Laboratorio de Fisiopatogenia, Departamento de Ciencias Fisiológicas, Instituto de Fisiología y Biofísica (IFIBIO) Houssay, Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina.
Laboratorio de Biología de la Reproducción, Departamento de Ciencias Fisiológicas, Instituto de Fisiología y Biofísica (IFIBIO) Houssay, Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina.
Reproduction. 2019 Mar;157(3):297-304. doi: 10.1530/REP-18-0581.
Shiga toxin (Stx2) producing Escherichia coli infections during early gestation may impair placentation through a Stx2 damage of extravillous trophoblast (EVT) cells. We have previously demonstrated that Stx2 injected in rats in the early stage of pregnancy causes spontaneous abortion by a direct cytotoxic effect in the highly perfused feto-uteroplacental unit. The main aim was to evaluate the effects of Stx2 on EVT in order to understand the possible adverse effects that the toxin may have on trophoblast cells during early pregnancy. Swan 71 and HTR-8 cell lines were used as human EVT models. The presence of Stx2 receptor, globotriaosylceramide (Gb3), on Swan 71 and HTR-8 cells was evaluated by thin layer chromatography. The effects of Stx2 on cell viability were evaluated by neutral red uptake, migration by wound-healing assay and invasion was determined by the 'transwell chamber' assay. Metalloproteinase activity (MMP-2) was evaluated by zymography and tubulogenesis was analyzed by counting the total tube length and the number of branch formation. We have demonstrated that Swan 71 expresses high levels of Gb3 compared to HTR-8 cells. Stx2 decreased significantly Swan 71 viability in a dose-dependent manner after 72 h of toxin exposure. Furthermore, Stx2 impaired migration, invasion and tube-like formation of Swan 71 cells and decreased the MMP-2 activity. These cytotoxic effects were partially prevented by aminoguanidine, an inducible nitric oxide synthase inhibitor. These studies demonstrate that the function and viability of EVT cells may be altered by Stx2 and suggest that NO overexpression may be involved in the detrimental effects.
志贺毒素（Stx2）产生的大肠埃希菌感染在妊娠早期可能通过 Stx2 对绒毛外滋养层（EVT）细胞的损伤来损害胎盘形成。我们之前已经证明，在妊娠早期向大鼠体内注射 Stx2 会通过高度灌注的胎-胎盘单位中的直接细胞毒性作用导致自然流产。主要目的是评估 Stx2 对 EVT 的影响，以了解该毒素在妊娠早期可能对滋养层细胞产生的潜在不利影响。Swan 71 和 HTR-8 细胞系被用作人类 EVT 模型。通过薄层色谱法评估 Swan 71 和 HTR-8 细胞上 Stx2 受体神经节苷脂（Gb3）的存在。通过中性红摄取评估 Stx2 对细胞活力的影响，通过划痕愈合测定评估迁移，通过“transwell 室”测定评估侵袭。通过明胶酶谱法评估金属蛋白酶活性（MMP-2），并通过计数总管长度和分支形成数来分析管形成。我们已经证明，与 HTR-8 细胞相比，Swan 71 表达高水平的 Gb3。Stx2 在毒素暴露 72 小时后以剂量依赖性方式显着降低 Swan 71 的活力。此外，Stx2 损害了 Swan 71 细胞的迁移、侵袭和管状形成，并降低了 MMP-2 活性。诱导型一氧化氮合酶抑制剂氨基胍部分预防了这些细胞毒性作用。这些研究表明，EVT 细胞的功能和活力可能会被 Stx2 改变，并表明 NO 过表达可能参与了有害作用。
