经典Wnt途径对间充质干细胞增殖、迁移和抗氧化应激损伤的作用

刘艾然; 杨毅; 郭凤梅; 陆晓旻; 邱海波

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经典Wnt途径对间充质干细胞增殖、迁移和抗氧化应激损伤的作用

首发时间：2013-07-12

刘艾然 ¹
刘艾然，（1982-），女，讲师，主要研究方向：急性呼吸窘迫综合征的病理生理和治疗。
杨毅 ² 郭凤梅 ² 陆晓旻 ² 邱海波 ²
邱海波（1966-），男，主任医师，教授，主要研究方向：急性呼吸窘迫综合征的病理生理和治疗

1、东南大学附属中大医院重症医学科，南京 210009
2、东南大学附属中大医院重症医学科，南京，210009

摘要：目的：探索经典Wnt途径对小鼠骨髓来源间充质干细胞（mMSCs）增殖、迁移和抗氧化应激损伤的作用。方法：（1）直接贴壁法原代分离C57BL/6小鼠骨髓MSCs（mMSCs），通过表面标志物流式细胞学检测和成脂、成骨、成软骨诱导分化以对其进行鉴定。（2）MTT法评估不同浓度LiCl及Wnt3a干预下mMSCs的数量变化，以评估经典Wnt途径对mMSCs增殖能力的影响。（3）LiCl、Wnt3a及DKK 1调节经典Wnt途径，划痕实验及Transwell小室迁移实验评估经典Wnt途径对mMSCs的迁移能力和向ALI小鼠肺组织迁移的能力的影响。（4）LiCl、Wnt3a及DKK 1调节经典Wnt途径，MTT法评估经典Wnt途径对H2O2诱导mMSCs损伤的作用, Western blot法检测mMSCs中凋亡相关蛋白Bax及Bcl-2的表达以评估经典Wnt途径对mMSCs抗氧化应激损伤的作用。结果：（1）LiCl及Wnt3a均能呈浓度依赖性的促进mMSCs增殖，且在LiCl 4mmol/L或Wnt3a 150ng/ml时最为明显。（2）LiCl及Wnt3a活化经典Wnt途径后能促进mMSCs的水平及垂直迁移能力，并且能促进mMSCs向ARDS小鼠肺组织的体外迁移。（3）H2O2能导致mMSCs的损伤及凋亡相关蛋白Bcl-2/Bax表达下调，Wnt3a或LiCl活化经典Wnt途径能抑制H2O2导致的细胞损伤和Bcl-2/Bax下调，而DKK 1则能部分增强H2O2的损伤作用。结论：（1）激活经典Wnt途径能促进mMSCs的增殖。（2）激活经典Wnt途径能增强mMSCs的迁移能力及向ARDS肺组织的迁移。（3）活化经典Wnt途径能抑制H2O2导致的mMSCs损伤。

关键词：重症医学经典Wnt信号通路间充质干细胞增殖迁移氧化应激

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Role of canonical Wnt pathway in the proliferaion, migration and anti-oxidative stress of mice bone marrow mesenchymal stem cells in vitro

LIU Airan ¹
刘艾然，（1982-），女，讲师，主要研究方向：急性呼吸窘迫综合征的病理生理和治疗。
YANG Yi ² GUO Fengmei ² LU Xiaomin ² QIU Haibo ²
邱海波（1966-），男，主任医师，教授，主要研究方向：急性呼吸窘迫综合征的病理生理和治疗

1、Department of Critical Care Medicine, Zhongda Hospital, Southeast University, Nanjing 210009
2、Department of Critical Care Medicine, Zhongda Hospital, Southeast University, Nanjing, 210009, China

Abstract：Objective: To investigate the role of canonical Wnt pathway in the proliferation of mMSCs (marrow mesenchymal stem cells), as well as migration or survival against oxidative stress induced injury in vitro. Methods: (1) mMSCs derived from bone marrow of C57BL/6 mice were identified by characterizing the cell surface phenotypes using fluorescence-activated cell sorting analysis and testing the capabilities of adipogenic, osteogenic and chondrogenic differentiation. (2) The effect of different concentrations of LiCl or Wnt3a to the proliferation of mMSCs was evaluated using MTT assay. (3) The scratch test and Transwell chamber test were used to analyze the migration ability of mMSCs treated with LiCl, Wnt3a or DKK 1 to higher concentration of FBS or conditioned medium of ARDS mice derived lung tissue. (4) The role of canonical wnt pathway in the survival of mMSCs injured by H2O2 was evaluated using MTT assay, and the expression of apoptosis related protein, Bax and Bcl-2 were evaluated through western blot. Results: (1) Both LiCl and Wnt3a could promote the proliferation of mMSCs in a dose dependent manner and the maximum effects were observed after 4mmol/L LiCl or 150ng/ml Wnt3a treatment. (2) The migration of mMSCs evaluated with either Transwell inserts assay or scratch test increased after treatment with Wnt3a or LiCl, which also promoted the migration of mMSCs towards the conditioned medium of ARDS mice lung tissue in the lower chambers of Tranwell inserts. (3) Wnt3a and LiCl could ameliorate the cell death and the reduction of Bcl-2/Bax induced by H2O2, which simultaneously caused the reduction of p-GSK 3β and nuclear β-catenin in mMSCs. Conclusion: (1) The proliferation of mMSCs was increased by activation of canonical Wnt pathway. (2) The migration of mMSCs was promoted by the activation of canonical Wnt pathway. (3) The activation of canonical Wnt pathway contributed to the protection of mMSCs against the injury of oxidative stress.

Keywords： Critical Care Medicine canonical Wnt pathway marrow mesenchymal stem cell proliferation migration oxidative stress

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刘艾然，杨毅，郭凤梅，等. 经典Wnt途径对间充质干细胞增殖、迁移和抗氧化应激损伤的作用[EB/OL]. 北京：中国科技论文在线 [2013-07-12]. https://www.paper.edu.cn/releasepaper/content/201307-186.

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论文编号	201307-186
论文题目	经典Wnt途径对间充质干细胞增殖、迁移和抗氧化应激损伤的作用
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