Role of calcium and reactive oxygen species in toxicant-induced immune failure

细胞内钙(Ca~(2+))和活性氧(ROS)动态平衡的控制对细胞至关重要 生存和功能。细胞内游离钙和ROS水平升高在淋巴细胞中起中心作用 对抗原刺激作出反应的激活。ROS和Ca~(2+)相互作用相互调节： 外源ROS诱导细胞内钙动员，增加细胞内钙离子 浓度诱导淋巴细胞产生ROS。在多环芳烃(PAH)诱导的 淋巴细胞免疫抑制，增强的ROS产生与基础水平的增加有关 细胞内钙离子。此外，这些高基础水平的细胞内钙离子不会进一步增加 对抗原受体刺激的反应。因此，我假设钙离子和钙离子之间的串扰 ROS对淋巴细胞的激活至关重要，这种串扰的破坏导致了异常的抗原 PAH免疫抑制中观察到的受体信号通路。因此，我计划调查 正常B细胞受体刺激过程中Ca~(2+)和ROS信号之间的串扰及其干扰 在PAH免疫抑制期间的这种串扰。具体来说，我会追求三个具体目标：(一) 利用电生理学和钙离子成像，确定钙离子的药理和生物物理性质 B细胞中由外源性ROS激活的通道，并决定是否有 其中涉及TRPC阳离子通道家族。根据初步实验，TRPC3和TRPC5 在HEK293细胞中表达时形成ROS敏感的阳离子通道；(2)确定ROS的作用 细胞内钙离子促进B淋巴细胞产生ROS及其膜结合的NADPH氧化酶 有丝分裂原诱导B细胞产生ROS所必需的；(3)检测细胞内ROS的慢性增加 PAH诱导的Ca~(2+)由ROS依赖的机制和/或膜去极化机制维持。 初步数据显示，PAH和ROS均能诱导B细胞内钙内流。我期待着一段牢固的关系 在PAH诱导的免疫衰竭中ROS和钙信号缺陷之间的关系。这一努力将有助于 更好地了解正常和病理的淋巴细胞免疫以及新的发现 治疗免疫缺陷和癌症等疾病。

Control of intracellular calcium (Ca2+) and reactive oxygen species (ROS) homeostasis is essential for cell survival and function. Elevation of intracellular free Ca2+ and ROS levels play a central role in lymphocyte activation in response to antigenic stimulation. ROS and Ca2+ exert a reciprocal regulation on each other: exogenous application of ROS leads to intracellular Ca2+ mobilization while increased cytoplasmic Ca2+ concentration induces ROS production by lymphocytes. In polycyclic aromatic hydrocarbons (PAH)-induced lymphocyte immune suppression, enhanced ROS production is associated with increased basal levels of intracellular Ca2+. Furthermore, these high basal levels of intracellular Ca2+ do not increase further in response to stimulation of the antigen receptor. Thus, I hypothesize that the crosstalk between Ca2+ and ROS is vital for lymphocyte activation and that the disruption of this crosstalk contributes to abnormal antigen receptor signaling pathways observed in immune suppression by PAH. Therefore, I plan to investigate the crosstalk between Ca2* and ROS signaling during normal B-cell receptor stimulation as well as the disruption of this crosstalk during immune suppression by PAH. Specifically, I will pursue three specific aims: (1) to define, using electrophysiology and Ca2+ imaging, the pharmacological and biophysical properties of the Ca2+ channels in B-cells that are activated by exogenous application of ROS, and determine whether members of the TRPC family of cation channels are involved. Based on preliminary experiments, TRPC3 and TRPC5 form ROS-sensitive cation channels when expressed in HEK293 cells; (2) to determine the role of intracellular Ca2+ rise in ROS production in B lymphocytes and which membrane-bound NADPH oxidase is necessary for mitogen-induced ROS production in B-cells; (3) to test if the chronic increase of intracellular Ca2+ induced by PAH is maintained by a ROS-dependent mechanism and/or by membrane depolarization. Preliminary data shows that both PAHand ROSinduce Ca2+ entry in B-cells. I excpect a strong relationship between ROS and Ca2+ signaling defects in PAH-induced immune failure. This effort will contribute to both a better understanding of normal and pathological lymphocyte immunity as well as to the discovery of new therapies for diseases such as immunodeficiency and cancer.