Ficolin-1 is present in a highly mobilizable subset of human neutrophil granules and associates with the cell surface after stimulation with fMLP
Ficolin-1 is present in a highly mobilizable subset of human neutrophil granules and associates with the cell surface after stimulation with fMLP
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DOI：
10.1189/jlb.1008606
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发表时间：
2009-12-01
影响因子：
5.5
通讯作者：
Borregaard, Niels
中科院分区：
文献类型：
--
作者：
Rorvig, Sara;Honore, Christian;Borregaard, Niels
纤维蛋白是一种结合微生物表面碳水化合物的可溶性分子，在凝集素补体途径中起识别分子的作用。在人类中已鉴定出三种ficolin: ficolin-1、ficolin-2和ficolin-3。Ficolin-1在单核细胞和II型肺泡上皮细胞中合成。Ficolin-1已被证明存在于人类中性粒细胞的分泌颗粒中，但尚不清楚中性粒细胞分泌颗粒的哪个亚群含有Ficolin-1。为了确定ficolin-1在中性粒细胞中的精确亚细胞定位，我们在中国仓鼠卵巢细胞中表达了重组ficolin-1，并用于产生多克隆抗体。这允许通过ELISA、Western blotting和免疫组织化学检测人中性粒细胞亚细胞组分中的ficolin-1。实时荧光定量PCR检测正常人骨髓显示，FCN1基因主要在髓细胞、超髓细胞和带状细胞中表达，其表达谱与明胶酶非常相似。与此相一致，中性粒细胞前体细胞的生物合成研究表明，ficolin-1主要在髓细胞、变髓细胞和带细胞中合成。免疫组织化学和亚细胞分离表明，ficolin-1主要定位于明胶酶颗粒中，但也存在于高度胞外化的明胶酶缺乏颗粒中，这是以前没有描述过的。Ficolin-1通过fMLP或PMA刺激从中性粒细胞颗粒中释放出来，大部分与细胞表面膜相关，可以通过流式细胞术检测到。我们的研究表明，中性粒细胞是ficolin-1的主要来源，它可以很容易地通过刺激排出细胞。j . Leukoc。生物学86:1439-1449；2009.
Ficolins are soluble molecules that bind carbohydrate present on the surface of microorganisms and function as recognition molecules in the lectin complement pathway. Three ficolins have been identified in humans: ficolin-1, ficolin-2, and ficolin-3. Ficolin-1 is synthesized in monocytes and type II alveolar epithelial cells. Ficolin-1 has been shown to be present in secretory granules of human neutrophils, but it is not known which subset of the neutrophils' secretory granules harbors ficolin-1. To determine the exact subcellular localization of ficolin-1 in neutrophils, recombinant ficolin-1 was expressed in Chinese hamster ovary cells and used for generation of polyclonal antibodies. This allowed detection of ficolin-1 in subcellular fractions of human neutrophils by ELISA, by Western blotting, and by immunohistochemistry. Real-time PCR examination of normal human bone marrow showed FCN1 gene expression largely in myelocytes, metamyelocytes, and band cells with a profile quite similar to that of gelatinase. In accordance with this, biosynthesis studies of neutrophils precursor cells showed that ficolin-1 was primarily synthesized in myelocytes, metamyelocytes, and band cells. Immunohistochemistry and subcellular fractionation demonstrated that ficolin-1 is primarily localized in gelatinase granules but also in highly exocytosable gelatinase-poor granules, not described previously. Ficolin-1 is released from neutrophil granules by stimulation with fMLP or PMA, and the majority becomes associated with the surface membrane of the cells and can be detected by flow cytometry. Our studies show that neutrophils are a major source of ficolin-1, which can be readily exocytosed by stimulation. J. Leukoc. Biol. 86: 1439-1449; 2009.