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Investigation of Macrophage Polarization Using Bone Marrow Derived Macrophages
Published on: June 23, 2013
Iron alters macrophage polarization status and leads to steatohepatitis and fibrogenesis
Priya Handa1, Sunil Thomas2, Vicki Morgan-Stevenson1
1Organ Care Research and Liver Care Network, Seattle, Washington, USA.
Abstract:
We have previously demonstrated that iron overload in hepatic reticuloendothelial system cells (RES) is associated with severe nonalcoholic steatohepatitis (NASH) and advanced fibrosis in patients with nonalcoholic fatty liver disease (NAFLD). Recruited myeloid-derived macrophages have gained a pivotal position as drivers of NASH progression and fibrosis. In this study, we used bone marrow-derived macrophages (BMDM) from C57Bl6 mice as surrogates for recruited macrophages and examined the effect of iron on macrophage polarization. Treatment with iron (ferric ammonium citrate, FAC) led to increased expression levels of M1 markers: CCL2, CD14, iNOS, IL-1β, IL-6, and TNF-α; it also increased protein levels of CD68, TNF-α, IL-1β, and IL-6 by flow cytometry. This effect could be reversed by desferrioxamine, an iron chelator. Furthermore, iron loading of macrophages in the presence of IL-4 led to the down-regulation of M2 markers: arginase-1, Mgl-1, and M2-specific transcriptional regulator, KLF4. Iron loading of macrophages with IL-4 also resulted in reduced phosphorylation of STAT6, another transcriptional regulator of M2 activation. Dietary iron overload of C57Bl6 mice led to hepatic macrophage M1 activation. Iron overload also stimulated hepatic fibrogenesis. Histologic analysis revealed that iron overload resulted in steatohepatitis. Furthermore, NAFLD patients with hepatic RES iron deposition had increased hepatic gene expression levels of M1 markers, IL-6, IL-1β, and CD40 and reduced gene expression of an M2 marker, TGM2, relative to patients with hepatocellular iron deposition pattern. We conclude that iron disrupts the balance between M1/M2 macrophage polarization and leads to macrophage-driven inflammation and fibrogenesis in NAFLD.
Insights
Iron overload in nonalcoholic fatty liver disease (NAFLD) promotes M1 macrophage activation, driving inflammation and fibrosis. Restoring iron balance may reverse these harmful effects in NAFLD.
Area of Science:
- Hepatology
- Immunology
- Cell Biology
Background:
- Hepatic iron overload in reticuloendothelial system (RES) cells is linked to severe nonalcoholic steatohepatitis (NASH) and fibrosis in nonalcoholic fatty liver disease (NAFLD).
- Myeloid-derived macrophages are key drivers of NASH progression and liver fibrosis.
Purpose of the Study:
- To investigate the impact of iron on macrophage polarization in the context of NAFLD.
- To determine if iron overload promotes M1 (pro-inflammatory) macrophage phenotypes and inhibits M2 (anti-inflammatory) phenotypes.
Main Methods:
- Bone marrow-derived macrophages (BMDM) from C57Bl6 mice were treated with iron (ferric ammonium citrate, FAC) or an iron chelator (desferrioxamine).
- Macrophage polarization markers (M1 and M2) were assessed by flow cytometry and gene expression analysis.
- Dietary iron overload was induced in C57Bl6 mice, and liver histology and macrophage polarization were analyzed.
- Gene expression of M1 and M2 markers in liver biopsies from NAFLD patients with different iron deposition patterns was compared.
Main Results:
- Iron treatment of BMDM increased M1 markers (CCL2, CD14, iNOS, IL-1β, IL-6, TNF-α) and protein levels (CD68, TNF-α, IL-1β, IL-6), effects reversed by desferrioxamine.
- Iron loading suppressed M2 markers (arginase-1, Mgl-1, KLF4) and STAT6 phosphorylation in IL-4 stimulated macrophages.
- Dietary iron overload in mice induced hepatic M1 macrophage activation, steatohepatitis, and liver fibrogenesis.
- NAFLD patients with hepatic RES iron deposition showed increased M1 markers (IL-6, IL-1β, CD40) and decreased M2 marker (TGM2) expression compared to those with hepatocellular iron deposition.
Conclusions:
- Iron disrupts the M1/M2 macrophage polarization balance, favoring pro-inflammatory M1 phenotypes.
- Iron overload drives macrophage-mediated inflammation and fibrogenesis in NAFLD.
- Targeting iron metabolism may be a therapeutic strategy for managing NAFLD-associated inflammation and fibrosis.
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