Brandi G, Fiorani M, Pierotti C, Albano A, Cattabeni F, Cantoni O
Instituto di Scienze Tossicologiche, Igienistiche e Ambientali, Università di Urbino, Italy.
Microbiol Immunol. 1989;33(12):991-1000. doi: 10.1111/j.1348-0421.1989.tb03157.x.
Escherichia coli cells challenged with low or high concentrations of hydrogen peroxide are killed via two different mechanisms and respond with morphological changes which are also dependent on the extracellular concentration of the oxidant. Treatment with low concentrations (less than 2.5 mM) of H2O2 is followed by an extensive cell filamentation which is dependent on the level of H2O2 or the time of exposure. In particular, addition of 1.75 mM H2O2 results in a growth lag of approximately 90 min followed by partial increase in optical density, which was mainly due to the onset of the filamentous response. In fact, microscopic analysis of the samples obtained from cultures incubated with the oxidant for various time intervals has revealed that this change in morphology becomes apparent after 90 min of exposure to H2O2 and that the length of the filaments gradually increases following longer time intervals. Analysis of the ability of these cells to form colonies has indicated a loss in viability in the first 90 min of exposure followed by a gradual recovery in the number of cells capable of forming colonies. Measurement of lactate dehydrogenase in culture medium (as a marker for membrane damage) has revealed that a small amount of this enzyme was released from the cells at early times (less than 150 min) but not after longer incubation periods (300 min). Cells exposed to high concentrations of H2O2 (greater than 10 mM) do not filament and their loss of viability is associated with a marked reduction in cell volume. In fact, treatment with 17.5 mM H2O2 resulted in a time-dependent decrease of the optical density, clonogenicity, and cellular volume. In addition, these effects were paralleled by a significant release in the culture medium of lactate dehydrogenase thus suggesting that the reduced cell volume may be dependent on membrane damage followed by loss of intracellular material. This hypothesis is supported by preliminary results obtained in electron microscopy studies. In conclusion, this study further demonstrates that the response of E. coli to hydrogen peroxide is highly dependent on the concentration of H2O2 and further stresses the point that low or high concentrations of the oxidant result in the production of different species leading to cell death via two different mechanisms and/or capable of specifically affecting the cell shape.
用低浓度或高浓度过氧化氢处理的大肠杆菌细胞通过两种不同机制被杀死，并伴随着形态变化，这些变化也取决于细胞外氧化剂的浓度。用低浓度（低于2.5 mM）的H₂O₂处理后，细胞会出现广泛的丝状化，这取决于H₂O₂的浓度或暴露时间。具体而言，添加1.75 mM H₂O₂会导致约90分钟的生长延迟，随后光密度部分增加，这主要是由于丝状化反应的开始。事实上，对在不同时间间隔与氧化剂一起培养的培养物中获得的样品进行显微镜分析表明，在暴露于H₂O₂ 90分钟后，这种形态变化变得明显，并且随着时间间隔延长，丝状体的长度逐渐增加。对这些细胞形成菌落能力的分析表明，在暴露的前90分钟内活力丧失，随后能够形成菌落的细胞数量逐渐恢复。测量培养基中的乳酸脱氢酶（作为膜损伤的标志物）表明，在早期（少于150分钟）有少量这种酶从细胞中释放出来，但在较长孵育期（300分钟）后则没有。暴露于高浓度H₂O₂（大于10 mM）的细胞不会出现丝状化，其活力丧失与细胞体积的显著减小有关。事实上，用17.5 mM H₂O₂处理导致光密度、克隆形成能力和细胞体积随时间下降。此外，这些效应伴随着培养基中乳酸脱氢酶的大量释放，因此表明细胞体积减小可能取决于膜损伤，随后细胞内物质流失。这一假设得到了电子显微镜研究初步结果的支持。总之，这项研究进一步证明大肠杆菌对过氧化氢的反应高度依赖于H₂O₂的浓度，并进一步强调了低浓度或高浓度氧化剂通过两种不同机制导致不同种类物质产生从而导致细胞死亡和/或能够特异性影响细胞形状这一点。
