Experimental Study On Treatment Of Early Steroid-induced Avascular Necrosis Of Femoral Head In Rabbit With Bmscs And RhBMP-2/rhVEGF Transplantation

Objective:The effection of recombinant human bone morphogenetic protein-2(rhBMP-2) and recombinant human vascular endothelial growth factor(rhVEGF)were investigated in avascular crosis of femoral head of rabbits by being associated with corecompression.The new method of treating avascular necrosis of femoral head in early stage was explored with rhVEGF and rhBMP-2 in clinics.Methods:1.cell culture:Bone marrow were obtained from rabbits posterior iliac crest and inoculated in 6-well dishes.Every well contained 5 milliliter DMEM supplemented with 20 percent of fetal bovine serum and 10^-8 molardexamethasone.These 6-well dishes were placed into a humidified incubator at 37 degrees Celsius.The medium were changed on the fifth day.When the cells were confluent on the bottom of well,these cells must be subcultured.The third generation of cultured cells were inoculated in gelatin sponge.Every piece of gelatin sponge to be prepared for transplantation contained 100,000 cells.2. Animal models were made and divided into six groups randomly as follows: group A:no treatment;group B:only decompression;group C:decompression with BMSCs transplantation; group D: decompression with BMSCs and rhBMP-2 transplantation; group E: decompression with BMSCs and rhVEGF transplantation; F: decompression with BMSCs and rhBMP-2/rhVEGF transplantation.Two femoral heads of every rabbits were studied in the experiment.The general information of animals were recorded,after 4 and 8 weeks of operation,3 rabbits of each group were chosen randomly to do radiograph and microscope.Angiogenesis of the repairing tissue in the femoral head and empty lacunae in the edge of the bone defect were compared with at 4 weeks and 8 weeks after operation.Finally,the statistical analysis was made.Result:1.When the medium were changed on the fifth day,some fibroblast-like unites were found growthing on the bottom of the well.The cells anchored in 2-4 hours after digesting,and were confluent on bottom of the well.The log phase was from 2 days to 7 days.The population doubling time was at 48 hours.2.X-ray examination: Group A,at 4 weeks,the density was higher,the disorder of the bone trabeculae could be observed.At 8 weeks,cystic degeneration,the density was higher than before.In other groups,the manifestation of the area outside of defect area was consistent with guoup A. In group B and group C, At 2 weeks,the density of defect area is low and the density of the brim of defect area was heightened.the bone density of defect area in group C was higher than those in group B.In group C,the trabeculae could be observed in the defect area. In group D,group E and group F the density of the brim of defect area also was heightened, the bone density of defect area in group F was highest.3.light microscope examination:In Group A, After two weeks chondrocyte and bone marrow were necrotic,and trabeculae kept intact,empty lacunae increased.New-formed bone either appeared in the intertrabecular space or covered the dead trabeculae.At 8 weeks,in the meantimes,a part of dead trabeculae became thin,irregular or broken.In group B,four weeks later after operation,Some inflammatory cells appeared in the center of the drilled hole. With the time going,only a few amount of osteoblasts were in the drilled hole. at 8 weeks,the drilled hole were full of marrow.In group C ,group D and group E,Four weeks later after operation, there were a large amount of osteoblasts appearing on the gelatin sponge in the bone defects,and some new-formed bone appearing around the drilled hole, With the time going,at 8 weeks,the drilled hole were full of new-formed trabeculae appearing ingroup D, the angiogenesis appearing in group E.In group F,Four weeks later after operation,there were osteoblasts and the angiogenesis appearing in the drilled hole. At 8 weeks,the drilled hole were full of new-formed trabeculae. And the trabeculae were mature and bone marrow formed in the intertrabecular space. In group F,the positive ratio of empty lacunae showing significant differences compared with other groups(P<0.05).The angiogenesis showing significant differences compared with other groups(P<0.05) except group E.Conclusion: 1. Avaseular neerosis of the femoral head model was sueeessfully established in rabbits.2.The femoral head of BMSCs and rhBMP-2/rhVEGF group can augment angiogenesis of bone tissue and improve microcirculation of necrotic femoral head.