The Chloride Channel CLC-2 is Involved in Organization of Murine Gastric Glands

Abstract

BACKGROUND: Primary cilia are necessary for Hedgehog (Hh) signaling and other pathways. Anterograde ciliary transport is mediated by kinesin-II, a heterotrimeric protein composed of two motor subunits, KIF3A, KIF3B, and a non-motor subunit KAP3. Intraflagellar transport protein 88 (IFT88) participates in protein transport along the cilia. Conventional null mice for KIF3A and IFT88 are not viable. Sonic hedgehog (Shh) is highly expressed in gastric epithelial cells of the corpus compared to the antrum. Therefore we examined the stomach of mice conditionally null for KIF3A and IFT88 to define the role of primary cilia in this organ. METHODS: KIF3A and IFT88 mice harboring one null allele and loxP sites on the other allele on a ROSA26 reporter background were crossed to Shh-Cre mice (KIF3A-/ FL and IFT88-/FL respectively.) KIF3A+/FL, IFT88+/FL, and wild type (WT) littermates were analyzed at 6 and 8 months. Cell lineages were identified by immunohistochemistry in paraffin-sections, and by β-galactosidase (β-gal) staining for the Cre-expressing cells. Acetylated-α-tubulin was used as the marker for cilia. RESULTS: Epithelial cells in the antrum and corpus expressed primary cilia. β-gal staining confirmed variegated glandular Cre expression in all epithelial cells. Gastric epithelial cells did not express cilia in the IFT88-/FL mice, while KIF3A-/FL mice showed reduced numbers of cilia in the corpus versus controls (0.59+0.04 cilia/gland, N=8 vs. 0.72+0.06 cilia/gland, N=14), but no changes in the antrum (0.94+0.05 vs. 1.08+0.09 cilia/gland.) KIF3A-/FL and IFT88-/FL mice exhibited different phenotypes. By 6 months, KIF3A-/FL mice showed mucous pit hyperplasia in the corpus. Mucous neck and zymogenic lineages were not expanded, but parietal cells were still present. The KIF3A-/FL mice phenotype resembled the mucous pit hyperplasia observed in the Shh conditionally null mice generated with H+K+ATPase-Cre. In contrast, the phenotype of the IFT88-/FL mice showed parietal cell atrophy, and proximal corpus hyperplasia due to expansion of the mucous neck cell compartment assessed by TFF2 expression (SPEM) by 4 months of age. We have previously observed that gastrinand ghrelin-secreting cells exhibit primary cilia, but none of the mouse models showed differences in the number of these endocrine cells.CONCLUSION:The IFT88-/FLmice exhibited amore severe phenotype than the KIF3A-/FL mice even though both molecules affect primary cilia function. The phenotype of these two mouse models suggests that Hh signaling is necessary for the maintenance of the corpus epithelium. Also, gastric epithelial cilia might be important for the function but not differentiation of gastrinand ghrelin-secreting cells.