Bergan Jonas, Skotland Tore, Lingelem Anne Berit Dyve, Simm Roger, Spilsberg Bjørn, Lindbäck Toril, Sylvänne Tuulia, Simolin Helena, Ekroos Kim, Sandvig Kirsten
Centre for Cancer Biomedicine, Faculty of Medicine, University of Oslo, Oslo, Norway.
Cell Mol Life Sci. 2014 Nov;71(21):4285-300. doi: 10.1007/s00018-014-1624-1. Epub 2014 Apr 18.
Shiga toxin-producing Escherichia coli bacteria cause hemorrhagic colitis and hemolytic uremic syndrome in humans. Currently, only supportive treatment is available for diagnosed patients. We show here that 24-h pretreatment with an ether lipid precursor, the alkylglycerol sn-1-O-hexadecylglycerol (HG), protects HEp-2 cells against Shiga toxin and Shiga toxin 2. Also the endothelial cell lines HMEC-1 and HBMEC are protected against Shiga toxins after HG pretreatment. In contrast, the corresponding acylglycerol, DL-α-palmitin, has no effect on Shiga toxicity. Although HG treatment provides a strong protection (~30 times higher IC₅₀) against Shiga toxin, only a moderate reduction in toxin binding was observed, suggesting that retrograde transport of the toxin from the plasma membrane to the cytosol is perturbed. Furthermore, endocytosis of Shiga toxin and retrograde sorting from endosomes to the Golgi apparatus remain intact, but transport from the Golgi to the endoplasmic reticulum is inhibited by HG treatment. As previously described, HG reduces the total level of all quantified glycosphingolipids to 50-70% of control, including the Shiga toxin receptor globotriaosylceramide (Gb3), in HEp-2 cells. In accordance with this, we find that interfering with Gb3 biosynthesis by siRNA-mediated knockdown of Gb3 synthase for 24 h causes a similar cytotoxic protection and only a moderate reduction in toxin binding (to 70% of control cells). Alkylglycerols, including HG, have been administered to humans for investigation of therapeutic roles in disorders where ether lipid biosynthesis is deficient, as well as in cancer therapy. Further studies may reveal if HG can also have a therapeutic potential in Shiga toxin-producing E. coli infections.
产志贺毒素大肠杆菌可导致人类患出血性结肠炎和溶血尿毒综合征。目前，对于确诊患者仅能进行支持性治疗。我们在此表明，用醚脂前体烷基甘油sn-1-O-十六烷基甘油（HG）进行24小时预处理，可保护人喉表皮样癌细胞（HEp-2细胞）免受志贺毒素和志贺毒素2的侵害。同样，内皮细胞系人微血管内皮细胞-1（HMEC-1）和人脑微血管内皮细胞（HBMEC）在HG预处理后也能免受志贺毒素的侵害。相比之下，相应的酰基甘油DL-α-软脂酸甘油酯对志贺毒素毒性没有影响。尽管HG处理对志贺毒素提供了强大的保护作用（半数抑制浓度[IC₅₀]约高30倍），但仅观察到毒素结合有适度降低，这表明毒素从质膜向胞质溶胶的逆行转运受到了干扰。此外，志贺毒素的内吞作用以及从内体到高尔基体的逆行分选保持完整，但HG处理会抑制从高尔基体到内质网的转运。如先前所述，HG可将人喉表皮样癌细胞中所有定量的糖鞘脂总水平降低至对照水平的50% - 70%，包括志贺毒素受体三糖基神经酰胺（Gb3）。据此，我们发现通过小干扰RNA（siRNA）介导的Gb3合酶敲低24小时来干扰Gb3生物合成，会产生类似的细胞毒性保护作用，且毒素结合仅适度降低（降至对照细胞的70%）。包括HG在内的烷基甘油已用于人体，以研究其在醚脂生物合成缺陷的疾病以及癌症治疗中的治疗作用。进一步的研究可能会揭示HG在产志贺毒素大肠杆菌感染中是否也具有治疗潜力。
