Lipid A heterogeneity within Porphyromonas gingivalis and other oral bacteria: effect of lipid A content on hTLR4 utilization and E-selectin expression

Thesis (Ph. D.)--University of Washington, 2005.Lipopolysaccharide (LPS) is an outer membrane molecule for virtually all Gram-negative bacteria. Specific components of the lipopolysaccharide (LPS) molecule, once thought to be highly conserved, are in some bacterial species surprisingly variable. One such portion of the LPS molecule displaying this surprising variability is the lipid A portion. Some examples of lipid A structural variability are seen within bacterial pathogens such as Yersina pestis, Salmonella typhimurium and Pseudomonas aeruginosa which have been linked to significant alteration in host biologic activity. However, what has yet to be determined is the presence, extent or mechanism behind lipid A heterogeneity in oral bacteria. This project was therefore developed to test the hypothesis that any structural changes observed within lipid A of oral bacteria would alter host responses. Results confirmed our initial hypothesis that lipid A structure from P. gingivalis was highly variable (heterogenic) as compared to other oral bacteria sampled. P. gingivalis lipid A structure was altered by growth phase, environmental pH, as well as by insertional inactivation of the PhoP transcriptional activator, which is part of a two component regulatory mechanism recently identified within the genome of P. gingivalis. It was also observed that application of phenol or base chemicals (especially in the presence of heat during extraction procedures) resulted in loss of structural components or degradation of the lipid A molecule in both oral and enteric bacteria. Reduction in the number of fatty acids and/or phosphate groups from the disaccharide backbone of oral lipid A, as a result of the experimental procedures, resulted in the reduction of E-selectin expression in HUVECs and hTLR4 utilization in transfected HEK 293 cells whereas unaltered lipid A structure retained full biologic activity. In addition, selective removal of fatty acids within different oral bacterial lipid A resulted in partial antagonism of both E. coli and P. gingivalis LPS. These results suggest that under specific environmental conditions, certain bacteria which naturally possess the ability to alter its own or neighboring bacterial lipid A structure, would be able to manipulate the local environment to potentially favor colonization, growth and survival within host tissues