| Objective1. To discuss the clinical effects of Sheng Mai Cheng Gu Tablet in the treatment of Steroid-induced Osteonecrosis of Femoral Head (SONFH)2. To screen the differential expression of microRNA in peripheral blood of patients with SONFH and explore the relationship between microRNA and the SONFH through bioinformatic analysis.3. To search for the differential expression of microRNA in the rabbit model of SONFH which Sheng Mai Cheng Gu Tablet intervented, and to analyze the possible mechanism of microRNA expression in the treatment of SONFH.Methods1. To observe 26 cases of SONFH in patients who will be taked oral Sheng Mai Cheng Gu Tablet (6 tablet once),3 times a day, and be treated for three months. SONFH patients’hip jiont Harris score and VAS score will be recorded and compared before treatment, after 1 month treatment, after 2 month treatment and after 3 month treatment. Then collect data for statistical analysis and observe the influence of SONFH patients with Sheng Mai Cheng Gu Tablet in Harris score and VAS score.2.6 cases of SONFH patients (observation group)in ARCO stage Ⅱ who were selected in the out-patient orthopedic department of the first affiliated hospital of TCM from July,2014 to February,2015. At the same time, Six healthy volunteers of Guangzhou University of TCM in the control group. Collecte the plasma of two groups and filtrate differential expression of micro RNA with using microRNA Real-time PCR microarray. Predict the target gene of differential expression microRNA and analyze bioinformatic according to TargetScan, mirBase and miRanda.3.30 adult New Zealand rabbits with weighing 2.5～3.0kg, were divided into 3 groups at random (Chinese medicine group, control group and blank group). The Chinese medicine group and control group were injected 2 times every day for 2 days through ear vein with lOug/kg LPS for the total. At the third time of LPS with MPS 40mg/kg with intragluteal injection every 24h, injection of MPS for 3 times. The blank group was injected with same dose saline at the same time. Every rabbit was injected with 40 thousand units penicillin to prevent infection. The Chinese medicine group will be taked Sheng Mai Cheng Gu Tablet 0.4/kg orally in the last injection of MPS, the control group and the blank group with the same dose saline by gavage for 8 weeks.To make sure the model success,we selecte 1 rabbit from the Chinese medicine group and 1 another from the control group randomly at the 4th week, slice the femoral head and conduct HE staining. Plasma was collected when complete gavage of rabbit to measure the microNRA relative expression by high-throughput sequencing. Screen the different microRNA between the observation group, the control group and the blank group, obtain the key microRNA through bioinformatic analysis.Results1. The Harris score was 38.17±4.02 befere the treatment,39.83±3.97 after treatment for 1 month,42.67±2.25 after treatment for 2 months,44.33±3.20 after treatment for 3 months,and VAS score was 6.83±0.41,6.33±0.52,5.00 ±0.89 and 4.33±0.52 in sequence. Before treatment compared to after treatment for 2 and 3 months were statistically different.2.752 microRNA were checked out with MicroRNA Real-time PCR microarray. 18 microRNA expressed differently in the two groups, of which in the observation group 13 microRNA are increased, and 5 microRNA are decreased.887 target genes were selected.GO analysis showed that the expression target gene function microRNA regulation included anatomical morphology, mesenchymal cell proliferation, cell metabolic processes in the observation group less than those in control group. Pathway analysis showed that the target gene was significantly enriched in multiple signaling pathways.3.1879 novel microRNA was predicted by miRDeep2.0. Between control group and blank group differences in microRNA expression by homology analysis revealed the corresponding human microRNA:hsa-miR-941 and hsa-miR-187. And hsa-miR-941 and hsa-miR-187 expression levels increased at first and then decline trend in expression between blank group, control group and Chinese medicine group. hsa-miR-187 can regulate apoptosis and angiogenesis. MAPK signaling pathways significantly enriched in the microRNA target gene which differences expression between Chinese medicine group and blank group.Conelusion1. Sheng Mai Cheng Gu Tablet can improve the patients with SONFH in Harris score and VAS score effectively.2. This study find that microRNA differentially expressed in the plasma between patients with SONFH and normal people, and indicate that these differentially expressed microRNAs may be as the screening of SONFH of biological markers.3. Sheng Mai Cheng Gu Tablet regulate of several signaling pathways, restrain apoptosis, promote angiogenesis by affecting microRNA expression, which may be the mechanism of it in the treatment of SONFH. |
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