Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, C027

Oral Communications

The apical Na+/H+ exchanger 2 regulates intestinal wound healing and barrier function

K. Nikolovska1, L. Cao1, A. K. Singh1, K. Zhou1, Y. Yu1, Z. Yuan1, A. Seidler1, G. di Stefano1, B. Riederer1, S. Yeruva1, U. E. Seidler1

1. Medical School Hannover, Hannover, Germany.


The apical Na+/H+ exchanger isoform NHE2 is highly expressed in the gastrointestinal tract. nhe2-/- mice display a delayed trefoil factor-stimulated gastric epithelial wound healing and impaired small intestinal barrier recovery after ischemia, but the molecular mechanisms behind are still not understood. Previous study on experimental wound healing in the gastric cell line RGM-1 showed that overexpression of NHE2 triggers increased cellular pHi, but decreased migratory speed (1). To understand how a reduced migratory speed may result in improved barrier function, we used models where the endogenous expression of NHE2 was deleted (nhe2-/- mice) or silenced by shRNA interference (Caco2Bbe cells) and studied enterocyte migration, proliferation and differentiation in vitro and in vivo. NHE2 gene was knocked-down using shRNA in Caco2Bbe cells, resulting in ~70±9.6% decrease of NHE2 expression on mRNA level. Colons from nhe2-/- and wt mice were used for the in vivo studies. All in vivo experiments were performed using 3 to 6 months old nhe2-/- and wt littermates, congenic on the FVB/N background. Mice were killed by cervical dislocation after light isoflurane anesthesia. The loss/downregulation of NHE2 triggered significant decrease of the pHi and alterations in cell proliferation. Colonocyte migration from the crypt base to the surface accessed at different time points after bromodeoxyuridine (BrdU) labeling of the mice was significantly faster in the colon of nhe2-/- mice compared to the control littermates. In vitro wound assay showed significantly faster restitution velocity of the NHE2 silenced Caco2Bbe cells, which was mechanistically supported by altered cell-cell adhesion and assembly of the E-cadherin/β-catenin complex. Further analysis of the nhe2-/- colon revealed elongated crypts with impaired differentiation. The expression and activity of alkaline phosphatase, as an enterocyte differentiation marker, was significantly reduced in cells lacking NHE2, both in vivo and in vitro. In contrast, a shift towards the secretory lineage was detected in the nhe2-/- colon identified by the increased number of MUC2 positive goblet cells. The results imply an important role of NHE2 in the lower/middle segment of the crypt where the progenitor intestinal cells emerge from the stem cell niche and enter enterocyte differentiation. The faster migratory speed in NHE2-deficient cells is associated with a dysregulated enterocyte differentiation pattern including junctional alterations and may explain the observed barrier disturbance previously observed in the gastrointestinal tract of nhe2-/- mice. The results suggest that NHE2 expression and activity are necessary for pHi regulation during enterocyte migration and differentiation.

Where applicable, experiments conform with Society ethical requirements