Yao Fei, Chen Wei, Gu Weiwei, Xu Heng, Hou Wenyue, Liang Guoqiang, Zhang Zhu Ruixian, Jiang Guorong, Zhang Lurong
Central Laboratory, Suzhou TCM Hospital Affiliated to Nanjing University of Chinese Medicine, Suzhou, Jiangsu, 215000, China.
Clinical Pharmaceutical Laboratory of Traditional Chinese Medicine, Suzhou Academy of Wumen Chinese Medicine, Suzhou, Jiangsu, 215000, China.
Comb Chem High Throughput Screen. 2024;27(6):834-844. doi: 10.2174/1386207326666230607155913.
The function of promoting bone regeneration of Moutan Cortex (MC), a traditional Chinese medicine, has been widely known but, the effective components of MC in promoting osteoblast-mediated bone regeneration were still unclear.
The method of osteoblast membrane bio-specific extraction conjugated with HPLC analysis was established to screen bone regeneration active components from MC.
The fingerprints, washing eluate and desorption eluate of MC extract were analyzed by the established HPLC-DAD method. The established MC3T3-E1 cells membrane chromatography method was used for the bio-specific extraction of MC. The isolated compounds were identified by MS spectrometry. The effects and possible mechanisms of the isolated compounds were evaluated by molecular docking, ALP activity, cell viability by MTT Assay and proteins expression by Western Blot Analysis.
The active compound responsible for bone regeneration from MC was isolated using the established method of osteoblast membrane bio-specific extraction conjugated with HPLC analysis, and it was identified as 1,2,3,4,6-penta-O-β-galloyl-D-glucose (PGG) by MS spectrometry. It was further demonstrated through molecular docking that PGG could fit well into the functional ALP, BMP2, and Samd1 binding pocket. The proliferation of osteoblasts was promoted, the level of ALP was increased, and the protein expression of BMP2 and Smad1 was increased as shown by further pharmacological verification.
It was concluded that PGG, the bone regeneration active compound from MC, could stimulate the proliferation of osteoblasts to promote osteoblast differentiation, and its mechanism might be related to the BMP/Smad1 pathway.
中药牡丹皮促进骨再生的作用已广为人知，但牡丹皮促进成骨细胞介导的骨再生的有效成分仍不清楚。
建立成骨细胞膜生物特异性提取结合高效液相色谱分析的方法，从牡丹皮中筛选骨再生活性成分。
采用建立的高效液相色谱-二极管阵列检测法分析牡丹皮提取物的指纹图谱、洗脱液和洗脱液。采用建立的MC3T3-E1细胞膜色谱法对牡丹皮进行生物特异性提取。通过质谱鉴定分离得到的化合物。通过分子对接、碱性磷酸酶活性、MTT法检测细胞活力以及蛋白质免疫印迹分析评估分离化合物的作用及可能机制。
采用建立的成骨细胞膜生物特异性提取结合高效液相色谱分析的方法，从牡丹皮中分离得到了具有骨再生活性的化合物，经质谱鉴定为1,2,3,4,6-五-O-β-没食子酰基-D-葡萄糖（PGG）。分子对接进一步表明，PGG能很好地结合到功能性碱性磷酸酶、骨形态发生蛋白2和Smad1的结合口袋中。进一步的药理验证表明，PGG能促进成骨细胞增殖，提高碱性磷酸酶水平，增加骨形态发生蛋白2和Smad1的蛋白表达。
牡丹皮中的骨再生活性化合物PGG可刺激成骨细胞增殖以促进成骨细胞分化，其机制可能与骨形态发生蛋白/Smad1信号通路有关。
