A molecular profile of the mouse gastric parietal cell with and without exposure to Helicobacter pylori

Abstract

The parietal cell (PC) plays an important role in normal gastric physiology and in common diseases of the stomach. Although the genes involved in acid secretion are well known, there is limited molecular information about other aspects of PC function. We have generated a comprehensive database of genes expressed preferentially in PCs relative to other gastric mucosal cell lineages. PCs were purified from FVB/N mouse stomachs by lectin panning. cRNA generated from PC-enriched (PC(+)) and PC-depleted (PC(-)) populations were used to query oligonucleotide-based microarrays. False-positive signals were filtered by using a new algorithm for noise reduction and selected results independently audited by real-time quantitative reverse transcription (RT)-PCR. The annotated database of 240 genes reveals previously unappreciated aspects of cellular function, including factors that may mediate PC regulation of gastric stem cell proliferation. PC(+) and PC(-) expression profiles were also prepared from germ-free mice 2 and 8 weeks after colonization with a clinical isolate of Helicobacter pylori (Hp)--the pathogen that produces acid-peptic disease (gastritis, ulcers) in humans. Whereas PC(+) gene expression was remarkably constant, the PC(-) fractions demonstrated a robust, evolving host response, with increased expression of genes involved in cell motility/migration, extracellular matrix interactions, and IFN responses. The consistency of PC(+) gene expression allowed identification of a cohort of 92 genes enriched in PCs under all conditions studied. These genes provide a molecular profile that can be used to define this epithelial lineage under a variety of physiologic, pharmacologic, and pathologic stimuli.

Figure 1

Histologic characterization of parietal cell-enriched and parietal cell depleted fractions prepared by magnetic bead-lectin panning. Cells from PC⁺ fraction (A) and PC⁻ fraction (B) were stained with rabbit antibodies to the noncatalytic β-subunit of mouse H⁺/K⁺-ATPase (labeled green with FITC-conjugated donkey anti-rabbit Ig). Nuclei were stained blue with bis-benzimide. Bars = 20 μm. (C) Transmission EM of two parietal cells in the PC⁺ fraction showing the integrity of the canalicular apparatus (closed arrow), the abundance of mitochondria (arrowhead), and binding of magnetic bead-DBA conjugates to the cell surface (open arrow). Magnification ×3300.

Figure 2

Expression of parietal cell-associated genes in PC⁺ vs. PC⁻ fractions, and in the intact stomachs of normal mice and tox176 transgenic animals with an engineered ablation of the PC lineage. For GeneChip and qRT-PCR comparisons of expression in PC⁺ vs. PC⁻ fractions, the level of each mRNA in the PC⁻ fraction was arbitrarily set at 1. For qRT-PCR comparisons of gene expression in the stomachs of conventionally raised normal and tox176 mice, mRNA levels in tox176 RNA were set at 1. Error bar = 1 SD (n = four 14- to 16-week-old mice per group; at this age, multi and oligo-potential lineage progenitors comprise 1% of the total epithelial cell population in gastric units of normal mice and ≥10% in tox176 units).

Figure 3

qRT-PCR analysis of the fold-difference in levels of selected mRNAs in PC⁺ fractions or the intact stomachs of H. pylori-infected vs. germ-free FVB/N mice. Error bar = 1 SD.