模拟微重力效应三维旋转培养促进造血干细胞向巨核细胞分化的机制研究

Megakaryocytes (MKs) are essential bone marrow cells responsible for generating platelets and cytokines which involve clotting and immune modulations. Generating magakaryocyte and platelet from human hematopoietic stem cells (hHSCs) holds great promise for drug screening and transfusion purpose in clinical. Currently, the efficiency of generating functional platelets from hHSC is poor and the mechanism underlying hHSCs differentiation into megakaryocyte is also far from clear. Studies have demonstrated that rotary cell culture system (RCCS), s an effective tool to simulate microgravity, provide an acceptable 3D dynamic environment to stimulate the proliferation and differentiation of stem cells. Our previous study has found that RCCS can potentiate megakaryopoiesis and significantly improve the efficiency of platelet generation. Base on this discovery, the present study is to investigate in detail a potential mechanism of 3D rotary culture regulating MKs differentiation. Whole-genome gene expression profile analysis by RNA-SEQ will be conducted to screen potential key genes regulating megakaryopoiesis. We also propose to explore the functions and regulation mechanisms of related genes that screened by RNA-SEQ during megakaryopoiesis in 3D rotary suspension culture. Our results will reveal the effect of 3D rotary suspension culture on megakaryopoiesis and thrombopoiesis, and our discovery will facilitate the strategy optimization for magakaryocytes and platelets generation from blood derived CD34+ cell in large-scale.

巨核细胞是一种重要的骨髓细胞，能够产生血小板和各种细胞因子，参与机体的促凝血及免疫调节等过程。利用人造血干细胞分化产生的巨核细胞和血小板在药物筛选、临床输注治疗等方面具有广阔的前景。目前体外分化产生巨核细胞和具有功能的血小板效率较低，且有关巨核细胞体外分化和调控机制知之甚少。我们前期研究中建立了模拟微重力效应3D旋转悬浮条件下造血干细胞向巨核细胞分化和血小板生成的体外3D模型，结果表明3D旋转培养促进巨核细胞分化和功能性血小板的产生。本课题在前期发现的基础上进一步深入探讨其作用机制，基于全基因组RNA测序挖掘能调控巨核分化的候选功能分子，并针对所发现的分子进一步分析其在巨核分化和血小板形成中的功能及作用机制。总之，本研究将进一步揭示模拟微重力效应3D旋转培养条件下巨核细胞分化、成熟和血小板产生的规律，为高效分化产生巨核细胞和血小板的策略提供理论依据和技术指导。

巨核细胞是一种重要的骨髓细胞，能够产生血小板和各种细胞因子，参与机体的促凝血及免疫调节等过程。利用人类干细胞分化产生的巨核细胞和血小板在药物筛选、临床输注治疗等方面具有广阔的前景。模拟微重力条件下细胞培养和组织构建研究表明, 微重力环境有利于细胞的三维生长, 形成具有功能的组织样结构, 培养后的三维组织无论从形态上还是基因表达上都更接近于正常的机体组织。我们前期发现采用模拟微重力旋转培养有利于提高脐带血来源的造血干细胞向巨核细胞分化及血小板的产生。本课题在前期发现的基础上进一步探讨了微重力和小分子化合物的联合作用，研究发现在一些特定时期联合旋转培养和小分子化合物可进一步促进巨核分化和血小板形成。此外，本课题还分别采用了EB分化模型和单层造血模型来探讨了模拟微重力在人类胚胎干细胞向早期造血前体细胞分化的影响，研究表明微重力对人胚胎干细胞向造血前体细胞分化具有一定的影响，主要影响分化阶段为中胚层向生血内皮及后期分化，但不会影响中胚层的分化。总之，本研究首次发现了微重力在人胚胎干细胞早期造血发生中的作用，为下一步的机制研究打下重要的基础。