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加味龟鹿二仙汤抑制顺铂诱导的人骨髓间充质干细胞损伤。

Flavored Guilu Erxian decoction inhibits the injury of human bone marrow mesenchymal stem cells induced by cisplatin.

作者信息

Ke Bin, Shi Lin, Xu Zhaozhong, Wu Gang, Gong Yuxin, Zhu Lixia, Wang Yanjing, Ouyang Huailiang, Wang Xilong

机构信息

Department of Traditional Chinese Medicine, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

Department of Traditional Chinese Medicine, Zhujiang Hospital of Southern Medical University, Guangzhou, China.

出版信息

Cell Mol Biol (Noisy-le-grand). 2018 May 15;64(6):58-64.

Abstract

To examine the exact role of flavored Guilu Erxian decoction, a Traditional Chinese Medicine (TCM) in the treatment of cisplatin-induced side-effects in bone marrow mesenchymal stem cells (BM-MSCs). BM-MSCs were isolated from bone marrow collected from SD rats and identified by flow cytometry. Cells were cultivated in MEM alpha medium containing 5% (TCM-L), 10% (TCM-M) and 20% (TCM-H) dosages of flavored Guilu Erxian decoction with or without cisplatin. Cell viability was determined through CCK-8 and thymidine analog 5-ethynyl-2'-deoxyuridine (EdU) staining assay. Flow cytometry was used to determine cell cycle and apoptosis. The expression of p21 and cleaved-caspase-3 were examined using Western blot assay. The PI3K-AKT-mTOR pathway associated proteins, including p-PI3K, p-AKT and p-mTOR, were also examined by Western blot assay. CCK-8 and EdU staining assay demonstrated that cisplatin could inhibit cell proliferation in BM-MSCs in a dose and time dependent manner. Further, cisplatin could induce apoptosis through increasing G0/G1 cell cycle arrest, p21 and cleaved-caspase-3 expression. However, these phenomena would be significantly alleviated when adding the serum containing flavored Guilu Erxian decoction. Furthermore, the PI3K-AKT-mTOR pathway activation could be inhibited by cisplatin in BM-MSCs, while flavored Guilu Erxian decoction treatment successfully abrogated this effect. Combination of flavored Guilu Erxian decoction and cisplatin could reduce the damage to BM-MSCs. This indicates that the flavored Guilu Erxian decoction can enhance the possibility of BM-MSCs repairing and rehabilitating the normal function of injured tissues induced by cisplatin, which could provide a new direction for therapeutic applications.

摘要

为研究中药加味龟鹿二仙汤在治疗顺铂诱导的骨髓间充质干细胞(BM-MSCs)副作用中的确切作用。从SD大鼠采集的骨髓中分离出BM-MSCs,并通过流式细胞术进行鉴定。细胞在含有5%(中药低剂量组,TCM-L)、10%(中药中剂量组,TCM-M)和20%(中药高剂量组,TCM-H)加味龟鹿二仙汤的MEMα培养基中培养,同时设置加或不加顺铂的条件。通过CCK-8和胸腺嘧啶类似物5-乙炔基-2'-脱氧尿苷(EdU)染色试验测定细胞活力。采用流式细胞术测定细胞周期和凋亡情况。使用蛋白质免疫印迹法检测p21和裂解型半胱天冬酶-3的表达。还通过蛋白质免疫印迹法检测PI3K-AKT-mTOR信号通路相关蛋白,包括p-PI3K、p-AKT和p-mTOR。CCK-8和EdU染色试验表明,顺铂可呈剂量和时间依赖性地抑制BM-MSCs的细胞增殖。此外,顺铂可通过增加G0/G1期细胞周期阻滞、p21和裂解型半胱天冬酶-3的表达诱导细胞凋亡。然而,添加含加味龟鹿二仙汤的血清后,这些现象将得到显著缓解。此外,顺铂可抑制BM-MSCs中PI3K-AKT-mTOR信号通路的激活,而加味龟鹿二仙汤治疗成功消除了这种作用。加味龟鹿二仙汤与顺铂联合使用可减少对BM-MSCs的损伤。这表明加味龟鹿二仙汤可提高BM-MSCs修复和恢复顺铂诱导的受损组织正常功能的可能性,这可为治疗应用提供新的方向。

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