Shape-Shifted Red Blood Cells: A Novel Red Blood Cell Stage?

Abstract

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright–Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells.

Rainbow Trout Erythrocytes ex vivo Transfection With a DNA Vaccine Encoding VHSV Glycoprotein G Induces an Antiviral Immune Response

“…Besides, we have not found any report showing nucleated RBCs expressing the protein encoded by a DNA vaccine in vivo . GVHSV-transfected RBCs showed a characteristic perinuclear expression of GVHSV protein that appeared in the perinuclear region of stressed RBCs or on the membrane of shape-shifted RBCs (shRBC) ( 33 ). shRBCs are small, round cells with a thin membrane derived from RBCs subjected to stressful conditions.…”

“…shRBCs are small, round cells with a thin membrane derived from RBCs subjected to stressful conditions. Translation of the GVHSV gene into protein and translocation to the cell membrane induces RBC transformation into stressed RBC and shRBC ( 33 ). Like RBCs, shRBCs have been shown to participate in roles related to immune response and homeostasis ( 33 ).…”

Immune Functions of Erythrocytes in Osteichthyes

“…In the rainbow trout (O. mykiss), immune responses also involve shRBCs (shape-shifted RBCs) whose morphology has been changed owing to thermal stress in the in vivo environment (5). They are characterized by, e.g., a round shape, hemoglobin loss, and expression of new, IgM-like markers (5). These cells show the ability to inhibit VHSV infection as well as antiviral activity, which is manifested by increased expression of IL8, IL1, IFNγ, and NKEF.…”

“…In Osteichthyes, RBCs can also contribute to elimination of pathogens/antigens, bound in the immune complexes containing the complement, through their recognizing and binding by the complement receptor 1 (CR1) (12). These cells show phagocytic properties (erythrophagocytosis) (12,13), are capable of reacting with various pathogen-associated molecular patterns via pattern recognition receptors, e.g., peptidoglycan recognition proteins, as well as Toll-like receptors (TLRs) and retinoic acid-inducible gene-I-like receptors (RIG-I-like receptors) (2,5,14). Moreover, erythrocytes of Osteichthyes are probably involved in the immune response as the antigen-presenting cells (APCs), via major histocompatibility complex (MHC) class II antigens, and they can contribute to formation of immune synapse with T cells and NK (natural killer) cells (15).…”

Nucleated Red Blood Cells Contribute to the Host Immune Response Against Pathogens

“…In addition, it has been shown that chicken RBCs stimulated with fungal species Candida albicans release cytokine-like factors with IFNγ-like activity [33]. The conditioned medium of shape-shifted RBCs (shRBCs), a new cell stage of rainbow trout RBCs, induced communication with rainbow trout stromal pronephros cells (TPS-2 cells) and resulted in the upregulation of IFNγ-activated genes in this cell line [34].…”