Jiaojiao Zhou1, Yajun Bai2, Yong Jiang3, Padamata Tarun4, Yuying Feng5, Rongshuang Huang5, Ping Fu5. 1. Division of Ultrasound, West China Hospital of Sichuan University, Chengdu, Sichuan, China. 2. Department of Nephrology, Nanchong Central Hospital, Nanchong, Sichuan, China. 3. Sichuan Center for Disease Control and Prevention, Chengdu, Sichuan, China. 4. West China Hospital of Sichuan University, Chengdu, Sichuan, China. 5. Kidney Research Institute, Department of Internal Medicine, Division of Nephrology, West China Hospital of Sichuan University, Chengdu, Sichuan, China.
Abstract
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.
Objective: The present study aimed to investigate whether recombinant humanerythropoietin (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.