Department of Molecular and Biomedical Sciences, University of Maine, Orono, Maine.
Graduate School of Biomedical Science and Engineering, University of Maine, Orono, Maine.
J Appl Toxicol. 2019 Dec;39(12):1672-1690. doi: 10.1002/jat.3884. Epub 2019 Aug 19.
Humans are exposed to the antimicrobial agent triclosan (TCS) through use of TCS-containing products. Exposed tissues contain mast cells, which are involved in numerous biological functions and diseases by secreting various chemical mediators through a process termed degranulation. We previously demonstrated that TCS inhibits both Ca influx into antigen-stimulated mast cells and subsequent degranulation. To determine the mechanism linking the TCS cytosolic Ca depression to inhibited degranulation, we investigated the effects of TCS on crucial signaling enzymes activated downstream of the Ca rise: protein kinase C (PKC; activated by Ca and reactive oxygen species [ROS]) and phospholipase D (PLD). We found that TCS strongly inhibits PLD activity within 15 minutes post-antigen, a key mechanism of TCS mast cell inhibition. In addition, experiments using fluorescent constructs and confocal microscopy indicate that TCS delays antigen-induced translocations of PKCβII, PKCδ and PKC substrate myristoylated alanine-rich C-kinase. Surprisingly, TCS does not inhibit PKC activity or overall ability to translocate, and TCS actually increases PKC activity by 45 minutes post-antigen; these results are explained by the timing of both TCS inhibition of cytosolic Ca (15+ minutes post-antigen) and TCS stimulation of ROS (45 minutes post-antigen). These findings demonstrate that it is incorrect to assume that all Ca -dependent processes will be synchronously inhibited when cytosolic Ca is inhibited by a toxicant or drug. The results offer molecular predictions of the effects of TCS on other mammalian cell types, which share these crucial signal transduction elements and provide biochemical information that may underlie recent epidemiological findings implicating TCS in human health problems.
人类通过使用含有三氯生 (TCS) 的产品而接触到这种抗菌剂。暴露的组织中含有肥大细胞，这些细胞通过脱颗粒过程分泌各种化学介质，参与许多生物功能和疾病。我们之前证明 TCS 抑制抗原刺激的肥大细胞内 Ca 流入和随后的脱颗粒。为了确定将 TCS 细胞溶质 Ca 抑制与抑制脱颗粒联系起来的机制，我们研究了 TCS 对 Ca 上升后激活的关键信号酶的影响：蛋白激酶 C（PKC；由 Ca 和活性氧 (ROS) 激活）和磷脂酶 D (PLD)。我们发现 TCS 在抗原后 15 分钟内强烈抑制 PLD 活性，这是 TCS 抑制肥大细胞的关键机制。此外，使用荧光构建体和共聚焦显微镜的实验表明，TCS 延迟抗原诱导的 PKCβII、PKCδ 和 PKC 底物豆蔻酰化丙氨酸丰富 C 激酶的易位。令人惊讶的是，TCS 不抑制 PKC 活性或整体易位能力，并且 TCS 实际上在抗原后 45 分钟增加 PKC 活性；这些结果可以通过 TCS 抑制细胞溶质 Ca 的时间（抗原后 15 分钟+）和 TCS 刺激 ROS 的时间（抗原后 45 分钟）来解释。这些发现表明，当细胞溶质 Ca 被毒物或药物抑制时，不能假设所有依赖 Ca 的过程都会被同步抑制。这些结果为 TCS 对其他哺乳动物细胞类型的影响提供了分子预测，这些细胞类型共享这些关键信号转导元件，并提供可能为最近将 TCS 与人类健康问题联系起来的流行病学发现提供生化信息。
