Mi Xiangquan, Zhou Xin, Zhu Shaochun, Mateus Andre, Backman Ludvig J, Danielson Patrik
Medical College, Shihezi University, Shihezi, China.
Department of Medical and Translational Biology, Umeå University, Umeå, Sweden.
FASEB J. 2025 Jun 30;39(12):e70740. doi: 10.1096/fj.202500502R.
The purpose of this study was to examine the nature of the underlying molecular mechanisms of cell death in human keratocytes treated with nigericin, a known pyroptosis inducer. Human keratocytes were exposed to nigericin, and cell death was assessed through morphological analysis and detection of related molecular markers. Proteomic profiling was performed to identify cell death-related proteins, with key findings validated by western blot. Additionally, organelle disruptions were examined using immunostaining techniques. Pyroptosis-like cell death was observed morphologically in cultured keratocytes. Moreover, an elevated release of IL-1beta was detected, accompanied by a significant loss of mitochondrial membrane potential. However, nigericin treatment induced a form of non-inflammatory cell death characterized by extensive vacuolation, resembling paraptosis. This was accompanied by the absence of caspase-3 activation and endoplasmic reticulum (ER) stress markers, along with increased accumulation of the autophagic marker LC3-II. Proteomic analysis revealed the absence of key components of the canonical pyroptosis pathway, including proteins involved in inflammasome assembly and the gasdermin (GSDM) family. These results were further confirmed by western blot. Significant alterations were also observed in the Golgi apparatus, mitochondria, and lysosomes following nigericin treatment. These findings suggest that nigericin triggers a paraptosis-like cell death in human keratocytes, rather than pyroptosis, as keratocytes lack the canonical executors of pyroptosis. This highlights an alternative mechanism of cell death in the cornea, warranting further exploration to understand its role and potential therapeutic implications.
本研究的目的是探讨用尼日利亚菌素（一种已知的焦亡诱导剂）处理人角膜细胞后细胞死亡的潜在分子机制的本质。将人角膜细胞暴露于尼日利亚菌素中，并通过形态学分析和相关分子标志物的检测来评估细胞死亡情况。进行蛋白质组分析以鉴定与细胞死亡相关的蛋白质，关键发现通过蛋白质印迹法进行验证。此外，使用免疫染色技术检查细胞器的破坏情况。在培养的角膜细胞中形态学上观察到类似焦亡的细胞死亡。此外，检测到白细胞介素 -1β 的释放增加，同时伴有线粒体膜电位的显著丧失。然而，尼日利亚菌素处理诱导了一种以广泛空泡化为特征的非炎症性细胞死亡，类似于副凋亡。这伴随着半胱天冬酶 -3 激活和内质网（ER）应激标志物的缺失，以及自噬标志物 LC3-II 的积累增加。蛋白质组分析显示经典焦亡途径的关键成分缺失，包括参与炎性小体组装的蛋白质和gasdermin（GSDM）家族。这些结果通过蛋白质印迹法进一步得到证实。在尼日利亚菌素处理后，高尔基体、线粒体和溶酶体也观察到显著变化。这些发现表明，尼日利亚菌素在人角膜细胞中触发了类似副凋亡的细胞死亡，而不是焦亡，因为角膜细胞缺乏焦亡的经典执行者。这突出了角膜中细胞死亡的另一种机制，值得进一步探索以了解其作用和潜在的治疗意义。
