Distribution of 3-hydroxy iC(17:0) in subgingival plaque and gingival tissue samples: Relationship to adult periodontitis

Abstract

Gram-negative organisms incorporate hydroxy fatty acids into the lipid A moiety of lipopolysaccharide (LPS), and in the case of some members of the family Enterobacteriaceae, hydroxy fatty acids are incorporated exclusively into lipid A. However, a limited number of Bacteroides species have been shown to incorporate several classes of 3-hydroxy fatty acids, particularly 3-hydroxy iC(17:0), into constitutive lipids as well as LPS. The present study examined the distribution of hydroxy fatty acids in two periodontal pathogens, Prevotella intermedia and Porphyromonas gingivalis, by employing a phospholipid extraction procedure (E. G. Bligh and W. J. Dyer, Can. J. Biochem. Physiol. 37:911-917, 1959) which partitioned constitutive lipids into the organic solvent phase and LPS into the aqueous phase. The distribution of hydroxy fatty acids within organic solvent and aqueous extracts of these bacterial species was then compared with the distribution in subgingival plaque samples isolated from either gingivitis or severe periodontitis sites as well as the distribution in gingival tissue samples. The organic solvent and aqueous extracts were hydrolyzed under strong alkaline conditions, and the free fatty acids were treated to form pentafluorobenzyl-ester, trimethylsilyl-ether derivatives. Hydroxy fatty acid levels were quantified by using gas chromatography-negative-ion chemical ionization-mass spectrometry. By using this approach, the mean values of the 3-hydroxy iC(17:0) recovered within organic solvent extracts of P. gingivalis strains ranged from 56 to 63% of total 3-hydroxy iC(17:0). Substantially less 3-hydroxy iC(17:0) (<5%) was recovered in organic solvent extracts of P. intermedia. By comparison, 75% of the 3-hydroxy iC(17:0) in periodontitis subgingival plaque samples was recovered in organic solvent extracts, while only 43% of the 3-hydroxy iC(17:0) in gingivitis plaque samples from the same patients was recovered in organic solvent extracts. However, 3-hydroxy iC(17:0) was recovered essentially only in organic solvent extracts of both healthy or mildly inflamed and periodontitis gingival tissue samples. The preferential recovery of 3-hydroxy iC(17:0) in tissue lipids indicates that gingival tissues do not harbor significant levels of subgingival plaque organisms which contain 3-hydroxy iC(17:0). Furthermore, these results indicate that LPS from these organisms is not prevalent in gingival tissues. Finally, these results indicate either selective penetration of certain bacterial lipids into gingival tissues or that 3-hydroxy iC(17:0) is metabolically transferred from bacterial lipids into gingival tissue lipids.