3D NICHE ENGINEERING FOR REGULATION OF STEM CELL FATE DECISIONS

骨髓中储存着造血干细胞(HSC)，这些干细胞负责在人的一生中不断再生血液组织。骨髓移植，也被称为造血干细胞移植(HSCT)，是用于治疗白血病患者和最近多发性硬化症等自身免疫性疾病的挽救生命的手术，可以导致持久的无病缓解。然而，HSCT受到供应有限、组织不匹配并发症和高死亡率(高达10%的患者无法在移植中存活)的困扰。如果每个捐献者样本中的HSC数量能够增加，更多的患者可以从更安全的HSCT中受益。为了实现这一进展，需要更详细地了解HSC与其原始3D骨髓环境或利基环境之间的相互作用。 当HSC和HSPC细胞从原生骨髓中移出并置于培养环境中时，它们很快就会失去干细胞功能(24-72小时)。越来越多的证据表明，骨髓细胞外基质(ECM)是一个关键的，但没有得到充分认识的利基成分，可能对干细胞调节有指导意义。为了使HSPC在体外扩增，需要发展概括本地ECM环境的培养条件。该计划旨在开发发现体外干细胞生态位条件的技术，并应用这一知识来阐明干细胞与细胞外基质的相互作用。将采用系统地组装和测试单个ECM组件以及ECM组件组合的组合化学方法，目的是阐明每个组件对HSPC功能的贡献。这些发现将在一种新的3D筛选试验中进一步实施，该试验使用能够识别调节干细胞功能的分子靶点的“非专利”药物。最后，将开发3D打印技术，以创建带有ECM的血管系统，并支持HSPC在厚厚的3D结构中存活。 这一计划导致了用于个性化造血干细胞移植药物的组织匹配工程骨髓组织的开发。加拿大白血病和自身免疫性疾病患者的等待时间更短，程序更安全，将受益最大。加拿大的医疗节省还可以通过加快患者的治疗和康复来实现，从而降低管理病人和病情恶化的患者的医疗成本。此外，在批准研究用新药之前，这项研究可以用于测试无意中导致骨髓抑制毒性的新药，这是一种潜在的致命副作用。这项额外的测试需要获得监管部门的批准，可以减少动物试验，推广更安全的新闻药物。 在该计划的头5年内，共有3名博士、3名硕士和10名本科生研究助理将接受干细胞和组织工程方面的培训，并为他们在再生医学和生物技术等高影响力领域的学术或行业就业机会做好准备。

Bone marrow houses hematopoietic stem cells (HSC) that are responsible for continuously regenerating blood tissue throughout one's lifetime. Bone marrow transplants, also called HSC transplants (HSCT), are life-saving procedures used to treat leukemia patients and more recently, autoimmune diseases like multiple sclerosis, which can lead to durable disease-free remission. However, HSCT are plagued by limited supply, tissue mismatching complications and high mortality rates (up to 10% of patients do not survive the transplant). Many more patients could benefit from safer HSCT if the number of HSC in each donor sample could be augmented. For this advancement to be engineered a more detailed understanding of the interplay between HSC and their native 3D bone marrow environment, or niche, is required. When removed from their native bone marrow niche and placed in culture, HSC and progenitor (HSPC) cells very quickly lose their stem cell function (24-72h). Mounting evidence suggests that the bone marrow extracellular matrix (ECM) is a critical, yet underappreciated, component of the niche that may be instructive to stem cell regulation. In order for HSPCs to be expanded ex vivo, culture conditions that recapitulate the native ECM environment need to be developed. This program aims to develop technology that discovers ex vivo stem cell niche conditions and apply this knowledge to elucidate stem cell-ECM interactions. Combinatorial chemistry methods that systematically assemble and test individual, and combinations of ECM components will be employed with the intent of elucidating the contribution of each component to HSPC function. These findings will further be implemented in a novel 3D screening assay using “off-patent” drugs able to identify molecular targets that regulate stem cell function. Finally, 3D Printing technology will be developed to create a vascular system with the ECM and support HSPC survival in thick 3D constructs. This program leads to the development of tissue-matched engineered bone marrow tissue for personalized HSCT medicine. Canadian leukemia and autoimmune disease patients stand to benefit the most with shorter waiting times and safer procedures. Canadian healthcare savings can also be realized by accelerating patient treatment and recovery, thereby reducing healthcare costs of managing sick and deteriorating patients. Furthermore, this research can be applied to test new drugs that unintentionally cause myelosuppression toxicity, a potentially lethal side-effect, before approving investigational new drugs. This additional test for regulatory approval can reduce animal testing and promote safer news drugs. Within the program's first 5 years, a total of 3 PhDs, 3 MASc and 10 undergraduate research assistants will be will be trained in stem cells and tissue engineering and position them for academic or industry employment opportunities in high-impact areas such as regenerative medicine and biotechnology.