The Alteration And Clinical Significance Of Th22/Th17/Th1 Cells In Patients With Chronic Myeloid Leukemia

Objective:Chronic myeloid leukemia (CML) is a malignant hematopoietic stem cell disease characterized by the presence of Philadelphia chromosome, t(9:22)(q34:q11), resulting in the BCR-ABL fusion gene which encodes for a constitutively activated tyrosine kinase. BCR-ABL gene is considered as the molecular basis of the pathogenesis of CML and as an effective indicator of diagnosis and prognosis. Although the disease can be readily controlled by the introduction of ABL tyrosine kinase inhibitors (TKI), approximately one-third of patients perform no response to this treatment, which seems to be associated with ABL mutation related drug resistance and the blast crisis.It is widely accepted that both genetic and immune factors play significant roles in the development of CML. Immune status in CML is very complex and ill-defined, and the roles of immune factors in CML have received increasing attention in recent years.However, little is known about the immunopathological events, especially the abnormal T-helper (Th) subsets, in the pathophysiology of CML.The number of T lymphocytes, CD4+ and CD8+ cells subtype, the ratio of both are within a certain range, maintain a state of dynamic balance, which is a central part of the immune stability. Abnormal changes of T lymphocyte number of CD4+and CD8+ cell subtype, up-regulation or down-regulation and both of two classes of subgroup ratio (CD4+/CD8+) change suggests that the body immune function disorder is closely related to the occurrence of certain clinical disease.Th cells play critical roles in the development and progression of inflammatory and autoimmune diseases as well as tumors. For many years, the heterogeneity of the CD4+T helper (Th) cells has been limited to Thl and Th2 cells due to the secretion of different cytokine patterns and immunity functions.Th1 cells mediate cellular immunity, activate CD8+ CTL cells and promote reproduction through secreting IL-2 and IFN-γ, then play an anti-bacterium, anti-virus and anti-tumor role. Also, IL-2 and IFN-γ can increase the anti-tumor effect of NK cell. They are thought to play a pathogenic role in organ-specific autoimmune and other autoimmune diseases.Th17 cells and Th22 cells are two newly described Th subsets, which have important roles in peripheral immune responses. Th17 is a unique CD4+ Th subset characterized by production of interleukin-17 (IL-17). Th17 cells may have evolved for host protection against microbes that Thl or Th2 immunity are not well suited for, such as extracellular bacteria and some fungi. Th17 cells, characterized by the expression of chemokine receptor CCR6 and its ligand CCL20, produceIL-17A, IL-17F, IL-21, and IL-22[9] They are important for extracellular pathogens including bacteria and fungi, and participate in inflammation and autoimmune diseases[10-11]. Recent data in humans and mice suggest that Th17 cells play an important role in the pathogenesis of a diverse group of immune-mediated diseases as well as tumor[12]. IL-17 is a highly inflammatory cytokine with robust effects on stromal cells in many tissues[13]. However, the role of Thl7 in cancer is still being intensively discussed, with conflicting reports related to the pro-versus anti-tumoral effects of these cells.Th22 subset is a recently identified CD4+ human T helper subset, first mentioned by Trifari et al. which is characterized by abundant secretion of IL-22 and TNF-α,but not IL-17 or IFN-y. Th22 cells express the chemokine receptor CCR6 and the skin-homing receptors CCR4 and CCR10, and have a high expression of the key transcription factor of aryl hydrocarbon receptor (AHR)[15-16]. the differentiation toward the Th22 phenotype from naive T cells is in the presence of IL-6 and TNF-α[17]. Th22 cells represent an independent and terminal differentiation T cell subtype[18-19].IL-22 belongs to the IL-10 family of cytokines and is primarily secreted by activated Th22 cells. Today, we know that in humans, adaptive immune cells of the immune system, Th22 is the major T cell subsets producing IL-22 [16-17,20]. IL-22 played either protective or pathogenic role in discrepant induction by naive and memory/effector cells [21-22]. However, considering IL-22 involved in the regulation of cell growth, proliferation, and cell cycle control[23-25], it is conceivable that IL-22 might play a role during tumor genesis.The discovery of Th17 and Th22 cells has opened up a new avenue for research into the etiology and treatment of a broad spectrum of diseases. Recent studies have implicated the roles of Th17 and Th22 cells and their effector cytokines in the pathogenesis of several autoimmune diseases and solid tumors in humans, such as hepatic cellular cancer[26],gastric cancer[27] and lung cancer [28]. However, little is known regarding the roles of Th17 and Th22 cells in hematological malignancy. To date, no previous study has reported data about the immunobiology of Th subsets especially Th22, Thl7 and Thl cells in CML.In this study, we measured the frequencies of Th cells (Th22, Thl 7 and Thl), the expression of transcription factors (AHR and RORC) and their related cytokines (IL-22, IL-17 and IFN-y) in PB and BM of CML patients. We also evaluated their correlations with clinical pathological characteristics.To further investigate the role of Th22 cells in t hematological tumor immune mechanisms.Methods:1. A total of 63 CML patients were enrolled in this study. Thirty-one of them were newly diagnosed (ND) CML patients and thirty-two were chronic-phase CP CML patients, Twenty-two age-matched healthy PB individuals, Eleven hematologically normal age-matched BM transplant donors. All cases of CML-CP were treated with imatinib mesylate.2. The proportions of Th22, Th17 and Thl Cells in PB or BM of CML were evaluated by Flow Cytometric Analysis.3. AHR and RORC mRNA expressions were examined by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR).4. The levels of cytokines IL-22, IL-17 and IFN-γ in PB and BM of each group were msasured by enzyme-linked immunoadsorbent assay (ELISA).Results:1. The frequencies of PB or BM Th22, Thl7and Thl cells were significantly decreased in ND CML patients than chronic phase (CP)-CML patients and healthy controls (P<0.05)2. AHR and RORC expression levels of the key transcription factor directing Th22 and Th17 lineage commitment: The expression levels of AHR were significantly decreased in PB or BM of ND CML patients compared with CP-CML patients and healthy controls(P< 0.05) The mRNA levels of AHR were increased in PB of CP patients compared with healthy controls (P<0.05), while no statistical significance of BM AHR mRNA levels were shown between CP-CML patients and controls. The mRNA levels of RORC were significantly decreased in PB or BM of ND CML patients compared with healthy controls (P<0.05). It was lower in ND than CP-CML patients (P< 0.05). No significant difference of RORC mRNA level was observed between CP-CML patients and healthy controls.3. The levels of PB IL-22 in CP-CML patients were slightly higher than controls (P< 0.05). The levels of PB IL-17 in ND patients and CP-CML patients were slightly higher than controls (P<0.05) As for the levels of BM IL-22 and IL-17, there were no significant difference between each group. A statistical increase of BM IFN-y level was found in ND patients compared with in controls. There was no significant difference regarding PB plasma IFN-y between each group.4. In CML patients, a significantly positive correlation were found between Th22 cells and Th17 cells both in PB and BM. Thl cells showed no significant correlations with Th22 or Th17 cells in CML patients.5. There were significantly negative correlations between Th22, Thl7 or Thl cells and peripheral white blood cell counts. There were also a statistical negative correlations between Th22, Th17 or Thl cells and BCR-ABL%(IS).Conclusion:The frequencies of Th22, Th17 and Th1 cells, along with the expression of specific transcription factors AHR and RORC were significantly decreased in ND patients compared with healthy controls, while all these abnormal recovered in chronic phase patients. In addition, there existed a significantly positive relationship between Th22 and Th17 cells in PB or BM. A significantly negative relationship was found between Th cells (Th22, Th17 or Th1) and BCR-ABL%(IS) or the number of peripheral blood white blood cells. All these results demonstrated that Th22, Th17 and Th1 cells might be important therapeutic targets in CML and could facilitate a better outcome for tumor immunotherapy.