Immune Reconstitution After Allogeneic Hematopoietic Stem Cell Transplantation And The Correlation Between CD4⁺CD25^highFOXP3⁺and GVHD

Background and purpose Background The incidence rate of hematological diseases shows a rising trend year by year and with more young age currently. The pathogenesis involves the pathological changes of hematopoietic stem cells and the microenvironment which the immune dysfunction plays the key role. In recent years, with the development of immunology,molecular biology and cytogenetics, the introduction of new treatment methods and new drugs, the mode of diagnosis and treatment of hematological diseases occurred fundamental changes, with these advances have significantly improved the quality of life, and prolong the survival time of the patients. Hematopoietic stem cell transplantation has become the effective mean for the benign and malignant hematological diseases. However, graft-versus-host disease（GVHD） is the important cause of death in patients after transplantation. Related studies have found that GVHD is often associated with delayed reconstruction of the immune function of patients with graft transplantation are closely.Purpose 1.To study the immune function of patients with hematologic diseases after allogeneic hematopoietic stem cell transplantation; 2.To analyse the correlation between CD4+CD25high FOXP3+regulatory T cells and graft versus host disease（GVHD） primarily.Research methods1. 20 cases received allogeneic hematopoietic stem cell transplantation in the department of Hematology of our hospital from April 2014 to April 2015, including 10 males and 10 females,average age of 21.7 years（3⁴6 years old）. Collecting the peripheral blood from 20 cases a month before transplantation and after transplantation in different periods（on +1, +3, +6, +12 month after transplantation） respectively.Monitoring the level of T lymphocyte subsets（CD3+, CD4+, CD8+, CD4+/CD8+, CD56+） a month before transplantation,and the level of T lymphocyte subsets(CD3+, CD4+, CD8+,CD4+/CD8+, CD56+, CD4⁺CD25^highFOXP3⁺) dynamicly by flow cytometry（FCM）. To study the early immune reconstruction of patients with allogeneic hematopoietic stem cell transplantation. T lymphocyte subsets of 20 healthy physical examinees were monitored during the same period as the healthy control group.Comparing the levels of T lymphocyte subsets in peripheral blood transplantation between the 20 cases before transplantation and the healthy control group, to reflect the immune function of patients with with hematological diseases. Dynamic observation the changes of T lymphocyte subsets in patients after transplantation, to evaluate the immune function recovery;2. Monitoring the level of CD4⁺CD25^highFOXP3⁺from 20 cases with flow cytometry after transplantation in the peripheral blood, and FOXP3 m RNA expression in peripheral blood mononuclear cells with QRT-PCR to analysis the correlation with GVHD.Results1.Compared with the healthy control group, before transplantation 20 cases occurred abnormal immune function which 15 cases of malignant hematologic diseases with significantly lower CD3+, CD4+, CD4+/CD8+and CD56+which was（54.12±5.41）%,（27.25±9.13）%,（0.91±0.74）%,（9.37±3.41）% respectively（P<0.01）, slightly higher CD8+which was（31.77±7.16）%（P<0.05）; 5 cases of aplastic anemia（AA） with slightly higher CD3+which was（66.79±7.35）%（P<0.05）, significantly higher CD8+which was（35.69±6.78）%, significantly decreased CD4+, CD4+/CD8+and CD56+which was（33.73±7.26）%,（1.23±0.56）%,（7.46±2.83）%（P<0.05） respectively;2.Follow up to December 2015, the levels of CD3+, CD4+, CD8+, CD4+/CD8+, CD56+and CD4⁺CD25^highFOXP3⁺in patients with different degrees of recovery after transplantation, which to the pretransplant level on +12 month. One month after the transplantation,the CD56+cells began to recover 3 months after transplantation, and return to the normal the firstly; CD8+cells recovered earlier than CD4+cells, CD4+cells decreased for more than 1 year; the CD4+/CD8+ratio was inverted for more than 1 year as well;3.Follow up to December 2015, 9 patients occured III degree or above acute GVHD（a GVHD）, which 2 cases died, and the remaining 18 cases survived; The lower level of CD4⁺CD25^highFOXP3⁺and FOXP3 m RNA expression in patients with a GVHD than the patients with no a GVHD and the healthy control group（P<0.05） at the same period which was（0.32±0.45）%,（1.92±0.70）%,（278.65±241.36）×106,（1123.45±1072.31）×106on +1 month respectively;（0.73±0.51）%,（2.35±0.72）%,（352.77±421.45）×106,（2052.09±2169.13）×106on +3 month respectively;（1.01±0.54）%,（2.63±0.77）%,（1085.67±954.32）×106,（2814.48±3076.82）×106on +6 month respectively;and（1.43±0.46）%,（3.23±057）%,（1721.47±1204.22）×106,（3179.25±4563.08）×106on +12month respectively. The level of CD4⁺CD25^highFOXP3⁺and a GVHD has a good linear regression relationship（r=0.882, r2=0.778）; The level of FOXP3 m RNA expression in peripheral blood mononuclear cells when a GVHD controlled（on +6 month） was significantly higher than a GVHD occurred（on +1, +3 month）（P=0.025）.Conclusions1.There is the abnormal immune function in patients with benign and malignant hematologic disease;2.The levels of T lymphocyte subsets with different degrees of recovery after transplantation, but the complete reconstruction of the immune function is a slow process;3.The level of CD4⁺CD25^highFOXP3⁺and the GVHD has the good correlation, which can be used to predict the occurrence of GVHD in clinical and is a guiding role of diagnosis and treatment for GVHD;4.It is important in clinical to monitor the T lymphocyte subsets dynamicly especially for CD4⁺CD25^highFOXP3⁺after transplantation and could forecast a GVHD before symptoms appeared, so as to reduce mortality after transplantation.