Li Juan, Wu Wei-kang, Sun Wei, Yu Ke-qiang
Institute of Integrated Traditional Chinese and Western Medicine, Sun Yet-sen University, Guangzhou 510000, China.
Di Yi Jun Yi Da Xue Xue Bao. 2004 Dec;24(12):1359-61, 1366.
To study the effects of kidney-tonifying Chinese herbal drugs on Ca2+ intake and mineralization of human osteoblasts in vitro.
Human osteoblasts were isolated from the iliac trabecular bone followed by purification and culture at 37 degrees Celsius with 5% CO2. The cells were identified by cell morphology, calcium nodule formation and alkaline phosphatase (ALP) activity assay. The third passage of the cultured osteoblasts were treated with 10% scrum from rat fed with the decoction of the kidney-tonifying Chinese herbal drugs of different concentrations for 30 min, 3 d and 28 d, respectively. The cells treated with 10% rat serum without the drugs served as the control. Flow cytometry was used to observe the changes in cell proliferation and intracellular Ca2+ concentration, and von Kossa staining employed for quantification of the mineral nodules.
The osteoblasts obtained were positive for ALP staining and could form calcium nodules in vitro. Flow cytometry showed that the drugs at different concentrations all increased Ca2+ influx, as compared with the control cells. The drugs also increased the relative proliferation index of the osteoblasts, and high concentration of the drugs resulted in greater number of the mineral nodules in the osteoblasts (P<0.05).
The kidney-tonifying Chinese herbal drugs may increase Ca2+ influx and stimulate proliferation and differentiation of adult osteoblasts in vitro.
研究补肾中药对体外培养的人成骨细胞钙摄取及矿化的影响。
从髂骨小梁骨分离人成骨细胞,经纯化后于37℃、5%二氧化碳条件下培养。通过细胞形态学、钙结节形成及碱性磷酸酶(ALP)活性测定对细胞进行鉴定。将培养的第三代成骨细胞分别用不同浓度补肾中药水煎剂灌胃大鼠的10%血清处理30分钟、3天和28天。用不含药物的10%大鼠血清处理细胞作为对照。采用流式细胞术观察细胞增殖及细胞内钙浓度变化,并用冯科萨染色法对矿化结节进行定量分析。
所获得的成骨细胞ALP染色呈阳性,且在体外能形成钙结节。流式细胞术显示,与对照细胞相比,不同浓度的药物均增加了钙内流。药物还增加了成骨细胞的相对增殖指数,高浓度药物使成骨细胞中的矿化结节数量增多(P<0.05)。
补肾中药可能增加钙内流,并在体外刺激成年成骨细胞的增殖和分化。