Objective: The present study aimed to investigate whether recombinant human erythropoietin (rHuEPO) plays an immunomodulatory function by regulating the TLR4/NF-κB signaling pathway.Materials and methods: C57BL/6 mice were intraperitoneally injected with rHuEPO and, half an hour later, with 50% glycerol at the dose of 7.5 ml/kg to induce crush syndrome (CS)-acute kidney injury (AKI). The levels of TNF-α, IL-1β, IL-6, serum creatinine (Scr), and creatine kinase (CK) were measured. The kidney tissues were analyzed by HE staining, and macrophage infiltration was detected by immunohistochemistry. Double immunofluorescence staining, RT-qPCR, and western blotting were conducted to analyze TLR4/NF-κB p65 expression. Ferrous myoglobin was co-cultured with RAW264.7 cells to mimic crush injury and the production of proinflammatory cytokines. The expression levels of TLR4 and NF-κB p65 were measured.Results: In vivo study results revealed that rHuEPO ameliorated renal function, tissue damage, production of proinflammatory cytokines, and macrophage infiltration in the kidneys. The protein and mRNA expression levels of genes involved in the TLR4/NF-κB signaling pathway in CS-induced AKI mice were upregulated (p < .05). Meanwhile, the expression levels of TLR4, NF-κB p65, and proinflammatory cytokines in RAW264.7 cells were downregulated in CS-AKI mice injected with rHuEPO (p < .05).Conclusions: Our results demonstrated the immunomodulatory capacity of rHuEPO and confirmed that rHuEPO exerts protective effects against CS-induced AKI by regulating the TLR4/NF-κB signaling pathway in macrophages. Therefore, our findings highlight the therapeutic potential of rHuEPO in improving the prognosis of CS-AKI patients.

中文翻译：

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重组人促红细胞生成素通过抑制巨噬细胞中的TLR4 /NF-κB信号通路在挤压综合征引起的急性肾损伤中的免疫调节作用。

目的：本研究旨在通过调节TLR4 /NF-κB信号通路来研究重组人促红细胞生成素（rHuEPO）是否具有免疫调节功能。材料与方法：C57BL / 6小鼠腹腔注射rHuEPO，半小时后，用7.5 ml / kg的剂量的50％甘油诱导急性挤压综合征（CS）-急性肾损伤（AKI）。测量了TNF-α，IL-1β，IL-6，血清肌酐（Scr）和肌酸激酶（CK）的水平。通过HE染色分析肾组织，并通过免疫组织化学检测巨噬细胞浸润。进行了双重免疫荧光染色，RT-qPCR和western印迹分析TLR4 /NF-κBp65表达。亚铁肌红蛋白与RAW264.7细胞共培养以模拟挤压损伤和促炎细胞因子的产生。结果：体内研究表明，rHuEPO改善了肾功能，组织损伤，促炎细胞因子的产生和巨噬细胞在肾脏的浸润。CS诱导的AKI小鼠中与TLR4 /NF-κB信号通路有关的基因的蛋白质和mRNA表达水平上调（p <.05）。同时，在注射rHuEPO的CS-AKI小鼠中RAW264.7细胞中TLR4，NF-κBp65和促炎细胞因子的表达被下调（p <.05）。结论：我们的结果证明了rHuEPO的免疫调节能力并得到证实rHuEPO通过调节巨噬细胞中的TLR4 /NF-κB信号通路对CS诱导的AKI发挥保护作用。因此，