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山东大学学报(医学版) ›› 2010, Vol. 48 ›› Issue (10): 9-13.

• 论文 • 上一篇    下一篇

胱抑素C对大鼠心肌成纤维细胞增殖的影响

张书敏1,2,王永梅1,杜贻萌1,魏峰涛1,蒋卫东1,王欣1,董兆强1,徐冬玲1,姜红2,鹿庆华1   

  1. 1. 山东大学第二医院心内科,济南  250033;2. 教育部和卫生部心血管重构和功能研究重点实验室,济南2500121. 山东大学第二医院心内科,济南  250033;2. 教育部和卫生部心血管重构和功能研究重点实验室,济南250012
  • 收稿日期:2010-05-17 出版日期:2010-10-16 发布日期:2010-10-16
  • 通讯作者: 鹿庆华(1964- ),男,教授,硕士生导师,主要从事临床冠状动脉介入治疗及术后再狭窄基础与临床干预的研究。 E-mail:lqhzhy@163.com
  • 作者简介:张书敏(1985- ),女,硕士研究生,主要从事压力负荷过重致心肌细胞外基质重塑及血管生成的基础与临床研究。

Effects of CysC on the proliferation of cultured rat cardiac fibroblasts

ZHANG Shu-min1,2, WANG Yong-mei1, DU Yi-meng1, WEI Feng-tao1, JIANG Wei-dong1,WANG Xin1, DONG Zhao-qiang1, XU Dong-ling1, JIANG Hong2, LU Qing-hua1   

  1. 1. Department of Cardiovasular disease, The Second Hospital of Shangdong University, Jinan 250033, China;
    2. Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and  Ministry of Public Health, Jinan 250012, China
  • Received:2010-05-17 Online:2010-10-16 Published:2010-10-16

摘要:

目的    探讨胱抑素C(CysC)高表达对大鼠心肌成纤维细胞(CFs)增殖能力的影响。方法   取出生1~3d的Sperague-Dawley(SD)清洁级大鼠10只,体质量10~15g,差速贴壁法培养乳鼠心肌成纤维细胞;采用Gateway技术构建重组CysC腺病毒高效表达载体(Ad-CysC)并体外转染大鼠CFs;将对数生长期的大鼠CFs随机分成正常细胞组(PBS组),携有绿色荧光蛋白(GFP)的腺病毒组(Ad-GFP组)及CysC腺病毒高表达组(Ad-CysC组)。采用蛋白免疫印迹技术(WB)检测CysC蛋白在CFs中的表达情况,通过5-溴脱氧尿嘧啶核苷(BrdU)掺入法检测CysC高表达对CFs增殖能力的影响。结果    经DNA测序及WB结果分析,CysC腺病毒高效表达载体构建成功,转染CFs 24h后(MOI=200),AdCysC的转染效率高达90%以上;用重组CysC腺病毒转染CFs后, CysC蛋白在24h即有明显的表达上调(P<0.05);与PBS组及Ad-GFP组相比,转染Ad-CysC的CFs在72h的 BrdU摄取率显著提高(P<0.05)。结论     大鼠CysC腺病毒高效表达能够促进CFs增殖,可能加速心肌纤维化进程。

关键词: 胱抑素C;心肌成纤维细胞;细胞增殖

Abstract:

Objective    To explore the  effects of cystatin(CysC)on the proliferation of rat cardiac fibroblasts. Methods    Cardiac fibroblasts from 1~3 days old Sprague-Dawley rats (clean grade, weight 10~15g) were cultured by the anchorage velocity-dependent separation method. The recombinant adenovirus vector for CysC gene(Ad-CysC)was constructed with the GatewayTM technique and transfected into rat cardiac fibroblasts in vitro. Rat cardiac fibroblasts were randomly divided into the normal control group, the Ad-GFP group and the Ad-CysC group. Expression of CysC at the protein level was examined by means of Western-blot .Cell proliferation rate was measured by bromodeoxyuridine(BrdU). Results    DNA sequencing results and Western blot analysis indicated that recombinant Ad-CysC was successfully constructed. The efficiency of Ad-CysC infecting rat cardiac fibroblasts was above 90% when multiplicity of infection(MOI) was 200. Furthermore,  protein expression of CysC was significantly increased at 24h after transfection(P<0.05). Compared to the normal control group and the Ad-GFP group, the BrdU incorporation rate of the Ad-CysC group was much higher at 72h after transfection(P<0.05). Conclusions    High expression of CysC can stimulate the proliferation of cardiac fibroblasts,which could promote the myocardial fibrosis.

Key words: CystatinC;Cardiac fibroblast;Cell proliferation

中图分类号: 

  • R363.1
[1] . 维甲酸诱导胚胎神经管畸形与βcatenin基因表达的关系[J]. 山东大学学报(医学版), 2009, 47(8): 45-49.
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