Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE, 68198, USA.
Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
Inflamm Res. 2024 Nov;73(11):1859-1873. doi: 10.1007/s00011-024-01936-y. Epub 2024 Sep 5.
RBC transfusions (RBCT) are life-saving treatment for premature and critically ill infants. However, the procedure has been associated with the development of systemic inflammatory response syndrome (SIRS) and potentially multiple organ dysfunction syndrome (MODS) in neonates. The present study aimed to investigate the mechanisms of RBCT-related SIRS in severely anemic murine neonates.
C57BL/6 (WT), TLR4 and myeloid-specific triggered myeloid receptor-1 (trem1) mouse pups were studied in 4 groups (n = 6 each): (1) naïve controls, (2) transfused control, (3) anemic (hematocrit 20-24%) and (4) anemic with RBC transfused using our established murine model of phlebotomy-induced anemia (PIA) and RBC transfusion. Plasma was measured for quantifying inflammatory cytokines (IFN-γ, IL-1β, TNF-α, IL-6, MIP-1α, MIP-1β, MIP2 and LIX) using a Luminex assay. In vitro studies included (i) sensitization by exposing the cells to a low level of lipopolysaccharide (LPS; 500 ng/ml) and (ii) trem1-siRNA transfection with/without plasma supernatant from stored RBC to assess the acute inflammatory response through trem1 by qRT-PCR and immunoblotting.
Anemic murine pups developed cytokine storm within 2 h of receiving stored RBCs, which increased until 6 h post-transfusion, as compared to non-anemic mice receiving stored RBCTs ("transfusion controls"), in a TLR4-independent fashion. Nonetheless, severely anemic pups had elevated circulating endotoxin levels, thereby sensitizing circulating monocytes to presynthesize proinflammatory cytokines (IFN-γ, IL-1β, TNF-α, IL-6, MIP-1α, MIP-1β, MIP2, LIX) and express trem1. Silencing trem1 expression in Raw264.7 cells mitigated both endotoxin-associated presynthesis of proinflammatory cytokines and the RBCT-induced release of inflammatory cytokines. Indeed, myeloid-specific trem1 murine pups had significantly reduced evidence of SIRS following RBCTs.
Severe anemia-associated low-grade inflammation sensitizes monocytes to enhance the synthesis of proinflammatory cytokines and trem1. In this setting, RBCTs further activate these monocytes, thereby inducing SIRS. Inhibiting trem1 in myeloid cells, including monocytes, alleviates the inflammatory response associated with the combined effects of anemia and RBCTs in murine neonates.
红细胞输注（RBCT）是治疗早产儿和危重症婴儿的救命治疗方法。然而，该操作与新生儿全身炎症反应综合征（SIRS）和潜在的多器官功能障碍综合征（MODS）的发生有关。本研究旨在探讨严重贫血的新生鼠 RBCT 相关 SIRS 的机制。
在 4 组（每组 6 只）中研究 C57BL/6（WT）、TLR4 和髓样特异性触发髓样受体 1（trem1）幼鼠：（1）未处理对照，（2）输注对照，（3）贫血（血细胞比容 20-24%）和（4）使用我们建立的放血诱导贫血（PIA）和 RBC 输血的鼠模型输血的贫血。使用 Luminex 测定法测量血浆以定量炎性细胞因子（IFN-γ、IL-1β、TNF-α、IL-6、MIP-1α、MIP-1β、MIP2 和 LIX）。体外研究包括：（i）通过暴露细胞于低水平的脂多糖（LPS；500ng/ml）来敏化细胞，以及（ii）用储存的 RBC 血浆上清液转染 trem1-siRNA，通过 qRT-PCR 和免疫印迹评估通过 trem1 的急性炎症反应。
与接受储存 RBC 的非贫血小鼠（“输血对照”）相比，接受储存 RBC 的贫血幼鼠在 2 小时内发生细胞因子风暴，在输血后 6 小时内增加，这是一种 TLR4 不依赖的方式。尽管如此，严重贫血的幼鼠循环内毒素水平升高，从而使循环单核细胞预先合成促炎细胞因子（IFN-γ、IL-1β、TNF-α、IL-6、MIP-1α、MIP-1β、MIP2、LIX）并表达 trem1。在 Raw264.7 细胞中沉默 trem1 表达可减轻内毒素相关的促炎细胞因子的预先合成以及 RBCT 诱导的炎性细胞因子的释放。事实上，髓样特异性 trem1 幼鼠在接受 RBCT 后，SIRS 的证据明显减少。
严重贫血相关的低度炎症使单核细胞易于增强促炎细胞因子和 trem1 的合成。在这种情况下，RBCT 进一步激活这些单核细胞，从而诱导 SIRS。在骨髓细胞（包括单核细胞）中抑制 trem1 可减轻与贫血和 RBCT 联合作用相关的炎症反应。
