title: Bacterial phospholipase C upregulates matrix metalloproteinase expression by cultured epithelial cells.
abstract: Phospholipase C (PLC) is a putative virulence factor of several pathogenic bacteria. We studied if exogenous PLC would perturb epithelial behavior in infected tissues. Gelatin and casein zymography of cell culture medium indicated that the broad-spectrum PLC of Bacillus cereus induced matrix metalloproteinase (MMP) production in epithelial cells of human skin (NHEK), human gingiva (HGE), and porcine periodontal ligament (PLE). In all three cell types, the strongest increase (ninefold) at 0.1 U/ml was seen in the MMP-9 (92-kDa gelatinase) activity, and the effect was dose dependent in the range of 0.1 to 1.0 U/ml. A relatively weaker increase (twofold) in MMP-2 (72-kDa gelatinase) was also observed in each cell type. PLC induction of MMP-3 (48-kDa stromelysin) was also seen in NHEK and HGE on gelatin and more sensitively for PLE by casein zymography (fivefold). Total gelatinolytic activity as measured by degradation of 14C-labeled denatured type I collagen increased by about 18-fold (NHEK), 12-fold (HGE), and 14-fold (PLE). Northern analysis showed a clear increase in the MMP-9, and a minor increase in MMP-3 mRNA levels but no significant increase in MMP-2 mRNA levels. Further studies with PLE revealed that MMP-9 induction by PLC progressively increased with the length of cell culture time in the absence of serum. PLC induction of MMPs was polar, with MMP-9 and MMP-3 secreted primarily in the apical direction and MMP-2 secreted mainly in the basal direction. The PLC effect was blocked by neomycin, an inhibitor of the phosphoinositol signal pathway. No significant effects were observed in MMP expression with the calcium ionophore A23187 or phospholipase A2. Morphologically, PLC treatment resulted in reduced contacts between the cultured cells and loss of the cell surface microvilli. These results suggest that PLC secreted by bacterial pathogens may disrupt epithelium of infected tissue and increase the subepithelial tissue destruction through induction of MMPs.
authors.affiliation: Department of Oral Biology, University of British Columbia, Vancouver, Canada.
authors.forename: J D
authors.initials: JD
authors.lastname: Firth
authors.forename: E E
authors.initials: EE
authors.lastname: Putnins
authors.forename: H
authors.initials: H
authors.lastname: Larjava
authors.forename: V J
authors.initials: VJ
authors.lastname: Uitto
mesh_terms: D000818:Animals; D001409:Bacillus cereus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000494:pharmacology; D002478:Cells, Cultured; D017364:Collagenases / Q000096:biosynthesis*; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology*; D018093:Gelatinases / Q000096:biosynthesis*; D006801:Humans; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D008666:Metalloendopeptidases / Q000096:biosynthesis*; D013552:Swine; D010738:Type C Phospholipases / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D001426:Bacterial Proteins; D010738:Type C Phospholipases; D017364:Collagenases; D018093:Gelatinases; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9