Inflammatory cytokines downregulate the Na+/H+ exchanger 3 (NHE3) interacting PDZ-domain protein PDZK1 in ulcerative coltis patients, colitis mice and in Caco-2bbe cells: Link to inflammation-associated NHE3 dysfunction

Physiology 2012 (Edinburgh) (2012) Proc Physiol Soc 27, C43

Oral Communications: Inflammatory cytokines downregulate the Na+/H+ exchanger 3 (NHE3) interacting PDZ-domain protein PDZK1 in ulcerative coltis patients, colitis mice and in Caco-2bbe cells: Link to inflammation-associated NHE3 dysfunction

S. Yeruva1, G. Chodisetti1, L. Min1, J. Goldstein1, M. Lünnemann1, A. Singh1, A. Bleich2, M. Gereke3, D. Bruder3, C. C. Yun4, U. Seidler1

1. Dept. of Gastroenterology, Hannover Medical School, Hannover, Germany. 2. Animal Research Institute, Hannover Medical School, Hannover, Germany. 3. Helmholtz Center for Infection Biology, Braunschweig, Germany. 4. Emory University, Atlanta, Georgia, United States.

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Introduction: We have previously reported a dysfunction of the major intestinal sodium absorptive transporter, the Na+/H+ exchanger NHE3, in the surface colonocytes of patients with ulcerative colitis, despite normal expression and membrane localization. Objective: In this study, we searched for potential mechanism for this inflammation-associated functional defect in NHE3 transport. Material and Methods: Realtime PCRs, Western blotting, immunohistochemistry, ileal loop experiments and pH fluorometry using pH sensitive BCECF dye. Results: In three immunologic mouse models for colitis (Rag2 KO CD45RBhigh and IL-10 KO ) and Crohn’s disease (TNF ARE +/-), and in biopsies from patients with active UC, we found no change in the mRNA expression of NHE3 or NHERF1, but a dramatic decrease in PDZK1, in the respective inflamed segments. This was confirmed at the protein level. Fluid absorption in isoflurane-anesthetized mice in vivo, and acid-activated NHE3 activity in vitro decreased significantly in the inflamed segments, despite normal immunohistochemical NHE3 staining in the brush border membrane. In PDZK1 heterozygote mice, where enterocyte PDZK1 protein content was reduced by > 50%, acid-activated NHE3 activity decreased to a similar degree in colonocytes compared to inflamed mice. PZDK1 knockdown in stably NHE3-tranfected Caco-2bbe similarly coaused a strong reduction in NHE3 activity, suggesting a causal relationship between PDZK1 downregulation and NHE3 dysfunction. In Caco-2bbe cells treated with cytokines, significant down regulation in PDZK1 mRNA and protein expression was observed. Conclusion: The data demonstrate a marked decrease in the PDZ-adaptor protein, PDZK1, in inflamed murine and human intestine, and a dysfunction of NHE3 activity that is similar in inflamed enterocytes and in those with genetic downregulation of PDZK1. PDZK1 downregulation during inflammation may thus be one factor responsible for inflammation-associated NHE3 dysfunction



Where applicable, experiments conform with Society ethical requirements.

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