中药骨康调控成骨细胞核内结合因子α1的表达  

Chinese medicine Gukang prescription modulates core binding factor alpha 1 expressing in osteoblasts

作　　者：赵可伟[1] 邱俊林 潘旭枫[1] 梁秀珍[1] 陈小华[1] 

机构地区：[1]广州中医药大学附属骨伤科医院,广东省广州市510240 [2]丰顺中医院,广东省梅州市514300

出　　处：《中国组织工程研究》2013年第33期5929-5935,共7页Chinese Journal of Tissue Engineering Research

摘　　要：背景:中药骨康在临床上治疗骨质疏松疗效明确,但其具体作用途径尚不清晰。目的:假设中药骨康通过调节核内结合因子 a1 水平,控制其下游基因核因子κB受体活化因子配体和骨保护素表达,起到调控成骨细胞生长发育的作用。方法:新生 24 h 内的 SD 乳鼠用于成骨细胞的分离培养。成年 SD 雌性大鼠用于制备药物血清,随机分为正常血清组和中药骨康组。2 组大鼠按体表面积的方法给予中药骨康方的提取药物和生理盐水灌胃,连续给药 1周。最后一次用药后 2 h 行心脏采血,分离血清。原代培养并传至第 3 代经碱性磷酸酶鉴定取得的大鼠成骨细胞,消化计数铺板并分为 2 组,以上述血清处理 72 h,MTT 法检测成骨细胞的增殖率,ELISA 方法检测碱性磷酸酶分泌量并以相应吸光度值进行纠正,运用 RT-PCR 检测 2 组成骨细胞核内结合因子α1 及其下游核因子κB受体活化因子配体和骨保护素 mRNA 表达情况。结果与结论:中药骨康组成骨细胞骨保护素和核内结合因子 a1 mRNA 水平显著高于正常血清组,核因子κB受体活化因子配体蛋白和 mRNA 水平显著低于正常血清组(P < 0.01)。实验结果证实,中药骨康可通过影响核内结合因子 a1 表达,调控其下游基因核因子κB受体活化因子配体和骨保护蛋白表达和分泌,进而发挥治疗骨质疏松的作用。BACKGROUND： Chinese medicine Gukang prescription has a clear effect on clinical treatment of osteoporosis, but the therapeutic pathway is still unclear. OBJECTIVE： To investigate the effects of Chinese medicine Gukang on the expression of receptor activator of nuclear factor kappa B ligand and osteoprotegerin by regulating core binding factor alpha 1 expression to control the growth and development of osteoblasts. METHODS： Sprague-Dawley neonatal rats within 24 hours after delivery were used for the separation and culture of osteoblasts. Adult Sprague-Dawley rats were used to prepare drug-containing serum, and then divided into two groups randomly： normal control group and Gukang group. Rats in the normal control and Gukang groups were intragastrically administrated with extract of Gukang prescription and normal saline based on rat’s body surface area, for 1 consecutive week. Two hours after the last administration, blood samples were taken from the heart. Then the serum was collected. Osteoblasts at passage 3 were confirmed with alkaline phosphatase assay and digested. After counting and planting, all osteoblasts were divided into two groups and treated with collected serum for 72 hours. Proliferative rate of osteoblasts was detected by using 3-（4,5-dimethylthiazol-2-yl）-2,5- diphenyltetrazolium bromide test. Secretion of alkaline phosphatase was detected by using enzyme linked immunosorbent assay and corrected with the corresponding absorbance value. mRNA expression of core binding factor alpha 1, receptor activator of nuclear factor kappa B ligand and osteoprotegerin were detected by using reverse transcription-PCR in all groups. RESULTS AND CONCLUSION： mRNA expression of osteoprotegerin and core binding factor alpha 1 in the Gukang group was significantly higher than that in the normal control group, but protein and mRNA expression of receptor activator of nuclear factor kappa B ligand were dramatically lower in the Gukang group compared with the normal control group （P 0.01）. These findin

关 键 词：组织构建 骨组织构建 骨质疏松 中药骨康 核内结合因子 a1 核因子 ΚB 受体活化因子配体 骨保护蛋白 成骨细胞 细胞培养 中药血清学 

分 类 号：R318[医药卫生—生物医学工程]