Inducible expression of human beta-defensin 2 by Fusobacterium nucleatum in oral epithelial cells: Multiple signaling pathways and role of commensal bacteria in innate immunity and the epithelial barrier

Human gingival epithelial cells (HGE) express two antimicrobial peptides of the beta-defensin family, human beta-defensin 1 (hBD-1) and hBD-2, as well as cytokines and chemokines that contribute to innate immunity, In the pre sent study, the expression and transcriptional regulation of hBD-2 was exam ined. HBD-2 mRNA was induced by cell wall extract of Fusobacterium nucleatu m, an oral commensal microorganism, but not by that of Porphyromonas gingiv alis, a periodontal pathogen, HBD-2 mRNA was also induced by the proinflamm atory cytokine tumor necrosis factor alpha (TNF-alpha) and phorbol myristat e acetate (PMA), an epithelial cell activator, HBD-2 mRNA was also expresse d in 14 of 15 noninflamed gingival tissue samples. HBD-2 peptide was detect ed by immunofluorescence in HGE stimulated with F. nucleatum cell wall, con sistent with induction of the mRNA by this stimulant. Kinetic analysis indi cates involvement of multiple distinct signaling pathways in the regulation of hBD-2 mRNA; TNF-alpha and F. nucleatum cell wall induced hBD-2 mRNA rap idly (2 to 4 h), while PMA stimulation was slower (similar to 10 h). In con trast, each stimulant induced interleukin 8 (IL-8) within 1 h, The role of TNF-alpha as an intermediary in F. nucleatum signaling was ruled out by add ition of anti TNF-alpha that did not inhibit hBD-2 induction, However, inhi bitor studies show that F. nucleatum stimulation of hBD-2 mRNA requires bot h new gene transcription and new protein synthesis, Bacterial lipopolysacch arides isolated from Escherichia coli and F, nucleatum were poor stimulants of hBD-2, although they up-regulated IL-8 mRNA. Collectively, our findings show inducible expression of hBD-2 mRNA via multiple pathways in HGE in a pattern that is distinct from that of IL-8 expression. We suggest that diff erent aspects of innate immune responses are differentially regulated and t hat commensal organisms have a role in stimulating mucosal epithelial cells in maintaining the barrier that contributes to homeostasis and host defens e.