The Study Of Reconstitution In Bone Marrow Immune Niche After Allogeneic Stem Cell Transplantation

Objective The immune function of patients with hematopoietic stem cell transplantation is an important factor which influence the prognosis of patients.The speed of immune reconstruction and the balance between different T lymphocyte subsets,especially of Th1、Th2、Th17、Treg,play an important role in the development of such as acute graft-versus-host disease,infection,and relapse,so immune reconstruction is the focus of attention.Previous researches on immune reconstruction pay more attention to the changes of immune cells and related cytokines in peripheral blood,and there are few studies on the immune changes in the bone marrow.As bone marrow is an important immune organ,the bone marrow immune niche is made up by a variety of immune cells and the cytokines they secreted,and the signal pathwaies they mediated.Different subsets are accelerate or inhibited by each other during proliferation and differentiation,to keep balance of the immune function of the body.So,our study is purpos to explore the immune reconstitution in the bone marrow after allogeneic stem cell transplantation,and its diffrences with the peripheral blood,and its relationship with patients’ s age、CVM infection、HLA zygosity and a GVHD。Methods In our study,we chose 52 patients diagnosed of hematologic malignancies who had undergone allo-HSCT in our hospital and 7 volunteers.The bone marrow and peripheral blood samples were got from all the objects of study.Flow cytometric immunophenotyping were used to evaluate T lymphocyte subsets,such as CD4⁺T cells,CD8⁺T cells,Th1 cells,Th2 cells,Th17 cells,Treg cells.What’s more,all the samles that from patients were got before transplantation and on day 15,30,60,90,180 after allo-HSCT.Results: 1.Changes in bone marrow after allo-HSCT: Within day 180 after transplantation,the percentage of CD4⁺T cells was significantly lower than the control group(（6.7±1.5）%^10.1±1.6%),the changes of CD4⁺T cells lead to the ratio of CD4⁺/CD8⁺ cells was inverted and significantly lower than the control group(（0.299±0.07）^{0.796 ±0.128}).The percentage of Th1 was significantly higher than the control group from the day pretransplantation to day 180 after transplantation(（4.4±0.4）%^9.0±0.6%),and it got the highest level（（9.0±0.6）%）on day 15 after transplantation,while there was no significant difference between the patients who undergone allo-HSCT and the control group on each node from the day pretransplantation to day 180 after transplantation of the percentage of Th2 cells.Relevantly,the ratio of Th1/Th2 cells up to the highest level on day 15 after transplantation and then keep falling trend,and was significantly higher than the control group on day pretransplantation to day 90 after transplantation（2.215±0.225³.928±0.299）,and it fall to normal on day 180 after transplantation successfully.As the percentage of Th17 cells were all significantly higher than the control group within 180 days after transplantation(（2.5±0.2）%^6.2±0.7%),and the percentage of Treg cells wasn’t change significantly at the same time,so the ratio of Th17/Treg cells increased rapidly after transplantation and was ignificantly higher than the control group within 90 days after transplantation（2.147±0.264².518±0.343）,and could fall to normal on day 180 after transplantation.2.Changes in peripheral blood after allo-HSCT: Within 180 days after transplantation,the proportion of CD4⁺T cells was significantly decreased after transplantation(（11.8±1.7）%^13.5±2.3%)and the proportion of CD8⁺T cells was significantly higher than that of control group after transplantation（13.1±2.5）%^42.7±3.1%),these changes lead to the ratio of CD4⁺/CD8⁺ cells was inverted and significantly lower than the control group（0.342±0.06⁰.441±0.083）.At the same time,the proportion of Th1 cells after transplantation wassignificantly higher than that of the control group(（3.5±0.3）%^7.2±0.5%),but the percentage of Th2 cells after transplantation had no significant changes,so the ratio of Th1/Th2 cells significantly increased（1.960±0.153³.816±0.267）compared with control group.On the other hand,the proportion of Th17 cellswas significantly higher after transplantation,and there was no significant difference in the proportion of Treg cells after transplantation,sothe ratio of Th17/Treg was significantly higher than that of control group but since the day 90 after transplantation,there was no significant difference in the ratio of Th17/Treg between experimental group and control group.3.The diffrences between bone marrow and peripheral blood immune reconstruction after allo-HSCT: The change trend of the proportion of CD4⁺T cells,CD8⁺ T cells,and the ratio of CD4⁺/CD8⁺ cells,Th1/Th2 cells,Th17/Treg cells between bone marrow and peripheral blood were similar on each date after transplantation.But the percentage of CD4⁺ T cells and CD8⁺ T cells in the bone marrow was significantly lower than in peripheral blood。And on day 15 after transplantation,the proportion of Th1 cells,Th2 cells,Th17 cells and Treg cells in bone marrow was significantly higher than it in peripheral blood.4.Patients undergon allo-HSCT were divided into two groups by median age（36 years old）: There were no significant differences of the proportion of CD4⁺ T cells、 CD8⁺ T cells,Th1 cells Th2 cells,Th17 cells and Treg cells,and the ratio Th1/Th2 cells in bone marrow between these two groups on each date after transplantation.On day 90 after transplantation,the proportion of Treg cells in bone marrow of ≥36 years group is significantly lower than that of <36 years group,and leading to the ratio of Th17/Treg cells in bone marrow of ≥36 years group were significantly higher than those of <36 years group.In both these two groups,the proportions of CD4⁺ T cells、 CD8⁺ T cells in bone marrow are significantly lower than those in peripheral blood.5.Patients undergon allo-HSCT were divided into two groups by HLA matches: There were no significant differences of the proportion of Treg cells,and the ratio Th1/Th2 cells in bone marrow between these two groups on each date after transplantation.Within the early days after transplantation,the proportion of CD4⁺T cells in bone marrow of HLA mismatched group was significantly lower than that of human leukocyte antigen（HLA）matched group,.the proportion of CD8⁺T cells in bone marrow of HLA mismatched group was significantly higher than that of HLA matched group,so The ratio of CD4⁺/CD8⁺ cells in bone marrow of HLA mismatched group（0.265±0.062）was significantly lower than that of HLA matched group（0.939±0.142）.On day 60 after transplantation,the proportion of Th1 cells in bone marrow of HLA mismatched group was significantly higher than that of HLA matched group,the proportion of Th17 cells in bone marrow of HLA mismatched group（5.5±0.5）% was significantly higher than that of HLA matched group（3.6±0.3）%.6.Patients undergon allo-HSCT were divided into two groups by the conditioning with or without anti-thymocyte globulin（ATG）:Within the early time after transplantation,the proportion of CD4⁺T cells in bone marrow of ATG-conditioned group was significantly lower than that of non-ATG-conditioned group,and the proportion of CD8⁺T cells in bone marrow of ATG-conditioned group was significantly higher than that of non-ATG-conditioned group,so the ratio of CD4⁺/CD8⁺ cells in bone marrow of ATG-conditioned group was significantly lower than that of non-ATG-conditioned group.On the other hand,the percentage of Th1 cells,Th2 cells,Th17 cells and Treg cells in bone marrow of ATG-conditioned group was significantly higher than that of non-ATG-conditioned group.7.Patients undergon allo-HSCT were divided into two groups by human cytomegalovirus（CMV）infected or not: No significant differences of the proportion of CD4⁺T cells,Th2 cells,Th17 cells and Treg cells,and the ratio of CD4⁺/CD8⁺ cells,Th1/Th2 cells and Th17/Treg cells in bone marrow were discovered between these two groups on each date after transplantation.But the percentage of CD8⁺T cells（63.1±8.4）% and Th1 cells（11.9±1.2）% in bone marrow of CMV+ group was significantly higher than that of CMV-group(（46.6±3.7）%,（8.2±0.70%）on day 30 after transplantation.8.All patients undergon allo-HSCT were divided into two groups by the occurrence of acute graft-versus-host disease or not: No significant differences were foun of the percentage of CD4⁺T cells,CD8⁺T cells and Th2 cells,and the ratio of Th1/Th2 cells in bone marrow between these two groups on each date after transplantation.On the early days after transplantation,the percentage of Th1 cells,Th17 cells and Treg cells,and the ratio of Th17/Treg cells in bone marrow of a GVHD group was significantly higher than that of nona GVHD group But the ratio of CD4⁺/CD8⁺ cells in bone marrow of a GVHD group was only significantly lower than that of non-a GVHD groupConclusions: 1.After allo-HSCT,the percentage of CD4⁺T cells and the ratio of CD4/CD8 cells in bone marrow were significantly reduced,and the percentage of CD8⁺T cells,Th1 cells,Th2 cells and Th17 cells in both bone marrow and peripheral blood were significantly increased,wich lead to the ratio of Th1/Th2 cells and Th17/Treg cells were significantly increased at the early time after transplantation.2.At the early time after allo-HCST,the percentage of Th1 cells,Th2 cells,Th17 cells and Treg cells in bone marrow significantly higher than that in peripheral blood,but the change rule is roughly similar with that in peripheral blood.Although both the ratio of Th17/Treg cells in bone marrow and peripheral blood could fall to normal within 180 days after transplantation,it’s later in bone marrow.3.After allo-HCST,the percentage of different T lymphocyte subgroups and ratio between them were effected by the patients’ ag,the match degree of HLA,ATG conditioned or not,CMV infection and a GVHD more or less.Our study provides important basic data for the further study the reconstruction of the bone marrow after hematopoietic stem cell transplantation immune microenvironment law and its relationship with early complications after transplantation...