摘要
背景:补肾中药密骨片可有效防治骨质疏松,但其具体的药理学机制仍不是很清楚。转化生长因子β1是调节骨吸收与骨形成的重要耦联因子。目的:探讨补肾方对成骨细胞转化生长因子β1mRNA表达的影响。设计:随机分组设计,对照实验。单位:华中科技大学同济医学院附属协和医院骨伤科研究室。材料:实验于2003-05/2004-04在华中科技大学同济医学院附属协和医院中西结合骨代谢实验室完成。动物:选择出生一两天清洁级SD大鼠16只。实验药:密骨片药液在华中科技大学同济医学院附属协和医院骨伤科研究室制备,主要成分为淫羊藿、杜仲、胡桃肉、干地黄、淮牛膝等;阳性对照药:重组碱性成纤维细胞生长因子由北京邦定公司提供。阴性对照组分为探针阴性对照和抗体阴性对照。方法:将体外分离培养的新生SD大鼠的颅骨成骨细胞加入100,1000,5000,10000mg/L补肾中药密骨片药液及阳性对照药5μg/L重组碱性成纤维细胞生长因子。24h后,利用自行制备的地高辛标记的鼠转化生长因子β1cDNA探针进行成骨细胞的核酸分子原位杂交分析,以其细胞内杂交阳性颗粒的平均光密度值代表转化生长因子β1mRNA的表达水平。在200倍视野下,每组随机选取40个成骨细胞,用TJTY-300型全自动图像分析仪测定细胞内杂交颗粒的平均光密度值(A)。主要观察指标:各组成骨细胞转化生长因子β1mRNA表达。结果:全自动图像分析显示,5000,10000mg/L密骨片药液组的细胞转化生长因子β1mRNA表达分别是阴性对照组的1.18倍和1.30倍,差异有显著性[5000,10000mg/L密骨片药液组、阴性对照组的细胞内杂交颗粒的平均光密度值(A)分别为0.21367±0.01500,0.23703±0.02173,0.18127±0.01528,P<0.05和P<0.01],5μg/L重组碱性成纤维细胞生长因子组的细胞内杂交颗粒的平均光密度值(A)(0.25445±0.02081)虽高于5000,10000mg/L密骨片药液组,但差异无显著性(P>0.05)。结论:补肾中药密骨片可刺激成骨细胞转化生长因子β1的分泌和合成,从而促进骨形成和抑制骨吸收。
BACKGROUND： Migu tablet, a Chinese drug for kidney invigorating, is effective on preventing and treating osteoporosis, but the concrete mechanism of pharmacology is still not clear. Transforming growth factor-β1 （TGF-β1） is an important cytokine, which can regulate bone resorption and formation.
OBJECTIVE： To investigate the effect of kidney invigorating recipe on mRNA expression of TGF-β1 of osteoblast.
DESIGN： A completely randomized controlled study was conducted. SETTING： Department of Traumatic Orthopedics, Union Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology.
MATERIALS： This experiment was conducted at the laboratory for bone metabolism of integration of Chinese and western medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology from May 2003 to April 2004. Experimental rats： Totally 16 newborn SD rats of clean degree were involved. Experimental drug： Medical liquor of Migu tablet was prepared in the Department of Traumatic Orthopedics, Union Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology. The subscription was mainly composed of Chinese herbs such as Herba Epimedii, Cortex Eucommiae, Semen Juglandis, Radix Rehmanniae, Radix Achyranthis Bidentatae. and so on. Positive control drug： which was recombinant basic fibroblastic growth factor （rbFGF）, was purchased from Beijing Banding Company. Negative control group was subdivided into negative control of probe and antibody.
METHODS： 100,1 000,5 000,10 000 mg/L Chinese herb Migu tablet liquor for kidney invigorating and positive control drug 5 p,g/L rbFGF were added into the osteoblasts of cranial bones of newly born SD rats separated and cultured in vitro. 24 hours later, nuclear acid ,molecular in situ hybridization of osteoblasts were analyzed by self-made digoxin-labeled TGF-β1 cDNA probe . The mean absorbance of positive particles representing the mRNA expression of TGF-β1, A total of 40 osteoblasts were randomly chosen from each group under 200-fold amplification. The average absorbance of hybridized particles of the cells was measured with TJTY-300 automatic image analyzer,
MAIN OUTCOME MEASURES： mRNA expression of TGF-β1 in osteoblasts of each group,
RESULTS： Automatic image analyzer showed that the TGF-β1 mRNA expressions of Migu liquor groups whose concentration were 5 000 mg/L and 10 000 mg/L were respective 13, 1.18 times and 1.30 times that of control group, with a significant difference, [The mean absorbance＇s of hybridized particles of the cells in the 5 000,10 000 mg/L Migu liquor groups and negative control were 0.213 67±0,015 00,0,237 03±0,021 73,0.181 27 ±0.015 28,respectively , P 〈 0.05 and P 〈 0.01].Ahhough the mean absorbance （ 0.254 45±0,020 81）of the hybridized particles of the cells in the 5 μg/L recombinant rbFGF was higher than those of 5 000,10 000 mg/L Migu liquor groups, but there was no significant difference（P 〉 0.05）. CONCLUSION： Migu tablet for kidney invigorating can stimulate the secretion and synthesis of TGF-β1 in osteoblasts, thus promote bone formation and. inhibit bone resorption,
出处
《中国临床康复》
CSCD
北大核心
2006年第11期177-179,共3页
Chinese Journal of Clinical Rehabilitation
基金
卫生部科学基金资助(961153)~~
