| Background:Sepsis is a syndrome in which the host develops a dysregulated immune response to pathogenic infection,resulting in life-threatening systemic multi-organ dysfunction.Sepsis-associated thrombocytopenia is considered to be one of the risk factors for poor prognosis in patients with sepsis.Correction of sepsis-associated thrombocytopenia can effectively improve the prognosis of patients with sepsis.However,current treatments for sepsis-associated thrombocytopenia are limited and associated with side effects.Therefore,there is a need to further search for effective alternative therapeutic agents.Previous studies have shown that interleukin-21(IL-21)promotes thrombopoiesis by stimulating megakaryocyte development and that IL-21 levels were elevated in neonates with sepsis.Furthermore,the mechanism of IL-21 action on megakaryocytes in sepsis is unclear.Therefore,the aims of this study are to:1)investigate whether IL-21 levels are elevated in sepsis patients and whether it is related to platelet indices and inflammatory cytokines;2)investigate whether IL-21 treatment can promote megakaryocyte production and improve thrombocytopenia in sepsis mice;and 3)investigate how IL-21 regulates megakaryopoiesis and its potential mechanisms.Methods:1.A retrospective study was used to compare whether there were differences in clinical and laboratory indices between septic patients and non-infected patients,and Pearson’s and Spearman’s correlation analyses were performed to evaluate whether there was a correlation between IL-21 and platelet indices and inflammatory factors.2.Sepsis mice were used to study the role of IL-21 on thrombopoiesis and death protection in vivo.3.Human megakaryoblastic cell lines(Dami and Meg-01)were treated with recombinant IL-21 to determine the influences of IL-21 on cell proliferation and apoptosis by CCK-8,EdU,Tunel,Annexin V/PI apoptosis assays.The expression of IL-21 receptor(IL-21R)and apoptosis-associated proteins were measured by PCR,Western Blot,and immunofluorescent staining.The production of CD41a+/CD42b+ megakaryocytes were detected by flow cytometry.The expression and phosphorylation of PI3K/AKT pathway-related proteins were analyzed in the presence or absence of PI3K/AKT inhibitors to study the mechanism of IL-21 action in magakaryopoiesis by Western Blot.Results:1.The IL-21 levels and mean platelet volume(MPV)were significantly higher in septic patients compared with non-infected patients,and platelet count was lower compared with controls;IL-21 was negatively correlated with platelet count,but positively correlated with TPO and IL-6(P<0.05).2.Bone marrow megakaryocyte proportion and circulating platelet count were higher in IL-21-treated sepsis mice compared with non-IL-21-treated sepsis mice(P<0.05),and prolonged median survival time was observed in IL-21-treated sepsis mice.3.IL-21 treatment increased megakaryocytes viability and proliferation,reduced cells apoptosis,and promoted CD41a+/CD42b+megakaryocytes production.Treatment of Dami and Meg-01 cells with IL-21 decreased the expression levels of p21 and Bax,but enhanced the expression of Bcl-2,PI3K,and the phosphorylation of AKT.Knockdown of IL-21 R expression and treatment of Dami and Meg-01 cells with PI3K inhibitor partially abolished the effect of IL-21 on megakaryocytes proliferation,anti-apoptotic protection and CD41a+/CD42b+ megakaryocytes production.Conclusion:IL-21 levels are elevated in patients with sepsis;IL-21 can improve thrombocytopenia in sepsis mice by stimulating bone marrow megakaryopoiesis;IL-21 regulates megakaryocyte proliferation,differentiation and apoptosis by activating the PI3K/AKT signaling pathway. |
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