The Effect of Bee Venom Peptides Melittin, Tertiapin, and Apamin on the Human Erythrocytes Ghosts: A Preliminary Study

Red blood cells (RBCs) are the most abundant cells in the human blood that have been extensively studied under morphology, ultrastructure, biochemical and molecular functions. Therefore, RBCs are excellent cell models in the study of biologically active compounds like drugs and toxins on the structure and function of the cell membrane. The aim of the present study was to explore erythrocyte ghost’s proteome to identify changes occurring under the influence of three bee venom peptides-melittin, tertiapin, and apamin. We conducted preliminary experiments on the erythrocyte ghosts incubated with these peptides at their non-hemolytic concentrations. Such preparations were analyzed using liquid chromatography coupled with tandem mass spectrometry. It was found that when higher concentrations of melittin and apamin were used, fewer proteins were identified. Moreover, the results clearly indicated that apamin demonstrates the greatest influence on the RBCs ghosts proteome. Interestingly, the data also suggest that tertiapin exerted a stabilizing effect on the erythrocyte membrane. The experiments carried out show the great potential of proteomic research in the projects focused on the toxin’s properties as membrane active agents. However, to determine the specificity of the effect of selected bee venom peptides on the erythrocyte ghosts, further proteomic research should be focused on the quantitative analysis.

Endocytosis and exocytosis processes of gold nanoparticle with erythrocyte ghosts

The interaction of spherical gold nanoparticles (AuNPs) of 20nm elaborated by Turkevich method with the erythrocytes ghosts (7-8 μm) cell membrane was evaluated. The AuNPs-Membrane interaction was determined using confocal microscope, Uv-Vis spectroscopy and SEM analysis. The result show that nanoparticles larger than 20nm are adhered to the erythrocyte ghost membrane due their size and surface modification. Smaller AuNPs enter onto the cell by simple diffusion through the plasmatic membrane voids, these data may favor the best design and application in the treatments applied in biomedicine.

Single molecule distribution of RhD binding epitopes on ultraflat erythrocyte ghosts

The Rh blood group system plays a key role in transfusion and organ transplant medicine. AFM-based recognition imaging on ulraflat erythrocyte ghost makes it possible to determine the molecular distribution of Rh epitopes over the entire erythrocyte.

Hemolysis in the Spleen Drives Erythrocyte Turnover

Erythrocytes circulate for an average of 120 days before they are removed from the circulation. Various processes and factors have been identified that may contribute to degradation of senescent erythrocytes, but this complex process is still not completely understood. Accumulation of removal signals such as phosphatidylserine exposure, changes in CD47 expression and oxidation of proteins and lipids that render them susceptible to complement deposition, may contribute to recognition and degradation by red pulp macrophages (RPM) of the spleen. However, many questions remain on the exact mechanisms that determine the fate of aged erythrocytes. This is well exemplified in a mouse study in which physiologically aged erythrocytes were found to undergo phagocytosis by RPM in vivo but not in vitro. This finding suggested that the splenic architecture may play an important role in facilitating erythrocyte turnover. Loss of membrane deformability may lead to the initial trapping of aged or damaged erythrocytes in the spleen, an event that precedes their degradation by macrophages. Loss of deformability can explain why certain genetic diseases that affect erythrocyte membrane deformability, such as is the case in sickle cell disease and spherocytosis, result in trapping in the spleen, giving rise to anaemia. Next to loss of deformability, activation of adhesion molecules, such as Lu/BCAM and CD44, specifically on aged erythrocytes has been proposed to contribute to retention of erythrocytes within the spleen, leading to their turnover. In this study we provide evidence that the splenic environment is of key importance in facilitating erythrocyte turnover through induction of hemolysis. Upon isolating human spleen RPM we noted that only a small proportion of the macrophages were in the process of phagocytosing intact erythrocytes. Based on a range of variables, including the number of erythrocytes that are cleared daily, the number of RPM present in the spleen, the degradation rate of erythrocytes as well as differential contribution of spleen and liver to erythrocyte turnover, conservative estimates approximate that at least a 30-fold fewer erythrophagocytic events are observed in RPM than anticipated. Detailed characterization of erythrocyte and macrophage subpopulations from human spleen tissue led to the identification of a large population of erythrocytes that are devoid of hemoglobin, so-called erythrocyte ghosts. By in vivo imaging of the spleen and transfusion experiments we further confirmed that senescent erythrocytes that are retained in the spleen are subject to hemolysis, thereby forming erythrocyte ghosts. Of note, we found that the levels of haptoglobin and hemopexin, two plasma proteins that are involved in scavenging of haemoglobin and heme, respectively, correlate well with the rate of hemolysis that was observed in the spleen. Additionally, we show that the erythrocyte adhesion molecules which are specifically activated on aged erythrocytes, Lu/BCAM and CD44, cause senescent erythrocytes to interact with the extracellular matrix of the spleen. This adhesion molecule-driven retention of senescent erythrocytes, under low shear conditions, was found to result in steady shrinkage of the erythrocytes and ultimately resulted in hemolysis and ghost formation. In contrast to intact senescent erythrocytes, the remnant erythrocyte ghosts were found to be immediately recognized and rapidly degraded (1-3 hours) by RPM, thereby explaining the lack of phagocytosis of intact erythrocytes in the spleen. Together, these data identify hemolysis and ghost formation as key events in the turnover of senescent erythrocytes, which alters our current understanding of how erythrocyte degradation is regulated. Disclosures No relevant conflicts of interest to declare.