Effect of multiple-point injection of the mixture of pBLAST49-mVEGF plasmid and liposomes into the subperiosteum on both sides of the fractured bone of rats on fracture healing: Expression of vascular endothelial growth factor at the local part of the fractured bone at different time points through a randomized blank control trial

Gang ZHONG; Fuxing PEI; Yubo FAN; Shengfu LI

ABSTRACT

BACKGROUND:

Research has shown that the vascular endothelial growth factor (VEGF) of the ends of the fractured bone is heavily expressed 72 hours to 3 weeks after the fracture and it is supposed that it has a promoting effect on fracture healing. Inducing angiogenesis through VEGF gene transfection has gradually attracted the attention of the researches.

OBJECTIVE:

To find an efficient way of exogenous VEGF gene in vivo transfection through injecting the mixture of positive ion liposome transfection agent and plasmid and to study the promoting effect of extra VEGF gene expression on bone fracture healing.

DESIGN:

A randomly grouping, blank control trial.

SETTING:

Animal Laboratory of Huaxi Medical Center of Sichuan University MATERIALS Totally 40 adult male SD rats, weighing 230 to 250 g,were involved. All the animals were randomly divided into the experiment group and the control group with 20 rats in each group.

METHODS:

The experiment was conducted at the Animal Laboratory of Huaxi Medical Center of Sichuan University from April to December 2003.Altogether 40 rats were involved to establish fractured models of right shaft of femur. Cut the bone in the middle of bone stem, retroplanted a Kirsh' nail with 1 mm diameter through intercondylar part and the fractured bone was fixed. In the experimental group, a mixture of 100 μL of liposome transfection agent and 100 μg of pBLAST49-mVEGF plasmid was injected in multiple points into the subperiosteum of the both sides of the ends of the fractured bone. The same volume of normal saline was injected into the rats in the control group. Then, 2 rats in each group were put to death 3,7,14,28,42,56,70 days after the operation and femoral bone specimen was collected.MAIN OUTCOME

MEASURES:

Observation of right femoral fractured staining results of VEGF, with the apperance of brown granules as positive.

RESULTS:

Two rats were selected at 7 time points separately, and altogether 28 rats entered the stage of result analysis. The other 12 rats were fracture at different time points For the experimental group, 28 days after the operation, cartilage callus appeared and replaced fibrocallus gradually,and the fracture line disappeared. Fifty-six days after the operation, the bone healed completely. For the control group, 28 days after the operation , fibrocallus was observed, and the fracture line was still clear. 56days after the operation, much callus appeared, and the fracture line beof fractured bone was stained with hemotoxylin eosin (HE). In the experiment group, 56 days after the operation, the bone healed completely and trabecular like bones were rebuilt. The bone marrow cavity of the fractured region was open again. In the control group, Fifty-six days after the operation, no mature bone was formed, and the bone marrow cavity was not different time points The expression in the two group reached to the peak on day 14 and began to decrease on day 28. The expression of VEGF in the experimental group was obviously higher than that in the control group.

CONCLUSION:

Injection of the mixture of positive ion liposome transfection agent into the subperiosteum of rats is an effective approach for in vivo transfection and pBLAST49-mVEGF gene transfection can effectively facilitate the bone fracture healing of rats.