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. 2021 Feb 18:12:632682.
doi: 10.3389/fphys.2021.632682. eCollection 2021.

Human Red Blood Cells Modulate Cytokine Expression in Monocytes/Macrophages Under Anoxic Conditions

Antonella Antonelli  1 , Emanuele Salvatore Scarpa  1 , Mauro Magnani  1
Affiliations

Human Red Blood Cells Modulate Cytokine Expression in Monocytes/Macrophages Under Anoxic Conditions

Antonella Antonelli et al. Front Physiol. .

Abstract

In the bone marrow (BM) hematopoietic niche, the oxygen tension is usually very low. Such condition affects stem and progenitor cell proliferation and differentiation and, at cellular level regulates hematopoietic growth factors, chemokines and adhesion molecules expression. In turn, these molecules affect the proliferation and maturation of other cellular components of the niche. Due to the complexity of the system we started the in vitro investigations of the IL-6, IL-8, TNFα cytokines expression and the vascular endothelial growth factor (VEGF), considered key mediators of the hematopoietic niche, in human macrophages and macrophage cell line. Since in the niche the oxygen availability is mediated by red blood cells (RBCs), we have influenced the anoxic cell cultures by the administration of oxygenated or deoxygenated RBCs (deoxy RBCs). The results reported in this brief paper show that the presence of RBCs up-regulates IL-8 mRNA while IL-6 and VEGF mRNA expression appears down-regulated. This does not occur when deoxy RBCs are used. Moreover, it appears that the administration of RBCs leads to an increase of TNFα expression levels in MonoMac 6 (MM6). Interestingly, the modulation of these factors likely occurs in a hypoxia-inducible factor-1α (HIF-1α) independent manner. Considering the role of oxygen in the hematopoietic niche further studies should explore these preliminary observations in more details.

Keywords: cytokine expression modulation; hypoxia; macrophages; pO2 variation; red blood cells.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
mRNA levels of IL-8, IL-6, and vascular endothelial growth factor (VEGF) in cell extracts of MonoMac 6 (MM6) cells after anoxia (95% N2, 5%CO2; A–C) and 100 μM CoCl2 (D–F) treatment at different times (hours). Values are expressed as mean ± SD; n = 3, ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; n.s., not significant. Western blotting analysis shows the hypoxia-inducible factor-1α (HIF-1α) protein in total extracts after anoxia (G) and CoCl2 (H) treatment of MM6 cells at the indicated times.
Figure 2
Figure 2
mRNA levels of IL-8, IL-6 and VEGF in cell extracts of human macrophages from peripheral blood after anoxia (95% N2, 5%CO2; A–C) and 100 μM CoCl2 (D–F) treatment at different times (hours). Values are expressed as mean ± SD; n = 3, ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; n.s. not significant. Western blotting analysis shows the HIF-1α protein in total extracts after anoxia (G) and CoCl2 (H) treatment of human macrophages at the indicated times.
Figure 3
Figure 3
mRNA levels of IL-8, IL-6, VEGF and TNFα in cell extracts of MM6 cells (A–D) and human macrophages (E–H) after the incubation with red blood cells (RBCs; 10% Ht) deoxygenated or not. Values are expressed as mean ± SD; n = 3; ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 when compared with normoxic samples. §p < 0.05, §§p < 0.01 when compared with anoxic samples. #p < 0.05 when anoxia + RBCs samples were compared with anoxia + deoxy RBCs samples. Western blotting analysis shows the HIF-1α protein in total extracts of MM6 cells (I, 1. Control 2. Anoxia 21 h 3. Anoxia 21 h + RBCs 3 h 4. Anoxia 21 h + deoxy RBCs 3 h 5. Anoxia 21 h + air 3 h; and 6. 100 μM CoCl2 21 h) or human macrophages (J, 1. Control 2. Anoxia 6 h + air 3 h 3. Anoxia 6 h 4. Anoxia 6 h + RBCs 3 h 5. Anoxia 6 h + deoxy RBCs 3 h; and 6. 100 μM CoCl2 6 h) after incubation with RBCs (10% Ht) deoxygenated or not.
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