Home  |  About  |  Contact  |  Join MCACS  Donate to MCACS
Massachusetts Chapter of the American College of Surgeons Massachusetts Chapter of the American College of Surgeons Massachusetts Chapter of the American College of Surgeons Massachusetts Chapter of the American College of Surgeons Massachusetts Chapter of the American College of Surgeons
Intestinal alkaline phosphatase protects against TLR-mediated inflammation

Kathryn Chen, MD, Madhu Malo, MD, Golam Mostafa, MD, Sayeda Alam, MD, Sundaram Ramasamy, MD, Elizabeth Hohmann, MD, Richard Hodin, MD
Massachusetts General Hospital- Department of Gastrointestinal Research; Boston, MA


PURPOSE OF STUDY
Intestinal alkaline phosphatase (IAP) is recognized as an antimicrobial gut defense factor, and we sought to examine the cellular mechanisms by which IAP functions.

METHODS USED:
Varying amounts of TLR ligands (LPS, CpG) and non-TLR ligand (TNF-alpha) were incubated +/- calf intestinal alkaline phosphatase (CIP) and free phosphate measured by malachite green assay. Ligands +/- CIP were applied to HT29 cells and IL-8 secretion measured by ELISA. HT29 cells stably transfected with an IAP expression plasmid (HT29-IAP), parent cells and TLR-deficient HEK-293 cells were incubated with Escherichia coli or Salmonella typhimurium and IL-8 secretion measured.

SUMMARY OF RESULTS:
HT29 cells showed dose-dependent Il-8 increases following exposure to LPS, TNF, and CpG. IL-8 production was blocked when LPS or CpG, but not TNF, were pre-incubated with CIP, and this inhibition was CIP dose-dependent. Increased free phosphate was detected when LPS and CpG, but not TNF, were incubated with CIP. Parent cells showed increased IL-8 production when incubated with E. coli and S. typhimurium as compared with HT29-IAP or HEK-293 cells.

CONCLUSIONS:
CIP inhibited IL-8 induction by TLR ligands LPS and CpG, but not by TNF-alpha, and concomitantly manifested increased free phosphate, suggesting that ligand dephosphorylation was responsible for inhibition of target cell effects. HT29-IAP and TLR-deficient HEK-293 cells, but not parent cells, were hypo-responsive in terms of IL-8 production when incubated with E. coli and S. typhimurium. These data indicate that the gut defense factor IAP may down-regulate TLR mediated inflammatory pathways by dephosphorylation of their cognate TLR ligands.

 

 Home | About | Contact