Adult hematopoietic stem cells (HSCs) produce the body’s full complement of blood and immune cells. They reside in specialized microenvironments, or niches, within the bone marrow. The perivascular niche near blood vessels is believed to help maintain primitive HSCs in an undifferentiated state but demonstration of this effect is difficult. In vivo studies make it challenging to determine the direct effect of the endosteal and perivascular niches as they can be in close proximity, and two-dimensional in vitro cultures often lack an instructive extracellular matrix environment. We describe a tissue engineering approach to develop and characterize a three-dimensional perivascular tissue model to investigate the influence of the perivascular secretome on HSC behavior. We generate 3D endothelial networks in methacrylamide-functionalized gelatin hydrogels using human umbilical vein endothelial cells (HUVECs) and mesenchymal stromal cells (MSCs). We identify a subset of secreted factors important for HSC function, and examine the response of primary murine HSCs in hydrogels to the perivascular secretome. Within 4 days of culture, perivascular conditioned media promoted maintenance of a greater fraction of hematopoietic stem and progenitor cells. This work represents an important first-generation perivascular model to investigate the role of niche secreted factors on the maintenance of primary HSCs.

成体造血干细胞（HSC）产生人体的全部血液和免疫细胞。它们存在于骨髓内特殊的微环境或生态位中。血管附近的血管周围生态位被认为有助于维持原始造血干细胞处于未分化状态，但证明这种作用很困难。体内研究使得确定骨内膜和血管周围微环境的直接影响具有挑战性，因为它们可能非常接近，并且二维体外培养物通常缺乏指导性的细胞外基质环境。我们描述了一种组织工程方法来开发和表征三维血管周围组织模型，以研究血管周围分泌组对 HSC 行为的影响。我们使用人脐静脉内皮细胞 (HUVEC) 和间充质基质细胞 (MSC) 在甲基丙烯酰胺功能化明胶水凝胶中生成 3D 内皮网络。我们确定了对 HSC 功能重要的分泌因子子集，并检查了水凝胶中原代小鼠 HSC 对血管周围分泌组的反应。在培养 4 天内，血管周围条件培养基促进了大部分造血干细胞和祖细胞的维持。这项工作代表了重要的第一代血管周围模型，用于研究生态位分泌因子对维持原代 HSC 的作用。