Enhanced suicidal erythrocyte death in mice carrying a loss‐of‐function mutation of the adenomatous polyposis coli gene

Loss‐of‐function mutations in human adenomatous polyposis coli (APC) lead to multiple colonic adenomatous polyps eventually resulting in colonic carcinoma. Similarly, heterozygous mice carrying defective APC ( apc Min/+ ) suffer from intestinal tumours. The animals further suffer from anaemia, which in theory could result from accelerated eryptosis, a suicidal erythrocyte death triggered by enhanced cytosolic Ca 2+ activity and characterized by cell membrane scrambling and cell shrinkage. To explore, whether APC‐deficiency enhances eryptosis, we estimated cell membrane scrambling from annexin V binding, cell size from forward scatter and cytosolic ATP utilizing luciferin–luciferase in isolated erythrocytes from apc Min/+ mice and wild‐type mice ( apc +/+ ). Clearance of circulating erythrocytes was estimated by carboxyfluorescein‐diacetate‐succinimidyl‐ester labelling. As a result, apc Min/+ mice were anaemic despite reticulocytosis. Cytosolic ATP was significantly lower and annexin V binding significantly higher in apc Min/+ erythrocytes than in apc +/+ erythrocytes. Glucose depletion enhanced annexin V binding, an effect significantly more pronounced in apc Min/+ erythrocytes than in apc +/+ erythrocytes. Extracellular Ca 2+ removal or inhibition of Ca 2+ entry with amiloride (1 mM) blunted the increase but did not abrogate the genotype differences of annexin V binding following glucose depletion. Stimulation of Ca 2+ ‐entry by treatment with Ca 2+ ‐ionophore ionomycin (10 μM) increased annexin V binding, an effect again significantly more pronounced in apc Min/+ erythrocytes than in apc +/+ erythrocytes. Following retrieval and injection into the circulation of the same mice, apc Min/+ erythrocytes were more rapidly cleared from circulating blood than apc +/+ erythrocytes. Most labelled erythrocytes were trapped in the spleen, which was significantly enlarged in apc Min/+ mice. The observations point to accelerated eryptosis and subsequent clearance of apc Min/+ erythrocytes, which contributes to or even accounts for the enhanced erythrocyte turnover, anaemia and splenomegaly in those mice.