Red Blood Cell Extracellular Vesicles Assist Macrophages in Atherosclerotic Plaque
Red blood cells lack a nucleus, but still undertake a range of interesting activities. For example, they release extracellular vesicles. Researchers here note that these vesicles are protective for macrophages that ingest them, protecting the macrophages from being overwhelmed by excess cholesterol in the environment of an atherosclerotic plaque. Macrophages are responsible for cleaning up excess deposition of cholesterol in blood vessel walls, and when they falter at this task in later life, a tipping point is reached at which atherosclerotic plaque begins to form. Evidently red blood cell extracellular vesicles are not enough to prevent this issue from occurring, but scientists are in search of mechanisms that might be enhanced to protect macrophages and allow them to prevent and repair plaque before it grows to the point of causing narrowing of blood vessels, heart attack, and stroke.
Extracellular vesicles (EVs) can be produced from red blood cells (RBCs). The EVs that originate from red blood cells (RBCEVs) have favourable characteristics for serving as an effective drug delivery platform. They are devoid of DNA and inherit their allogenic transfusion compatibility traits from RBCs, hence potentially providing safe, 'off-the-shelf' medication. In addition, RBCs can be readily collected from volunteers and stimulated with calcium ionophore to release large amounts of RBCEVs.
Since EVs are complex entities which act as carriers of biological agents that can modulate their target cells, applying them for therapeutic purposes requires an in-depth understanding of their interactions with these cells and the potential effects of their various components. In the case of RBCEVs, haemoglobin is the most abundant protein present. In human adults, haemoglobin is mainly present in the form of haemoglobin A. Haemoglobin is safe when carried by RBCs but it is toxic when released from RBCs into the bloodstream and interstitial space due to hemolysis. However, this toxicity of free haemoglobin can be neutralized by haptoglobin, a protein secreted from liver cells. This is because haemoglobin and haptoglobin form a complex that is rapidly processed by macrophages through the CD163 receptor. Upon internalization, the haemoglobin component of the complex is broken down, and the heme groups are processed by an enzyme called heme oxygenase 1 (HO-1).
HO-1 plays a protective role against atherosclerosis. This protective effect is speculated to stem from the catalytically enzymatic degradation of heme by HO-1. During the process, heme is broken down into ferrous ions, CO (which inhibits inflammation), and biliverdin (which has antioxidant properties). Thus we hypothesize that in RBCEVs, haemoglobin is protected in enclosed vesicles, preventing cytotoxicity. In addition, we speculate that the haemoglobin carried by RBCEVs exerts both anti-inflammatory and anti-atherosclerosis effects mediated via the HO-1 pathway when the EVs are taken up by macrophages.
In this study, we investigated the uptake of RBCEVs by macrophages. We also monitored the intracellular trafficking of RBCEVs and the fate of haemoglobin, their most abundant protein cargo. We found that RBCEVs were preferentially taken up by macrophages in the liver and spleen. The EVs then released heme into the cytoplasm via the heme transporter HRG1, which promoted the differentiation of the macrophages to a phenotype characterized by upregulated HO-1 expression, and prevented the accumulation of oxidized low-density lipoproteins (oxLDL) in these cells. This natural therapeutic characteristic of RBCEVs suggests their potential benefits in atherosclerosis treatment, especially when combined with other drug cargoes that can be loaded into and carried by these EVs. In addition, the anti-inflammatory properties of RBCEVs might be effective for the treatment of other inflammatory conditions.