The Effect Of Electro-acupuncture Treatment On Expression Of HIF-1α And VEGF In Synovial Cells Of Rabbit With Knee Osteoarthritis

Objective: Knee osteoarthritis(knee osteoarthritis, KOA), a condition characterized by degenerative joint, is a common complaint among the elderly. Knee joint maintains a physiological hypoxia environment according to the study. Hypoxia inducible factor-1(HIF-1) is a kind of nuclear transcription factor(NF) as well as DNA-binding protein with the function of regulating other gene transcription in lowoxygen environment, and it plays an important role in that condition. HIF- 1 composes of HIF-1α and HIF-1β, and the former is a unique regulatory subunit in low oxygen condition. The expression of HIF-1α increases in low oxygen condition, and it transfers from cytoplasm to nucleus. HIF-1α combines with the DNA of VEGF, which leads to the high expression of VEGF. The research on osteoarthritis transfers from the study of cartilage and subchondral bone to the study of chronic synovitis with the constantly deepening of study. Treatment of KOA can be divided into operation therapy and non-operation treatment, and the electro-acupuncture therapy is clinically used as the non-operation treatment. The traditional acupuncture and electrical stimulation are combined in electro-acupuncture. The acupuncture stimulates the acupoints of KOA patients, which leads to the release of analgesicsubstance, and electrical stimulation causes muscle contraction, which improves local blood circulation. Those all can help damage repair. Therefore, this article tries to explore the mechanism of electroacupuncture treatment of rabbit knee KOA model through observing the gross morphology changes of articular morphology, the pathological changes of synovial cells and the expression of HIF-1α and VEGF in the synovial cells. Methods:(1) The object and intervention measures: Purchasing 35 male New Zealand rabbits with health(5 spare rabbits), 30 rabbits were divided into three groups randomly, 10 rabbits in each group. 10 rabbits were in control group without any treatment. All rabbits in model group and EA group group were gave plaster cast to fix their left knees straightly for 6 weeks to build OA model according to the Videman method. After 6 weeks following modeling, electro-acupuncture treatment group accepted acupuncture treatment with acupoints selection of left lower limb of rabbits such as Zu Sanli, Liang Qiu, Yang Lingquan and Xue Hai for 30 minutes at the same time every day, and the treatment was continued for 15 days, however, the remaining two groups were fixed for 30 minutes at the same time every day. Three animal groups were killed through air embolism after 15 days, and opened the left knee joint for gross observation, then used the sharp blade to cut synovium specimens, which was making HE staining, immunohistochemistry and light microscopy observing. Scoring was according to the assessment standardof synovial pathology, then the scores and the testing data were statistically analyzed.(2)Observation index: ?Visual observed three groups of rabbit knee joint gross morphological changes.?Cut a little rabbit knee medial femoral condyle synovium and fixed with 10% neutral Formaldehyde Solution. Paraffin embeding, slicing, and HE staining were made, and observed the change of morphology of synovial cells, and recorded the synovial cells score of synovial pathology.?Analysis of immunohistochemical expression: Used Rabbit anti-HIF-1α kit for detecting the HIF-1α expression of left knee joint medial condyle synovium, and observed the expression in the light microscope.(3)Statistical methods: statistical analysis was performed by using SPSS19.0. Mean and standard deviation were used to express the result. One-way ANOVA, LSD-t test and rank sum test were used to analyze all experimental data, and the correlation test was undertaken with the linear correlation analysis. The significance of difference was represent by P <0.05. Results:(1) Joint gross observation: there was no joint cavity swelling or congestion in control group. Synovial fluid was transparent, and synovium surface had the smooth appearance without edema or hyperaemia. Cartilage was translucent without defect or new biological. Articular surface of the model group was rough and most obvious effusion in articular cavity. Synovial membrane was obvious thickening. Synovial fluid was pale yellow, however, gloss and transparency ofcartilage decreased, whose color was gray, and partial cartilage surface had partial cracks and abnormal protuberance. Electro-acupuncture treatment group had only 3 rabbits with visible intra-articular effusion, and the amount was less than the model group. There were a little ulcer on part of the cartilage and synovium surface, however, cartilage and synovium surface gloss were smooth. Compared with the model group, those improved on the structure of cartilage and synovium but were poorer than control group.(2) Synovium HE stain observation: Total scores and scores of synovial cells hyperplasia, synovial inflammation, fibrous tissue hyperplasia and blood vessels hyperplasia were differentia significantly in three groups(P<0.05).(3)Immunohistochemistry HIF-1α and VEGF testing results: Under the microscope, brown particles within the cells are the expression of HIF-1α and VEGF. The control group had HIF-1α and VEGF weakly positive expression, and this experiment was extremely rare brown granules. Model group had HIF-1α and VEGF strongly positive expression, and full-thickness synovium were visible dark brown particles, such as synovial cells, synovial macrophages, vascular endothelial cell, fibroblast and so on. HIF-1α expresses mainly in cell nucleus and cytoplasm, and the expression of VEGF is in cytoplasm. Electro-acupuncture treatment group decreased than those in the model group about the expression of HIF-1α and VEGF, only in synovial tissue surface. Positive rate and IOD of HIF-1α and VEGF weredifferentia significantly in three groups(P<0.05). Conclusion:(1) Using tube type plaster fixation method can set up the knee osteoarthritis model successfully, which keeps the rabbit knee in extension position for six weeks.(2) The expression of HIF-1α and VEGF in articular synovium is positively correlated with the damage of joint, and the expression of VEGF is regulated by HIF-1α.(3)Electro-acupuncture therapy can reduce the degree of damage of articular synovium, and the mechanism may be that electro-acupuncture could decrease the expression of HIF-1α and VEGF in synovial cells.