). In the plateau group, some organs were pathologically damaged in the rats, fracture union was delayed, and the callus differentiated and matured slowly with the chondrocytes still dominant at the 8th week. The bone mineral density and the maximum load of the femur in the plateau group were significantly lower than those in the control group ( P< 0.05). Angiography showed that the rats in the plateau group had microvascular proliferation which did not penetrate the fracture end at the 8th week. The bone formation indexes like osteocalcin, procollagen type Ⅰ N-terminal propeptide (PⅠNP), and osteoprotegerin of the rats in the plateau group were significantly lower than those in the control group at the 4th week ( P<0.05). The bone resorption indexes like tartrate resistant acid phosphatase 5b (TRACP-5b) and receptor activator for nuclear factor-κB ligand (RANKL) in the plateau group were significantly higher than those in the control group ( P<0.05). Conclusion:A simulated plateau environment at an altitude of 5,000 m may lead to delayed fracture healing in rats.
RESUMO
It is known that low-frequency pulsed electromagnetic fields (PEMFs) can promote the differentiation and maturation of rat calvarial osteoblasts (ROBs) cultured in vitro. However, the mechanism that how ROBs perceive the physical signals of PEMFs and initiate osteogenic differentiation remains unknown. In this study, we investigated the relationship between the promotion of osteogenic differentiation of ROBs by 0.6 mT 50 Hz PEMFs and the presence of polycystin2 (PC2) located on the primary cilia on the surface of ROBs. First, immunofluorescence staining was used to study whether PC2 is located in the primary cilia of ROBs, and then the changes of PC2 protein expression in ROBs upon treatment with PEMFs for different time were detected by Western blotting. Subsequently, we detected the expression of PC2 protein by Western blotting and the effect of PEMFs on the activity of alkaline phosphatase (ALP), as well as the expression of Runx-2, Bmp-2, Col-1 and Osx proteins and genes related to bone formation after pretreating ROBs with amiloride HCl (AMI), a PC2 blocker. Moreover, we detected the expression of genes related to bone formation after inhibiting the expression of PC2 in ROBs using RNA interference. The results showed that PC2 was localized on the primary cilia of ROBs, and PEMFs treatment increased the expression of PC2 protein. When PC2 was blocked by AMI, PEMFs could no longer increase PC2 protein expression and ALP activity, and the promotion effect of PEMFs on osteogenic related protein and gene expression was also offset. After inhibiting the expression of PC2 using RNA interference, PEMFs can no longer increase the expression of genes related to bone formation. The results showed that PC2, located on the surface of primary cilia of osteoblasts, plays an indispensable role in perceiving and transmitting the physical signals from PEMFs, and the promotion of osteogenic differentiation of ROBs by PEMFs depends on the existence of PC2. This study may help to elucidate the mechanism underlying the promotion of bone formation and osteoporosis treatment in low-frequency PEMFs.