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骨质疏松症和阿尔茨海默病均为退行性疾病,越来越多的研究表明它们的发病机制具有关联[1]。骨质疏松症往往伴随着认知缺陷,Aβ沉积是阿尔茨海默病的典型症状,而阿尔兹海默病小鼠(APPswe/PS1dE9)的骨组织中也会出现Aβ沉积,并出现骨密度降低,骨强度减弱等骨质疏松症状[2]。另有研究表明,氧化应激会导致 Aβ沉积,这种氧化损伤状态可被抗氧化剂改善[3]。因此, Aβ沉积偶联氧化损伤可视为阿尔茨海默病及骨质疏松症的共同发病机制。巴戟天丸收载于明代《古今医统大全》,由君药巴戟天,臣药远志、石菖蒲、茯苓、人参以及地骨皮、茯神组成[4]。前期课题组研究已证实,巴戟天丸组方可以从体内外水平上改善D-半乳糖(D-gal)引发的骨丢失,但巴戟天丸组方治疗Aβ沉积所致的骨丢失及具体作用机制有待进一步阐明[4, 5]。因此,本研究拟以Aβ1-42寡聚体损伤成骨细胞模型,对巴戟天丸组方的抗氧化能力及对骨形成干预作用进行探究,并通过网络药理学方法对潜在的作用机制进行预测。
The roles of Bajitianwan formula on Aβ-injured osteoblasts and the mechanism based on network pharmacology
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摘要:
目的 探讨巴戟天丸组方对Aβ损伤成骨细胞的骨形成作用及其机制。 方法 以新生24 h Wistar大鼠所分离的成骨细胞为研究对象,用 Aβ1-42 寡聚体对成骨细胞进行损伤,并用巴戟天丸组方水提物进行药物干预。分别采用MTT法、碱性磷酸酶(ALP)活性检测、过氧化氢酶(CAT)活性检测、超氧化物歧化酶(SOD)活性检测、谷胱甘肽(GSH)活性检测以及丙二醛(MDA)活性检测。采用蛋白质印迹法检测骨形成相关蛋白骨形态发生蛋白2(BMP2)、成骨特异性转录因子(RUNX-2)、骨保护蛋白(OPG)的表达水平;明确巴戟天丸组方对Aβ损伤成骨细胞的作用后,采用网络药理学方法对潜在的作用机制进行预测。 结果 巴戟天丸组方可显著促进Aβ损伤成骨细胞的增殖,提高 ALP、SOD、GSH 活性,抑制MDA活性,并促进骨形成相关蛋白BMP2、RUNX-2、OPG的表达。网络药理学分析显示,巴戟天丸组方发挥改善Aβ损伤成骨细胞的作用主要与AGE-RAGE信号通路、PI3K-Akt信号通路、MAPK信号通路以及神经活性配体与受体的相互作用通路等有关。 结论 明确巴戟天丸组方具有改善Aβ损伤成骨细胞的作用,并通过网络药理学方法探究其相关作用通路,为传统方剂巴戟天丸抗骨质疏松的临床应用提供借鉴。 Abstract:Objective To explore the effect of Bajitianwan(BJTW)formula on bone formation of Aβ-injured osteoblasts and its mechanism. Methods Osteoblasts isolated from neonatal 24-hour Wistar rats were used for the study, and osteoblasts were subjected to damage with Aβ1-42 oligomers, and pharmacological intervention was performed with the aqueous extract of BJTW formula. The MTT assay, alkaline phosphatase(ALP)activity assay, catalase(CAT)activity assay, superoxide dismutase(SOD)activity assay, glutathione(GSH)activity assay and malondialdehyde(MDA)activity assay were carried out respectively. The expression levels of bone morphogenetic protein 2(BMP2), osteogenic specific transcription factor(RUNX-2)and osteoprotective protein(OPG)were detected by Western blotting. After confirming the effect of BJTW formula on Aβ-injured osteoblasts, the network pharmacology method was used to predict the potential pathways. Results The BJTW formula significantly promoted the proliferation of Aβ-injured osteoblasts, increased ALP, SOD and GSH activity, inhibited MDA activity, and promoted the expression of bone formation-related proteins BMP2, RUNX-2 and OPG. Network pharmacological analysis showed that the effect of ameliorating of Aβ-injured osteoblasts by BJTW formula was mainly mediated by AGE-RAGE, PI3K-Akt, MAPK and neuroactive ligand-receptor interaction signaling pathways. Conclusion In this study, the effect of BJTW formula on improving the osteoblasts damaged by Aβ was confirmed for the first time, and its related mechanism was explored based on network pharmacology method. The results lay a strong foundation for the clinical application of traditional formula BJTW against osteoporosis. -
Key words:
- osteoblasts /
- Bajitianwan /
- Aβ deposition /
- network pharmacology
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