Cyclic mechanical stresses alter erythrocyte membrane composition and microstructure and trigger macrophage phagocytosis

Abstract

Red blood cells (RBCs) are cleared from the circulation when they become damaged or display aging signals targeted by macrophages. This process occurs mainly in the spleen, where blood flows through submicrometric constrictions called inter-endothelial slits (IES), subjecting RBCs to large-amplitude deformations. In this work, we circulated RBCs through microfluidic devices containing microchannels that replicate the IES. The cyclic mechanical stresses experienced by the cells affected their biophysical properties and molecular composition, accelerating cell aging. Specifically, RBCs quickly transitioned to a more spherical, less deformable phenotype that hindered microchannel passage, causing hemolysis. This transition was associated with the release of membrane vesicles, which self-extinguished as the spacing between membrane-cytoskeleton linkers became tighter. Proteomics analysis of the mechanically aged RBCs revealed significant losses of essential proteins involved in antioxidant protection, gas transport, and cell metabolism. Finally, we show that these changes made mechanically aged RBCs more susceptible to macrophage phagocytosis. These results provide a comprehensive model to explain how physical stress induces RBC clearance in the spleen.

Supplementation with uric and ascorbic acid protects stored red blood cells through enhancement of non-enzymatic antioxidant activity and metabolic rewiring

“…Despite the −not clinically significant− slight increase in the osmotic fragility of AA samples, the same supplementation resulted in resistance to mechanical lysis, a finding consistent with the bibliography [ 22 ]. Mechanical fragility reflects the existence of sublethal injuries in stored cells that render them prone to removal post-transfusion, as recently shown by using microfluidic devices [ 44 ]. The unaltered Ca 2+ homeostasis and control levels of calpain recruitment to the membrane are in line with the (mostly) unchanged mechanical stability and the control levels of discocytes in the treated sub-groups, at least in late storage.…”

The Post-Storage Performance of RBCs from Beta-Thalassemia Trait Donors Is Related to Their Storability Profile

“…The preferential linkage of fragility indices with RBC survival in the group of heterozygotes is extremely fascinating. It is known that poorly deformable RBCs are retained in the spleen [50], while replication of the inter-endothelial slits in vitro via microfluid devices suggested that the mechanically aged RBCs are susceptible to phagocytosis, showing in parallel loss of proteins involved in cytoskeleton architecture, cell metabolism, antioxidant protection, and microvesiculation of the membrane [51]. Of note, rapid clearance of nonreversible micro-erythrocytes decreases transfusion recovery [52]; a finding consistent with the higher levels of circulating βThal + RBCs in the blood stream of our transfused mice, since these cells are reported to resist non-reversible transformation during middle/late storage [6].…”

Physical mechanisms of red blood cell splenic filtration

“…It is therefore essential that our body regularly tests the deformability of RBCs in order to eliminate damaged ones. The RBC selection occurs in the sinusoids of the red pulp of the spleen that each RBC visits approximatively every 200 minutes (9, 10). To return to the vascular system, RBCs must squeeze through slits of 0.25-1.2 μm in width, 0.9 −3.2 μm in length, and around 5 μm in depth (11, 12) between neighboring cells of the vein endothelium where they undergo extreme deformation as shown in electron microscopy images of RBCs in splenic slits (12, 13).…”