title: Enhanced suicidal erythrocyte death in mice carrying a loss-of-function mutation of the adenomatous polyposis coli gene.
abstract: 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.
authors.affiliation: Department of Physiology, University of Tübingen, Tübingen, Germany.
authors.forename: Syed M
authors.initials: SM
authors.lastname: Qadri
authors.forename: Hasan
authors.initials: H
authors.lastname: Mahmud
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Lang
authors.forename: Shuchen
authors.initials: S
authors.lastname: Gu
authors.forename: Diwakar
authors.initials: D
authors.lastname: Bobbala
authors.forename: Christine
authors.initials: C
authors.lastname: Zelenak
authors.forename: Kashif
authors.initials: K
authors.lastname: Jilani
authors.forename: Alexandra
authors.initials: A
authors.lastname: Siegfried
authors.forename: Michael
authors.initials: M
authors.lastname: Föller
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics*; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D002118:Calcium / Q000378:metabolism; D061207:Calcium Ionophores / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000187:drug effects / Q000378:metabolism; D004912:Erythrocytes / Q000187:drug effects / Q000502:physiology*; D005260:Female; D017491:Genes, APC; D015759:Ionomycin / Q000494:pharmacology; D008297:Male; D051379:Mice; D009154:Mutation; D010718:Phosphatidylserines / Q000378:metabolism; D026941:Sodium Channel Blockers / Q000494:pharmacology; D013154:Spleen / Q000502:physiology
chemical_list: D025601:Adenomatous Polyposis Coli Protein; D017304:Annexin A5; D061207:Calcium Ionophores; D010718:Phosphatidylserines; D026941:Sodium Channel Blockers; D015759:Ionomycin; D000584:Amiloride; D002118:Calcium