| Objective: To explore the intervention mechanism of Sheng Gu Zai Zao pill on the steroid-induced femoral head osteonecrosis of Rabbit by investigating its effect on vascular endothelial growth factor(VEGF)and its receptor FMS-like tyrosine kinase-1(VEGFR-1/FLT-1).Method:10 Japanese rabbits were randomly selected as blank normal group(A group).The remaining 70 Japanese rabbits were chosen to establish SANFH model,and dealed with low doses of Escherichia coli endotoxin(LPS)combined with high dose of Hormone.Two weeks after establishing model,9 Japanese rabbits were randomly selected and sacrificed by air vein injection.HE staining was applied to the pathological sections,and observe the pathological changes of femoral head tissue under high power(×400)optical microscope.When it is confirmed that SANFH model was established successfully,randomly dividing the remaining Japanese rabbits into5 groups: model group(B Group),and low-,medium-and high-dose of Sheng Gu Zai Zao pill(C,D,E group),and core decompression group(F group),10 Japanese rabbits in each group.According to the experimental design,observe and record the behavior change of Japanese rabbits.At the 4th week and the 8th week,respectively,performing the X-ray and collecting the bilateral femoral head.On the other hand,detecting the quantity of VEGF and its receptor FLT-1 gene in femoral head tissue by RT-PCR.On the other hand,placing the femoral head in 4% paraformaldehyde for the preparation of making tissue sections.Dealing with HE staining and observing the pathological change of the femoral head tissue of the rabbits under light microscope;Detecting the expression of VEGF and the receptor FLT-1 in the femoral head tissue by immunohistochemistry.Results:(1)Change in the behavior of the Japanese rabbits: Before establishing model,all Japanese rabbits were under well condition,and the limb of flexion and extension is normal.After model was established,the Japanese rabbits were apathetic and unresponsive,the limb of flexion and extension decreased and the limb of muscle strength weaken,some Japanese rabbits appeared limping.After medicating,the joint flexion and mobility of Japanese rabbits in C,D,E and F group were respectively improved,compared with that in group B.The symptom of E and F group improved obviously.With the time passing,the symptom of of Japanese rabbits in C,D,E and F group was recovery in different degree,E and F group recover more obviously than that in other groups.(2)X-ray finding:Agroup femoral head shape is integrity,joint space is normal,articular surface is smooth,femoral head bone density is uniform;B group of joint space narrowing,bone density is uneven,the point of cystic degeneration was found under cartilage,trabecular bone is blurred.At the same time,the range change of cystic was significantly lower in C,D,E and F group than that in group B,and the decreasing of range in E and F group was more significant than that in group C,D,E and F group.At 8th week,the range of cystic reduced more significantly than that at 4th week.the reduce in group E and F is more significant.(3)HE staining result: There is no pathological change in Agroup.The HE staining sections in B,C,D,E and F group appear empty bone lacunae.Compared with B group,the number of osteoblasts in the C,D,E,and F group was significantly decreased(P <0.05).The number of cavities in the E group was the most significant(P <0.05).The number of osteoblasts in the C,D,E and F group at 8th week was significantly less than that in the 4th week,and the change in group F was significant(P<0.05).(4)Immunohistochemical result: the expression of VEGF:Alittle of positive particle can be seen in Agroup;The positive particle in B group was not obvious;The number of positive particle in group C,D,E and F group was significantly more than that in group B,and the positive particle in group E is the most significant(P <0.05).the expression of FLT-1:Alittle of positive particle can be seen in Agroup;The positive particle in B group was not obvious;The expression of positive particles in C,D,E and F group was significantly higher than that in group B,and the expression of positive particle in group E was the most significant,especially in the necrotic area(P <0.05).At the 8th week,the positive particle of C,D,E and F group was increased compared with that at the4 th week.The change of group E was the most obvious(P <0.05).(5)The result of RT-PCR:?The expression of VEGF:the expression of VEGF in C,D,E and F group was significantly higher than that in group Aand B(P <0.05).The expression of VEGF in group E was significantly higher than that in group C,D and F(P <0.05);?The expression of FLT-1 in group C,D,E and F was significantly higher than that in group Aand B(P<0.05).The expression of FLT-1 in C,D and F group was significantly increased(P <0.05).Conclusion:(1)The experiment proved that high dose of hormone can successfully induce the occurrence of rabbit SANFH.(2)The effect of Sheng Gu Zai Zao pill on the early SANFH in rabbits was clear,and it can control the change in early SANFH condition,and promote the healing of necrotic bone tissue.Compared with the core decompression,which is recent effect,its effect lasting more,(3)VEGF and its receptor FLT-1 relate with SANFH lesion closely.Upregulating the expression of VEGF and receptor FLT-1 is one of the mechanisms of Sheng Gu Zai Zao pill in the treatment of SANFH. |
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