兔骨髓基质干细胞的培养与鉴定体系的建立

摘要
图/表
参考文献(22)
相关文章 (2)

全文: PDF (709 KB) HTML (1 KB)
输出: BibTeX | EndNote (RIS)

摘要目的探讨兔骨髓基质干细胞的培养与鉴定体系的建立方法与效果。方法空气栓塞的方式处死SPF级新西兰大白兔6只，分离骨髓组织，利用贴壁培养分离纯化兔骨髓基质干细胞，多次传代后以流式细胞术检测细胞表面抗原；以BrdU标记骨髓基质干细胞，将骨髓基质干细胞注入兔颅内，分别于注射1、3周后取脑组织制作石蜡切片，进行免疫荧光染色。结果通过贴壁法培养幼年兔骨髓细胞可获得较纯化的骨髓基质干细胞。免疫细胞化学染色显示骨髓基质干细胞中CD44+细胞率超过90%，细胞核未着色；流式细胞仪测定培养骨髓基质干细胞的CD90+细胞率为94.5%，CD34+细胞率为0.24%。BrdU标记显示得到了能够掺DNA合成期的骨髓基质干细胞，标记细胞所占的比例在40.0%以上。结论通过贴壁法培养幼年兔骨髓细胞呈多角短梭形，可获得骨髓基质干细胞，BrdU可作为体内示踪骨髓基质干细胞的标志物之一。

	服务

	把本文推荐给朋友
	加入我的书架
	加入引用管理器
	E-mail Alert
	RSS
	作者相关文章
	邓辰红1 王飞2 王文兰1

关键词 ：骨髓基质干细胞, 培养与鉴定体系, BrdU, 免疫组化, 流式细胞仪

Abstract：Objective To investigate the methods and effects of the establishment and identification of rabbit bone marrow stromal stem cells. Methods 6 New Zealand rabbits of SPF grade were selected and sacrificed, and the bilateral femoral bone were separated and the bone marrow was given to aspiration, the rabbit bone marrow stromal stem cells were separated and purified by adherent culture method, after several passages, the cell surface antigen was detected by flow cytometry. And the BrdU was used to label bone marrow stromal stem cells. And the bone marrow stromal stem cells were injected into the rabbit brain,1 and 3 weeks after the brain for paraffin sections and immunofluorescence staining. Results The purified bone marrow stromal cells could be obtained by adherent culture method. Immunocytochemical staining showed that the CD44+ cell rate in bone marrow stromal stem cells was over 90%, and the nuclei were not stained. The CD90+ cell rate and CD34+ cell rate in cultured bone marrow stromal cells by flow cytometry were 94.5% and 0.24%. BrdU markers showed that bone marrow stromal cells could be synthesized into DNA, and the percentage of labeled cells was above 40%. Conclusion The cultured rabbit bone marrow cells are polygonal, juvenile short spindle, it can obtain purified bone marrow stromal stem cells, BrdU can be used as one of the ideal markers of bone marrow stromal stem cells in vivo.

Key words： Bone marrow stromal cells Culture and identification system BrdU Immunohistochemistry Flow cytometry

基金资助:内蒙古自治区自然科学基金项目（2012MS1157）。

通讯作者: 王飞（1971-），男，博士，主任医师；研究方向：脊柱微创。

作者简介: 邓辰红（1972-），女，硕士，副主任医师；研究方向：小儿神经。

引用本文:

邓辰红1 王飞2 王文兰1. 兔骨髓基质干细胞的培养与鉴定体系的建立[J]. 中国医药导报, 2017, 14(20): 20-22.
DENG Chenhong1 WANG Fei2 WANG Wenlan1. Culture and identification of rabbit bone marrow stromal stem cells. 中国医药导报, 2017, 14(20): 20-22.

链接本文:

http://www.yiyaodaobao.com.cn/CN/ 或 http://www.yiyaodaobao.com.cn/CN/Y2017/V14/I20/20

[1] 李东，毛豪丽，白珊珊，等.种植体联合骨组织工程技术修复犬下颌骨节段缺损的实验研究[J].组织工程与重建外科杂志，2016，12（5）：285-288.
[2] Shen LH，Chen J，Shen HC，et al. Possible mechanism of therapeutic effect of 3-Methyl-1-phenyl-2-pyrazolin-5-one and bone marrow stromal cells combination treatment in rat ischemic stroke model [J].Chin Med J，2016，129（12）：1471-1476.
[3] 王婷婷，何爱娟，刘豫，等.骨髓基质干细胞扩增体系研究[J].组织工程与重建外科杂志，2016，12（3）：152-155，159.
[4] Di Rosa F. Commentary：memory CD8+ T cells colocalize with IL-7+ stromal cells in bone marrow and rest in terms of proliferation and transcription [J].Front Immunol，2016，31（7）：102-109.
[5] 陈晨，张刚，谷铭勇，等.骨细胞网络结构对骨形成和骨吸收的影响[J].中国骨质疏松杂志，2016，22（1）：120-124.
[6] Ma Q，Cai M，Shang JW，et al. In vitro neural differentiation of bone marrow stromal cells induced by hepatocyte growth factor and glial cell derived neurotrophic factor [J].Eur Rev Med Pharmacol Sci，2016，20（22）：4654-4663.
[7] 崔晓萍，陈建梅，穆军山，等. PI3K-AKT通路对脑缺血损伤神经干细胞的增殖作用[J].中国康复医学杂志，2016， 31（2）：154-159.
[8] Wang H，Zhao Z，Du J，et al. MiRNA-378 controls cell proliferation in rabbit umbilical cord mesenymal stem cells [J]. Cell Mol Biol，2016，62（12）：97-101.
[9] 张黎龙，路磊，张学利，等.富血小板血浆诱导兔骨髓基质干细胞向成骨细胞的分化[J].中国组织工程研究与临床康复，2010，32（2）：5891-5896.
[10] Fayyad-Kazan H，Faour WH，Badran B，et al. The immunomodulatory properties of human bone marrow-derived mesenchymal stromal cells are defined according to multiple immunobiological criteria [J]. Inflamm Res，2016， 65（6）：501-510.
[11] Cortes-Dericks L，Froment L，Kocher G，et al. Human lung-derived mesenchymal stem cell-conditioned medium exerts in vitro antitumor effects in malignant pleural mesothelioma cell lines [J]. Stem Cell Res Ther，2016，9（7）：25-32.
[12] Huang H，Yang X，Bao M，et al. Ablation of the sox11 gene results in clefting of the secondary palate resembling the pierre robin sequence [J]. J Biol Chem，2016，291（13）：7107-7118.
[13] 张欣，贺晓生. Toll样受体4表达对创伤性颅脑损伤后小鼠海马区神经干细胞增殖的影响[J].中华神经外科杂志，2016，32（11）：1156-1161.
[14] Deane JA，Ong YR，Cain JE，et al. The mouse endometrium contains epithelial， endothelial and leucocyte populations expressing the stem cell marker telomerase reverse transcriptase [J]. Mol Hum Reprod，2016，22（4）：272-284.
[15] Song L，Zhu J，Zhang X，et al. BDNF-hypersecreting human umbilical cord blood mesenchymal stem cells promote erectile function in a rat model of cavernous nerve electrocautery injury [J]. Int Urol Nephrol，2016，48（1）：37-45.
[16] Kusuma GD，Menicanin D，Gronthos S，et al. Ectopic bone formation by mesenchymal stem cells derived from human term placenta and the deciduas [J]. PLoS One，2015， 10（10）：e0141246.
[17] 杨晓红，王胜男，刘庆余，等. Nrf2通过上调自噬促进C17.2神经干细胞的增殖[J].中山大学学报：医学科学版，2016，37（4）：481-489.
[18] Gu J，Gu W，Lin C，et al. Human umbilical cord mesenchymal stem cells improve the immune-associated inflammatory and prothrombotic state in collagen type-Ⅱ-induced arthritic rats [J]. Mol Med Rep，2015，12（5）：7463-74670.
[19] Bickford PC，Kaneko Y，Grimmig B，et al. Nutraceutical intervention reverses the negative effects of blood from aged rats on stem cells [J]. Age，2015，37（5）：103-109.
[20] Pirzad Jahromi G，Shabanzadeh Pirsaraei A，Sadr SS，et al. Multipotent bone marrow stromal cell therapy promotes endogenous cell proliferation following ischemic stroke [J]. Clin Exp Pharmacol Physiol，2015，42（11）：1158-1167.
[21] Li H，Li X，Zhang M，et al. Three-dimensional co-culture of BM-MSCs and eccrine sweat gland cells in Matrigel promotes transdifferentiation of BM-MSCs [J]. J Mol Histol，2015，46（4-5）：431-438.
[22] 王盼，单伟.局部联合移植hUCMSCs与VEGF对MCAO大鼠神经功能的影响[J].中国现代医生，2016，54（11）：17-19.