唑来膦酸对破骨细胞分化中钙调蛋白依赖性激酶Ⅱ和Ⅳ基因表达的影响

doi:10.13481/j.1671-587x.20160105

吉林大学学报(医学版) ›› 2016, Vol. 42 ›› Issue (01): 19-23.doi: 10.13481/j.1671-587x.20160105

唑来膦酸对破骨细胞分化中钙调蛋白依赖性激酶Ⅱ和Ⅳ基因表达的影响

刘娟娟¹, 李鹏¹, 董伟¹, 冯晓洁¹, 温黎明¹, 孙红², 戚孟春¹

1. 华北理工大学口腔医学院口腔颌面外科教研室, 河北唐山 063000;
2. 华北理工大学基础医学院病理学教研室, 河北唐山 063000

收稿日期:2015-05-23 发布日期:2016-01-26
通讯作者: 戚孟春,教授,硕士研究生导师(Tel:0315-3721508,E-mail:qimengchun@163.com) E-mail:qimengchun@163.com
作者简介:刘娟娟(1988-),女,山西省朔州市人,医学硕士,主要从事骨组织工程学方面的研究。
基金资助:
国家自然科学基金资助课题(81270965)

Effects of zoledronate on gene expressions of calmodulin- dependent kinase Ⅱ and Ⅳ in osteoclast differentiation

LIU Juanjuan¹, LI Peng¹, DONG Wei¹, FENG Xiaojie¹, WEN Liming¹, SUN Hong², QI Mengchun¹

1. Department of Oral and Maxillofacial Surgery, College of Stomatology, North China University of Science and Technology, Tangshan 063000, China;
2. Department of Pathology, College of Basic Medical Science, North China University of Science and Technology, Tangshan 063000, China

Received:2015-05-23 Published:2016-01-26

摘要/Abstract

摘要：

目的: 研究唑来膦酸(ZOL)对破骨细胞(OC)分化及信号分子钙调蛋白依赖性激酶 (CaMK)Ⅱ、Ⅳ基因表达的影响,阐明其作用机制。方法: 应用小鼠单核巨噬细胞株RAW264.7向破骨细胞分化。细胞分为对照组和ZOL组,用核因子κB受体激活蛋白配体(RANKL)诱导5 d,ZOL组同时加用ZOL处理2 d,第5天收集细胞后检测OC生成及CaMKⅡ、CaMKⅣ基因表达情况。结果: ZOL组中多核OC数、骨吸收陷窝数和面积分别为33.0±1.0、46.0±3.5和(4 125.9±674.8)μm²,明显低于对照组的66.6±3.2、86.0±9.2和(9 418.3±1 260.8)μm²(P<0.05或P<0.01)。与对照组比较,ZOL组细胞中CaMKⅡ和CaMKⅣ基因表达水平明显降低(P<0.05),CaMKⅡ mRNA和蛋白表达水平分别下降了39.3%和58.9％,CAMKⅣ mRNA和蛋白表达水平分别下降了51.7%和46.8％(P<0.01)。免疫荧光化学,与对照组比较,ZOL组细胞中CaMKⅡ、Ⅳ蛋白荧光强度明显下降(P<0.05或P<0.01)。结论: ZOL在OC多核化阶段可以明显抑制OC的生成和骨吸收功能,并下调CaMKⅡ和CaMKⅣ基因表达。信号分子CaMKⅡ和CaMKⅣ可能参与了唑来膦酸对OC形成和功能的抑制作用,并在该过程中发挥重要的作用。

关键词: 唑来膦酸, 破骨细胞, 钙调蛋白依赖性激酶

Abstract:

Objective: To determine the effects of zoledronate (ZOL)on the osteoclast differention and gene expressions of Ca²⁺/calmodulin-dependent kinase (CaMK)Ⅱ and Ⅳ,and to clarify their mechanisms. Methods: The mouse RAW264.7 cells were used for osteoclast differentiation and the cells were divided into control and ZOL group. The cells in two groups were induced with receptor activatior of nuclear factor κB ligand(RANKL)(50 mg·L^-1)for 5 d,while the cells in ZOL group were also treated with ZOL (1×10^-6mol·L^-1)for 2 d. 5 d later,the cells were collected and the osteoclastogenesis and the gene expressions of CaMKⅡand CaMKⅣ were detected. Results: The number of osteoclasts,the number and size of resorption lacunaes in ZOL group were 33.0±1.0,46.0±3.5,and (4 125.9±674.8)μm², respevtively,which were significantly lower than those in control group (66.6±3.6,86.0±9.2, and 9 418.3μm²±1 260.8μm²)(P<0.05 or P<0.01). Compared with control group,the mRNA and protein levels of CaMKⅡ and CaMKⅣ in ZOL group were also significantly down-regulated (P<0.05); the expression levels of CaMK Ⅱ mRNA and protein were decreased by 39.3% and 58.9％,the expression levels of CaMK Ⅳ mRNA and protein were decreased by 51.7% and 46.8％.The immunofluorescence examination showed that the fluorescence intensities of CaMKⅡ and CaMKⅣ protein in ZOL group were decreased compared with control group(P<0.05 or P<0.01). Conclusion: At stage of multinuclear mature of osteoclasts,ZOL could significantly inhibit the final formation and resorption function of osteoclasts and down-regulate the gene expressions of CaMKⅡ and CaMKⅣ.CaMKⅡ and CaMKⅣ may play an important role in ZOL-induced inhibition of osteoclastogenesis.

Key words: zoledronate, osteoclasts, Ca²⁺/calmodulin-dependent kinase

中图分类号:

Q813

刘娟娟, 李鹏, 董伟, 冯晓洁, 温黎明, 孙红, 戚孟春. 唑来膦酸对破骨细胞分化中钙调蛋白依赖性激酶Ⅱ和Ⅳ基因表达的影响[J]. 吉林大学学报(医学版), 2016, 42(01): 19-23.

LIU Juanjuan, LI Peng, DONG Wei, FENG Xiaojie, WEN Liming, SUN Hong, QI Mengchun. Effects of zoledronate on gene expressions of calmodulin- dependent kinase Ⅱ and Ⅳ in osteoclast differentiation[J]. Journal of Jilin University Medicine Edition, 2016, 42(01): 19-23.

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唑来膦酸对破骨细胞分化中钙调蛋白依赖性激酶Ⅱ和Ⅳ基因表达的影响

Effects of zoledronate on gene expressions of calmodulin- dependent kinase Ⅱ and Ⅳ in osteoclast differentiation

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