Erythrocyte Plasmalemma and Its Changes During the Cell Lifespan

The article reviews literature on the organization of the erythrocyte plasmalemma and its rearrangements at different periods of the cell lifespan. In the absence of a nucleus and organelles, the plasmalemma is the only structural element of erythrocytes involved in all processes of their vital activity. The plasmalemma supports the disk-like shape of the erythrocyte, provides its ability to reversible deformation, maintains intracellular homeostasis, participates in gas transport and energy metabolism, also transfers hormones, enzymes, antibodies, medicines and other substances on its surface. The polyfunctionality of the plasmalemma is provided by the peculiarities of its lipid, protein, and carbohydrate composition, as well as by the presence of a unique cytoskeleto n, morphologically associated with the erythrocyte membrane. The plasmalemma has the substantial modifications during the erythrocyte lifespan, namely, in maturation of reticulocytes, in the processes of functioning, aging, and cell death. Biochemical rearrangements of the plasmalemma serve as triggers for events such as membrane vesiculation, eryptosis, and elimination of senescent erythrocytes by macrophages. Age-related changes in the erythrocyte plasmalemma are adoptive in nature and aimed at maintaining cellular homeostasis and functional activity of these formed elements during a four-month stay in the bloodstream.

Hemolysis in the spleen drives erythrocyte turnover

Red pulp macrophages of the spleen mediate turnover of billions of senescent erythrocytes per day. However, the molecular mechanisms involved in sequestration of senescent erythrocytes, their recognition and their subsequent degradation by red pulp macrophages remain unclear. In this study we provide evidence that the splenic environment is of substantial importance in facilitating erythrocyte turnover through induction of hemolysis. Upon isolating human spleen red pulp macrophages we noted a substantial lack of macrophages that were in the process of phagocytosing intact erythrocytes. Detailed characterization of erythrocyte and macrophage subpopulations from human spleen tissue led to the identification of erythrocytes that are devoid of hemoglobin, so-called erythrocyte ghosts. By in vivo imaging and transfusion experiments we further confirmed that senescent erythrocytes that are retained in the spleen are subject to hemolysis. Additionally, we show that erythrocyte adhesion molecules, which are specifically activated on aged erythrocytes, cause senescent erythrocytes to interact with extracellular matrix proteins that are exposed within the splenic architecture. Such adhesion molecule-driven retention of senescent erythrocytes, under low shear conditions, was found to result in steady shrinkage of the cell and ultimately resulted in hemolysis. In contrast to intact senescent erythrocytes, the remnant erythrocyte ghost shells were prone to recognition and breakdown by red pulp macrophages. These data identify hemolysis as a key event in the turnover of senescent erythrocytes, which alters our current understanding of how erythrocyte degradation is regulated.

Stress Erythropoiesis is a Key Inflammatory Response

Bone marrow medullary erythropoiesis is primarily homeostatic. It produces new erythrocytes at a constant rate, which is balanced by the turnover of senescent erythrocytes by macrophages in the spleen. Despite the enormous capacity of the bone marrow to produce erythrocytes, there are times when it is unable to keep pace with erythroid demand. At these times stress erythropoiesis predominates. Stress erythropoiesis generates a large bolus of new erythrocytes to maintain homeostasis until steady state erythropoiesis can resume. In this review, we outline the mechanistic differences between stress erythropoiesis and steady state erythropoiesis and show that their responses to inflammation are complementary. We propose a new hypothesis that stress erythropoiesis is induced by inflammation and plays a key role in maintaining erythroid homeostasis during inflammatory responses.

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.

Resolution metabolomes activated by hypoxic environment

Targeting hypoxia-sensitive pathways in immune cells is of interest in treating diseases. Here, we demonstrate that physiologic hypoxia (1% O2), as encountered in bone marrow and spleen, accelerates human M2 macrophage efferocytosis of apoptotic-neutrophils and senescent erythrocytes via lipolysis-dependent biosynthesis of specialized pro-resolving mediators (SPMs), i.e. resolvins, protectins, maresins and lipoxin. SPM-production was enhanced via hypoxia in M2 macrophages interacting with neutrophils and erythrocytes enabling structural elucidation of a novel eicosapentaenoic acid (EPA)–derived resolvin, resolvin E4 (RvE4) that stimulates efferocytosis of senescent erythrocytes and more potently than aspirin in mouse hemorrhagic exudates. In hypoxia, glycolysis inhibition enhanced neutrophil RvE4-SPM biosynthesis. Human macrophage-erythrocyte co-incubations in physiologic hypoxia produced RvE4-SPM from erythrocyte stores of omega-3 fatty acids. These results indicate that hypoxic environments, including bone marrow and spleen as well as sites of inflammation, activate SPM-biosynthetic circuits that in turn stimulate resolution and clearance of senescent erythrocytes and apoptotic neutrophils.

Comparative hematology of wild Anguilliformes (Muraena helena, L. 1758, Conger conger, L. 1758 and Anguilla anguilla L. 1758)

The objective of this study was to compare circulating blood cell counts and morphology of three eel species: Muraena helena (moray), Conger conger (European conger) and Anguilla anguilla (European common eel). Moray and conger were collected from the Adriatic Sea at the Elaphite Islands near Dubrovnik, Croatia; common eels were collected in the Neretva River, Croatia. Hematological comparison was conducted using Natt-Harrick’s and May-Grünwald Giemsa staining methods. The hematocrit of moray and conger were similar, while common eel had higher values by 60%. Manual cell count showed that common eel had the highest erythrocyte count. Conger had a higher erythrocyte count than moray, with a higher percentage of proerythrocytes and senescent erythrocytes compared to moray and common eel. The leukocyte count was similar in common eel and moray and slightly lower in conger. The thrombocyte count was highest in conger and lowest in moray. In all three species, the neutrophil (heterophil) nuclei appeared as either circular or bi-lobed. Moray had the highest neutrophil (heterophil) percentage and a subtype with intensively basophilic cytoplasm appearing in a similar percentage as the normal type. In common eel, neutrophils (heterophils) were the only detected granulocytes. Basophils were detected in conger eels. Eosinophils were not detected in any of the sampled fish. The size of all cell types in moray was slightly larger than in other two species. In conclusion, our findings reveal major differences in the cell count and diversity in cell subtypes between three kin species of eels.