Use of synthetic derivatives to determine the minimal active structure of cytokine-inducing lipoteichoic acid
Use of synthetic derivatives to determine the minimal active structure of cytokine-inducing lipoteichoic acid
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DOI：
10.1128/cvi.00007-07
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发表时间：
2007-12-01
影响因子：
--
通讯作者：
von Aulock, Sonja
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文献类型：
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作者：
Deininger, Susanne;Figueroa-Perez, Ignacio;von Aulock, Sonja
来自革兰氏阳性菌的脂磷壁酸（LTA）是来自革兰氏阴性菌的脂多糖的对应物。LTA激活Toll样受体2（TLR 2），诱导独特的细胞因子和趋化因子模式。LTA的化学合成证明了其免疫刺激特性。为了确定LTA的最小活性结构，我们将合成的LTA以多个步骤还原为合成的锚，并使用这些分子刺激人全血中的白细胞介素-8（IL-8）释放。诱导IL-8释放需要比天然LTA制剂多十倍的具有四到六个D-丙氨酸取代的聚甘油磷酸酯单元（50 nM）的合成结构。进一步减少至具有两个或没有D-丙氨酸残基的三个骨架单元导致仅从500 nM诱导细胞因子。合成的锚即使在5 μ M时也不能诱导IL-8释放。当LTA衍生物以500 nM使用时，它们诱导IL-8和肿瘤坏死因子α的水平随着主链的延长而增加。腹膜巨噬细胞对合成结构的反应不如人血。因此，对于两个最大的合成结构，仅用来自TLR 2缺陷小鼠的细胞可以显示TLR 2依赖性。这通过使用TLR 2转染的HEK 293细胞证实。总之，这些数据表明，尽管合成的锚（与天然的锚不同，其仅含有肉豆蔻酸）不能诱导细胞因子释放，但添加三个骨架单元，即使没有D-丙氨酸取代基，也赋予这种能力。用D-丙氨酸取代的主链单元延长链导致增加的精氨酸诱导效力和更敏感的反应。
Lipoteichoic acid (LTA) from gram-positive bacteria is the counterpart to lipopolysaccharide from gram-negative bacteria. LTA, which activates Toll-like receptor 2 (TLR2), induces a unique cytokine and chemokine pattern. The chemical synthesis of LTA proved its immunostimulatory properties. To determine the minimal active structure of LTA, we reduced synthetic LTA in a number of steps down to the synthetic anchor and employed these molecules to stimulate interleukin-8 (IL-8) release in human whole blood. Ten times more of the synthetic structures with four to Six D-alanine-substituted polyglycerophosphate units (50 nM) than of the native LTA preparation was required to induce IL-8 release. A further reduction to three backbone units with two or no D-alanine residues resulted in cytokine induction only from 500 nM. The synthetic anchor was not able to induce IL-8 release even at 5 mu M. When the LTA derivatives were used at 500 nM, they induced increasing levels of IL-8 and tumor necrosis factor alpha with increasing elongation of the backbone. Peritoneal macrophages were less responsive than human blood to the synthetic structures. Therefore, TLR2 dependency could be shown only with cells from TLR2-deficient mice for the two largest synthetic structures. This was confirmed by using TLR2-transfected HEK 293 cells. Taken together, these data indicate that although the synthetic anchor (which, unlike the native anchor, contains only myristic acid) cannot induce cytokine release, the addition of three backbone units, even without D-alanine substituents, confers this ability. Lengthening of the chain with D-alanine-substituted backbone units results in increased cytokine-inducing potency and a more sensitive response.