Traini Leonardo, Negueruela Javier, Elvira Bernat, St-Pierre-Wijckmans Wadsen, Vandenbempt Valerie, Buss Carlos E, Li Ao, Pérez-Chávez Israel, Ribeiro-Costa Francisco, Nunes Mariana, Messens Joris, Ezeriņa Daria, Hay David C, Bansal Mayank, Gurzov Esteban N
Signal Transduction and Metabolism Laboratory, Université Libre de Bruxelles, Route de Lennik 808, B-1070, Brussels, Belgium.
VIB-VUB Center for Structural Biology, Vlaams Instituut Voor Biotechnologie, B-1050, Brussels, Belgium.
Stem Cell Res Ther. 2025 May 4;16(1):225. doi: 10.1186/s13287-025-04314-5.
Thioredoxin-interacting protein (TXNIP) plays a role in regulating endoplasmic reticulum (ER) and oxidative stress, which disrupt glucose homeostasis in diabetes. However, the impact of TXNIP deficiency on the differentiation and functionality of human stem cell-derived somatic metabolic cells remains unclear.
We used CRISPR-Cas12a genome editing to generate TXNIP-deficient (TXNIP) H1 human embryonic stem cells (H1-hESCs). These cells were differentiated into hepatocyte-like cells (HLCs) and stem-cell-derived insulin-producing islets (SC-islets). The maturation and functionality TXNIP and TXNIP SC-islets were assessed by implantation under the kidney capsule of male or female NOD-SCID mice.
TXNIP deficiency significantly increased H1-hESC proliferation without affecting pluripotency, viability, or differentiation potential into HLCs and SC-islets. Bulk RNA-sequencing of thapsigargin-treated TXNIP and TXNIP hESCs revealed differential expression of stress-responsive genes, with enriched apoptosis-related pathways in TXNIP cells, but minimal transcriptional changes specific to TXNIP deficiency. In HLCs, TXNIP deletion reduced albumin secretion and insulin signalling, as indicated by decreased AKT phosphorylation, while showing no differences in glycolytic activity or lipid metabolism markers. Under thapsigargin-induced ER stress, TXNIP HLCs exhibited transiently reduced eIF2α phosphorylation and lower BiP expression, suggesting compromised adaptive responses to prolonged stress. SC-islets derived from TXNIP hESCs showed comparable viability, endocrine cell composition, and cytokine responses to TXNIP islets. Following IFNα or IFNγ treatment, STAT1 phosphorylation was increased in TXNIP SC-islets, indicating that IFN signalling remained intact despite TXNIP deficiency. Upon implantation into NOD-SCID mice, both TXNIP and TXNIP SC-islets produced human C-peptide and responded to glucose stimulation. However, TXNIP SC-islets did not demonstrate enhanced glycaemic control or glucose-stimulated insulin secretion compared to controls.
Our study demonstrates that TXNIP deficiency does not improve the differentiation or functionality of HLCs and SC-islets. We present the generation and characterisation of TXNIP and TXNIP H1-hESCs, HLCs, and SC-islets as valuable models for future studies on the role of TXNIP in metabolic cell biology.
硫氧还蛋白相互作用蛋白（TXNIP）在调节内质网（ER）和氧化应激中发挥作用，而内质网和氧化应激会破坏糖尿病患者的葡萄糖稳态。然而，TXNIP缺乏对人干细胞来源的体细胞代谢细胞的分化和功能的影响仍不清楚。
我们使用CRISPR-Cas12a基因组编辑技术生成了TXNIP缺陷型（TXNIP）H1人胚胎干细胞（H1-hESCs）。这些细胞被分化为肝细胞样细胞（HLCs）和干细胞来源的胰岛素分泌胰岛（SC-胰岛）。通过将其植入雄性或雌性NOD-SCID小鼠的肾包膜下，评估TXNIP和TXNIP SC-胰岛的成熟度和功能。
TXNIP缺乏显著增加了H1-hESC的增殖，而不影响其多能性、活力或向HLCs和SC-胰岛的分化潜能。对毒胡萝卜素处理的TXNIP和TXNIP hESCs进行的RNA测序显示，应激反应基因存在差异表达，TXNIP细胞中凋亡相关途径富集，但TXNIP缺乏特异性的转录变化最小。在HLCs中，TXNIP缺失减少了白蛋白分泌和胰岛素信号传导，AKT磷酸化降低表明了这一点，而在糖酵解活性或脂质代谢标志物方面没有差异。在毒胡萝卜素诱导的内质网应激下，TXNIP HLCs表现出eIF2α磷酸化短暂降低和BiP表达降低，表明对长期应激的适应性反应受损。源自TXNIP hESCs的SC-胰岛在活力、内分泌细胞组成和细胞因子反应方面与TXNIP胰岛相当。在IFNα或IFNγ处理后，TXNIP SC-胰岛中STAT1磷酸化增加，表明尽管存在TXNIP缺乏，IFN信号传导仍然完整。植入NOD-SCID小鼠后，TXNIP和TXNIP SC-胰岛均产生人C肽并对葡萄糖刺激作出反应。然而，与对照组相比，TXNIP SC-胰岛并未表现出更好的血糖控制或葡萄糖刺激的胰岛素分泌。
我们的研究表明，TXNIP缺乏并不能改善HLCs和SC-胰岛的分化或功能。我们展示了TXNIP和TXNIP H1-hESCs、HLCs和SC-胰岛的生成和特性，作为未来研究TXNIP在代谢细胞生物学中作用的有价值模型。
