Case report: Simultaneous occurrence of primary pulmonary lymphoma and opportunistic infections in a patient with chronic myeloid leukemia.

Background: The occurrence of primary pulmonary lymphoma (PPL) as a secondary malignancy in patients diagnosed with chronic myeloid leukemia (CML) is extremely rare. As the clinical manifestations are atypical, most patients with PPL tend to be misdiagnosed with pneumonia. When the radiographic features of PPL and pulmonary infection overlap, clinicians can be confused about the diagnosis. Here, we report the first case of coexistence of PPL and opportunistic infections in a patient with CML in chronic phase (CML-CP). Case presentation: A 55-year-old woman presented with three weeks of hemorrhage of the oral mucosa at the Department of Hematology. After undergoing various examinations, she was diagnosed with CML-CP and was started on imatinib (400 mg/daily). Due to sudden respiratory distress, the patient was admitted to the respiratory intensive care unit 11 months later. Chest computed tomography (CT) revealed ground-glass opacities, patchy shadows, and multiple nodules in both lungs and enlarged mediastinal lymph nodes. The combination of biapenem and voriconazole antibiotic treatments was effective. The patient's respiratory distress was relieved, but there was intermittent coughing. In the following time, the patient developed a fever, and the imaging findings indicated progression of the disease in both lungs. Bronchoalveolar lavage (BAL) identified pathogens of multiple opportunistic infections. The coexistence of lymphomatoid granulomatosis (LYG) was not confirmed in this patient until a second CT-guided biopsy was performed. Ultimately, the patient underwent chemotherapy in time and is currently alive today. Conclusions: When the patient's recurrent respiratory symptoms and imaging findings do not coincide, secondary tumors should be considered in addition to infection as a diagnosis. In these cases, multiple pathological tissue biopsies should be performed.

CircRNA CircEHBP1 Regulates the Maturation of MiR-129 to Increase the Chemoresistance of Cancer Cells to Adriamycin in Acute Myeloid Leukaemia.

Background: MicroRNA-129 (miR-129) is known to promote chemosensitivity of many types of cancer cells. However, the role of miR-129 in acute myeloid leukaemia (AML) is unclear. We predicted that premature miR-129 might interact with circEHBP1, a well-characterized oncogene in bladder cancer, and analyzed the interaction between circEHBP1 and miR-129 in AML. Methods: Expression of circEHBP1 and miR-129 in AML patients before and after adriamycin (ADR) treatment was determined by RT-qPCR. CircEHBP1 distribution in nuclear and cytoplasm fractions of AML cells was determined using a cellular fractionation assay. The direct interaction of circEHBP1 with premature miR-129 was explored with an RNA-RNA pull-down assay. Finally, the role of circEHBP1 in regulating miR-129 maturation was analyzed in overexpression cells by RT-qPCRs. Results: Compared to the controls, AML patients exhibited increased circEHBP1 and premature miR-129 levels but decreased mature miR-129 levels. Altered gene expression was more obvious in ADR resistant group than in ADR sensitive group. CircEHBP1 was detected in both nuclear and cytoplasm fractions of AML cells and directly interacted with premature miR-129. CircEHBP1 overexpression increased premature miR-129 level but decreased mature miR-129 level. In AML cells, circEHBP1 suppressed ADR-induced cell apoptosis and attenuated the enhancing effects of miR-129 on cell apoptosis. More importantly, the role of circEHBP1 in regulating cell apoptosis was more obvious in ADR resistance cells. Conclusion: CircEHBP1 may suppress miR-129 maturation to increase the chemoresistance of cancer cells to ADR in AML.

Abnormal GRHL2 Methylation Confers Malignant Progression to Acute Leukemia.

Purpose: Abnormal methylation of Grainyhead-like 2 (GRHL2) is associated with a substantial role in the malignant phenotype of tumor patients. Our present research is aimed at studying the abnormal expression of GRHL2 and the association of methylation in patients with acute leukemia and its relationship with prognosis. Materials and Methods: We used quantitative real-time polymerase chain reaction (qRT-PCR) for detecting the aberrant expression level of GRHL2 in 60 patients with acute leukemia and 60 normal controls. We analyzed the significant correlation between the expression level of GRHL2 with clinicopathological features and patients' prognosis in acute leukemia using the corresponding statistical methods. Secondly, we employed qRT-PCR and Western blotting to detect the mRNA and protein levels of GRHL2 in leukemia cell lines. Next, we used methylation-specific polymerase chain reaction (MSP) technology for detecting the methylation of GRHL2 in clinical samples with acute leukemia and cell lines. Then we investigated the demethylating effect of arsenic trioxide and 5-azacitidine on the mRNA and protein expression levels of GRHL2 in cell lines of acute leukemia. Finally, we studied the effects of arsenide trioxide and 5-azacitidine on the proliferation of leukemia cells and the TGF-ß signaling pathway. Results: We found a lower level of GRHL2 expression not only in acute leukemia patients but also in cell lines when compared with normal controls. At the same time, the expression level of GRHL2 in patients with acute leukemia was significantly correlated with leukocyte count, platelet count, and cytogenetic risk grouping. In addition, the lower GRHL2 expression group showed a significantly lower overall survival rate in acute leukemia patients than that of patients with a higher GRHL2 expression group. Univariate and multivariate analyses revealed that the expression of GRHL2 is an independent risk factor in acute leukemia patients. The methylation level of the GRHL2 promoter region in acute leukemia patients and cell lines was significantly higher than the normal control group, and we found the elevated mRNA and protein levels of GRHL2 in acute leukemia cell lines after the use of the demethylation drug arsenic trioxide and 5-azacitidine. At the same time, arsenide trioxide and 5-azacitidine are associated with the inhibition of cellular proliferation of acute leukemia cells and also promote the elevated expression of TGF-ß signaling pathway-linked proteins, including TGF-ß, Smad2, Smad3, and Smad4. Conclusion: Increased expression and methylation level of GRHL2 are closely associated with the prognosis and malignant phenotype of acute leukemia patients and play an irreplaceable role in the occurrence and development of patients with acute leukemia.

Dynamic trajectory of platelet counts after the first cycle of induction chemotherapy in AML patients.

BACKGROUND: Platelet counts varied over time after induction chemotherapy. We aimed to investigate the different trajectories of platelet counts after the first cycle of induction chemotherapy in patients newly diagnosed with acute myeloid leukemia. METHODS AND RESULTS: In total, 149 individuals were included in this study. We identified four distinct trajectories using a group-based trajectory model: low- stability group (n = 27, 18.12%), low-level decrease-medium elevation group (n = 42, 28.19%), low-level decrease-high elevation group (n = 60, 40.27%), and high-level decrease-medium elevation group (n = 20, 13.42%). The baseline characteristics of the high-level decrease-medium elevation group included higher platelet count, lower white blood cell count, lower percentage of bone marrow blasts, and lower rates of complete remission after the first cycle of induction chemotherapy. Compared with the low-stability group, the hazard ratios were 0.32 (95% confidence interval, 0.15-0.68) for the low-level decrease-medium elevation group, 0.31 (95% confidence interval, 0.15-0.63) for the low-level decrease-high elevation group, and 0.35 (95% confidence interval, 0.13-0.89) for the high-level decrease-medium elevation group after adjustment for age and gender by Cox proportional hazard regression. Compared with the low-stability group, the hazard ratios were 0.33 (95% confidence interval, 0.14-0.77) for the low-level decrease-medium elevation group and 0.31 (95% confidence interval, 0.14-0.67) for the low-level decrease-high elevation group after adjustment for age, gender, white blood cell count, and bone marrow blasts. These associations persisted after adjusting for age, gender, white blood cell count, bone marrow blasts, and platelet count. CONCLUSION: The dynamic trajectory of platelet counts after the first cycle of induction chemotherapy is a significant predictor of all-cause mortality in patients with acute myeloid leukemia. Timely intervention should be considered for the low-stability group. The low-level decrease-medium elevation and low-level decrease-high elevation groups were independent protective factors for all-cause mortality.

MiR-140 Targets lncRNA DNAJC3-AS1 to Suppress Cell Proliferation in Acute Myeloid Leukemia.

OBJECTIVES: MiR-140 and DNAJC3-AS1 have been demonstrated to play critical roles in cancer biology, while their participation in acute myeloid leukemia (AML) is unclear. This study aimed to explore the role of miR-140 and DNAJC3-AS1 in AML. METHODS: The expression of DNAJC3-AS1 and miR-140 were detected by RT-qPCR. Then, the role of DNAJC3-AS1 and miR-140 in regulating each other was explored by overexpression assay. Next, the direct interaction between DNAJC3-AS1 and miR-140 was analyzed using an RNA pull-down assay. Next, the subcellular location of DNAJC3-AS1 was explored using cellular, subcellular fractionation assay. Finally, cell proliferation analysis was evaluated with BrdU assay. RESULTS: Increased expression levels of DNAJC3-AS1 and decreased expression levels of miR-140 were observed in AML patients. DNAJC3-AS1 was detected in nuclear and cytoplasm samples and direct interaction between DNAJC3-AS1 and miR-140 was observed. DISCUSSION: Reduced expression levels of DNAJC3-AS1 were observed after overexpression of miR-140 in AML cells. DNAJC3-AS1 increased cell proliferation and inhibited the role of miR-140 in suppressing cell proliferation. CONCLUSION: In conclusion, miR-140 may target DNAJC3-AS1 to suppress cell proliferation in AML.

CircRNA circ_POLA2 is Upregulated in Acute Myeloid Leukemia (AML) and Promotes Cell Proliferation by Suppressing the Production of Mature miR-34a.

PURPOSE: CircRNA circ_POLA2 has been reported as an oncogene in lung cancer, while its role in other malignancies is unknown. This study aimed to explore the role of circ_POLA2 in acute myeloid leukemia (AML). METHODS: The expression levels of circ_POLA2, mature miR-34a and miR-34a precursor in bone marrow mononuclear cells (BMMNCs) from AML patients (n = 50) and healthy controls (n = 50) were determined by RT-qPCR. Correlations among them were analyzed by Pearson's correlation coefficient. Overexpression of circ_POLA2 was achieved in AML cell lines, followed by the measurement of the expression levels of mature miR-34a and miR-34a precursor. The role of circ_POLA2 and miR-34a in regulating AML cell proliferation was assessed by CCK-8 assay. RESULTS: Circ_POLA2 was upregulated in AML and inversely correlated with mature miR-34a, but not miR-34a precursor. In AML cells, overexpression of circ_POLA2 decreased the expression levels of mature miR-34a, but not miR-34a precursor. Cell proliferation analysis showed that the overexpression of miR-34a attenuated the effects of overexpression of circ_POLA2 on cell proliferation. CONCLUSION: Circ_POLA2 is upregulated in AML and promotes cell proliferation by suppressing the production of mature miR-34a.

Cyclosporine enhances the sensitivity to lenalidomide in MDS/AML in vitro.

Our previous study revealed that expression of G protein-coupled receptor 68 (GPR68) was upregulated in MDSL cells, a cell line representing myelodysplastic syndromes (MDS), in response to lenalidomide (LEN), and mediated a calcium/calpain proapoptotic pathway. Isx, a GPR68 agonist, enhanced the sensitivity to LEN in MDSL cells. The fact that Isx is not a U.S. Food and Drug Administration-approved drug prompts us to look for alternative candidates that could enhance the sensitivity of LEN in MDS as well as other hematologic malignancies, such as acute myeloid leukemia (AML). In the study described here, we found that regulator of calcineurin 1 (RCAN1), an endogenous inhibitor of calcineurin (CaN), was upregulated in MDSL cells in response to LEN, possibly through degradation of IKZF1. Consistently, cyclosporin (Cys), a pharmacological inhibitor of CaN, inhibited the activity of CaN and induced apoptosis in MDSL cells, indicating that CaN provided a prosurvival signal in MDSL cells. In addition, Cys enhanced the cytotoxic effect of LEN in MDS/AML cell lines as well as primary bone marrow cells from MDS patients and AML patient-derived xenograft models. Intriguingly, pretreatment with LEN reversed the suppressive effect of Cys on T-cell activation. Our study suggests a novel mechanism of action of LEN in mediating cytotoxicity in MDS/AML via upregulation of RCAN1, thus inhibiting the CaN prosurvival pathway. Our study also suggests that Cys enhances the sensitivity to LEN in MDS/AML cells without compromising T-cell activation.

G protein-coupled receptor 68 increases the number of B lymphocytes.

G protein-coupled receptor 68 (GPR68) is a proton sensor that is activated upon binding to extracellular protons. We have previously found that GPR68 induces a proapoptotic pathway in bone marrow (BM) cells from the patients with myelodysplastic syndromes (MDS) after treated with lenalidomide. However, the function of GPR68 in normal hematopoietic cells remains unclear. With genetic loss of function approach, we found reduced frequency and number of B lymphocytes in the peripheral blood (PB) of whole body Gpr68-/- mice compared to control littermates upon aging. During hematopoietic regeneration, such as in response to fluorouracil (5-FU), we also found reduced frequency and number of B lymphocytes in Gpr68-/- mice compared to wild type mice. Mechanism studies revealed that Gpr68 expression was upregulated in B lymphocytes of BM during aging and in hematopoietic progenitor cells after treatment with 5-FU. In addition, activation of Gpr68 by its activators increased the frequency and number of B lymphocytes. Our studies indicate that Gpr68 expression is upregulated in hematopoietic cells upon aging and during hematopoietic regeneration that ends up with increased number of B lymphocytes.

Nephrogenic diabetes insipidus in initial stage of acute lymphoblastic leukemia and relapse after haploidentical hematopoietic stem-cell transplantation: A case report.

RATIONALE: Nephrogenic diabetes insipidus (NDI) rarely presents in the initial stage of acute lymphoblastic leukemia (ALL) and relapse due to renal infiltration is also rare. PATIENT CONCERNS: A 19-year-old man presented with weakness, polydipsia, and polyuria for 1 month. DIAGNOSES: NDI was diagnosed with insignificant response to a water deprivation test after stimulation with vasopressin injection. Bone marrow examination combined with immunophenotypic analysis, cerebrospinal cytology, and abdominal ultrasonography confirmed the diagnoses of precursor B cell ALL with renal infiltration. INTERVENTIONS: The patient accepted standardized combination chemotherapy and ultimately had sustained remission, and his polydipsia and polyuria disappeared after 3 days of treatment. The ALL relapsed 1 year later and he received haploidentical stem cell transplantation (haplo-SCT) from his father. OUTCOMES: One year later, he again developed NDI, with bilateral renal enlargement because of extramedullary relapse, leading to subsequent death. LESSONS: This case demonstrates unusual early renal involvement in ALL presenting with initial NDI. Interestingly, the NDI returned with the relapse of renal infiltration 1 year after haplo-SCT. This case suggests that NDI was probably secondary to renal leukemic infiltration.

Arsenic trioxide increases expression of secreted frizzled-related protein 1 gene and inhibits the WNT/ß-catenin signaling pathway in Jurkat cells.

The aim of the present study was to investigate the demethylation effect of arsenic trioxide (As2O3) on the secreted frizzled-related protein 1 (SFRP1) gene and its ability to inhibit the Wingless-type MMTV integration site family (WNT) pathway in Jurkat cells. Methylation-specific polymerase chain reaction was used to examine the CpG island methylation status of the SFRP1 gene in leukemia cell lines. In addition, the effects on Jurkat cells of treatment with different concentrations of As2O3 for 48 h were investigated. Reverse transcription-quantitative polymerase chain reaction was employed to measure the expression of mRNAs, while western blot analysis was used to examine protein expression in cells. The SFRP1 gene was methylated in Jurkat cells. However, both methylated and unmethylated SFRP1 genes were detected in HL60 and K562 cells. In normal bone marrow mononuclear cells, the SFRP1 gene was unmethylated. Following treatment with As2O3 for 48 h, the SFRP1 gene was demethylated, and the mRNA and protein expression levels of the SFRP1 gene were increased. By contrast, the mRNA and protein expression levels of ß-catenin and cyclin Dl were downregulated. The protein expression of c-myc was also downregulated, but As2O3 exhibited no significant effect on the mRNA expression of c-myc. Abnormal methylation of the SFRP1 gene was detected in Jurkat cells. These results suggest that As2O3 activates SFRP1 gene expression at the mRNA and protein levels in Jurkat cells by demethylation of the SFRP1 gene. Furthermore, they indicate that As2O3 regulates WNT target genes and controls the growth of Jurkat cells through the WNT/ß-catenin signaling pathway.

Suppression of graft-versus-host disease and retention of graft-versus-tumour reaction by murine genetically engineered dendritic cells following bone marrow transplantation.

The effect of infusion of lentiviral vector­mediated, genetically engineered dendritic cells (DCs) following allogeneic bone marrow transplantation (allo­BMT) on graft­versus­host disease (GVHD) and graft­versus­leukemia (GVL) was investigated in a mouse model. Lentivirus­mediated expression of soluble tumor necrosis factor receptor 1 (sTNFR1) converted immature DCs (imDCs) from BABL/c mice into engineered DCs in vitro. An EL4 leukemia allo­BMT model of BABL/c to C57BL/6 mice was established. Engineered DCs with donor bone marrow cells and splenocytes were subsequently transplanted into myeloablatively irradiated recipients. The average survival duration in the sTNFR1­ and pXZ9­imDC groups was significantly prolonged compared with that of the allo­BMT group (P<0.05). Mild histological changes in GVHD or leukemia were observed in the recipients in the sTNFR1­imDC group and clinical GVHD scores in this group were significantly decreased compared with those of the transplantation and pXZ9­imDC groups. Serum interferon­Î³ levels were decreased in the pXZ9­imDC and sTNFR1­imDC groups compared with those in the allo­BMT group (P<0.05), with the reduction being more significant in the sTNFR1­imDC group (P<0.05). Serum interleukin­4 expression levels were decreased in the allo­BMT group, but gradually increased in the pXZ9­imDC and sTNFR1­imDC groups (P<0.05). Co­injection of donor genetically­engineered imDCs was able to efficiently protect recipient mice from lethal GVHD while preserving GVL effects during allo­BMT.

Matrix metalloproteinase-2 and -9 secreted by leukemic cells increase the permeability of blood-brain barrier by disrupting tight junction proteins.

Central nervous system (CNS) involvement remains an important cause of morbidity and mortality in acute leukemia, the mechanisms of leukemic cell infiltration into the CNS have not yet been elucidated. The blood-brain barrier (BBB) makes CNS become a refugee to leukemic cells and serves as a resource of cells that seed extraneural sites. How can the leukemic cells disrupt this barrier and invasive the CNS, even if many of the currently available chemotherapies can not cross the BBB? Tight junction in endothelial cells occupies a central role in the function of the BBB. Except the well known role of degrading extracellular matrix in metastasis of cancer cells, here we show matrix metalloproteinase (MMP)-2 and -9, secreted by leukemic cells, mediate the BBB opening by disrupting tight junction proteins in the CNS leukemia. We demonstrated that leukemic cells impaired tight junction proteins ZO-1, claudin-5 and occludin resulting in increased permeability of the BBB. However, these alterations reduced when MMP-2 and -9 activities were inhibited by RNA interference strategy or by MMP inhibitor GM6001 in an in vitro BBB model. We also found that the disruption of the BBB in company with the down-regulation of ZO-1, claudin-5 and occludin and the up-regulation of MMP-2 and -9 in mouse brain tissues with leukemic cell infiltration by confocal imaging and the assay of in situ gelatin zymography. Besides, GM6001 protected all mice against CNS leukemia. Our findings suggest that the degradation of tight junction proteins ZO-1, claudin-5 and occludin by MMP-2 and -9 secreted by leukemic cells constitutes an important mechanism in the BBB breakdown which contributes to the invasion of leukemic cells to the CNS in acute leukemia.

[Disruption of blood brain-barrier by leukemic cells in central nervous system leukemia].

OBJECTIVE: To observe the effect of leukemic cells on blood-brain barrier (BBB) in mice with central nervous system leukemia (CNSL) by establishing mice CNSL model and an in vitro BBB model and explore the mechanism of leukemic cell infiltrating central nervous system (CNS). METHODS: After splenectomy, cytoxan intraperitoneal injection, and sublethal irradiation, 10 BALB/c nu/nu mice were transplanted intravenously with 1.2 × 10(7) of SHI-1 human monocytic leukemic cells. Mice were monitored for survival and clinical manifestation of nerve palsy. The leukemic cells engrafted were examined by RT-PCR, histopathology and bone marrow (BM) smears. Immunofluorescence analysis with laser scanning fluorescence confocal microscopy was used to determine the expression of fibrinogen and tight-junction protein ZO-1. An in vitro BBB model composed of human brain microvascular endothelial cells (BMVECs) was developed on a Matrigel-based insert. Different leukemic cell lines were seeded onto the upper compartment of transwell insert. After incubated for 24 h with BMVECs, cells that had migrated into the lower compartment were counted and analyzed. RESULTS: (1) Paralysis with or without sight loss was developed in half the mice 30-35 d after innoculated with SHI-1 cells. Leukemic cells infiltrates were observed in BM and in different part of brain tissues including brain parenchyma. The transcriptions of human MLL/AF6 fusion gene were also detected in BM and brain tissues in paralysis mice. The fibrinogen expression and ZO-1 disruption were detected in the infiltrated tissue. (2) After 24 h incubation with leukemic cells, the BMVECs sheets were disrupted and grew singly and ZO-1 expression was down-regulated markedly. SHI-1 cells showed more injurious to BMVECs and higher invasive rate \[(40.33 ± 1.53)% vs (11.83 ± 1.44)%, P < 0.05\] than HL-60 cells did. CONCLUSION: One of the mechanisms of leukemic cells infiltrates CNS in CNSL is injure to the BBB.

Efficacy of pretreatment of allografts with methoxypolyethylene glycol-succinimidyl-propionic acid ester in combination with an anti-OX40L monoclonal antibody in relieving graft-versus-host disease in mice.

We designed to determine whether pretreatment of allografts with methoxypolyethylene glycol-succinimidyl-propionic acid ester (mPEG-SPA) and an anti-OX40L monoclonal antibody (McAb) can relieve acute graft-versus-host disease in allogeneic bone marrow transplantation recipients. Responder splenocytes from C57BL/6 donor mice were incubated with stimulator splenocytes from BALB/c recipient mice for 7 days in the presence or absence of anti-OX40L McAb followed by mPEG-SPA modification. Donor BM cells plus mixed culture T cells were then transplanted into myeloablatively irradiated BALB/c mice. The signs of GVHD were less evident in mice of groups B (mPEG-SPA modification group), C (anti-OX40L McAb pretreated group) and D (dual-treated group), with average survival durations all longer than those in group A (non-treated BMT group) (P < 0.05). The survival rates on day 60 post-BMT in groups B, C and D were 50, 41.7 and 66.7%, respectively. After BMT, serum IL-4 and IL-10 levels elevated in groups B, C (P < 0.05) and even more significantly increased in group D (P < 0.01), while serum IFN-gamma levels decreased in these three groups (P < 0.01). In conclusion, the combination of mPEG-SPA and anti-OX40L McAb can block T cell-activated antigens, co-stimulatory pathways and induce the immune shift of Th cells toward Th2 cells; their effects in ameliorating GVHD are synergistic.

[Prophylaxis of graft-versus-host disease in mice by chemical modification of graft and OX40-OX40L costimulatory pathway.].

OBJECTIVE: To explore the prophylaxis effect of pretreatment of allograft with methoxypolyethylene glycol-succinimidyl-propionic acid ester (mPEG-SPA) and anti-OX40L monoclonal antibody (McAb) on acute graft-versus-host disease (aGVHD) after allogeneic bone marrow transplantation (allo-BMT) in mice. METHODS: Responder splenocytes from C57BL/6 donor mice (H-2(b)) were co-cultured with stimulator splenocytes from BALB/c recipient mice (H-2(d)) for 7 days in the presence or absence of anti-OX40L McAb followed by mPEG-SPA chemical modification. Donor bone marrow cells plus the mixed culture of T-cells were then transplanted into lethally irradiated BALB/c mice. The BALB/c recipient mice were divided into four groups: group A (allo-BMT control group), group B(mPEG-SPA modification group), group C (anti-OX40L McAb pretreated group) and group D (mPEG-SPA and anti-OX40L McAb dual-treated group). Survival time and survival rate of the recipients were observed after allo-BMT. GVHD was assessed by clinical signs and histological changes of skin, liver and small intestines. Enzyme-linked immunosorbent assay (ELISA) was used to detect cytokines (IL-4, IL-10 and INF-gamma) production. Flow cytometry (FCM) analysis was used to detect allogeneic chimerism. RESULTS: (1) The mice in group A developed typical clinical signs of aGVHD and all mice died within 17 days after BMT with an average survival time (AST) of (12.1 +/- 5.5) days. The signs of aGVHD were less evident in mice of groups B, C and D, and their AST (36.2 +/- 24.9, 32.0 +/- 24.8 and 44.3 +/- 23.2 days, respectively) were all longer than that in group A (P < 0.05). AST of group D being the longest (P < 0.05). The survival rates at day 60 post-BMT in groups B, C and D were 50%, 41.7% and 66.7%, respectively. (2) Serum IFN-gamma level was increased after BMT in group A, and peaked in day 10 to day 15 post-BMT, while the level was decreased in groups B, C and D, reached the nadir on the day 10 post-BMT, with the lowest in group D (P < 0.01). After BMT, IL-4 and IL-10 levels were slightly decreased in group A, their levels were elevated in groups B and C (P < 0.05) and even more significantly increased in group D (P < 0.01). IL-4 and IL-10 levels peaked between day 10 and 15 post-BMT. (3) The average proportion of H-2(b) positive cells in recipient mice was 95% - 100% on day 60 post-BMT, with complete donor-type implantation. CONCLUSION: Combination of mPEG-SPA and anti-OX40L McAb can block T-cell activated antigens and co-stimulatory pathway, regulate the T cells differentiation and induce the immune shift of Th0 cells toward Th2 cells. The immune tolerance induced by this method can significantly relieve aGVHD after allo-BMT.