Acute graft-versus-host disease (aGVHD) is a serious complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, the mechanisms of aGVHD are not well understood. We aim to investigate the roles of the three angiogenic factors: angiopoietin-1 (Ang-1), Ang-2 and vascular endothelial growth factor (VEGF) in the development of aGVHD. Twenty-one patients who underwent allo-HSCT were included in our study. The dynamic changes of Ang-1, Ang-2 and VEGF were monitored in patients before and after allo-HSCT. In vitro, endothelial cells (ECs) were treated with TNF-β in the presence or absence of Ang-1, and then the Ang-2 level in the cell culture medium and the tubule formation by ECs were evaluated. After allo-HSCT, Ang-1, Ang-2 and VEGF all exhibited significant variation, suggesting these factors might be involved in the endothelial damage in transplantation. Patients with aGVHD had lower Ang-1 level at day 7 but higher Ang-2 level at day 21 than those without aGVHD, implying that Ang-1 may play a protective role in early phase yet Ang-2 is a promotion factor to aGVHD. In vitro, TNF-β promoted the release of Ang-2 by ECs and impaired tubule formation of ECs, which were both weakened by Ang-1, suggesting that Ang-1 may play a protective role in aGVHD by influencing the secretion of Ang-2, consistent with our in vivo tests. It is concluded that monitoring changes of these factors following allo-HSCT might help to identify patients at a high risk for aGVHD.
Acute Disease ; Adolescent ; Adult ; Angiogenesis Inducing Agents ; immunology ; metabolism ; pharmacology ; Angiopoietin-1 ; genetics ; immunology ; pharmacology ; Angiopoietin-2 ; genetics ; immunology ; pharmacology ; Antineoplastic Agents ; therapeutic use ; Female ; Gene Expression Regulation, Neoplastic ; Graft vs Host Disease ; genetics ; immunology ; pathology ; Hematopoietic Stem Cell Transplantation ; Human Umbilical Vein Endothelial Cells ; cytology ; drug effects ; immunology ; Humans ; Leukemia, Myeloid ; genetics ; immunology ; pathology ; therapy ; Lymphoma, Non-Hodgkin ; genetics ; immunology ; pathology ; therapy ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; immunology ; pathology ; therapy ; Retrospective Studies ; Signal Transduction ; Transplantation, Homologous ; Tumor Necrosis Factor-alpha ; pharmacology ; Vascular Endothelial Growth Factor A ; genetics ; immunology

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an effective method for the treating of malignant diseases of hematopoietic system or non-malignant proliferative diseases, but the occurrence of graft-versus-host disease (GVHD) limits the success rate of hematopoietic stem cell transplantation. Moreover, chronic graft-versus-host disease (cGVHD) is the main factor affecting the long-term survival rate and life quality of recipient after hematopoietic stem cell transplantation. In this article, the latest research progress of the pathogenesis of cGVHD and related problems are reviewed from the thymus, cytokines, T lymphocyte subsets, B lymphocytes and its secreted antibody.
Chronic Disease ; Graft vs Host Disease ; immunology ; pathology ; Hematopoietic Stem Cell Transplantation ; adverse effects ; Humans ; Transplantation, Homologous

Animals ; Graft vs Host Disease ; immunology ; metabolism ; Intestines ; immunology ; pathology ; Lymph Nodes ; cytology ; Lymphocyte Activation ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; T-Lymphocytes ; immunology ; metabolism

OBJECTIVETo investigate the association of the ratio of regulatory and effector T cells with recurrence and chronic graft-versus-host disease (cGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT).

METHODSThirty patients with hematological malignancies who underwent allo-HSCT were classified as recurrence with cGVHD (n=4), non-recurrence with cGVHD (n=14), recurrence without cGVHD (n=5) and non-recurrence without cGVHD (n=7). The different percentage of CD4⁺CD25⁻CD69⁺ regulatory T cells in bone marrow and CD4⁺CD25⁺FoxP3⁺ regulatory T cells, Th1 cells and Th17 cells in peripheral blood were analyzed by flow cytometry.

RESULTSThere were no significant differences in all these T-cell subsets among different groups (P>0.05). While the ratio of CD4⁺CD25⁻CD69⁺ regulatory T cells and Th1 cells (0.211±0.177) in 9 recurrence patients was significant higher than that (0.133±0.160) in 21 non-recurrence patients (P=0.033). The ratio were also significance between recurrence without cGVHD and non-recurrence without cGVHD patients (0.167±0.073 vs 0.073±0.057, P=0.048), and between recurrence with cGVHD and non-recurrence without cGVHD patients (0.218±0.113 vs 0.073±0.057, P=0.024). Furthermore, the ratio of CD4⁺CD25⁺FoxP3⁺ regulatory T cells and Th17 cells was significant lower (1.975±2.045) in 18 cGVHD patients than that of 12 without cGVHD patients (3.198±1.132, P=0.010), and the ratio was also significant lower in non-recurrence patients with cGVHD (1.695±1.178) than that of without cGVHD (3.446±1.376, P=0.028).

CONCLUSIONOur results show that the ratio of CD4⁺CD25⁻CD69⁺ regulatory T cells and Th1 cells raise in recurrence patients, and the ratio of CD4⁺CD25⁺FoxP3⁺ regulatory T cells and Th17 decrease in cGVHD patients, which suggest that the ratio of regulatory and effector T cells had association with recurrence and cGVHD in patients with allo-HSCT.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Female ; Graft vs Host Disease ; immunology ; pathology ; Hematologic Neoplasms ; immunology ; therapy ; Hematopoietic Stem Cell Transplantation ; Humans ; Male ; Middle Aged ; Recurrence ; T-Lymphocytes, Regulatory ; cytology ; immunology ; Transplantation, Homologous ; Young Adult

Graft-versus-host disease (GVHD) is a prevalent and potential complication of hematopoietic stem cell transplantation. An animal model, xenogeneic GVHD (X-GVHD), that mimics accurately the clinical presentation of GVHD would provide a tool for investigating the mechanism involved in disease pathogenesis. Murine models indicated that inhibiting IL-21 signaling was a good therapy to reduce GVHD by impairing T cell functions. We sought to investigate the effect of exogenous human IL-21 on the process of X-GVHD. In this study, human IL-21 was expressed by hydrodynamic gene delivery in BALB/c-Rag2⁻/⁻ IL-2RΓc⁻/⁻ (BRG) immunodeficient mice which were intravenously transplanted human peripheral blood mononuclear cells (hPBMCs). We found that human IL-21 exacerbated X-GVHD and resulted in rapid fatality. As early as 6 days after hPBMCs transplanted to BRG mice, a marked expansion of human CD19⁺ B cells, but not T cells, was observed in spleen of IL-21-treated mice. Compared with control group, IL-21 induced robust immunoglobulin secretion, which was accompanied by increased accumulation of CD19⁺ CD38(high) plasma cells in spleen. In addition, we demonstrated that B-cell depletion was able to ameliorate X-GVHD. These results are the first to find in vivo expansion and differentiation of human B cells in response to IL-21, and reveal a correlation between the expansion of B cells and the exacerbation of xenogeneic GVHD. Our findings show evidence of the involvement of B cells in X-GVHD and may have implications in the treatment of the disease.
Animals ; B-Lymphocytes ; immunology ; metabolism ; pathology ; Cell Differentiation ; Cell Proliferation ; DNA-Binding Proteins ; deficiency ; Female ; Graft vs Host Disease ; blood ; genetics ; immunology ; metabolism ; Heterografts ; immunology ; Humans ; Immunoglobulin G ; metabolism ; Immunoglobulin M ; metabolism ; Interleukins ; genetics ; immunology ; Male ; Mice ; Mice, Inbred BALB C ; Plasmids ; genetics

This study was aimed to explore the influence of excessive complement activation on the pathological process of acute graft-versus-host disease (aGVHD) in mice. A murine model with aGVHD was established by injecting cell mixture containing splenocytes and bone marrow cells at 2:1 ratio from donor C57BL/6(H-2K(b)) mice into recipient BALB/c (H-2K(d)) mice within 4-6 hours after 8 Gy (60)Co γ-ray total body irradiation. The mice received syngeneic bone marrow transplantation were used as control group. After transplantation, the mice were monitored daily for body weight and mortality. At day 14, all mice were sacrificed and each liver was freshly dissociated for histological analysis. The hepatic mRNA abundance for complement components C3a and C5a as well as receptors for these two anaphylatoxin were tested by real-time quantitative PCR method. And the levels of C3a and C5a production in liver were detected by ELISA. The deposition of complement C3 in liver was determined by immunofluorescence staining using frozen section. The results indicated that as compared with syngeneic bone-marrow transplantation control group, experimental animals underwent aGVHD characterized by weight loss, depilation, diarrhea and lassitude. Interestingly, the hepatic mRNA expression for complement anaphylatoxin family member C3a and C5a as well as their receptors C3aR and C5aR1 in mice with aGVHD were significantly up-regulated in comparison with control group (p < 0.05). Consistently, the content of C3a and C5a in liver increased markedly in mice with aGVHD (p < 0.01). For animals ongoing aGVHD, complement component C3 depositions were observed in hepatic portal areas, around which massive inflammatory cell infiltration was also observed. It is concluded that in aGVHD animals, excessive complement activation occurs, and the activated complement components participate in pathological process of the aGVHD.
Animals ; Bone Marrow Transplantation ; Complement Activation ; Female ; Graft vs Host Disease ; immunology ; pathology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL

OBJECTIVETo evaluate the impact of luteinizing hormone-releasing hormone (LHRH) on the protection of thymic function after allogenic hematopoietic stem cell transplantation (allo-HSCT).

METHODSMurine model of MHC mismatched allogeneic HSCT (C57BL/6-->BALB/c) was established. The severity of acute graft-versus-host-disease (GVHD) was assessed according to a clinical scoring system. The intra-cellular levels of IFN gamma, TNFalpha and IL-1 beta in thymocyte were analyzed by protein array and thymic function by quantification of signal-joint TCR rearrangement excision circles (sjTRECs).

RESULTSAll recipients in group A (allogeneic mice), B (allogeneic LHRH castrated-mice) and C (syngenic mice) achieved hematopoietic reconstitution. White blood cell (WBC) over 1.0 x 10(9)/L in groups A, B and C were on day (11.2 +/- 1.4), day (9.8 +/- 0.6) and day (9.7 +/- 0.7), respectively (P = 0.003, 0.002). The onset of acute GVHD in group B was (14.1 +/- 0.7) d and in group A was (11.4 +/- 1.2) d (P = 0.000). All mice in groups A and B developed acute GVHD. No mice occurred aGVHD in group C. The average scores of acute GVHD in groups A and B were (9.1 +/- 0.7) and (5.1 +/- 1.0), respectively (P = 0.000). The levels of IFN gamma, TNFalpha and IL-1 beta in control group were (2.3 +/- 2.5) ng/ml, (1.7 +/- 1.1) pg/ml and (1.8 +/- 1.2) pg/ml, respectively. The IFN gamma levels in groups A, B and C were (10.5 +/- 2.1) ng/ml, (6.7 +/- 2.1) ng/ml and (5.2 +/- 3.3) ng/ml, TNFalpha levels were (7.0 +/- 2.6) pg/ml, (4.3 +/- 0.8) pg/ml and (3.0 +/- 1.8) pg/ml, and IL-1 beta levels were (24.9 +/- 9.0) pg/ml, (17.4 +/- 3.9) pg/ml and (10.8 +/- 3.1) pg/ml, respectively. There were significant differences in the levels of cytokines between group A and the control group (P = 0.000, 0.000, 0.000). The levels of cytokines in group B were significantly higher than those in control group (P = 0.000, 0.003, 0.000). The levels of IFN gamma and IL-beta in group C were significantly higher than those of in control group (P = 0.015, 0.013), and so did in group A than in group B (P = 0.002, 0.002, 0.004), and in group A than in group C (P = 0.000, 0.000, 0.000). The analysis of linear regression showed that the average levels of IFN gamma and TNFalpha paralleled with aGVHD scores (r(2) = 0.359, P = 0.045; r(2) = 0.228, P = 0.019). The average sjTRECs copies/1000 PBMNCs were (39.4 +/- 44.7) in the control group and (12.3 +/- 13.0), (58.0 +/- 71.8) and (19.6 +/- 14.6) in groups A, B and C, respectively. There was no significant difference in the multiple comparisons of peripheral blood levels of sjTRECs among these four groups (P = 0.468).

CONCLUSIONIFN gamma, TNFalpha and IL-1 beta might be involved in the damage to the thymus by acute GVHD. Sex steroid inhibitor can not only reduce the severity of thymic damage after allo-HSCT, but also reduce the severity of aGVHD and the mechanism might be associated with the reduction of intra-cellular levels of IFN gamma in thymocyte.

Animals ; Castration ; methods ; Female ; Gonadotropin-Releasing Hormone ; therapeutic use ; Graft vs Host Disease ; pathology ; prevention & control ; Hematopoietic Stem Cell Transplantation ; Interferon-gamma ; metabolism ; Interleukin-1beta ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Thymus Gland ; immunology ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo explore the effects of graft composition on hematopoietic reconstitution and graft-versus-host disease (GVHD) in HLA-identical related donor peripheral blood stem cell transplantation (PBSCT) for hematological malignancies.

METHODSThe relationship between the number of graft composition and their hematopoietic reconstitution and GVHD in 107 patients with hematological malignancies undergoing HLA-identical related donor PBSCT was retrospectively analyzed.

RESULTSNone of the graft composition numbers had correlation with the time of neutrophil reconstitution. There was a negative correlation between the number of mononuclear cells (MNCs) or CD34+ cells and the time of platelet reconstitution (P < 0.05) with the absolute correlation coefficients below 0.4, and so did between the number of CD34+ or CD34+ CD38- cells and the development of acute GVHD (P < 0.05) and their absolute correlation coefficients. Each lymphocyte subset number had no correlation with acute GVHD. There was a positive correlation between the number of CD25+ CD4+, CD3+ or CD4+ CD3+ cells or CD4+ /CD8+ ratio and the development of chronic GVHD (P < 0.05). And the correlation coefficients all exceeded 0.4, more over, CD25+ CD4+ cells number reached up to 0.78. CD34+, CD34+ CD38- cells number had no correlation with chronic GVHD.

CONCLUSIONIn HLA-identical related donor PBSCT, further increasing infused MNC, CD34+, CD34+ CD38- cells can no more accelerated hematopoietic reconstitution after these cells attained their threshold values, otherwise the incidence of cGVHD might increase due to the increase of the accompanied lymphocytes.

Adolescent ; Adult ; Child ; Female ; Graft vs Host Disease ; immunology ; pathology ; Hematologic Neoplasms ; therapy ; Humans ; Male ; Middle Aged ; Peripheral Blood Stem Cell Transplantation ; Retrospective Studies ; Tissue Donors ; Transplantation, Homologous ; Young Adult

This study was purposed to investigate the change of Th cell subsets in the patients with acute graft-versus host disease (aGVHD) and to explore its role in the pathogenesis of aGVHD after allogeneic hematopoietic stem cell transplantation (allo-HSCT). 23 patients underwent allo-HSCT were selected for analysis. The aGVHD in patients was diagnosed according to clinical features, and was confirmed by skin biopsy in some patients. The peripheral blood from 23 patients was collected before and after allo-HSCT. The quantitative chenges of Th1 and Th2 cells in peripheral blood samples were detected by using flow cytometry. The results showed that out of 23 patieats the aGVHD occured in 8 patients including 1 case of grade I, 2 case of grade II, 3 cases of grade III; no aGVHD occured in 15 patients. The flow cytometry analysis revealed that the amount of Th1 cells in patients with aGVHD was much higher than that in patients without aGVHD (p < 0.01), the IFN-gamma expression of Th1 cells in patients with aGVHD of grad II - III significantly was higher than that in patients without aGVHD (p < 0.01), meanwhie the IL-4 expression of Th2 cells in patients with aGVHD of grade II - III was significantly lower than that in patients without aGVHD (p < 0.05). Dynamical detection indicated that the Th1 obviously increased before occurrence of aGVHD and before treatment of aGVHD, while the Th1 cells obviously decreased after treatment of aGVHD. The Th1 cells not changed significantly in patients without aGVHD before and after allo-HSCT. It is concluded that Th1 cells obviously increase in patients with aGVHD, this increase is related to aGVHD pathogenesis. Detecting the changes of Th cell subsets in the early stage after allo-HSCT may be contributed to early diagnosis and therapy of aGVHD.
Graft vs Host Disease ; blood ; immunology ; Hematologic Neoplasms ; immunology ; therapy ; Hematopoietic Stem Cell Transplantation ; adverse effects ; Humans ; Th1 Cells ; immunology ; pathology

This study was aimed to investigate the migration and distribution processes of allogeneic donor T lymphocytes in the organs of recipient mice. GVHD model was established by transfusion of the splenocytes of eGFP transgeneic C57BL/6 mice together with born marrow cells harvested from C57BL/6 mice into BALB/c mice underwent 8.0 Gy total body irradiation. The migration and homing of eGFP(+) cells were tracked by stereo-fluorescent microscopy or inverted fluorescent microscopy and flow cytometry. The enzyme linked immunosorbent assay (ELISA) was performed on supernatants from the tissue homogenates to detect the amount of MIP-1alpha. The results indicated that GVHD clinical manifestation and pathological changes of organs appeared on day 8 post transplantation. eGFP-positive donor T cells in recipient organs were observed by inverted fluorescence microscope in frozen section, or by stereo-fluorescence microscopy in living organs, such as liver, spleen, skin, lungs, bowels, and tongue. The highest expression of MIP-1alpha was on day 7 post transplantation in the liver (491.3 +/- 32.1 pg/ml), and day 3 post transplantation in the spleen (881.5 +/- 45.2 pg/ml), respectively (P < 0.05). It is concluded that GVHD was induced by splenocytes of eGFP transgeneic C57BL/6 mice. eGFP(+) cells in the organs can be observed by fluorescent microscopy. In this GVHD model, donor T cells proliferate and infiltrate in liver, skin, bowels, as well as lungs and tongue. MIP-1alpha may be in relation with the infiltration of T lymphocytes in liver and spleen.
Animals ; Bone Marrow Transplantation ; adverse effects ; Cell Movement ; Female ; Graft vs Host Disease ; immunology ; pathology ; Green Fluorescent Proteins ; Liver ; immunology ; pathology ; Lung ; immunology ; pathology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Skin ; immunology ; pathology ; Spleen ; cytology ; T-Lymphocytes ; immunology