Two-compartment microfluidic bioreactor with functionalized PEG hydrogels to promote platelet production in culture

1265029 -Miller从培养的巨核细胞中产生3000 - 6000亿个血小板的许多单位将需要在造血（血液）干细胞和祖细胞扩增、巨核细胞产生和成熟以及血小板释放和恢复方面取得进展。 窦中血流的剪切对于前血小板和血小板释放是重要的。受骨髓窦状隙的启发，该项目将采用具有两个隔室的微生物反应器，所述两个隔室由涂覆有微孔聚（乙二醇）水凝胶的多孔支持物隔开，所述微孔聚（乙二醇）水凝胶用粘合剂配体官能化。为了模拟跨窦状隙内皮的前血小板形成，将巨核细胞接种在上隔室的水凝胶上，并将培养基灌注通过下隔室。微生物反应器将用于研究剪切应力、水凝胶刚度和粘附配体类型和密度对前血小板和血小板产生的影响。将研究增加pH和趋化因子的梯度以进一步增加前血小板形成。灌注系统将允许血小板在释放时收获，这很重要，因为血小板质量在20-24°C储存期间会恶化，在37°C储存时甚至更快。将通过激动剂诱导的活化来验证培养衍生血小板的质量。微生物反应器可以使用中空纤维生物反应器和螺旋缠绕膜盒等形式进行规模化以用于血小板的临床生产，以产生所需的表面积。 在美国和欧洲，每年输注数百万单位的血小板。 通过来源于患者的巨核细胞或匹配的造血干细胞在培养物中生产血小板将增加供应并降低污染风险和不良免疫应答，但目前培养物中的血小板生产效率非常低。这项研究的成功完成将使培养血小板的大规模生产更接近于实现。 从事该项目的研究生和本科生将在跨学科的环境中接受培训。 该项目的一个关键要素是为研究生开发针对高中生的微流体、聚合物水凝胶、巨核细胞和血小板的教育模块，包括为Oncofertility Saturday Academy开设两个半天课程，并在当地高中进行演示。CBET部门的生物工程项目由材料研究部门的生物材料项目共同赞助。

1265029 - MillerGenerating many units of 300-600 billion platelets from megakaryocytic cells in culture will require advances in hematopoietic (blood) stem and progenitor cell expansion, megakaryocyte production and maturation, and platelet release and recovery. Shear from blood flow in the sinuses is important for proplatelet and platelet release. Inspired by the bone marrow sinusoid niche, this project will employ a microbioreactor with two compartments separated by a porous support coated with microporous poly(ethylene glycol) hydrogel functionalized with adhesive ligands. To mimic proplatelet formation across the sinusoidal endothelium, megakaryocytes will be seeded on the hydrogel in the upper compartment and medium will be perfused through the lower compartment. The microbioreactor will be used to investigate effects of shear stress, hydrogel stiffness, and adhesive ligand types and density on proplatelet and platelet production. Increasing gradients of pH and chemokines will be investigated to further increase proplatelet formation. The perfusion system will allow platelets to be harvested as they are released, which is important because platelet quality deteriorates during storage at 20-24°C and even faster at 37°C. The quality of culture-derived platelets will be verified via agonist-induced activation. The microbioreactor could be scaled for clinical production of platelets using formats such as hollow-fiber bioreactors and spiral-wound membrane cartridges to generate the required surface area. Several million units of platelets are transfused each year in the United States and Europe. Production of platelets in culture by megakaryocytes derived from patient or matched blood stem cells would increase the supply and decrease the contamination risk and adverse immune responses, but platelet production in culture is currently very inefficient. Successful completion of the proposed research will bring the large-scale production of culture-derived platelets closer to fruition. Graduate students and undergraduates working on this project will be trained in an interdisciplinary environment. A key element of the project is for the graduate students to develop educational modules on microfluidics, polymer hydrogels, megakaryocytic cells, and platelets for high school students, including two half-day classes for the Oncofertility Saturday Academy and presentation at local high schools.The award from the Biotechnology, Biochemical, and Bioengineering Program of the CBET Division is co-sponsored by the Biomaterials Program of the Division of Materials Research.