Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu 610041, China.
J Ethnopharmacol. 2011 May 17;135(2):553-60. doi: 10.1016/j.jep.2011.03.056. Epub 2011 Apr 2.
The seeds of Cuscuta chinensis (Tu-Si-Zi, TSZ) have long been used for the treatment of osteoporosis in China and some Asian countries. The compounds in TSZ responsible for the antiosteoporotic activity are still poorly understood.
The present study was designed to investigate the osteogenic compounds in TSZ, and to evaluate their antiosteoporotic effects in osteoblastic cells.
Osteoblast-like UMR-106 cells were used for bioactivity-guided isolation of the active compounds. The activity of alkaline phosphatase (ALP) in UMR-106 cells was measured by p-nitrophenyl sodium phosphate assay. The proliferation of UMR-106 cells was assayed by Alamar-Blue method. Estrogenic activity of the extracts and isolated compounds was evaluated by activation of estrogen response element (ERE) luciferase reporter expression in HeLa cells co-transfected with human estrogen receptor subtypes (ERα or ERβ) expression vectors and 5×ERE luciferase reporter plasmid. Antiestrogenic activity of the extracts and isolated compounds were evaluated by activation of activator protein-1 (AP-1) luciferase reporter expression in HeLa cells co-transfected with human estrogen receptor subtypes (ERα or ERβ) expression vectors and 6×AP-1 luciferase reporter plasmid.
ALP-guided fractionation led to the isolation of five known flavonoids, quercetin, kaempferol, isorhamnetin, hyperoside and astragalin from the crude ethanolic extract of TSZ. Further study showed that kaempferol and hyperoside significantly increased the ALP activity in UMR-106 cells. Astragalin promoted the proliferation of UMR-106 cells whereas other compounds had no such effect. The isolated compounds showed estrogenic activity but quercetin, kaempferol and isorhamnetin showed more potent ERβ agonist activity. However, compared with their ER agonist activity, only quercetin and kaempferol showed potent ER antagonist activity by activating ERα/β-mediated AP-1 reporter expression.
Our findings validated the clinical use of TSZ in the treatment of osteoporosis, and demonstrated that kaempferol and hyperoside are the active compounds in TSZ for the osteogenic effect.
菟丝子（菟丝子，TSZ）的种子在中国和一些亚洲国家长期以来一直被用于治疗骨质疏松症。负责抗骨质疏松活性的 TSZ 化合物仍知之甚少。
本研究旨在研究 TSZ 中的成骨化合物，并评估其在成骨细胞中的抗骨质疏松作用。
使用成骨样 UMR-106 细胞进行活性化合物的生物活性指导分离。通过对硝基苯磷酸钠测定法测量 UMR-106 细胞碱性磷酸酶（ALP）的活性。通过 Alamar-Blue 法测定 UMR-106 细胞的增殖。通过激活人雌激素受体亚型（ERα或 ERβ）表达载体和 5×ERE 荧光素酶报告质粒共转染的 HeLa 细胞中的雌激素反应元件（ERE）荧光素酶报告表达来评估提取物和分离化合物的雌激素活性。通过激活人雌激素受体亚型（ERα或 ERβ）表达载体和 6×AP-1 荧光素酶报告质粒共转染的 HeLa 细胞中的激活蛋白-1（AP-1）荧光素酶报告表达来评估提取物和分离化合物的抗雌激素活性。
ALP 指导的分步分离导致从 TSZ 的粗乙醇提取物中分离出五种已知的类黄酮，槲皮素，山奈酚，异鼠李素，高圣草素和芦丁。进一步的研究表明，山奈酚和高圣草素显着增加了 UMR-106 细胞中的 ALP 活性。芦丁促进 UMR-106 细胞的增殖，而其他化合物则没有这种作用。分离的化合物表现出雌激素活性，但槲皮素，山奈酚和异鼠李素表现出更强的 ERβ激动剂活性。然而，与它们的 ER 激动活性相比，只有槲皮素和山奈酚通过激活 ERα/β 介导的 AP-1 报告表达表现出有效的 ER 拮抗剂活性。
我们的研究结果验证了菟丝子在骨质疏松症治疗中的临床应用，并表明山奈酚和高圣草素是 TSZ 中成骨作用的活性化合物。
