Open AccessFree Lipid A Isolated from Porphyromonas gingivalis Lipopolysaccharide Is Contaminated with Phosphorylated Dihydroceramide Lipids: Recovery in Diseased Dental Samples
Author(s) -
Frank C. Nichols,
Bekim Bajrami,
Robert B. Clark,
William Housley,
Xudong Yao
Publication year - 2012
Publication title -
infection and immunity
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.508
H-Index - 220
eISSN - 1070-6313
pISSN - 0019-9567
DOI - 10.1128/iai.06180-11
Subject(s) - lipid a , porphyromonas gingivalis , lipopolysaccharide , biology , periodontal pathogen , microbiology and biotechnology , tlr2 , biochemistry , chemistry , tlr4 , bacteria , immunology , receptor , genetics
Recent reports indicate thatPorphyromonas gingivalis mediates alveolar bone loss or osteoclast modulation through engagement of Toll-like receptor 2 (TLR2), though the factors responsible for TLR2 engagement have yet to be determined. Lipopolysaccharide (LPS) and lipid A, lipoprotein, fimbriae, and phosphorylated dihydroceramides ofP. gingivalis have been reported to activate host cell responses through engagement of TLR2. LPS and lipid A are the most controversial in this regard because conflicting evidence has been reported concerning the capacity ofP. gingivalis LPS or lipid A to engage TLR2 versus TLR4. In the present study, we first preparedP. gingivalis LPS by the Tri-Reagent method and evaluated this isolate for contamination with phosphorylated dihydroceramide lipids. Next, the lipid A prepared from this LPS was evaluated for the presence of phosphorylated dihydroceramide lipids. Finally, we characterized the lipid A by the matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) and electrospray-MS methods in order to quantify recovery of lipid A in lipid extracts from diseased teeth or subgingival plaque samples. Our results demonstrate that both the LPS and lipid A derived fromP. gingivalis are contaminated with phosphorylated dihydroceramide lipids. Furthermore, the lipid extracts derived from diseased teeth or subgingival plaque do not contain free lipid A constituents ofP. gingivalis but contain substantial amounts of phosphorylated dihydroceramide lipids. Therefore, the free lipid A ofP. gingivalis is not present in measurable levels at periodontal disease sites. Our results also suggest that the TLR2 activation of host tissues attributed to LPS and lipid A ofP. gingivalis could actually be mediated by phosphorylated dihydroceramides.