The Relationship Between The Changes Of Th17 Cells In The Peripheral Blood And Disease Stage And Risk Stratification In Patients With Acute Myeloid Leukemia

Objective:To investigate the relationship between the changes of Th17 cells and related lymphocyte subsets in the peripheral blood and disease stage and risk stratification in patients with acute myeloid leukemia(AML),to provide experimental basis for prognosis and clinical immunotherapy treatment.Methods:Sixty-five patients with acute myeloid leukemia were enrolled in the study between August 2015 and March 2017 in the Affiliated Hospital of Nanjing University medical school.AML patients were divided into 3 groups including newly diagnosed,incomplete remission,and complete remission according to disease stage,and divided into low risk,medium risk and high risk according to the risk stratification respectively.Twenty-four healthy controls were recruited.Flow cytometry was used to detect CD4+T lymphocyte subsets including Th17,Treg,Thl and Th2 and the proportion of lymphocyte subsets in the peripheral blood of AML patients and healthy control group;Luminex was applied to detect the plasma concentration of Th17 related cytokines(IL-17A,IL-6,and IL-113);Real-time-PCR was used to detect the mRNA expression of RORyt which was major transcription factor in Th17 differentiation and FOXP3 that was a specific marker of Tregs in peripheral blood mononuclear cells.The changes of the indexes were analyzed to evaluate the cellular immune status of the peripheral blood and bone marrow microenvironment in AML patients.Results:1)In newly diagnosed patients with AML,we can see that Th17 and Tregs of PBMCs were increased significantly(P<0.0001),however there was no statistical difference in Thl cells,and Th2 cells were decreased significantly(P=0.0092).When patients received standardized chemotherapy but not yet reached complete remission,Thl7 cells were significantly increased than the control group,however no significant difference with newly diagnosed group,but no significant difference between Treg and Thl cells compared with newly diagnosed and the control groups,and Th2 cells were significantly higher than that of newly diagnosed.When patients reached complete remission,the percentage of Thl 7 cells were decreased significantly than newly diagnosed and incomplete remission group(P=0.0231,P=0.0044),the level of Tregs returned to a normal level,and Thl cell levels were significantly higher than the other three groups,in addition Th2 cells were increased significantly compared with newly diagnosed group no matter whether achieved CR(P=0.032);2)AML patients were grouped according to the risk stratification,the levels of Thl 7%in medium and high risk group were significantly higher than that of controls(P=0.0071,P=0.0197),and patients with low risk had increased trend,but there was no statistical difference;high levels of Tregs were mainly distributed in low and medium risk group,while high risk group had no significant difference compared with the other three groups.There was no significant difference in Thl cells no matter the risk stratification was;the levels of Th2 reduction is mainly distributed in the medium and high risk group,the changes similar to Thl 7 cells.3)The plasma concentration of Th17 related cytokines including IL-17A and IL-6 in AML patients with newly diagnosed group were significantly higher than those in healthy control group,and mainly in the medium and high risk group,the level would significantly reduced when reached complete remission.However the AML patients with incomplete remission had no statistical difference compared with the other three groups.The plasma concentration of IL-1β Th17 related cytokine in different stages of disease had no significantly different than that in healthy control group,but the AML patients of high-risk were significantly higher than those of the others.4)The mRNA expression of RORyt the main transcription factor in the differentiation of Th17 cell and FOXP3 which was Treg’s specific marker in the newly diagnosed patients with AML were significantly higher than those in the control group(P<0.0001,P=0.0087),and the levels were lower after chemotherapy.The RORyt mRNA expression in AML patients were significantly higher than the control group according to risk stratification(P=0.0154,0.0011,0.0243);among the three groups,suggesting that the levels of mediun and high risk groups were increased significantly than the low-risk group(P=0.0029 0.0426).The FOXP3 mRNA level of the mediun and high risk groups were higher than that in control group(P=0.0482,0.0243),and the medium risk group was significantly increased than the low risk group,however the high risk group had a increased trend,but no significant difference compared with low risk group(P=0.0646).5)The balance between Th17/Treg and Thl/Th2 in newly diagnosed AML patients were disorders,Th17/Treg imbalance was more significant in medium and high-risk groups combined with risk stratification,and after received standard chemotherapy can correct the patients’ Th17/Treg and Th1/Th2 imbalance.6)NK cells with anti-tumor effect were significantly lower in AML patients of newly diagnosed and incomplete remission group,and the level was significantly higher than that berore however not reached the normal lever after the complete remission.Conclusion:1)Th lymphocyte subsets and related cytokines and mRNA are abnormally expressed in peripheral blood of patients with acute myeloid leukemia,and T cell immune disturbances may be involved in the pathogenesis of AML.2)High level of Th17 cells in ND AML patients may in the medium and high-risk group,and the level of Th17 cells can be evaluated in patients whether reached complete remission receiving chemotherapy;quantitative detection of Th17 cells in vivo may contribute to the stratified treatment and prognosis.In addition,Th17 cells may served as a potential therapeutic target in the medium and high-risk AML patients,and provided a guideline for novel cellular immunotherapy.3)Treg cells in peripheral blood of AML patients were significantly increased,and correlated with tumor prognosis and risk stratification.4)Th17/Treg and Th1/Th2 balance were changes in AML patients,and Th17/Treg was more significant in the medium and high risk group combined with the risk stratification;standard chemotherapy can make the patients’ Th17/Treg and Th1/Th2 to achieve new balances.