Effect of local injection of nerve growth factor and basic fibroblast growth factor on the healing of tibial fracture in rats (2020)

Objective
To investigate the effects and mechanism of local injection of nerve growth factor (NGF) and basic fibroblast growth factor (BFGF) on tibial fracture healing in rats.


Methods
Forty-eight clean mature male SD rats were randomly divided into four groups: NGF group, BFGF group, NGF+ BFGF group and control group. Each group was randomly divided into two subgroups (2 weeks after operation and 4 weeks after operation) according to different observation time points, six rats in each subgroup. The left tibial fracture model was established in 48 rats, and intramedullary fixation with 1 mm Kirschner wire was performed. On the third day after surgery, local percutaneous injection was given around the fracture end. The NGF group was injected with 0.8 μg+ 0.3 mL normal saline, the BFGF group received local injection of BFGF 1.2 μg + 0.3 mL normal saline, the NGF+ BFGF group received local injection of NGF 0.8 μg+ BFGF 1.2 μg+ 0.3 mL normal saline, and the control group received 0.3 mL normal saline. Each group was injected once a day for 7 consecutive days.X-ray films were taken at 2 weeks and 4 weeks after the operation, and blood samples of femoral artery of rats in each group were taken, then the rats were killed by cervical vertebra disconnection.The content of alkaline phosphatase (ALP) was detected by enzyme-linked immunosorbent assay (ELISA). The callus tissue was made into pathological sections, and HE staining was performed to observe the histological characteristics. Osteoblastic index (IOB), trabecular width (TW) and trabecular area percentage (TV) of bone tissue were calculated by multifunctional true-color cell image analysis and management system. The contents of bone morphogenetic protein-2 (BMP-2), vascular endothelial growth factor (VEGF) and insulin-like growth factor-1 (IGF-1) in serum of rats were detected by protein chip technique.


Results
(1) X-ray film: the fracture healing of NGF, BFGF and NGF+ BFGF group was faster than that of the control group, and that of NGF+ BFGF group was the fastest. (2) ELISA: the content of ALP at 2 weeks after surgery in the callus of the control group, NGF group, BFGF group and NGF+ BFGF group were(110.02±1.92), (140.87±2.22), (136.12±1.23) and (187.44±0.90)ng/mL, respectively. The ALP contents in callus were (91.23±1.47), (106.62±1.64), (104.83±1.05) and (130.59±1.18)ng/mL, respectively, at 4 weeks after surgery. The content of ALP in the callus of the NGF+ BFGF group was the highest, and the difference between the groups was statistically significant (all P values 0.05), and all other indicators had statistical significance between the two groups (all P values 0.05). The VEGF content at 2 weeks after surgery, and the BMP-2 and IGF-1 content at 2 weeks and 4 weeks after surgery of NGF group, BFGF group and NGF+ BFGF group were all higher than those of the control group, and the content of NGF+ BFGF group was the highest. Pairwise comparison between groups showed statistically significant differences (F=198.285, 368.060, 2 006.017, 33.332, 292.643, all P values<0.05).


Conclusions
Both NGF and BFGF can promote the healing of tibial fracture in rats, and the combination of them has synergistic effects. The mechanism may be related to the better promotion of VEGF, BMP-2 and IGF-1 expression in the process of fracture healing.


Key words: 
Fracture healing; Bony callus; Rats; Tibial fractures; Nerve growth factor; Basic fibroblast grouth factor; Local injection; Protein chip