Loss of gastrokine-2 drives premalignant gastric inflammation and tumor progression
Trevelyan R. Menheniott, Louise O’Connor, Yok Teng Chionh, Jan Däbritz, Michelle Scurr, Benjamin N. Rollo, Garrett Z. Ng, Shelley Jacobs, Angelique Catubig, Bayzar Kurklu, Stephen Mercer, Toshinari Minamoto, David E. Ong, Richard L. Ferrero, James G. Fox, Timothy C. Wang, Philip Sutton, Louise M. Judd, Andrew S. Giraud
Trevelyan R. Menheniott, Louise O’Connor, Yok Teng Chionh, Jan Däbritz, Michelle Scurr, Benjamin N. Rollo, Garrett Z. Ng, Shelley Jacobs, Angelique Catubig, Bayzar Kurklu, Stephen Mercer, Toshinari Minamoto, David E. Ong, Richard L. Ferrero, James G. Fox, Timothy C. Wang, Philip Sutton, Louise M. Judd, Andrew S. Giraud
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Research Article Oncology

Loss of gastrokine-2 drives premalignant gastric inflammation and tumor progression

Abstract

Chronic mucosal inflammation is associated with a greater risk of gastric cancer (GC) and, therefore, requires tight control by suppressive counter mechanisms. Gastrokine-2 (GKN2) belongs to a family of secreted proteins expressed within normal gastric mucosal cells. GKN2 expression is frequently lost during GC progression, suggesting an inhibitory role; however, a causal link remains unsubstantiated. Here, we developed Gkn2 knockout and transgenic overexpressing mice to investigate the functional impact of GKN2 loss in GC pathogenesis. In mouse models of GC, decreased GKN2 expression correlated with gastric pathology that paralleled human GC progression. At baseline, Gkn2 knockout mice exhibited defective gastric epithelial differentiation but not malignant progression. Conversely, Gkn2 knockout in the IL-11/STAT3-dependent gp130F/F GC model caused tumorigenesis of the proximal stomach. Additionally, gastric immunopathology was accelerated in Helicobacter pylori–infected Gkn2 knockout mice and was associated with augmented T helper cell type 1 (Th1) but not Th17 immunity. Heightened Th1 responses in Gkn2 knockout mice were linked to deregulated mucosal innate immunity and impaired myeloid-derived suppressor cell activation. Finally, transgenic overexpression of human gastrokines (GKNs) attenuated gastric tumor growth in gp130F/F mice. Together, these results reveal an antiinflammatory role for GKN2, provide in vivo evidence that links GKN2 loss to GC pathogenesis, and suggest GKN restoration as a strategy to restrain GC progression.

Authors

Trevelyan R. Menheniott, Louise O’Connor, Yok Teng Chionh, Jan Däbritz, Michelle Scurr, Benjamin N. Rollo, Garrett Z. Ng, Shelley Jacobs, Angelique Catubig, Bayzar Kurklu, Stephen Mercer, Toshinari Minamoto, David E. Ong, Richard L. Ferrero, James G. Fox, Timothy C. Wang, Philip Sutton, Louise M. Judd, Andrew S. Giraud

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Figure 2

Targeted deletion of the mouse Gkn2 locus.

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Targeted deletion of the mouse Gkn2 locus. (A) Strategy used to target t...
(A) Strategy used to target the mouse Gkn2 locus. Red arrowheads denote primer pairs used to amplify WT and targeted alleles by PCR. Scale bar: 1 kbp. (B) PCR genotyping strategy. WT and targeted alleles generate 442-bp and 214-bp bands, respectively. Upper and lower bands show 500 and 200 bp, respectively. M, molecular weight marker of the 100-bp ladder. (C) Immunoblot showing GKN2 protein expression in WT, Gkn2+/–, and Gkn2–/– gastric corpus tissues. Molecular weights (kDa) of protein bands are shown. Histograms show densitometry of protein bands/GAPDH (n = 3 per group). P values were determined using a 2-tailed Student’s t test: ***P < 0.001. (D–K) β-Gal reporter activity in Gkn2+/– and WT stomachs stained with X-gal (bright blue stain). (D and E) Macroscopic images of X-gal–stained stomachs. Boxed areas are shown at higher magnification in F and G. Scale bar: 5 mm. (F and G) High-magnification images showing (F) proximal and (G) distal extents of β-gal staining in Gkn2+/– stomachs. Scale bar: 500 μm. Original magnification, ×2 (inset). (H–K) Histological images of (H and I) corpus and (J and K) antral β-gal staining in Gkn2+/– stomachs (dark blue), with hematoxylin counterstain (pale blue). Scale bar: 50 μm (H and J); 20 μm (I and K). (L–N) Immunofluorescent colocalization of GKN2 and β-gal proteins in Gkn2+/– corpus mucosa. Boxed areas are shown at higher magnification in M and N. Scale bar: 50 μm (L); 20 μm (M); 5 μm (N).