Detection of β-defensins secreted by human oral epithelial cells
Detection of β-defensins secreted by human oral epithelial cells
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DOI：
10.1016/s0022-1759(01)00442-2
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发表时间：
2001-10-01
影响因子：
2.2
通讯作者：
Dale, BA
中科院分区：
文献类型：
--
作者：
Diamond, DL;Kimball, JR;Dale, BA
人p -防御素是抗菌肽，可能在感染的先天免疫反应中起关键作用。hBD1和hBD2在口腔上皮细胞中表达，并在口腔组织表面附近检测到，与参与上皮保护屏障功能的作用一致。在本报告中，我们使用ProteinChip（(R)）阵列、表面增强激光解吸/电离（SELDI）技术结合飞行时间质谱法检测了p -防御素在体外和生物体液中的分泌。我们发现47个氨基酸形式的hBD1和41个氨基酸形式的hBD2是主要的分泌形式。这些形式在先前p -防御素mrna分析预测的条件下表达和分泌；具体来说，hBD1在未刺激细胞和刺激细胞的培养上清中均可检测到，而hBD2仅在刺激细胞中检测到。在ProteinChip表面的免疫捕获证实了hBD1和hBD2的同源性。这两种肽也存在于积聚在组织和牙齿表面之间的龈沟液中，尽管hBD1也以几种较小的形式存在，提示细胞外蛋白水解。该方法为生物液体中防御素的检测提供了几个技术优势，包括筛选容易和速度快，不需要HPLC初步处理，样本量小。(C) 2001 Elsevier Science B.V.版权所有
Human P-defensins are antimicrobial peptides that may be critical in the innate immune response to infection. hBD1 and hBD2 are expressed in oral epithelial cells and are detected near the surface of oral tissue, consistent with a role in the epithelial protective barrier function. In this report, we examine secretion of P-defensins in vitro and in biological fluid using ProteinChip((R)) Array, surface enhanced laser desorption/ionization (SELDI) technology combined with time-of-flight mass spectrometry. We show that the 47-amino acid form of hBD1 and the 41-amino acid form of hBD2 are the major secreted forms. These forms are both expressed and secreted under conditions anticipated from previous analysis of P-defensin mRNAs; specifically, hBD1 is detected in culture supernatant from both unstimulated and stimulated cells, and hBD2 is detected only in stimulated cells. Identity of hBD1 and hBD2 was confirmed by immunocapture on the ProteinChip surface. Both peptides are also present in gingival crevicular fluid that accumulates between the tissue and tooth surface, although hBD1 is also found in several smaller forms suggesting extracellular proteolysis. This methodology offers several technical advantages for detection of defensins in biological fluids, including ease and speed of screening, no need for HPLC preliminary processing, and small sample size. (C) 2001 Elsevier Science B.V. All rights reserved.