Engineered cardiac tissue microsphere production through direct differentiation of hydrogel-encapsulated human pluripotent stem cells
Engineered cardiac tissue microsphere production through direct differentiation of hydrogel-encapsulated human pluripotent stem cells
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DOI：
10.1016/j.biomaterials.2021.120818
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发表时间：
2021-05-21
期刊：
影响因子：
14
通讯作者：
Lipke, Elizabeth A.
中科院分区：
文献类型：
--
作者：
Finklea, Ferdous B.;Tian, Yuan;Lipke, Elizabeth A.
人类诱导多能干细胞（hiPSCs）可在可扩展的悬浮培养系统中直接产生工程化心脏组织，以满足心脏再生医学的需求。在这里，我们展示了通过水凝胶包膜hipsc的直接分化成功生产功能性心脏组织微球。为了形成微球，hiPSCs悬浮在可光交联的生物材料peg -纤维蛋白原（2500万个细胞/mL）中，并使用定制的微流体系统以42万个细胞/分钟的速度封装。即使在这种高细胞密度和快速生产速率下，也实现了高批内和批间重复性。微球形成后，hiPSCs保持高细胞活力，并在原始peg纤维蛋白原基质内外继续生长。这些最初的软微球（250 Pa）支持有效的心脏分化；分化第8天开始自发收缩，微球中含有75%的心肌细胞（CMs）。CMs对药理学刺激有适当的反应，并且在电节奏时表现出高达6.0 Hz的1:1捕获。随着时间的推移，细胞形成细胞间连接并排列肌原纤维；工程心脏微球在培养中保存3年以上。快速生成均匀心脏微球组织的能力对于推进下游应用至关重要，包括生物制造、多孔板药物筛选和注射再生治疗。
Engineered cardiac tissues that can be directly produced from human induced pluripotent stem cells (hiPSCs) in scalable, suspension culture systems are needed to meet the demands of cardiac regenerative medicine. Here, we demonstrate successful production of functional cardiac tissue microspheres through direct differentiation of hydrogel encapsulated hiPSCs. To form the microspheres, hiPSCs were suspended within the photocrosslinkable biomaterial, PEG-fibrinogen (25 million cells/mL), and encapsulated at a rate of 420,000 cells/minute using a custom microfluidic system. Even at this high cell density and rapid production rate, high intra-batch and batchto-batch reproducibility was achieved. Following microsphere formation, hiPSCs maintained high cell viability and continued to grow within and beyond the original PEG-fibrinogen matrix. These initially soft microspheres ( 250 Pa) supported efficient cardiac differentiation; spontaneous contractions initiated by differentiation day 8, and the microspheres contained 75% cardiomyocytes (CMs). CMs responded appropriately to pharmacological stimuli and exhibited 1:1 capture up to 6.0 Hz when electrically paced. Over time, cells formed cell-cell junctions and aligned myofibril fibers; engineered cardiac microspheres were maintained in culture over 3 years. The capability to rapidly generate uniform cardiac microsphere tissues is critical for advancing downstream applications including biomanufacturing, multi-well plate drug screening, and injection-based regenerative therapies.