Xiang Li-Xin, Peng Bo, Dong Wei-Ren, Yang Zai-Feng, Shao Jian-Zhong
College of Life Sciences, Zhejiang University, Hangzhou, PR China.
Dev Comp Immunol. 2008;32(8):992-1001. doi: 10.1016/j.dci.2008.01.009. Epub 2008 Feb 20.
Lipopolysaccharide (LPS), the endotoxin of Gram-negative bacteria, is capable of eliciting a wide variety of pathophysiological effects, including endotoxin shock, tissue injury and lethality in both humans and animals. It is also a potent stimulant to initiate the proliferation, differentiation and activation of B lymphocytes and macrophages, resulting in changes of inflammatory cytokines, such as TNF-alpha, IL1-beta, IL6, IL-8 and IL-12, and enhancement of immune responses. However, little is known about its effect on the induction of apoptosis in lymphocytes. In the present study, the lymphocytes from Carassius auratus were employed for this purpose. The cells were exposed to LPS at various doses for different time periods. By careful apoptotic characteristic analysis, such as condensation of nuclear chromatin, fragmentation of genomic DNA and formation of apoptotic bodies, it provided the first evidence that LPS had apoptotic-inducing effect on fish lymphocytes in a time- and dose-dependent manner. LPS exposure induced significant increase of intracellular reactive oxygen species (ROS), loss of mitochondrial transmembrane potential (DeltaPsi), depletion of ATP production, down-regulation of Bcl-2 expression, up-regulation of Bax and mitochondrial NO-synthase (mNOS) expression, and selective activation of caspase-9 rather than caspase-8. Each of these observations suggests that the LPS-induced apoptosis in C. auratus lymphocytes occurs largely via the mitochondrial apoptotic pathway. This observation was different from the mechanism behind the LPS-induced apoptosis in mammalian macrophages/thymocytes that occurs via the TNF-alpha-mediated death-receptor pathway. Our study suggested the existence of a possible novel role in the pathogenesis of Gram-negative bacterial infection in fish and even in mammals, which may contribute to the therapy of bacterial diseases. Also, it will help to gain more insights into the mechanisms of septic shock and of LPS-induced immunosuppression and autoimmunity.
脂多糖（LPS）是革兰氏阴性菌的内毒素，能够引发多种病理生理效应，包括内毒素休克、组织损伤以及人和动物的致死性。它也是启动B淋巴细胞和巨噬细胞增殖、分化及激活的强效刺激物，导致炎性细胞因子如肿瘤坏死因子-α（TNF-α）、白细胞介素1-β（IL1-β）、白细胞介素6（IL6）、白细胞介素-8（IL-8）和白细胞介素-12（IL-12）发生变化，并增强免疫反应。然而，关于其对淋巴细胞凋亡诱导作用的了解甚少。在本研究中，为此选用了鲫鱼的淋巴细胞。将细胞暴露于不同剂量的LPS中不同时间段。通过仔细的凋亡特征分析，如核染色质凝聚、基因组DNA片段化和凋亡小体形成，首次证明LPS对鱼类淋巴细胞具有时间和剂量依赖性的凋亡诱导作用。LPS暴露导致细胞内活性氧（ROS）显著增加、线粒体跨膜电位（ΔΨ）丧失、ATP生成减少、Bcl-2表达下调、Bax和线粒体一氧化氮合酶（mNOS）表达上调，以及半胱天冬酶-9（caspase-9）而非半胱天冬酶-8（caspase-8）的选择性激活。这些观察结果均表明，LPS诱导鲫鱼淋巴细胞凋亡主要通过线粒体凋亡途径发生。这一观察结果与LPS诱导哺乳动物巨噬细胞/胸腺细胞凋亡通过TNF-α介导的死亡受体途径的机制不同。我们的研究表明，在鱼类甚至哺乳动物革兰氏阴性菌感染的发病机制中可能存在一种新的作用，这可能有助于细菌性疾病的治疗。此外，它将有助于更深入地了解脓毒症休克以及LPS诱导的免疫抑制和自身免疫的机制。
