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Abstract：
Inducible pluripotent stem cell-derived human β-like cells (BLCs) hold promise for both therapy and disease modeling, but their generation remains challenging and their functional analyses beyond transcriptomic and morphological assessments remain limited. Here, we validate an approach using multicellular and single-cell electrophysiological tools to evaluate function of BLCs from pioneer protocols that can be easily adapted to more differentiated BLCs. The multi-electrode arrays (MEAs) measuring the extracellular electrical activity revealed that BLCs, like primary β-cells, are electrically coupled and produce slow potential (SP) signals that are closely linked to insulin secretion. We also used high-resolution single-cell patch clamp measurements to capture the exocytotic properties, and characterize voltage-gated sodium and calcium currents, and found that they were comparable with those in primary β- and EndoC-βH1 cells. The KATP channel conductance is greater than in human primary β-cells, which may account for the limited glucose responsiveness observed with MEA. We used MEAs to study the impact of the type 2 diabetes-protective SLC30A8 allele (p.Lys34Serfs50*) and found that BLCs with this allele have stronger electrical coupling activity. Our data suggest that BLCs can be used to evaluate the functional impact of genetic variants on β-cell function and coupling.
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摘要：
iPSC衍生的人β样细胞（BLC）有望用于治疗和疾病建模，但是它们的产生仍然具有挑战性，并且它们在转录组学和形态学评估之外的功能分析仍然有限。这里,我们验证了一种使用多细胞和单细胞电生理工具评估先驱方案BLC功能的方法，该方法可以轻松适应更多分化的BLC。测量细胞外电活动的多电极阵列(MEAs)表明BLC是电耦合的,产生慢电位（SP）信号，如与胰岛素分泌密切相关的原代β细胞。我们还使用高分辨率单细胞膜片钳测量来捕获胞吐特性，并表征电压门控钠和钙电流，发现它们与原代β和EndoC-βH1细胞相当。KATP通道电导大于人原代β细胞，这可能是MEA观察到的有限葡萄糖反应性的原因。我们使用MEAs研究了2型糖尿病保护性SLC30A8等位基因的影响（p。Lys34Serfs*50），并发现具有该等位基因的BLC具有更强的电耦合活性。我们的数据表明，BLC可用于评估遗传变体对β细胞功能和偶联的功能影响。
