NLRP3 inflammasome activation and interleukin-1β release in macrophages require calcium but are independent of calcium-activated NADPH oxidases.
NLRP3 inflammasome activation and interleukin-1β release in macrophages require calcium but are independent of calcium-activated NADPH oxidases.
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DOI：
10.1007/s00011-014-0756-y
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发表时间：
2014-10
影响因子：
6.7
通讯作者：
Leto, Thomas L.
中科院分区：
文献类型：
--
作者：
Rada, Balazs;Park, Jonathan J.;Sil, Payel;Geiszt, Miklos;Leto, Thomas L.
我们研究了钙和钙活化的NADPH氧化酶在NLRP3炎性体激活和IL-1β释放中的作用，以更好地了解巨噬细胞中炎性体的信号传导。招募志愿献血者分离单核细胞并将其分化为巨噬细胞。采用野生型或duox1缺失型C57/B6小鼠制备骨髓源性巨噬细胞。在含钙或无钙培养基中，用NLRP3炎性小体激动剂（ATP、二氧化硅晶体）或钙激动剂（thapsigargin、离子霉素）体外处理小鼠或人巨噬细胞。通过测量基于fura2的荧光来跟踪细胞内钙的变化。采用实时荧光定量PCR检测基因表达变化。western blotting检测蛋白表达。采用荧光caspase-1活性测定法测定酶活性。ELISA法测定IL-1β释放量。采用方差分析和Tukey事后检验对ELISA数据进行分析。我们的数据表明，钙对于人巨噬细胞中IL-1β的释放至关重要。单独增加胞质钙可导致IL-1β分泌。钙的去除阻断了caspase-1的激活。人巨噬细胞表达Duox1，一种钙调节的NADPH氧化酶，产生活性氧。然而，duox1缺陷小鼠巨噬细胞显示正常的IL-1β释放。人巨噬细胞炎性体的激活和IL-1β的分泌需要钙，但不涉及NADPH氧化酶。
We studied the involvement of calcium and calcium-activated NADPH oxidases in NLRP3 inflammasome activation and IL-1β release to better understand inflammasome signaling in macrophages. Human volunteer blood donors were recruited to isolate monocytes to differentiate them into macrophages. Wild-type or DUOX1-deficient C57/B6 mice were used to prepare bone marrow-derived macrophages. Murine or human macrophages were treated in vitro with NLRP3 inflammasome agonists (ATP, silica crystals) or calcium agonists (thapsigargin, ionomycin) in calcium-containing or calcium-free medium. Intracellular calcium changes were followed by measuring FURA2-based fluorescence. Gene expression changes were measured by quantitative real-time PCR. Protein expression was assessed by western blotting. Enzymatic activity was measured by fluorescence caspase-1 activity assay. IL-1β release was determined by ELISA. ELISA data were analyzed by ANOVA and Tukey’s post-hoc test. Our data show that calcium is essential for IL-1β release in human macrophages. Increases in cytosolic calcium alone lead to IL-1β secretion. Calcium removal blocks caspase-1 activation. Human macrophages express Duox1, a calcium-regulated NADPH oxidase that produces reactive oxygen species. However, Duox1-deficient murine macrophages show normal IL-1β release. Human macrophage inflammasome activation and IL-1β secretion requires calcium but does not involve NADPH oxidases.
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影响因子：
8.7
作者：
Broz P;Monack DM
通讯作者：
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影响因子：
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作者：
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DOI：
10.1074/jbc.m110.203398
发表时间：
2011-08-05
期刊：
The Journal of biological chemistry
影响因子：
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作者：
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作者：
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影响因子：
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作者：
Martinon, F;Burns, K;Tschopp, J
通讯作者：
Tschopp, J