Z Gastroenterol 2016; 54 - KV024
DOI: 10.1055/s-0036-1586801

Endogenous intestinal alkaline phosphatase controls inflammatory pathways and cellular homeostasis in the gut

F Kühn 1, 2, SR Hamarneh 1, JM Ramirez 1, AR Munoz 1, SA Morrison 1, SS Gul 1, F Adiliaghdam 1, RA Hodin 1
  • 1Massachusetts General Hospital, Harvard Medical School, Department of Surgery, Boston, USA
  • 2Universitätsmedizin Rostock, Allgemein-, Thorax-, Gefäß- und Transplantationschirurgie, Rostock, Deutschland

Objective: The inability of epithelial cells to cope with various stresses and cellular damage plays a crucial role in the development of many diseases in the gut such as colitis, Crohn's disease and carcinoma. The exact functions of alkaline phosphatases in various cellular pathways are not well understood. We propose that intestinal alkaline phosphatase (IAP) plays a critical role in intracellular homeostasis and stress responses in the gut.

Methods: IAP protein complexes were immuno-precipitated using Anti-RFP antibody followed by masses identification using liquid chromatography-tandem mass spectrometry (LC-MS/MS) from Caco-2 cells transfected with RFP-IAP plasmid. Peptides and proteins were identified using Peaks program and Mascot server, which use the MS and MS/MS spectra of peptide ions to search against the publicly available SwissProt, NCBInr and contaminant data bases followed by functional analysis. Furthermore, IAP knockout Caco-2 cells were developed using the CRISPR/Cas9 gene editing technique to study the effect of IAP deletion on cellular functions.

Results: Functional analysis demonstrated an extended role of IAP in intracellular homeostasis: Pathways in cancer (p < 0.0001), MAPK signaling pathway (p < 0.0001), cell cycle (p < 0.0001), bacterial invasion of epithelial cells (p < 0.0001), intestinal permeability and junction formation (p < 0.0001), apoptosis (p < 0.0001), inflammatory pathways (p < 0.001), p53 signaling pathway (p = 0.0282) and adipocytokine signaling pathway (p = 0.0373). Furthermore, deletion of IAP in Caco-2 cell line increased the expression levels of TNF-α, IL-1β and IL-8 genes. Over-expression of IAP resulted in significantly less inflammation and cytokine production in Caco-2 cells when incubated with multiple inflammatory mediators such as TNF-α, LPS and bacterial contents.

Conclusions: IAP plays a crucial role in gut epithelial lining function and homeostasis. IAP pathways in the gut may represent an important therapeutic target to prevent or treat disorders of the gut.