大鼠肺细小动脉平滑肌细胞培养新方法的建立及血小板衍生因子对其增殖迁移的影响

doi:10.13459/j.cnki.cjap.2016.04.015

中国应用生理学杂志 ›› 2016, Vol. 32 ›› Issue (4): 343-346.doi: 10.13459/j.cnki.cjap.2016.04.015

大鼠肺细小动脉平滑肌细胞培养新方法的建立及血小板衍生因子对其增殖迁移的影响

朱宁, 赵旭勇, 向贻佳, 曾春来

温州医科大学附属第五医院心内科, 浙江丽水 323000

收稿日期:2015-09-28 修回日期:2016-05-16 出版日期:2016-07-28 发布日期:2018-06-20
通讯作者: 曾春来,Tel:0578-2681018;E-mail:zengchunlai@medmail.com.cn E-mail:zengchunlai@medmail.com.cn
基金资助:
曾春来,Tel:0578-2681018;E-mail:zengchunlai@medmail.com.cn

A method of primary cell culture of rat pulmonary artery smooth muscle cells and effects of platelet-derived growth factor induced proliferation and migration

ZHU Ning, ZHAO Xu-yong, XIANG Yi-jia, ZENG Chun-lai

Cardiovascular Department, the Fifth Affiliated Hospital, Wenzhou Medical University, Lishui 323000, China

Received:2015-09-28 Revised:2016-05-16 Online:2016-07-28 Published:2018-06-20
Contact: 曾春来,Tel:0578-2681018;E-mail:zengchunlai@medmail.com.cn E-mail:zengchunlai@medmail.com.cn
Supported by:
曾春来,Tel:0578-2681018;E-mail:zengchunlai@medmail.com.cn

摘要/Abstract

摘要： 目的：建立一种操作简单、实验仪器要求低的大鼠肺细小动脉平滑肌细胞（PASMCs）分离和培养的方法，并且探索血小板衍生因子（PDGF）介导的增殖、迁移的情况。方法：向右心注射铁及琼脂糖，利用琼脂糖能同时粘附血管内皮细胞、平滑肌细胞及铁粉，再结合胶原酶I的消化，通过磁力架吸引铁，特异性地分选出带血管的肺组织，经过3~4周左右的培养及纯化，得到肺细小动脉平滑肌细胞。用倒置相差显微镜观察细胞形态，免疫细胞化学法和免疫荧光染色法进行α-平滑肌肌动蛋白鉴定。MTT实验和划痕实验检测PDGF诱导的肺动脉细小平滑肌细胞的增殖和迁移。结果：分离后第14天、第21天及传代后进行鉴定，均表明分离培养的细胞为PASMCs。MTT结果表明，与不加PDGF组相比，PDGF增殖明显增加（P<0.05）。划痕实验结果显示PDGF刺激组比不刺激组迁移显著增多。结论：本方法分离培养大鼠的PASMCs，操作方便，实验仪器要求低。PDGF能够促进肺细小动脉平滑肌细胞的增殖、迁移。

关键词: 肺动脉平滑肌细胞, 大鼠, 分离, 原代培养。

Abstract: Objective:To establish an easy, not depending on advanced laboratory apparatus method to isolate and culture rat pulmonary artery smooth muscle cells (PASMCs), and to explore the effects of platelet-derived growth factor (PDGF) on cell proliferation and migration. Methods:The right ventricle was perfused with the mixture of iron, agarose, and the PASMCs and iron could adhere to agarose. The iron-con-taining tissue would move to side of the tube next to the magnet and could be digested by collagenase I. By the method, vessel-containing tissue could be attained. With 3-4 weeks' purification, the PASMCs could be obtained. The PASMCs morphology was observed by an inverted micro-scope, and identified by immunocytochemistry and immunofluorescence. The effects of PDGF on cell proliferation and migration was detected by MTT assay and scratch wound assay. Results:14 days、21 days and primary culture after isolation, the PASMCs was identified, and the re-sult showed that isolation and primary culture of the cells were PASMCs. Compared with the cells with no stimulation, the proliferation of PASMCs exposed to PDGF was increased significantly(P<0.05), and scratch wound assay demonstrated that PDGF induced the significant increase of migration of PASMCs. Conclusion:This method to isolate and culture rat PASMCs is simple, not depending on advanced laborato-ry. PDGF can promote the proliferation and migration of PASMCs.

Key words: pulmonary arterial smooth muscle cells, rat, isolation, primary cell culture

朱宁, 赵旭勇, 向贻佳, 曾春来. 大鼠肺细小动脉平滑肌细胞培养新方法的建立及血小板衍生因子对其增殖迁移的影响[J]. 中国应用生理学杂志, 2016, 32(4): 343-346.

ZHU Ning, ZHAO Xu-yong, XIANG Yi-jia, ZENG Chun-lai. A method of primary cell culture of rat pulmonary artery smooth muscle cells and effects of platelet-derived growth factor induced proliferation and migration[J]. CJAP, 2016, 32(4): 343-346.

参考文献

[1] Michelakis ED. Pulmonary arterial hypertension:yesterday, today, tomorrow[J]. Circ Res, 2014, 115(1):109-114.
[2] Hoeper MM, Mayer E, Simonneau G, et al. Chronic throm-boembolic pulmonary hypertension[J]. Circulation, 2006, 113(16):2011-2020.
[3] Courboulin A, Paulin R, Giguere NJ, et al. Role for miR-204 in human pulmonary arterial hypertension[J]. J Exp Med, 2011, 208(3):535-548.
[4] Rabinovitch M. Molecular pathogenesis of pulmonary arterial hypertension[J]. J Clin Invest, 2012, 122(12):4306-4313.
[5] 钱国清, 王良兴, 陈婵, 等. 大鼠细小肺动脉平滑肌细胞原代培养和鉴定方法的研究[J]. 中国应用生理学杂志, 2010, 26(1):125-128.
[6] 虞晓明, 郭瑞, 唐江锋, 等. 小鼠肺细小动脉平滑肌细胞原代培养和鉴定以及低氧对其增殖与凋亡的影响[J]. 中国病理生理杂志, 2014, 30(9):1724-1728.
[7] Owens GK, Kumar MS, Wamhoff BR. Molecular regulation of vascular smooth muscle cell differentiation in development and disease[J]. Physiol Rev, 2004, 84(3):767-801.
[8] Owens GK. Regulation of differentiation of vascular smooth muscle cells[J]. Physiol Rev, 1995, 75(3):487-517.
[9] Savai R, Al-Tamari HM, Sedding D, et al. Pro-proliferative and inflammatory signaling converge on FoxO1 transcription fa ctor in pulmonary hypertension[J]. Nat Med, 2014, 20(11):1289-1300.
[10] Roque M, Kim WJ, Gazdoin M, et al. CCR2 deficiency de-creases intimal hyperplasia after arterial injury[J]. Arte-rioscler Thromb Vasc Biol, 2002, 22(4):554-559.
[11] Li P, Zhu N, Yi B, et al. MicroRNA-663 Regulates human vascular smooth muscle cell phenotypic switch and vascular neointimal formation[J]. Circ Res, 2013, 113(10):1117-1127.
[12] 孟立平, 周昌钻, 郭艳, 等. 黄酒中抑制同型半胱氨酸诱导的大鼠血管平滑肌细胞增殖和迁移成分探讨[J]. 中国应用生理学杂志, 2015, 31(5):437-442.
[13] Tallquist M, Kazlauskas A. PDGF signaling in cells and mice[J]. Cytokine Growth Factor Rev, 2004, 15(4):205-213.

[1]	扈敬, 李法东, 邵小婷, 李思葳, 张欣颖, 赵冰冰, 徐长庆, 魏璨. 精胺对高糖引起的大鼠心肌纤维化的影响及其机制[J]. 中国应用生理学杂志, 2020, 36(3): 193-196.
[2]	朱建忠, 赵灿, 隋月林, 刘媛媛, 乔跃兵. 瘦素对糖尿病大鼠糖脂代谢及炎性因子的影响[J]. 中国应用生理学杂志, 2020, 36(3): 197-201.
[3]	孙晓娟, 冯武龙, 侯娜, 李娜, 姜蔚丽. 有氧运动和白藜芦醇对2型糖尿病大鼠肾脏JAK2及TGF-β1的影响[J]. 中国应用生理学杂志, 2020, 36(3): 202-206.
[4]	石慧, 梁晓珊, 黄丽文, 罗之刚, 谭龙. 一种高尿酸血症大鼠模型诱导方法的改良和效果评价研究[J]. 中国应用生理学杂志, 2020, 36(3): 223-227.
[5]	杨琼, 潘娅, 陈粲, 戴桃李. 阿尼西坦对血管性痴呆大鼠的治疗作用[J]. 中国应用生理学杂志, 2020, 36(3): 232-234.
[6]	韩洁, 刘子怡, 方振, 田蕾, 杨丹凤, 袭著革, 刘晓华. 重复制动应激对雌性大鼠下丘脑-垂体-卵巢轴的影响[J]. 中国应用生理学杂志, 2020, 36(3): 245-249.
[7]	孙景然, 樊祢祢, 张桢, 吴瑾, 韩殿鹏, 白家磊, 杜连群, 房彦军. 有机磷阻燃剂TDCIPP对雌性大鼠甲状腺的潜在毒性作用[J]. 中国应用生理学杂志, 2020, 36(3): 250-254.
[8]	陈宇峰, 董凡赫, 楼云玮, 寿今豪, 张慧婷, 周一超, 严明, 毛红娇, 张云. 补骨脂素对TCP磨损颗粒所致大鼠成骨细胞损伤的干预作用及其机制[J]. 中国应用生理学杂志, 2020, 36(3): 255-260.
[9]	李家卉, 张敏, 付子豪, 李勇枝, 王佳平, 高峰, 董玲. 经皮穿刺抽取大鼠脑脊液方法的改良[J]. 中国应用生理学杂志, 2020, 36(3): 265-267.
[10]	徐昌顺, 孙培, 林春. 改良型大鼠鞘内穿刺置管针及导管末端注药装置在大鼠鞘内置管术中的应用研究[J]. 中国应用生理学杂志, 2020, 36(3): 283-288.
[11]	王凌星, 洪姗燕, 杨美丽, 黄红红. 丁苯酞对睡眠剥夺大鼠额叶小胶质细胞活化的影响^*[J]. 中国应用生理学杂志, 2020, 36(2): 106-110.
[12]	完建永, 张勇, 刘世强, 华田苗, 孙庆艳. 运动对胰岛素抵抗大鼠肝脏BIM信号通路的影响^*[J]. 中国应用生理学杂志, 2020, 36(2): 115-118.
[13]	姚良雪, 韩家鑫, 魏培韵, 侯思博, 何潇剑, 张瑞岭, 杜爱林. 丁苯酞对慢性酒精中毒大鼠海马、杏仁核H₂S/CBS通路及学习记忆能力的影响^*[J]. 中国应用生理学杂志, 2020, 36(2): 138-142.
[14]	窦丽. 游泳训练对高血压大鼠血压及血栓前状态分子的影响^*[J]. 中国应用生理学杂志, 2020, 36(2): 161-164.
[15]	苏艳红, 袁乾坤, 肖蓉, 陈娟, 李强, 张世超. 抗阻训练对增龄大鼠骨骼肌线粒体功能的影响^*[J]. 中国应用生理学杂志, 2020, 36(2): 165-170.

大鼠肺细小动脉平滑肌细胞培养新方法的建立及血小板衍生因子对其增殖迁移的影响

A method of primary cell culture of rat pulmonary artery smooth muscle cells and effects of platelet-derived growth factor induced proliferation and migration

PDF (PC)

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价