Abstract
The present studies were designed to examine the effects of ClC-2 ablation on cellular morphology, parietal cell abundance, H/K ATPase expression, parietal cell ultrastructure and acid secretion using WT and ClC-2-/- mouse stomachs. Cellular histology, morphology and proteins were examined using imaging techniques, electron microscopy and western blot. The effect of histamine on the pH of gastric contents was measured. Acid secretion was also measured using methods and secretagogues previously established to give maximal acid secretion and morphological change. Compared to WT, ClC-2-/- gastric mucosal histological organization appeared disrupted, including dilation of gastric glands, shortening of the gastric gland region and disorganization of all cell layers. Parietal cell numbers and H/K ATPase expression were significantly reduced by 34% (P<0.05) and 53% (P<0.001) respectively and cytoplasmic tubulovesicles appeared markedly reduced on electron microscopic evaluation without evidence of canalicular expansion. In WT parietal cells, ClC-2 was apparent in a similar cellular location as the H/K ATPase by immunofluorescence and appeared associated with tubulovesicles by immunogold electron microscopy. Histamine-stimulated [H+] of the gastric contents was significantly (P<0.025) lower by 9.4 fold (89%) in the ClC-2-/- mouse compared to WT. Histamine/carbachol stimulated gastric acid secretion was significantly reduced (range 84–95%, P<0.005) in ClC-2-/- compared to WT, while pepsinogen secretion was unaffected. Genetic ablation of ClC-2 resulted in reduced gastric gland region, reduced parietal cell number, reduced H/K ATPase, reduced tubulovesicles and reduced stimulated acid secretion.
Highlights
ClC-2 is a broadly expressed Cl− channel activated by hyperpolarization, extracellular acidic pH, fatty acids including lubiprostone, amidation, acid-activated omeprazole and in some species protein kinase A [1,2,3,4,5,6,7,8,9,10]
There appeared to be reduced numbers of parietal cells in the glands (Fig 1A) and the height of the gastric gland region of the gastric mucosa was significantly reduced by 24.4% (P
The aim of the present study was to investigate whether genetic ablation of ClC-2 has any effects on the gastric mucosa with a focus on parietal cell abundance, H/K ATPase expression, morphology and acid secretion using WT and ClC-2-/- mouse gastric mucosa
Summary
ClC-2 is a broadly expressed Cl− channel activated by hyperpolarization, extracellular (luminal) acidic pH, fatty acids including lubiprostone, amidation, acid-activated omeprazole and in some species (rabbit and human, but not mouse) protein kinase A [1,2,3,4,5,6,7,8,9,10]. Secretagogue stimulation results in a rapid, major morphological transformation in parietal cells, essential for maximal acid secretion to occur wherein cytoplasmic tubulovesicles containing H/K ATPase (and perhaps Cl- and K+ channels/transporters) fuse with the apical membrane to form a greatly expanded secretory canaliculus with increased elongated microvilli which are are recycled back during the resting stage [20,26,27]. Experimental conditions needed to obtain maximal parietal cell acid secretion and morphological transformation in the mouse stomach described in [27], were followed in the present studies
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