This work was supported in part by grants from the NIH (R01-098014 to SLS, K08-HL103756 to EAH) and a Louis V. Gerstner Scholars Award (to EAH).
Macrophages clear refrigerator storage–damaged red blood cells and subsequently secrete cytokines in vivo, but not in vitro, in a murine model
Version of Record online: 20 JUL 2014
© 2014 AABB
Volume 54, Issue 12, pages 3186–3197, December 2014
How to Cite
Wojczyk, B. S., Kim, N., Bandyopadhyay, S., Francis, R. O., Zimring, J. C., Hod, E. A. and Spitalnik, S. L. (2014), Macrophages clear refrigerator storage–damaged red blood cells and subsequently secrete cytokines in vivo, but not in vitro, in a murine model. Transfusion, 54: 3186–3197. doi: 10.1111/trf.12755
- Issue online: 11 DEC 2014
- Version of Record online: 20 JUL 2014
- Manuscript Accepted: 24 APR 2014
- Manuscript Revised: 22 APR 2014
- Manuscript Received: 2 NOV 2013
- NIH. Grant Numbers: R01-098014, K08-HL103756
- Louis V. Gerstner Scholars Award
In mice, refrigerator-stored red blood cells (RBCs) are cleared by extravascular hemolysis and induce cytokine production. To enhance understanding of this phenomenon, we sought to model it in vitro.
Study Design and Methods
Ingestion of refrigerator-stored murine RBCs and subsequent cytokine production were studied using J774A.1 mouse macrophage cells and primary murine splenic macrophages. Wild-type and Ccl2-GFP reporter mice were used for RBC clearance in vivo.
Although J774A.1 cells and primary macrophages preferentially ingested refrigerator-stored RBCs in vitro, compared to freshly isolated RBCs, neither produced increased cytokines after erythrophagocytosis. In contrast, phagocytosis of refrigerator-stored RBCs in vivo induced increases in circulating monocyte chemoattractant protein-1 (MCP-1) and keratinocyte chemoattractant (KC) and correspondingly increased mRNA levels in mouse spleen and liver. In the spleen, these were predominantly expressed by CD11b+ cells. Using Ccl2-GFP reporter mice, the predominant splenic population responsible for MCP-1 mRNA production was tissue-resident macrophages (i.e., CD45+, CD11b+, F4/80+, Ly6c+, and CD11clow cells).
J774A.1 cells and primary macrophages selectively ingested refrigerator-stored RBCs by phagocytosis. Although cytokine expression was not enhanced, this approach could be used to identify the relevant receptor–ligand combination(s). In contrast, cytokine levels increased after phagocytosis of refrigerator-stored RBCs in vivo. These were primarily cleared in the liver and spleen, which demonstrated increased MCP-1 and KC mRNA expression. Finally, in mouse spleen, tissue-resident macrophages were predominantly involved in MCP-1 mRNA production. The differences between cytokine production in vitro and in vivo are not yet well understood.