Effects of JAK1/2 Inhibitor Ruxolitinib on the Survival and Immune Function of Neutrophils
JIANG Shan, SUN Lu, ZHANG Shiyue, WANG Tong, XU Yuanfu*
Ruxolitinib is an immunosuppressant, which has been used to treat myelofibrosis-related diseases. Recently, it has been clinically used to inhibit the occurrence of cytokine storm in acute GVHD (graft-versus-host disease) and COVID-19. This article aimed to investigate the effects of Ruxolitinib on immunological functions such as neutrophil survival and inflammatory factor secretion. In this study, vehicle control group, 0.1 μmol/L Ruxolitinibtreatment group, 1 μmol/L Ruxolitinib-treatment group and 5 μmol/L Ruxolitinib-treatment group were set up. Mouse bone marrow neutrophils isolated by Percoll method were analyzed for bone marrow neutrophil purity and apoptosis by flow cytometry after treatment with Ruxolitinib for 0 h, 24 h, 48 h, and 72 h. After treatment with Ruxolitinib for 1 h, neutrophils were detected by immunofluorescence for the ability to phagocytose zymosan bioparticles, TAXIScanFL chemokine for the migration ability of neutrophils towards fMLP, and coating plate method for the ability of neutrophils to kill E. coli. After treatment with Ruxolitinib for 24 h, the ability of neutrophils pretreated and activated by LPS (1 μg/mL) to produce cytokines was detected by ELISA and qPCR. The results showed that treatment with 0.1 μmol/L, 1 μmol/L, and 5 μmol/L Ruxolitinib for 24 h could accelerate the early apoptosis of neutrophils (P<0.05), and enhance the phagocytic index (P<0.05). There was no significant differences in chemotaxis ability and bactericidal ability compared with the control group. The expression levels of TNF-α and IL-6 were significantly increased in neutrophils treated with Ruxolitinib (0.1 μmol/L, 1 μmol/L) after 1 h stimulation with 1 μg/mL LPS (P<0.05). This article mainly demonstrated that Ruxolitinib could accelerate the early apoptosis of neutrophils, and promote the release of inflammatory factors such as TNF-α and IL-6 in neutrophils. This study revealed that the JAK1/2 inhibitor Ruxolitinib could not inhibit the production of major inflammatory factors in neutrophils, and provided theoretical and experimental basis for the prognostic analysis of patients treated with Ruxolitinib in the future, and also enriched the therapeutic targets and strategies for the clinical treatment of cytokine storm.
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