Extracellular calcium-sensing receptor/PTH knockout mice colons have increased Wnt/β-catenin signaling, reduced non-canonical Wnt signaling, and increased susceptibility to azoxymethane-induced aberrant crypt foci

Abstract

Epidemiological evidence suggests increased dietary calcium and dairy products reduce the onset of colon cancer. To understand a role of the colonic extracellular calcium-sensing receptor (CaSR) in calcium-mediated chemoprevention of colon cancer, we induced formation of aberrant crypt foci (ACF) caused by azoxymethane (AOM) injection in ‘rescued’ CaSR−/PTH− (C−/P−) double knockout colons compared with colons from control CaSR+/PTH+ (C+/P+) mice. C−/P− colonic epithelia had increased Wnt/β-catenin signaling as evidenced by 3–8-fold increases in Wnt3a, CyclinD1, and MMP-7 proteins compared with C+/P+ colonic epithelia. The C−/P− colonic epithelia had reduced Wnt5a and Ror2, and a three-fold increase in TNFR1 compared with C+/P+ epithelia. The C−/P− colons and small intestine had extensive neutrophil infiltration with myeloperoxidase (MPO) levels 18-fold higher then C+/P+ small intestine and colon. Saline-injected C−/P− colons had the same number of ACF/cm2 as C+/P+ colons, which were injected with AOM. However, there were eight times more ACF/cm2 in the C−/P− injected with AOM compared with C+/P+ colons, which received AOM. Together our results suggest both inflammation and Wnt/β-catenin signaling are increased in the epithelia of ‘rescued’ CaSR/PTH double knockout colons, and the capacity for non-canonical Wnt signaling through Wnt5a/Ror2 engagement is reduced. The loss of the colonic CaSR increased the number of ACF/cm2 in response to AOM injection, suggesting colonic CaSR may mediate the chemoprotective effect of increased dietary calcium against colorectal cancer observed in humans.