Immunological Functions of Cholesteryl α -glucosides in Helicobacter pylori-associated Inflammation

Helicobacter pylori (H. pylori) uniquely contains cholesteryl α-glucosides (αCGs) as the major components of its cell wall. Cholesteryl-α-glucoside (αCGL) is synthesized by cholesterol α-glycosyltransferase (αCgT), and αCGL is a precursor for cholesteryl-acyl-α-glucoside (αCAG) and cholesteryl-phosphatidyl-a-glucoside (αCPG). We reported that mucin-type core 2-branched O-glycans expressing α1,4-N-acetylglucosamine residues suppress bacterial growth by inhibition of αCGs synthesis (Kawakubo et al Science 2004, Lee et al BBRC 2006).

To determine roles of αCGs, we analyzed amino acid substitutions in αCgT of H. pylori from 24 Japanese clinical isolates. Recombinant αCgT showed diverse activity from 15 to 174% compared to the αCgT in the wild-type H. pylori 26695. Analysis of the mutated αCgT activity and histological features revealed that the enzyme activity is associated with the progression of gastric atrophy. We generated αCgT deficient H. pylori (αCgT^Δ), and αCgT^high and αCgT^low, which expressed higher and lower αCgT activity than wild-type, respectively. αCgT^Δ exhibited significantly delayed growth and morphologic abnormality caused by loss of all of αCGs. These results suggested that αCGs are essential for H. pylori growth.

We investigated further functions of αCGs in immune response. Phagocytosis and activation of dendritic cell were observed at the same degrees in the wild-type H. pylori and all mutants, regardless of αCgT activity. However, response by invariant natural killer T (iNKT) cells was positively correlated with the activity of αCgT. In synthetic compounds of αCGs, αCPG most activated iNKT cells. Moreover, monoacyl-αCPG was the most potent antigen in vivo assay. More H. pylori strain was retrieved from the stomach of iNKT deficient (Jα18^−/-) mice than wild-type mice. Furthermore, expression level of cytokines in αCgT^high infected Jα18^−/- mice was significantly decreased than αCgT^high infected wild-type mice. Our findings suggested that recognition of αCGs by iNKT cells are necessary for bacterial clearance and stimulation immune cell response in various cell lineages in Th1/Th2/Th17 cells. Further analysis is currently in progress. Supported by NIH grants PO1CA71432, CA33000 and Toyobo Biotechnology Foundation Fellowship.