In vitro erythrophagocytosis model to study alcohol-mediated heme turnover

Background & Aims The accumulation of hepatic iron still remains poorly understood in alcoholic liver disease (ALD) patients. Our preliminary clinical data suggest that ethanol promotes red blood cell (RBC) turnover. The overexpression of the erythrophagocytosis marker CD163 and other findings also suggest that ethanol causes enhanced sequestration of RBCs rather than direct hemolysis.

Methods RBCs were taken from healthy donors and incubated with oxidizing CuSO4 or other ALD mimicking conditions for 2 hours following co-incubation with differentiated THP-1 macrophages. Heme oxygenase-1 (HO-1), hepcidin and CD163 levels were measured by q-PCR or/and western blotting. Hemin was used or hypo-osmotically lysed RBCs. Finally, we also tested the effects of heme-scavenging haptoglobin (Hp) and hemopexin (Hpx).

Results 24 hours 0.01% to 1% hematocrit (Htc) of lysed RBCs significantly induced HO-1 in THP-1. Hepcidin and CD163 were induced in parallel by lysed RBCs in a dose-dependent manner. In addition, 1% oxidized RBCs induced HO-1 in a time-dependent manner. Hepcidin and CD163 were up-regulated in line with HO-1 induction. Furthermore, HO-1 was gradually induced by different dosages of hemin for 24 hours. Heme degradation production bilirubin and bile acids also enhanced erythophagocytosis while induction of HO-1 was blunted both by Hpx ad Hp.

Conclusion We here establish an in vitro erythrophagocytosis model using differentiated THP-1 macrophages to study in detail enhanced RBC turnover by ALD-mimicking conditions. First data suggest enhanced erythrophagocytosis by bilirubin and bile acids and protection of uncontrolled HO1-mediated heme degradation by Hpx and Hp.

Publikationsverlauf

Artikel online veröffentlicht:
26. Januar 2022