Abstract
Red blood cell (RBC) transfusion is associated with systemic inflammation and immune suppression as adverse outcomes. To investigate the immunomodulatory function of the transfused autologous RBC in altering pro-inflammatory and immunosuppressive effects. A total of 24 Sprague Dawley male rats were randomly divided into 3 groups (n = 8 in each group). Group 1 did not receive blood transfusions, while the other 2 groups of rats separately received transfusion of RBC stored for 14 days (group 2) and 35 days (group 3). The rats were treated with HO-1 inhibitor, HO-1 inducer and nuclear factor erythroid 2-related factor 2 (Nrf2) activator after they separately received autologous transfusion of RBC that were cryopreserved for 14 days or 35 days. The blood samples of the rats were collected 12 h after the transfusion, and the macrophage phenotype of M1 and M2 were analyzed with flow cytometry (FCM). Also, the surface protein expression of CD68 and CD200R in macrophages were analyzed and the inflammatory signals in the serum were measured with enzyme-linked immunosorbent assay (ELISA). Moreover, the location and expression of proteins heme oxygenase 1 (HO-1), arginine 1 (Arg-1) and nitric oxide synthase 2 (NOS2) in macrophage were detected with immunofluorescence (IF). Autologous transfusion of long-time stored ("old") RBC promoted macrophage polarization to M2 phenotype and upregulated the expression of its surface proteins CD68 and CD200R. The pro-inflammatory cytokines tumor necrosis factor α (TNF-α), interleukin (IL)-6, IL-1β, and IL-18 were inhibited, and the secretion of NOS isoforms (iNOS) in serum was reduced with blood transfusion; contrarily, the production of IL-10 and CCL22 was increased. Additionally, HO-1, Arg-1 and NOS2 proteins were located in the cytoplasm, and HO-1 and Arg-1 proteins were highly expressed in macrophage, while the expression of protein NOS2 was low. Moreover, Nrf2, HO-1 and Arg-1 proteins were upregulated in macrophage after receiving "old" RBC transfusion. Autologous transfusion of "old" RBC drove the macrophage phenotype toward M2 macrophages and induced immunosuppressive effects through the IL-10-NRF2-HO-1 signals.
Highlights
Blood transfusion is widely used in clinical practice; it is estimated that the transfusion of red blood cells (RBC) is the major type, accounting for nearly 50% of transfusions.[1]
heme oxygenase 1 (HO-1), arginine 1 (Arg-1) and nitric oxide synthase 2 (NOS2) proteins were located in the cytoplasm, and HO-1 and Arg-1 proteins were highly expressed in macrophage, while the expression of protein NOS2 was low
Nrf[2], HO-1 and Arg-1 proteins were upregulated in macrophage after receiving “old” Red blood cell (RBC) transfusion
Summary
Blood transfusion is widely used in clinical practice; it is estimated that the transfusion of red blood cells (RBC) is the major type, accounting for nearly 50% of transfusions.[1]. Even though low-temperature storage slows down the metabolism of RBC, studies have revealed that the structure and physiological and immunological characteristics of the long-time stored RBC ( named “old” RBC) have changed. The stored RBC induced hemolysis and the release of free heme, hemoglobin and free iron, which play important roles in immunomodulation.[8]. The free heme act as a pro-inflammatory factor engaged in the immune response of monocytes, macrophages, Tregs, and endothelial cells.[12,13]. The microvesicles were found in the stored “old” RBC, which participated in the activity of endothelial activation,[15] blood coagulation[16] and immunomodulation.[17]. The variability of stored RBC is closely associated with systemic proinflammatory and immunosuppressive effects in blood post-transfusion. Red blood cell (RBC) transfusion is associated with systemic inflammation and immune suppression as adverse outcomes
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