锁阳含药血清对MC3T3-E1成骨细胞增殖分化及矿化的影响
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摘要
目的:研究锁阳含药血清对体外培养小鼠成骨细胞的增殖、分化以及矿化的影响。方法:小鼠灌胃14天后摘眼球取血制备含药血清,分别将10%浓度的各组含药血清及对照组血清(生理盐水组血清)分别加入细胞培养液中培养成骨细胞。采用CCK8法检测成骨细胞增殖能力,钙钴法检测其碱性磷酸酶活性,0.2%茜素红染色法观察成骨细胞钙化结节情况。结果:同一时间点,与生理盐水血清组比较,锁阳含药血清各组均能促进MC3T3-E1成骨细胞增殖,其中,锁阳含药血清中剂量组与高剂量组在三个时间点均能够显著促进细胞增殖(p<0.01);锁阳含药血清能够增强MC3T3-E1成骨细胞分泌碱性磷酸酶(ALP),且有一定的浓度依赖性;随着锁阳含药血清浓度的增加,钙化结节数量也呈现增加的趋势。结论:锁阳含药血清能够促进MC3T3-E1成骨细胞增殖分化与矿化,且在一定浓度范围内,浓度越高,促进作用愈明显。
Objective: To study the effects of cynomorium-containing serum on the proliferation, differentiation and mineralization of mouse osteoblasts in vitro. Method: The drugwas intragastrically administered to mice for 14 days, and the blood was collected from eyeballs to prepare the drug-containing serum. The 10% serum of each group and the serum of the control group(serum of saline group) were separately added to the cell culture medium to culture osteoblasts. The CCK8 method was used to detect the proliferation of osteoblasts. The alkaline phosphatase activity was detected by calcium and cobalt method, and the calcified nodules of osteoblasts were observed by 0.2% alizarin red staining. Result: Compared with the saline serum group,each group of cynomorium-containing serum could promote the proliferation of MC3 T3-E1 osteoblasts at the same time point.Among them, the medium dose group of cynomorium-containing serum and high dose group of cynomorium-containing serum could significantly promote cell proliferation at three time points.(p<0.01). Cynomorium-containing serum could enhance alkaline phosphatase(ALP) secreted by MC3 T3-E1 osteoblasts in a concentration-dependent manner. With the increase of the concentration of cynomorium-containing serum, the number of calcified nodules also increased. Conclusion: Cynomorium-containing serum can promote the proliferation, differentiation and mineralization of MC3 T3-E1 osteoblasts. Within a certain concentration range, the promotion effect is related to concentration.
Objective: To study the effects of cynomorium-containing serum on the proliferation, differentiation and mineralization of mouse osteoblasts in vitro. Method: The drugwas intragastrically administered to mice for 14 days, and the blood was collected from eyeballs to prepare the drug-containing serum. The 10% serum of each group and the serum of the control group(serum of saline group) were separately added to the cell culture medium to culture osteoblasts. The CCK8 method was used to detect the proliferation of osteoblasts. The alkaline phosphatase activity was detected by calcium and cobalt method, and the calcified nodules of osteoblasts were observed by 0.2% alizarin red staining. Result: Compared with the saline serum group,each group of cynomorium-containing serum could promote the proliferation of MC3 T3-E1 osteoblasts at the same time point.Among them, the medium dose group of cynomorium-containing serum and high dose group of cynomorium-containing serum could significantly promote cell proliferation at three time points.(p<0.01). Cynomorium-containing serum could enhance alkaline phosphatase(ALP) secreted by MC3 T3-E1 osteoblasts in a concentration-dependent manner. With the increase of the concentration of cynomorium-containing serum, the number of calcified nodules also increased. Conclusion: Cynomorium-containing serum can promote the proliferation, differentiation and mineralization of MC3 T3-E1 osteoblasts. Within a certain concentration range, the promotion effect is related to concentration.
引文
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[10]程婉,汤小康,应航,等.牵张应力环境下六味地黄汤含药血清对成骨细胞增殖分化影响的实验研究[J].中国骨伤,2013,26(02):142.
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[12]肖亚平,曾杰,焦琳娜,等.补肾中药对骨质疏松症的治疗及其信号通路调节作用的研究进展[J].中国中药杂志,2018,43(01):21.
[13]Zhang JF,Li G,Chan CY,et al.Flavonoids of Herba Epimedii regulate osteogenesis of human mesenchymal stem cells through BMP and Wnt/beta-catenin signaling pathway.[J].Mol.Cell.Endocrinol,2010,314(1):70.
[14]Tang DZ,Yang F,Yang Z,et al.Psoralen stimulates osteoblast differentiation through activation of BMP signaling.[J].Biochem.Biophys.Res.Commun,2011,405(2):256.
[2]李振华,郭静霞,崔占虎,等.锁阳的研究进展与资源保护[J].中国现代中药,2014,16(10):861.
[3]Cui Z,Guo Z,Miao J,Wang Z,et al.The genus Cynomorium in China:an ethnopharmacological and phytochemical review[J].J Ethnopharmacol,2013,147(1):1.
[4]张霞,徐力生,许晓雪,等.锁阳80%乙醇提取物对去卵巢大鼠骨质疏松防治作用研究[J].中药药理与临床,2017,33(03):101.
[5]史平,朱薇,李晓鸣.锁阳多糖对去卵巢大鼠骨质疏松的改善作用[J].第三军医大学学报,2015,37(23):2360.
[6]冯方.锁阳对兔激素性股骨头坏死低氧状态下Wnt信号通路的影响[D].河南中医药大学,2016.
[7]Quarles LD,Yohay DA,Lever LW,et al.Distinct proliferative and differentiated stages of murine MC3T3-E1 cells in culture:an in vitro model of osteoblast development.[J].J.Bone Miner.Res,1992,7(6):683.
[8]Wang D,Christensen K,Chawla K,et al.Isolation and characterization of MC3T3-E1 preosteoblast subclones with distinct in vitro and in vivo differentiation/mineralization potential[J].J.Bone Miner.Res,1999,14(6):893.
[9]张会鲜,何琪杨.CCK-8法检测药物影响肿瘤细胞增殖的优化研究[J].药学研究,2016,35(02):63.
[10]程婉,汤小康,应航,等.牵张应力环境下六味地黄汤含药血清对成骨细胞增殖分化影响的实验研究[J].中国骨伤,2013,26(02):142.
[11]王勇平,廖燚,蒋垚.成骨细胞的体外培养与鉴定[J].中国组织工程研究与临床康复,2011,15(33):6231.
[12]肖亚平,曾杰,焦琳娜,等.补肾中药对骨质疏松症的治疗及其信号通路调节作用的研究进展[J].中国中药杂志,2018,43(01):21.
[13]Zhang JF,Li G,Chan CY,et al.Flavonoids of Herba Epimedii regulate osteogenesis of human mesenchymal stem cells through BMP and Wnt/beta-catenin signaling pathway.[J].Mol.Cell.Endocrinol,2010,314(1):70.
[14]Tang DZ,Yang F,Yang Z,et al.Psoralen stimulates osteoblast differentiation through activation of BMP signaling.[J].Biochem.Biophys.Res.Commun,2011,405(2):256.