Ramezankhani Roya, De Smedt Jonathan, Toprakhisar Burak, van der Veer Bernard K, Tricot Tine, Vanmarcke Gert, Balaton Bradley, van Grunsven Leo, Vosough Massoud, Chai Yoke Chin, Verfaillie Catherine
Stem Cell Institute, Department of Development and Regeneration, KU Leuven, O&N IV Herestraat 49, Leuven, 3000, Belgium.
Department of Applied Cell Sciences, Faculty of Basic Sciences and Advanced Medical Technologies, Royan Institute, Academic Center for Education, Culture and Research, Tehran, Iran.
Stem Cell Rev Rep. 2025 Apr;21(3):744-766. doi: 10.1007/s12015-025-10842-7. Epub 2025 Jan 24.
Reliable models of the blood-brain barrier (BBB), wherein brain microvascular endothelial cells (BMECs) play a key role in maintenance of barrier function, are essential tools for developing therapeutics and disease modeling. Recent studies explored generating BMEC-like cells from human pluripotent stem cells (hPSCs) by mimicking brain-microenvironment signals or genetic reprogramming. However, due to the lack of comprehensive transcriptional studies, the exact cellular identity of most of these cells remains poorly defined. In this study we aimed to identify the most likely master transcription factors (TFs) for inducing brain endothelial cell (EC) fate and assess the transcriptomic changes following their introduction into immature ECs. Therefore, we first generated PSC-derived immature ECs by transient overexpression of the TF, ETV2. Subsequently, by performing an extensive meta-analysis of transcriptome studies of brain and non-brain ECs, 12 candidate TFs were identified, which might fate immature ECs towards cells with brain EC features. Following combinatorial overexpression of these 12 TFs tagged with unique barcodes, single cell transcriptomics identified a subset of transduced cells that resembled mid-gestational human brain ECs. Assessment of the TF barcodes present in these cells revealed significant enrichment of the TFs ZIC3, TFAP2C, TFAP2A, and DLX2. These TFs might be useful to fate PSC-EC to BMEC-like cells, which could be incorporated in human in vitro BBB models.
血脑屏障（BBB）的可靠模型对于开发治疗方法和疾病模型至关重要，其中脑微血管内皮细胞（BMECs）在维持屏障功能中起关键作用。最近的研究探索了通过模拟脑微环境信号或基因重编程从人多能干细胞（hPSCs）中生成类BMEC细胞。然而，由于缺乏全面的转录研究，这些细胞中大多数的确切细胞身份仍不清楚。在本研究中，我们旨在确定诱导脑内皮细胞（EC）命运最可能的主转录因子（TFs），并评估将它们引入未成熟EC后转录组的变化。因此，我们首先通过短暂过表达TF ETV2生成了PSC来源的未成熟EC。随后，通过对脑EC和非脑EC的转录组研究进行广泛的荟萃分析，确定了12个候选TF，它们可能使未成熟EC向具有脑EC特征的细胞分化。在用独特条形码标记这12个TF进行组合过表达后，单细胞转录组学确定了一部分转导细胞类似于妊娠中期人脑EC。对这些细胞中存在的TF条形码的评估显示，TF ZIC3、TFAP2C、TFAP2A和DLX2显著富集。这些TF可能有助于将PSC-EC诱导为类BMEC细胞，可用于构建人体体外血脑屏障模型。
