Karahashi H, Amano F
Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Tokyo, Japan.
Biol Pharm Bull. 1998 Oct;21(10):1102-5. doi: 10.1248/bpb.21.1102.
The structure-activity relationships of lipopolysaccharide (LPS) in tumor necrosis factor-alpha (TNF-alpha) production and induction of macrophage cell death in the presence of cycloheximide (CHX) were examined in a murine macrophage-like cell line, J774.1. TNF-alpha production is one of the characteristic phenotypes of LPS-activated macrophages, and is observed upon incubation with LPS. On the contrary, macrophage cell death, which had been found in our laboratory (Amano F., Karahashi H., J. Endotoxin Res., 3, 415423 (1996)) and was examined as the release of lactate dehydrogenase (LDH) from cells into the culture supernatant, was observed 2.5 h after the addition of LPS in the presence of CHX. However, both TNF-alpha production and macrophage cell death were similarly dependent on the structures of LPS and lipid A. At more than 10 ng/ml, wild-type LPS from E.coli and S. minnesota exhibited the highest activity, and LPS as well as diphosphoryl lipid A from S. minnesota rough mutants also exhibited activity, but E. coli LPS detoxified by alkaline treatment or monophosphoryl lipid A from S. minnesota exhibited no activity even at 100 ng/ml. These results suggest that LPS-induced macrophage cell death in the presence of CHX shows similar requirements for LPS and/or lipid A structures as for the macrophage activation at higher doses than 10 ng/ml, although the former was not observed at 1 ng/ml LPS, while the latter was. However, TNF-alpha does not seem to be involved in the induction of macrophage cell death, because a neutralizing anti-TNF-alpha antibody failed to block the macrophage cell death and because recombinant TNF-alpha had little effect on the extent of LDH release in the presence or absence of LPS and/or CHX, and also because TNF-alpha production by LPS was abolished in the presence of CHX.
在小鼠巨噬细胞样细胞系J774.1中，研究了脂多糖（LPS）在肿瘤坏死因子-α（TNF-α）产生以及在存在环己酰亚胺（CHX）的情况下诱导巨噬细胞死亡中的构效关系。TNF-α产生是LPS激活的巨噬细胞的特征性表型之一，在与LPS孵育时可观察到。相反，巨噬细胞死亡在我们实验室中已被发现（天野F.，唐桥H.，《内毒素研究杂志》，3，415 - 423（1996）），并作为乳酸脱氢酶（LDH）从细胞释放到培养上清液来检测，在存在CHX的情况下，添加LPS后2.5小时观察到巨噬细胞死亡。然而，TNF-α产生和巨噬细胞死亡同样依赖于LPS和脂质A的结构。在浓度超过10 ng/ml时，来自大肠杆菌和明尼苏达沙门氏菌的野生型LPS表现出最高活性，来自明尼苏达沙门氏菌粗糙突变体的LPS以及二磷酸化脂质A也表现出活性，但经碱性处理解毒的大肠杆菌LPS或来自明尼苏达沙门氏菌的单磷酸化脂质A即使在100 ng/ml时也无活性。这些结果表明，在存在CHX的情况下，LPS诱导的巨噬细胞死亡对LPS和/或脂质A结构的要求与高于10 ng/ml剂量时巨噬细胞激活的要求相似，尽管在1 ng/ml LPS时未观察到前者，而后者可观察到。然而，TNF-α似乎不参与巨噬细胞死亡的诱导，因为中和抗TNF-α抗体未能阻断巨噬细胞死亡，并且重组TNF-α在存在或不存在LPS和/或CHX的情况下对LDH释放程度几乎没有影响，还因为在存在CHX的情况下，LPS诱导的TNF-α产生被消除。
