Detection of beta-defensins secreted by human oral epithelial cells

Human P-defensins are antimicrobial peptides that may be critical in the in nate immune response to infection. hBD1 and hBD2 are expressed in oral epit helial cells and are detected near the surface of oral tissue, consistent w ith a role in the epithelial protective barrier function. In this report, w e examine secretion of P-defensins in vitro and in biological fluid using P roteinChip((R)) Array, surface enhanced laser desorption/ionization (SELDI) technology combined with time-of-flight mass spectrometry. We show that th e 47-amino acid form of hBD1 and the 41-amino acid form of hBD2 are the maj or secreted forms. These forms are both expressed and secreted under condit ions anticipated from previous analysis of P-defensin mRNAs; specifically, hBD1 is detected in culture supernatant from both unstimulated and stimulat ed cells, and hBD2 is detected only in stimulated cells. Identity of hBD1 a nd hBD2 was confirmed by immunocapture on the ProteinChip surface. Both pep tides are also present in gingival crevicular fluid that accumulates betwee n the tissue and tooth surface, although hBD1 is also found in several smal ler forms suggesting extracellular proteolysis. This methodology offers sev eral technical advantages for detection of defensins in biological fluids, including ease and speed of screening, no need for HPLC preliminary process ing, and small sample size. (C) 2001 Elsevier Science B.V. All rights reser ved.