| Objective: The avascular necrosis of the femoral head (ANFH) remains to be one of the greatest challenges to the orthopaedics surgeons. The incidence rate of steroid-induced femoral head necrosis (SIFHN) increases gradually with the widespread use of glucocorticosteroid nowadays, but the pathogenesies of this disease remains unclear dispite extensive studies during the past years. As apoptosis has been realised to be one of the new patterns of cell death, it provided a new method for investigation of SIFHN .Preliminary studies have already revealed that apoptosis may be intimately related to SIFHN. Via intervention of Levodopa on rabbits ANFH model induced by a single low dose lipopolysaccharide (LPS) injection followed by 3 consecutive high dose of methylprednisolone (MPS) injections, the present study was aimed to explore the effects of Levodopa (L-Dope)on cell apoptosis in rabbits ANFH induced by methylprednisolone which is a member of glucocorticosteroid, and to provide a new method for prevention and early treatment on SIFHN.Methods: 44 Chinese albino rabbits weighed 2.5±0.5 Kg were randomly divided into 3 groups. Group A was the model group with 15 rabbits were treated by injection of 10ug/Kg LPS via ear vein, 3 consective MPS sodium succinate injections (20mg/Kg) via right buttock was then carried out 24h later with 24hs'intervals. Group B was the group with Levodopa intervention which contained 15 rabbits, the animals of this group were firstly treated by the same method as to the rabbits in Group A, then oral intake of Levodopa (0.4g/Kg/day) at the same day of the last injection of MPS sodium succinate was carried out. Group C was the control group contained 14 rabbits which were injected with Sodium Chloride in a same way as to the animals of the other two groups. During the experiment, all the rabbits were injected with Penicilinium (800,000 U/rabbit) twice a week for infection prophylaxis. Radiographic and MRI images of both hip joints were taken in 7 rabbits from every group 6 weeks and 8 weeks later, respectively, then the rabbits were sacrificed by aeroembolism. Hip joint samples were taken and immersed in 10% formalin for 24h, then the samples were decalcified, embedded, sliced for HE as well as apoptosis staining and observed microscopically. At the same time, 8 weeks later, blood samples were taken from three group rabbits for IGF-I detection.Results: There were 2 rabbits (1 in group A, 1 in group B, respectively) dead because of anaphylactic shock 48h after injection of LPS. The x-ray of group A showed a little collapse of the femoral head shape, partial disruption of trabecular bone, bone mineral density in non-uniform, with cystis degeneration or cresent signs, joint space narrowing slightly. In bilateral femoral head of 11 rabbits, MRI examination revealed: T1WIshowed small strip and low flake signal; T2WI showed thin strip, sheet high signal or low signal. The incidence of osteonecrosis of the femoral head up to 78.57% (11/14). HE staining of the samples of group A showed that bone trabeculae thinned and sparse with discontinuity of the integrity; the transitional rate from stromal cells to fat cells increased in this group was found.A lot of bone lacuna could be observed, the average empty rate was 33.86%±8.38%, compared with the group C, there was a significant difference (P <0.05). Apoptosis detection by TUNEL method showed a large number of apoptic bone cells could be observed in group A, the apoptic rate is 202.02‰±18.99‰, compared with the group C (45.27‰±12.11‰), there was a significant difference (P <0.01).Serum IGF-1 contents of group B is 14.78±2.37 ng/ml, which is statistically higher than that of group A(10.12±2.49 ng/ml). Radiographically, hip joint spaces and the shape of femoral heads of rabbits in group B were normal, and bone density increased uniformly. HE staining of the samples of group B showed that bone trabeculae aligned regularly with no discontinuity of the integrity. Some empty bone lacuna could be observed, the average empty rate was 25.97%±6.29%, lower than that of group A(33.86%±8.38%). Fibrous repair could be observed in necrotic areas with increasement of osteoblasts and capillary regeneration. And obvious osteogenicity also could be observed. Apoptosis detection by TUNEL method showed that sporatic brown particles in nucleus of osteocytes which is indicative of positive staining of apoptic cells could be observed in group B, the apoptic rate is 120.67‰±13.13‰which is statistically lower than that of group A (202.02‰±18.99‰).Conclusions: Single dose of LPS followed by 3 consective methylprednisolone injections successfully established rabbit ANFH model with a high success rate and low animal death rate. Existence of ponderosus apoptic osteocytes indicated that apoptosis of osteocytes contributed to the pathogenetic processes of SIFHN. Levodopa effectively intervened the progress of SIFHN, which may resulted from the release of growth hormone from pituitary adenoma stimulated by Levodopa, and then the growth hormone stimulated the differentiation of stromal cells osteogenetically. At the same time, the effect of Levodopa may indirectly increased the content of IGF-1 which is helpful to the osteoblastic proliferation and thus prevented the collapse of femoral head by augmention of osteogenetic capability. IGF-1 could prevent the incidence of SIFHN via inhibit apoptosis of osteocytes and osteoblasts. |
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