The differentiation of HSCs to erythroid cells is regulated by various cytokines and growth factors ( Table 2 ). The first stage of erythropoiesis is regulated by hematopoietic cytokines, such as stem cell factor (SCF; also known as c-Kit ligand), granulocyte-macrophage colony-stimulating factor, interleukin-3 (IL-3), thrombopoietin, IL-11, and transforming growth factor β (TGF-β). Further erythropoiesis is mainly regulated by erythropoietin (EPO), and iron metabolism is essential for hemoglobin synthesis. GATA1, GATA2, KLF1, and TAL1 are key transcription factors involved in erythropoiesis, while the transcription factors, FOG1, and BCL11A, regulate the expression of genes encoding enzymes associated with heme biosynthesis and hemoglobin production (62). Factors involved in the erythropoiesis are listed in Table 2 .

Stress erythropoiesis is a stem cell-based tissue regeneration response that occurs in the spleen and fetal liver (76). Anemia or hypoxia accompanied by inflammation, which occur frequently during cancer development (77, 78), chronic infection (79), severe trauma (80), and chronic psychological stress (81, 82), disrupt the homeostasis between erythroid cell production through steady-state erythropoiesis and clearance of senescent or damaged erythroid cells by phagocytes, inducing stress erythropoiesis (79, 83); this process is regulated by bone morphogenetic protein 4 (BMP4), SCF, Hedgehog, EPO, growth-differentiation factor 15 (Gdf15), and glucocorticoids (GCs) ( Figure 2 ) (84). Under homeostatic conditions, low EPO levels support terminal differentiation of only the most EPO-sensitive progenitors, while other erythroid progenitors undergo apoptosis; however, during stress erythropoiesis, increased EPO levels induce massive and rapid terminal differentiation of all erythroid progenitors (56). In addition, BMP4 and Hedgehog signals restrict the transition of short-term-HSCs to EPO-sensitive stress erythroid progenitors. Immature stress-induced erythroid progenitors maintain stem cell properties, including self-renewal, and can be serially transplanted (84–87). Further, BMP4 and SCF are required for the expansion of stress BFU-E spleen cells under hypoxic conditions (88), while Gdf15 regulates murine stress erythroid progenitor proliferation and controls stress erythropoiesis niche development (89). GCs are also essential for immature erythroid cell expansion during stress erythropoiesis, and act by binding and modulating the transcriptional activity of their cognate nuclear receptor, glucocorticoid receptor (GR) (90).

Although biomarkers of BFU-E erythroid progenitors (Lin^-cKit⁺CD71^LowCD150⁺CD9⁺Sca-1^-) responsive to stress erythropoiesis in the murine spleen are the same as those detected during steady state (91), whole genome transcriptional analysis demonstrated that mouse stress-BFU-E gene signatures are more BMP4-responsive and associated with erythropoiesis and proliferation, relative to those detected in the steady-state (92). Single-cell RNA-seq analysis of human stress-induced erythroid progenitors also revealed a distinct sub-population to that observed under steady-state erythropoiesis (93). Furthermore, splenic BFU-E exhibit different growth properties to their bone marrow counterparts; splenic BFU-E require only EPO to form colonies, while bone marrow BFU-E require EPO and a second factor (94).

Factors upstream of stress erythropoiesis have fundamental immunomodulatory effects. EPO is the principal cytokine regulating erythropoiesis through EPOR; however, EPOR is expressed not only on erythroid cells, but also on immune cells, such as macrophages, dendritic cells (DCs), mast cells, and lymphocytes (40). EPO can bind to EPOR and tissue-protective receptor (TPR, an EPOR/CD131 heterodimer), which are important in tissue protection and immune regulation (95). EPO inhibits the induction of genes encoding proinflammatory factors, such as TNF-α and inducible nitric oxide (NO) synthase (iNOS), in activated macrophages by decreasing NF-κB p65 activation (96). In addition, EPO suppresses DC maturation through the Jak2/STAT-3/SOCS1 pathway (97). Furthermore, EPO directly promotes regulatory T cell (Treg) proliferation, while inhibiting the expansion of conventional T cells via molecular crosstalk with the IL-2 pathway (98). GCs are required for regulation of stress erythroid progenitor expansion (90); however, GR signaling also has potent anti-inflammatory effects (99). Stress erythropoiesis produces more RBCs and CECs, and both populations possess considerable immunomodulatory functions under various conditions (see below for further details).

RBCs modulate innate and adaptive immunity mainly through their surface molecules (proteins, lipids, and carbohydrates) and potent antioxidant capacity (102); they express large amounts of the key complement regulators, CD55, CD59, and complement receptor type 1 (CR1, also referred to as CD35), where CD55 inactivates C3 convertases generated by all three complement activation pathways, CD59 prevents membrane attack complex formation by preventing C9 incorporation, and CR1 recognizes collagen-like regions of C1q, mannose-binding lectin, C3b, and C4b, to remove complement-tagged inflammatory particles. For example, in patients with HIV, the virus binds to RBCs via C1q-CR1 interaction (103). Further, CR1 is decreased on RBCs in patients with coronavirus disease 2019 (COVID-19), resulting in consistent inflammatory responses and tissue damage (104, 105).

Toll-like receptor 9 (TLR9), a nucleic acid sensing receptor, is expressed on the surface of mammalian RBCs. Under basal conditions, RBCs bind cell-free mitochondrial DNA (mtDNA) through TLR9 and mediate DNA scavenging to prevent unnecessary inflammation (106). Further, in the context of inflammation, TLR9 binds to CpG-containing DNA derived from bacteria, plasmodia, and mitochondria, which drives innate immune activation and red cell clearance (107). Erythroid-specific TLR9 deletion blocks erythrophagocytosis and decreases local and systemic cytokine production (107). During viral pneumonia and sepsis secondary to COVID-19, RBCs also exhibit protein oxidation, together with decreased antioxidant capacity, increased glycolysis, an altered membrane lipidome, and elevated mtDNA binding, which may contribute to anemia and disease severity (102, 108).

Moreover, RBCs can induce DCs toward an immature/tolerogenic phenotype in response to lipopolysaccharide (LPS), through a CD47-dependent mechanism (109). Mechanosensing by RBCs also ensures exposure of splenic type-2 conventional DCs to blood flow, allowing them to capture circulating antigens, while retaining them in the spleen through CD55-CD97 signaling (110); the same mechanism is also important for marginal B cell retention and function (111). In addition, Duffy blood antigen, primarily expressed on the surface of RBCs, can potently bind to multiple chemokines (112, 113).

Transfusion of fresh RBCs under noninflammatory conditions will reduce RBC clearance and therefore lessen macrophage loading with heme, as well as up-regulating heme oxygenase (HO), shifting macrophages toward the anti-inflammatory M2 state (112). Protein factors released from RBCs, such as Hb and peroxiredoxin II, can sustain normal and leukemic T cell growth and survival (113). RBCs can also synergize with TCR/CD3-mediated activation signals and enhance T cell survival and proliferation through a calcineurin-dependent mechanism (111).

Programmed mitochondrial removal occurs during normal erythropoiesis (114). A hypoxia-inducible factor-mediated metabolic switch and consequent activation of the ubiquitin-proteasome system precede, and are necessary for, autophagic mitochondria removal, and disruption of this pathway leads to accumulation of RBCs carrying mitochondria (Mito⁺ RBCs) (115). This process is defective in patients with systemic lupus erythematosus (SLE) (115) and sickle cell disease (SCD) (116), as well as in aged mtDNA mutator mice (117). In patients with SLE, Mito⁺ RBC levels are correlated with disease activity, and antibody-mediated Mito⁺ RBC internalization by macrophages induces type I interferon (IFN-I) production through cGAS/STING activation (115), while Mito⁺ RBCs may contribute to SCD pathophysiology via high reactive oxygen species (ROS) production (118).

RBCs are frequently exposed to various stressful conditions during their lifespan, including oxidative stress, osmotic shock, and mechanical squeezing (119), and consequently accumulate damage which influences their functions. Senescent RBCs show pathologic properties, including decreased deformability (120), MP release (121), increased hemin-carrying Hb (122), and surface antigen modification (123). RBC senescence occurs alongside oxidative stress and in turn becomes a source of ROS, which serves as an important signal of RBC senescence (124). Accumulation of oxidized lipids, such as 4-hydroxynonenal, may induce vascular inflammation (125, 126). At the molecular level, the major features of senescent RBCs are Band 3 clustering or breakdown (127), PS externalization (128), loss of glycophorin A, and reduction of CD47 expression (124). Consequently, senescent RBCs lose the ability to control LPS-induced DC maturation (129).

Oxidized or senescent RBCs or RBC-derived MPs are potential modifiers of T cell responses, which enhance mitogen-driven T cell proliferation and apoptosis through an antigen presenting cell- and cell contact-dependent mechanism, and regulate IFN-γ production from T helper 1 cells (124). Moreover, oxidized RBCs release Hb, heme, and iron which are both sources of radicals and able to activate ECs (130) and innate immune cells, such as monocytes (131), in a proinflammatory manner, as detailed below (see section 3.4). Stored RBCs display senescence-related changes, such as reduced structural integrity, MP release, and iron overload, and the transfusion of stored RBCs exacerbates existing lung inflammation and promotes lung injury, due to loss of Duffy antigen expression and their chemokine scavenging function during storage (132). Further, rapid clearance of transfused stored RBCs by macrophages polarizes the macrophages toward the classical M1 phenotype, with a huge Hb iron load (112, 133). In addition, packed RBCs suppress T cell proliferation via cell-cell contact and inhibit T cell activation via ROS-dependent signaling (134).

RBCs are highly differentiated cells with an elegant structure that allows them to survive under continuous shear stress when transiting, making them ideal messengers between distant organs. The erythrocyte membrane skeleton is a polygonal 2D lattice structure, consisting of lipids, proteins, and carbohydrates (135, 136). The skeleton attaches to the cell membrane through the spectrin-actin junctional complex (adducin, dematin, and P4.1 interact with band 3, GPC/D, and Glut1) and the ankyrin complex (137). Disorders of the RBC cytoskeleton or dehydration cause hemolytic anemia, which is associated with altered immune regulation, as hemolysis breakdown products, including hemoglobin, heme, and iron, have remarkable effects on immunological functions (138).

Erythroid cells play a crucial role in immunological regulation during the neonatal period and in maternal-fetal tolerance. Mouse placental erythroid cells are mainly CD45⁺ and secrete the chemokines, CCL2, CCL3, CCL4, and CXCL1 (193). Further, CECs are abundant in mouse neonatal spleen and human cord blood, and possess unique immunosuppressive properties (167). CECs are abundant in the liver of children with biliary atresia (BA), and suppress the activation of hepatic mononuclear cells (172). Further, CECs are numerous in the peripheral blood of human newborns, but decline rapidly by 4 weeks of age (168).

CECs influence neonatal infections through various mechanisms. Bordetella pertussis is a common neonatal respiratory tract pathogen and CECs prevent the recruitment of immune cells to the mucosal infection site (167). CECs from human newborn peripheral blood mononuclear cells (PBMCs) suppress TNF-α production by CD14⁺ monocytes and IFN-γ production by T cells (168). Further, ablation of CECs enhances the innate immune response by increasing the production of protective cytokines, including IL-17, IFN-γ, TNF-α, and IL-12 in B. pertussis-infected lungs (169, 170).

L-arginine is essential for T cell proliferation and function (194). Arginase (ARG) depletes L-arginine, thereby inhibiting T cells, and is encoded by two recently-discovered genes, Arg1 and Arg2 (195). ARG1 is expressed in the cytosol, whereas ARG2 localizes to mitochondria. Neonatal and human cord blood CECs express ARG2 and ablation of CECs augments B. pertussis-specific T cell responses in the lung and spleen on re-infection or vaccination (170). In addition, ablation of CECs also induces enhanced systemic and mucosal B. pertussis-specific antibody responses (170). Accordingly, CECs in human cord blood can suppress T and B cell functions in vitro (170). Regarding innate immunity, CECs inhibit B. pertussis phagocytosis via ARG2 in vitro (169, 170). Such effects of CECs facilitate intestinal colonization with commensal microbes during the neonatal period (167). Depletion of CECs in neonatal mice renders them more resistant to infections by Listeria monocytogenes, Escherichia coli, and B. pertussis, indicating the protective effects against neonatal infectious diseases (167, 168); however, ablation of CD71⁺ cells failed to modify neonatal mortality in either a model of endotoxin challenge or a model of polymicrobial sepsis (173).

BA is a rare and progressive disease that develops in early infancy (196). Rhesus rotavirus (RRV) infection of neonatal mice induces an obstructive cholangiopathy, which is similar to BA (197). CECs expand in the liver of children with BA or RRV-infected mice and suppress the immune response by reducing TNF-α production. Preemptive depletion of hepatic CD71⁺ erythroid cells in neonatal mice augments the number of effector lymphocytes and delays RRV infection of the liver and extrahepatic bile duct, suppressing bile duct injury (172). Clearance of CECs before RRV infection renders mice resistant to RRV-induced BA, while repopulation of CD71⁺ erythroid cells after RRV inoculation promotes long-term survival (172).

CEC-mediated immunosuppression is crucial for fetomaternal tolerance. Both BALB/c and C57BL/6 female mice, and human women are enriched for CECs (198). Further, analysis of 155 umbilical cord blood samples showed that the proportion of CECs was 50-fold higher in cord blood than that in maternal blood (199). Erythropoiesis becomes active during pregnancy, and erythrocytes significantly expand in the peripheral blood (200). TGF-β has an important role in regulating the erythroid lineage differentiation potential of HSCs (201, 202). CECs in pregnant mice express more PD-L1/PD-L2 and suppress T cells expressing programmed cell death protein-1 (PD-1) at the fetomaternal interface (174). Maternal CECs inhibit IFN-γ and TNF-α production to protect the fetus against the allogeneic response. Further, fetal liver CECs also exhibit immunosuppressive activity. A recent transcriptional study demonstrated expression of high levels of galectin-9, galectin-1, and VISTA on the surface of neonatal splenic CECs. CD71⁺VISTA⁺ cells produce more TGF-β than CD71⁺VISTA⁻ cells, and can promote CD4⁺ T cell differentiation into Tregs (171); however, CECs in human cord blood express negligible amounts of VISTA. Indeed, VISTA expression levels are significantly higher in placental CECs than those in cord blood (171). Thus, both maternal and fetal CECs are essential for fetomaternal tolerance (175). Accordingly, depletion of CECs in pregnant mice induces a proinflammatory immune response, by reducing IL-4 and IL-10 production, while increasing TNF-α and IL-6 levels in placental tissues, which in turn results in fetal resorption (175, 203); however, in pregnant women with inflammatory bowel disease (IBD), CECs are decreased in the peripheral blood, cord blood, and placenta tissue, and express lower levels of inhibitory molecules, including VISTA, TGF-β, and ROS. Accordingly, pregnant women with IBD have lower levels of Tregs and increased immune-activation. Patients with IBD are more likely to have a pro-inflammatory environment in the gastrointestinal tract, which leads to impairment of CECs during pregnancy (176).

CECs not only function during neonatal infections, they participate in various infections throughout life.

Acquired immune deficiency syndrome is a systemic disease caused by human immunodeficiency virus (HIV), the genome of which comprises two copies of a 9749 nucleotide sequence packaged in each virion (204). CECs are expanded in the peripheral blood of patients with HIV and there is a positive correlation between CEC frequency and plasma viral load. When cocultured with CD4⁺ T cells, CECs exacerbate HIV-1 infection/replication, by enhancing NF-κB activation in CD4⁺ T cells to facilitate HIV infection (177). Meanwhile, CECs bind to HIV-1 via CD235a and subsequently transfer the virus to uninfected CD4⁺ T cells. Moreover, in the presence of antiretroviral therapy, CECs from HIV-infected individuals contain infective viral particles, which mediate HIV-1 trans-infection of CD4⁺ T cells (177). CECs are also significantly expanded and possess immunosuppressive properties in the blood of patients with COVID-19. With high levels of ARG2, ARG1, and ROS, CECs mediate immunosuppression by inhibiting CD4⁺ and CD8⁺ T cell production of TNF-α and IFN-γ in vitro (179). Furthermore, CD45⁺ CECs express ACE2, TMPRSS2, CD147, and CD26 and can be infected with SARS-CoV-2 (179).

CECs are also expanded in adult patients with sepsis and serve as predictors of 30-day mortality as well as nosocomial infection development. Low levels of RBCs and high levels of IL-6 and IFN-γ may contribute to the expansion of CECs in sepsis (178). During Salmonella infection, accumulation of CECs in the spleen and increased EPO production are dependent on Myd88/TRIF signaling (182); EPO neutralization reduces the population of CECs in the spleen and slightly improves the host immune response (182).

Malaria is an insect-borne infection caused by the bite of Anopheles mosquitoes, and a major global health problem, with approximately 247 million cases worldwide in 2021 and many more residents of endemic areas having asymptomatic parasitemia (chronic malaria) (205). Different species of malaria parasites exhibit distinct tropism (206). Plasmodium falciparum can invade all stages of erythrocytes while Plasmodium vivax and Plasmodium cynomolgi invade only reticulocytes (207, 208). P. vivax is the most widely distributed human malaria parasite and exhibits a strong preference for immature reticulocytes, with CD71 acting as an anchor receptor (209, 210). Reticulocytes have a more complex and enriched metabolic profile than mature erythrocytes, providing metabolic reservoirs for malaria parasites (206). P. vivax-infected reticulocytes express high levels of human leukocyte antigen class I (HLA-I), which can be specifically detected by cytotoxic CD8⁺ T cells to protect against intracellular parasites (180). In BALB/c mice, reticulocytes can secrete exosomes when infected by the reticulocyte-tropic non-lethal Plasmodium yoelii 17X strain (211). These reticulocyte-derived exosomes carry parasite proteins and are involved in antigen presentation. Mice immunized using purified exosomes produce IgG antibodies that can recognize P. yoelii-infected RBCs and show increased survival time and altered reticulocyte cell tropism of the parasite (181). Furthermore, during P. vivax infection, parasites invariably affect bone marrow CD71⁺ cells, inducing dyserythropoiesis and ineffective erythropoiesis (212). Identification and characterization of the reticulocyte receptor, metabolism, and the underlying mechanisms involved in malaria may provide insights to inform the development of novel antimalarial drugs and vaccines.

Inflammation is the automatic defense response to tissue injury, and can be classified as acute and chronic, according to its duration. Inflammation modifies bone marrow hematopoiesis towards innate immune effector cells at the expense of lymphoid cells and erythrocytes (79). Inflammatory cytokines, such as TNF-α, limit steady-state erythropoiesis and promote granulopoiesis. Further, mature granulocytes contact the central macrophage of EBIs and alter EBI structures, leading to increased numbers of maturing granulocytes and fewer erythroid precursors (213). In chronic inflammation resulting from sterile abscesses, erythropoiesis is impaired at Ter119⁺ stages of erythroid development (185). Although inflammation inhibits erythropoiesis in the bone marrow, inflammatory signals induce stress erythropoiesis in the spleen, to maintain erythroid homeostasis. Inflammatory signaling through TLRs enhances erythrophagocytosis by splenic macrophages and augments expression of the transcription factor, SPI-C. In turn, SPI-C couples with TLR signaling to promote the expression of Gdf15 and Bmp4, which encode ligands that initiate the expansion of stress erythroid progenitors in the spleen (79). The spleen is the largest secondary lymphoid organ, and has a wide range of immunologic functions alongside its roles in erythropoiesis, and splenic erythropoiesis alters the histological structure of spleen to become rich in granulomatous lesions and devoid of clear separation between red and white pulp (214).

Autoimmune diseases comprise a range of disorders in which the immune response to self-antigens results in tissue damage or dysfunction (215). In patients with autoimmune diseases, CECs can inhibit inflammatory responses to prevent excessive inflammation and injury. Experimental autoimmune encephalomyelitis (EAE) is an autoimmune disease mainly mediated by specific sensitized CD4⁺ T cells, which serves as the best experimental model reflecting the autoimmune pathogenesis of human multiple sclerosis (216), and iron-deficient mice fail to develop EAE (217). Management using EPO or its non-erythropoietic derivatives consistently decreases EAE-associated TNF-α, IL-1β, and IL-1Ra production in the spinal cord, and IFN-γ by peripheral lymphocytes, which ameliorates chronic murine EAE (218). IBD inflammation spreads systemically and can cause complications, such as arthritis, cachexia, and anemia. In a CD45-deficient Rag1-deficient mouse model of T cell-induced colitis, an increased number of erythroid progenitors are found in the spleen. These CECs can suppress TNF-α expression in red pulp macrophages in a phagocytosis-dependent manner (183). Further, erythropoiesis-related genes are upregulated in PBMCs of patients with systemic-onset juvenile idiopathic arthritis (SoJIA) (184), while active SoJIA-driven CECs co-cultured with healthy donor monocytes suppress IL-1β, IL-6, and IL-8 secretion. Although ARG2 is the top upregulated gene in SoJIA-driven CECs, cytokine production from monocytes remains suppressed when they are treated using an arginase inhibitor (184).

Tumors are complex ecosystems, comprising tumor cells and various non-neoplastic cells (219), where non-neoplastic cells in the tumor microenvironment play critical roles in cancer development. Targeting the tumor microenvironment is considered a promising approach for cancer intervention (220). CECs are abundant in both the tumor microenvironment and the circulation and their levels can be used to predict tumor recurrence (165).

Tumor-associated myeloid cells include myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), and neutrophils (221), which are important immune cell populations in the tumor microenvironment that are crucial for immune checkpoint blockade efficacy (222). MDSCs can be divided into at least two major subsets: mononuclear MDSCs (M-MDSCs, CD11b⁺Ly6G^-Ly6C^high) and polymorphonuclear MDSCs (PMN-MDSCs, CD11b⁺Ly6G⁺Ly6C^low) (223), where M-MDSCs exert more robust immunosuppression than PMN-MDSCs. Further, erythroid cells can differentiate into myeloid cells in tumors and mediate immunosuppression. Lineage tracking in patients with cancer and tumor-bearing mice revealed that > 30% of erythroid progenitor cells lose their developmental potential and switch to the myeloid lineage, and that these erythroid differentiated myeloid cells (EDMCs) are similar to their myeloid-originated counterparts at the transcription level (166). The phenotypes of EDMCs are CD45⁺CD235a⁺CD71⁺CD11b⁺CD33⁺HLA-DR^- in patients with cancer and CD45⁺Ter119⁺CD71⁺CD11b⁺Gr1⁺ in tumor-bearing mice. EDMCs express more immune inhibitory molecules, including PD-L1, PD-L2, iNOS, ARG1, and CD49, than MDSCs, which may endow EDMCs with the ability to inhibit CD8⁺ T cell proliferation and IFN-γ production. Accordingly, EDMCs reduce the efficacy of anti-PD-1/PD-L1 treatment (166).

In tumors, CD45⁺ CECs exert a strong immune suppressive function, mainly by regulating T cells. In Lewis lung cancer, CD45⁺ CECs are induced by tumor growth-associated extramedullary hematopoiesis (EMH) in the spleen and their transcriptome closely resembles that of MDSCs. As robust immunosuppressors, CD45⁺ CECs hinder both CD8⁺ T cell priming in the spleen and effector function in peripheral tissues (164). In hepatocellular carcinoma (HCC) tissues, CD45⁺ CEC numbers are higher than those of CD45^- CECs. Further, CD45⁺ CECs from patients with HCC inhibit CD4⁺ T cell proliferation and differentiation and suppress CD8⁺ T cell proliferation and cytotoxicity by generating factors including ROS, IL-10, and TGF-β (165). In patients with virus-associated solid tumors, substantially greater expansion of CECs occurs in the blood compared with that in healthy controls. CD45⁺ CECs have more immunosuppressive properties than their CD45^- counterparts, mediated by higher levels of ROS, PD-L1/PD-L2, and VISTA. Further, the abundance of CECs in the circulation may be associated with anemia (187). Moreover, CECs in mice with melanoma secrete artemin, while this is not the case for VISTA⁺ CECs in patients with virus-associated solid tumors (187).

CD45^- CECs have lower immunosuppression abilities than their CD45⁺ counterparts; however, they also play a crucial role in promoting tumor progression. One population of tumor-induced erythroblast-like cells (CD45^-Ter119⁺CD71⁺, Ter-cells) derived from MEPs (186, 224), accumulate in the spleen of patients with terminal cancer and secret artemin, where artemin is a neurotrophic factor with an important role in cancer progression through its induction of Caspase-9 Thr125 phosphorylation, to maintain cell survival, and upregulation of TRIOBP and ITGB5 expression, to promote invasion. Blocking artemin, or its receptor, GFRα3, signaling inhibits HCC growth in vivo (186). In this context, the phenotype of Ter-cells is CD45^-Ter119⁺ CD71⁺CD41⁺CD44⁺, and they mainly exist in the spleen of advanced-tumor bearing hosts; however, a few can also be found in the tumor. TGF-β and Smad3 activation contribute to Ter-cell generation. Moreover, serum artemin levels in patients with HCC are correlated with poor prognosis (186).

Anemia is a common blood disorder characterized by a decreased number of RBCs in the peripheral blood, which is defined as a hemoglobin level less than the 5th percentile for age (225). Anemia is the main cause of EPC expansion by increasing EPO concentration in response to oxygen deficit (226). In mice with anemia, CD45⁺ CECs expand in the spleen and express high levels of ARG2 and ROS. CEC expansion-induced L-arginine depletion suppresses T cell responses. In patients with anemia, CECs expand in the peripheral blood and express ARG1 and ARG2, which suppress IFN-γ production by T cells (189). Furthermore, human erythroleukemia-derived erythroid cell lines, including K562, HEL92.1.7, and TF-1, which express multiple erythroid-lineage markers, such as CD71 and CD235a, suppress T cells in an ARG- and ROS-dependent manner (189). Serum levels of IL-22 are increased in patients with anemia secondary to chronic kidney disease and myelodysplastic syndromes, and the IL-22 receptor, IL-22RA1, is present on erythroid precursors, with blockade of IL-22 signaling alleviating anemia in mice (192).

Anemia is also a common feature of sepsis (227). In patients with sepsis, RBC levels are negatively associated with CD45⁺ CEC frequency, suggesting that anemia may lead to CEC expansion through the EPO pathway (178). EPO can induce the expansion of CD45⁺ CECs, while EPO neutralization prevents infection-related CEC accumulation (188). In patients with COVID19, SARS-CoV-2 infection is associated with lower blood oxygen levels and the numbers of CECs in the blood are negatively correlated with hemoglobin levels; this may be due to the elimination of infected/damaged CECs by lysis and/or phagocytosis. Dexamethasone enhances the maturation of CECs to RBCs by downregulating ACE2 and TMPRSS in a dose-dependent manner (179).

In addition, anemia is very common among patients with cancer and tumor bearing animal models; approximately 30%–90% of patients with cancer have varying degrees of anemia, depending on the type of cancer (228). Immunosuppressive CECs can be detected in patients with cancer and anemia. Further, hematocrit, HGB levels, and mature RBC counts are decreased in the blood of mice after prolonged tumor-bearing, and HGB is negatively correlated with numbers of splenic CECs. Tumor-initiated anemia and EMH may act synergistically to cause splenic CEC accumulation (164): anemia induces EMH, whereas terminal differentiation is blocked in the presence of tumors. RNA sequencing of CD45⁺ and CD45^- CECs generated by anemia induced in different ways revealed that CD45⁺ CECs differ significantly from their CD45^- counterparts, particularly regarding signature genes defining the erythrocyte lineage and immunosuppression. Notably, ROS and NOX-2 are highly expressed in CD45⁺ CECs, particularly those from tumor-bearing individuals (164). EPO has been widely used to overcome hypoxia in patients with cancer. Recombinant human EPO and erythropoiesis-stimulating agents can promote EPC differentiation and maturation to RBCs, and thereby effectively treat anemia; however, these agents do not prolong the survival of patients with cancer (229–231). Immune checkpoint inhibitors (ICIs) targeting co-inhibitory molecules, including PD-1, PD-L1, and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), have been widely applied in the therapy of various tumors (232); however, EPO treatment in patients receiving anti-PD-L1 therapy reduce the therapeutic effects of this monoclonal antibody (187); the underlying mechanism involves EPO induction of continual differentiation of CECs into EDMCs, which mediate systemic immunosuppression against immune surveillance (166).

Erythroid cells participate in several immune conditions and have important roles in regulating immune responses. Further, CECs may have beneficial effects in fetomaternal tolerance and autoimmune diseases; however, in contexts including infection, tumor, and anemia, CECs appear to exert detrimental effects ( Table 6 ). Thus, further understanding of the immune regulatory mechanisms used by erythroid cells can provide new insights into pathogenic mechanisms, and CECs may serve as a novel target in immunological therapies ( Figure 5 ). CECs can have opposite effects in different diseases, and different measures could be selected to manipulate CECs, according to context; for example, promotion/inhibition of CEC expansion, inhibition/promotion of CEC differentiation, and inhibition/promotion of CEC immunosuppressive properties.

Under normal conditions, CECs suppress the immune response to protect tissue from immunologic injury; however, CECs may be impaired in autoimmune diseases and chronic inflammation. Thus, promoting CEC expansion may be beneficial in inflammatory disorders. In contrast, in patients with tumors, infection, or anemia, CECs expand and arrest at an early stage, to inhibit immune activity; therefore, preventing CEC expansion and promoting CEC differentiation may be promising therapies to attenuate immune evasion and enhance immune responses in these contexts.

Targeting RBCs is an underdeveloped therapeutic strategy; however, with their strong ability to contribute to material exchange and high immunocompatibility, RBCs have potential as drug delivery carriers (274, 275). RBC membrane-coated polymeric nanoparticles can effectively deliver doxorubicin in a mouse model of lymphoma (275). Further, RBC extracellular vesicles (RBCEVs) are taken up by leukemia cells with high efficiency, and may serve as a valid vehicle to deliver antisense oligonucleotides to leukemia cells (274). RBCEVs are also used as a vehicle for osteoclast-targeted delivery of anti-miR-214 oligonucleotides. TBP-CP05 is a functional peptide which binds to both CD63 on RBCEVs and receptors on osteoclasts, and TBP-CP05 binds with RBCEVs through CP05 and endows them with osteoclast-targeting ability. Intravenous injection of osteoclast-targeting RBCEVs significantly inhibits osteoclast activity, elevates osteoblast activity, and improves bone density in osteoporotic mice (276). Hence, RBCs have huge potential in cancer and clinical therapy as a novel type of nanoparticle-based RNA drug vehicle.

RBCs also have potential applications in disease diagnosis, prognosis, and monitoring (277). A considerable fraction of cell-free mtDNA, which is associated with trauma, autoimmune disease, sepsis, malignancy, cellular injury, and organ dysfunction, binds to the outer surface of RBCs and can serve as a biomarker (278–280). Furthermore, long DNA fragments which cover most nuclear and mitochondrial genome regions can be detected in RBCs from patients with cancer (281).