慢粒细胞外泌体选择性转运miR-711抑制BMSCs黏附功能

The adhesion deficiency of BMSCs to hematopoietic cells is an important cause of the progression of CML and the failure of bone marrow transplantation, but the exactly mechanisms have not been fully elucidated. Our previous studies have found that the CML exosomes selectively carry high concentrations of miR-711. Further bioinformatical analyses indicate that the target genes of miR-711 are include adhesion molecules, such as CD44, CXCL12, CCR9 and CDH5. Based on these, we put forward the hypothesis that CML cell derived-exosomes may selectively transport miR-711 to down-regulate the expression of adhesion molecules in BMSCs and inhibit their adhesion function. We propose to apply series experiments, such as confocal microscopy, cell adhesion assay, non-contact in vitro co-culture assay and co-transplantation to mouse model, to confirm that exosomes have inhibitory effects on adhesion functions of BMSCs; and utilize luciferase assay, over-expression or knock-down of miR-711 in BMSCs, RT-PCR and Western blot to verify that exosomes achieve their inhibition function through miR-711 reducing the expression of adhesion molecules in BMSCs. This study will make a meaningful exploration for verifying the theory that CML cells may reconstruct the bone marrow microenvironment by shedding exosomes,and understanding of CML progress mechanism and improving the curative effect of CML.

骨髓间充质干细胞（BMSCs）黏附造血细胞的能力低下是慢性粒细胞白血病(CML)进展及移植治疗失败的重要原因，但黏附缺陷具体机制未明。我们前期发现CML细胞外泌体携带高浓度miR-711，生物信息学分析显示其靶基因包括CD44、CXCL12、CCR9、CDH5等黏附相关分子。据此，我们提出"CML细胞通过外泌体携带转运miR-711抑制BMSCs黏附功能"的假说。我们拟通过激光共聚焦显微镜检查、细胞黏附实验、非接触共培养及动物模型共移植等体内外实验证实CML通过释放外泌体抑制BMSCs黏附功能；通过荧光素酶实验、过表达及抑制实验、RT-PCR及蛋白印迹等分子生物学实验证实外泌体的抑制作用是通过其携带的miR-711下调BMSCs黏附相关分子表达而实现的。最终证实CML可以通过释放外泌体对骨髓微环境进行"趋利避害"性改造，为深入理解CML进展机制及提高CML疗效提供理论基础。

骨髓间充质干细胞（ BMSCs）黏附造血细胞的能力低下是慢性粒细胞白血病(CML)进展及移植治疗失败的重要原因，但黏附缺陷具体机制未明。本项目中我们证实：慢性粒细胞白血病细胞能通过其分泌的外泌体作用于骨髓间充质干细胞，下调其黏附分子表达，从而降低其对造血干细胞的黏附能力，这可能与外泌体介导传递高浓度的miR-711至BMSCs细胞有关，为慢性粒细胞白血病的进展机制提供新思路。除此以外，我们还应用小RNA测序技术，通过全面地分析不同性质胸腔积液外泌体所携带的miRNA，并结合荧光定量PCR验证技术，筛选出特征性miRNA谱，为胸腔积液的鉴别诊断提供新的分子标志物。.通过本项目研究，课题组共发表中英文期刊论文9篇，申请专利3项，获批实用新型专利2项，参编大型专业书、医学本科教材3部，培养研究骨干2名、培养研究生5名。