The distinctive molecular structure of the prion protein, PrPsc, is established only in mammals with infectious prion diseases. Prion protein characterizes either the transmissible pathogen itself or a primary constituent of the disease. Our report suggested that prion protein-mediated neuronal cell death is triggered by the autophagy flux. However, the alteration of intracellular calcium levels, AMPK activity in prion models has not been described. This study is focused on the effect of the changes in intracellular calcium levels on AMPK/autophagy flux pathway and PrP (106–126)-induced neurotoxicity. Western blot and Immunocytochemistry was used to detect AMPK and autophagy-related protein expression. Flow cytometry and a TdT-mediated biotin-16-dUTP nick-end labeling (TUNEL) assay were used to detect the percentage of apoptotic cells. Calcium measurement was employed using fluo-4 by confocal microscope. We examined the effect of calcium homeostasis alterations induced by human prion peptide on the autophagy flux in neuronal cells. Treatment with human prion peptide increased the intracellular calcium concentration and induced cell death in primary neurons as well as in a neuronal cell line. Using pharmacological inhibitors, we showed that the L-type calcium channel is involved in the cellular entry of calcium ions. Inhibition of calcium uptake prevented autophagic cell death and reduction in AMP-activated protein kinase (AMPK) activity induced by human prion peptide. Our data demonstrated that prion peptide-mediated calcium inflow plays a pivotal role in prion peptide-induced autophagic cell death, and reduction in AMPK activity in neurons. Altogether, our results suggest that calcium influx might play a critical role in neurodegenerative diseases, including prion diseases.

中文翻译：

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朊病毒肽介导的钙水平改变通过 AMPK-自噬通量控制神经元细胞损伤。

朊病毒蛋白 PrPsc 的独特分子结构仅在患有传染性朊病毒疾病的哺乳动物中建立。朊病毒蛋白表征可传播病原体本身或疾病的主要成分。我们的报告表明朊病毒蛋白介导的神经元细胞死亡是由自噬通量引发的。然而，尚未描述朊病毒模型中细胞内钙水平、AMPK 活性的改变。这项研究的重点是细胞内钙水平的变化对 AMPK/自噬通量通路和 PrP（106-126）诱导的神经毒性的影响。Western印迹和免疫细胞化学用于检测AMPK和自噬相关蛋白的表达。流式细胞术和 TdT 介导的生物素-16-dUTP 缺口末端标记 (TUNEL) 测定用于检测凋亡细胞的百分比。通过共聚焦显微镜使用fluo-4进行钙测量。我们检查了人朊病毒肽诱导的钙稳态改变对神经元细胞自噬通量的影响。用人朊病毒肽处理增加了原代神经元以及神经元细胞系的细胞内钙浓度并诱导细胞死亡。使用药理抑制剂，我们发现 L 型钙通道参与钙离子的细胞进入。抑制钙摄取可防止自噬细胞死亡并降低由人类朊病毒肽诱导的 AMP 活化蛋白激酶 (AMPK) 活性。我们的数据表明朊病毒肽介导的钙流入在朊病毒肽诱导的自噬细胞死亡和神经元中 AMPK 活性的降低中起着关键作用。共，