IκB激酶复合物对系统性红斑狼疮患者骨髓间充质干细胞迁移及增殖的作用

Migration and proliferation of human mesenchymal stem cells from patients with systemic lupus erythematosus mediated by IKK-β

目的观察系统性红斑狼疮（SEE）患者骨髓间充质干细胞（BMSCs）迁移、增殖变化，并探讨IκB激酶复合物（IKK—β）对SLE患者BMSCs迁移及增殖的影响。方法采用密度梯度离心和贴壁分离法分离培养6例SEE患者及6名健康人BMSCs；通过细胞划痕实验和构建侵袭小室检测BMSC8迁移、活细胞计数（CCK-8）检测BMSCs增殖能力；实时荧光定量聚合酶链反应技术检测IKK-βmRNA表达水平；蛋白印迹法检测IKK—β、磷酸化IKK—β表达水平；观察加入IKK—β抑制剂对SLE患者BMSCs增殖、迁移的影响。采用t检验或Mann—Whitney秩和检验。结果①体外培养过程中，SLE患者BMSCs迁移率（5．2±3．8）‰与增殖能力（0．21±0．49）显著低于健康对照组BMSCs迁移率（7．0±2．9）‰及增殖能力（1．00±0．35），差异有统计学意义（P〈0．05）；②SLE患者BMSCsIKK—β基因表达水平（1．9±1．4）与健康人（1．9±2．4）相比差异无统计学意义（P〉0．05）；SEE患者BMSCs磷酸化IKK—B蛋白表达水平（1．41±0．19）显著高于健康人（0．93±1．24），差异有统计学意义（P〈0．05）；③IKK—β抑制剂能够增加SLE患者BMSCs的迁移率（3．3±1．6）％。和增殖能力（1．13±0．26），高于未加IKK—β抑制剂组迁移率（2．3±1．1）‰与增殖能力（0．81±0．17），差异有统计学意义（P〈0．05）。结论SEE患者BMSCs在体外培养过程中，迁移、增殖能力较健康人显著降低，其机制可能是SEE患者体内IKK—β的表达增加，通过活化核因子-κB信号通路，影响迁移与增殖相关基因的表达，从而抑制了BMSCs的迁移与增殖。

Objective To investigate the effect of IKB kinase （IKK-β） on migration and proliferation of bone marrow mesenchymal stem cells （BMSCs） from patients with systemic lupus erythematosus（SLE）. Methods Human bone marrow aspirates were coUected from iliac of six donors and six SLE patients and cultured in vitro. Migration of BMSCs were observed by wound healing and transwell migration assays. Proliferation of BMSCs was quantified by cell counting kit-8 assay. Total RNA was extracted and reverse transcribed into complementary DNA. Real-time PCR technique was used to determine the gene expression of IKK-β at transcription level. The expression of IKK-β and phospho-IKK-β （p-IKK-β） protein were determined by Western blotting analysis. Statistical analysis was conducted with or Mann-Whitney rank test. Results （1） The migration rate of BMSCs from SLE patients （5.2±3.8）‰ were significantly reduced as compared with normal controls （7.0±2.9）‰（P〈0.05）. The proliferation of BMSCs of SLE patients （0.21±0.49） was lower than that from healthy controls （1.00±0.35）（P〈0.05）. （2） No difference in IKK-13 mRNA expression between SLE （1.9±1.4） subjects and normal controls （1.9±2.4）（P〉0.05）, IKK-β protein expression of BMSCs from SLE patients （1.41±0.19） increased significantly compared with healthy controls （0.93±1.24） （P〈0.05）.IKK-β may be involved in migration and proliferation of BMSCs.