[Overexpression of eIF4E gene in acute myeloid leukemia and its relation with disease progression].

The aim of this study was to detect the expression level of eIF4E gene in patients with non-treated, remission and non-remission/relapse acute myeloid leukemia (AML), and other non-malignant haematologic diseases so as to analyze and reveal the relationship of eIF4E gene expression with AML progression. SYBR Green I RT-PCR was used to assay the expression level of eIF4E mRNA extracted from bone marrow mononuclear cells in 30 patients with AML (6 in M2, 5 in M3, 8 in M4, 10 in M5, 1 in M6) and 20 patients with non-malignant hematologic diseases. The ß2-microglubin(ß2M) was used as internal reference and the formula 2(-ΔCt)×100% was applied to calculate the expression level of eIF4E gene. The results showed that the eIF4E expression level (7.098 ± 5.544)% in patients with non-treated and non-remitted/relapsed AML was significantly higher than that in patients with remission (0.964 ± 0.312)% (P < 0.01) and non-malignant hematologic diseases (0.248 ± 0.163)% (P < 0.01). There was no difference between latter two group patients, even though the expression level of eIF4E gene in patients with M4 and M5 was higher. As compared with non-malignant hematologic diseases, the expression level of eIF4E gene of patients with remission patients showed no significant difference. It is concluded that the over-expression of eIF4E gene has been found in patients with AML, and its level obviously decreases along with remission of disease, thus the eIF4E gene may be a surveillance parameter for disease progression.

Incidence, risks, and outcome of idiopathic pneumonia syndrome early after allogeneic hematopoietic stem cell transplantation.

We retrospectively analyzed 23 cases with early-onset idiopathic pneumonia syndrome (IPS) of 192 patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) from April 1997 to October 2007. Risk factors for IPS development were evaluated using Cox proportional hazards model, including age, gender, underlying disease, disease status at transplant, transplant type, conditioning regimens, donor type, acute graft-vs.-host disease (GVHD), severity of acute GVHD (aGVHD), human leukocyte antigen (HLA) disparity, and organ involvement of aGVHD. Factors that were significant at the 0.1 level on univariate analysis were evaluated by multivariate analysis. Twenty-three of 192 patients developed IPS (12.0%). Median time to IPS onset after allogeneic HSCT was 76 d (range 32-120 d); median time to death after the diagnosis of IPS was 9 d (range 3-92 d); 20 patients with IPS died because of the rapid progression of respiratory failure (87.0%). Nineteen patients with IPS developed aGVHD (82.6%), with grade III-IV aGVHD in 11 patients (47.8%) and aGVHD of gut in 16 patients (69.6%). The following six factors were associated with an increased risk of IPS by univariate analysis: not in remission, unrelated donor, HLA disparity, occurrence of aGVHD, grade III-IV aGVHD and aGVHD of gut. These risk factors were entered into a multivariate analysis model. Only unrelated donor, grade III-IV aGVHD and aGVHD of gut are identified as being significantly associated with the occurrence of IPS, and among them, aGVHD of gut was associated with the largest risk of IPS, suggesting that the lung may be a target organ of aGVHD.

[Construction and expression of idiotype TCR Valpha1-pIRES-TCR Vbeta8 eukaryotic vectors in vitro].

AIM: To construct the eukaryotic vectors with idiotype TCR Valpha1-pIRES-TCR Vbeta8 of Jurkat cell line and investigate their expression in vitro after transferred into eukaryotic cells. METHODS: TCR Valpha and Vbeta subfamilies of Jurkat cells were analyzed by using RT-PCR and genescan technique. Then the monoclonal TCR Valpha1 and Vbeta8 genes of Jurkat cells were cloned into multiple clone site (MCS) A and MCS B of the eukaryotic vector pIRES respectively. Its sequences were identified by restriction enzyme cutting and sequence analysis. The expression of TCR mRNA and idiotypic protein in transferred A549 and Molt4 cells was tested by RT-PCR, indirect immunophenotyping fluorescein dyeing and flow cytometry, respectively. RESULTS: Two recombinavot eukaryotic vectors with idiotype TCR Valpha1-pIRES-TCR Vbeta8 were developed successfully and the expression of both idiotypic TCR mRNA and protein was detected in transferred A549 and Molt4 cells. CONCLUSION: The successful construction of the two eukaryotic vectors with idiotype TCR Valpha1-pIRES-TCR Vbeta8 will provide a basic method for further study of the specific TCR gene modified T cells.

[Low dose heparin for the prevention of hepatic veno-occlusive disease after allogeneic hematopoietic stem cell transplantation].

OBJECTIVE: To explore the safety and efficacy of low dose heparin for the prevention of hepatic veno-occlusive disease (VOD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) conditioned with busulfan and cyclophosphamide (BU-CY2) and the ideal time frame of the heparin administration. METHODS: From April 1997 to December 2005, 134 patients (75 male, 59 female; median age 25 years old) were transplanted in our bone marrow transplantation unit. All patients were conditioned with BU-CY2 or busulfan-based regimen and scheduled to receive a continuous injection of heparin at a dose of 100 IUxkg-1xd-1. The patients were divided into different groups: 87 patients received related donor transplantation and 47 patients unrelated donor transplantation; 40 patients had abnormal liver function prior to transplantation and the remaining 94 patients normal liver function; 68 patients received heparin from day -7 to day +21 post HSCT and the other 66 patients from day -7 to day +14. RESULTS: All 134 patients had sustained engraftment. The median time for neutrophils to reach 0.5 x 10(9)/L and for platelets to reach 20x10(9)/L were 12 days (range 9-28) and 20 days (range 6-65), respectively. The disease-free survival at day +100 for leukemia patients was 82.2% (97/118). None of 134 patients developed VOD (0%), including 10 patients with class 3 thalassemia major. The incidence and severity of clinical bleeding events in 15 patients monitored with prothrombin time and activated partial thromboplastin time is comparable with those not monitored. CONCLUSIONS: It is suggested that low dose heparin alone is effective and safe for the prophylaxis of VOD following allo-HSCT conditioned with BU-CY regimen. Shortening the duration of administering heparin from -7 d to +21 d to -7 d to +14 d does not affect the effectiveness of low dose heparin to prevent VOD.

Clinical features and risk factors of pure red cell aplasia following major ABO-incompatible allogeneic hematopoietic stem cell transplantation.

The objective of this paper was to study the incidence, risk factors, clinical outcome, management and prevention of pure red cell aplasia (PRCA) following major ABO-incompatible allogeneic hematopoietic stem cell transplantation (allo-HSCT). We retrospectively analyzed 11 cases of PRCA from a series of 42 patients undergoing major ABO-incompatible allo-HSCT from April 1997 to December 2005. Eleven out of the 42 patients developed PRCA (26.1%). All the 11 cases of PRCA were in blood group O recipients of grafts from blood group A donor (n = 9) or blood group B donor (n = 2). The following factors were associated with an increased risk of PRCA: (1) blood group O recipient; (2) blood group A donor; and (3) blood group O/A in recipient/donor pair. Only blood group O/A in recipient/donor pair was identified as being significantly associated with the occurrence of PRCA by multivariate analysis. Six patients who received donor-type plasma exchange did not develop PRCA and among them 5 cases were the blood group O recipients. Eight patients obtained spontaneous remission and in the remaining 3 patients 2 with long-lasting PRCA were successfully treated with plasma exchange with donor-type plasma replacement and the other one who was also complicated by EBV-associated lymphoproliferative disorder (EBV-PTLD) responded rapidly to anti-CD20 monoclonal antibody and achieved complete resolution of clinical finding and symptom of both EBV-PTLD and PRCA. We conclude that blood group A/O in donor/recipient pair is identified as being significantly associated with the occurrence of PRCA by multivariate analysis. Donor-type plasma exchange and anti-CD20 monoclonal antibody is an effective approach for the treatment of PRCA. PRCA could be prevented by plasma exchange prior to transplantation.

[Expressions of transcription factor GATA-1 and GATA-2 genes in bone marrow stromal cells from patients with leukemia].

In order to investigate expressions of transcription factor GATA-1 and GATA-2 genes in the bone marrow stromal cells (BMSCs) from patients with leukemia or normal controls, bone marrow stromal cells from 34 normal cases and 42 cases with leukemia were cultured long-term in vitro. Nonadherent cells (bone marrow hematopoietic cells) and amplified adherent cells (BMSC) were collected separately. Expressions of GATA-1 and GATA-2 genes were analyzed by using RT-PCR-ELISA; the semi-quantitative expression levels of GATA genes in the BMSCs from patients with leukemia were compared with normal controls. The results showed that expressions of GATA-1 and GATA-2 genes could be detected in the BMSCs and the bone marrow hematopoietic cells from both normal controls and the cases of leukemia. The expression ratio of GATA-1 in the BMSCs from acute lymphocytic leukemia (ALL) (85.7%) was similar to the normal controls (88.2%), whereas the expression ratios in BMSCs from acute myelocytic leukemia (AML) (55.6%) and chronic myelocytic leukemia (CML) (41.2%) were significant lower than the normal controls (P < 0.05). The rank of expression level of GATA-1 gene in the BMSCs was "ALL>AML>normal>CML". There was no difference in the expression level of GATA-2 gene within the BMSCs from normal controls and patients with leukemia. The ranks of expression levels of GATA-1 and GATA-2 genes in bone marrow hematopoietic cells were "AML>normal>ALL>CML" and "AML>CML>ALL>normal". The dominant expression of GATA-2 gene was found in the BMSCs from AML, CML or normal controls. It is inferred that the expressions of GATA-1 and GATA-2 genes in the BMSCs of normal controls and patients with leukemia may influence the regulation of hematopoiesis in the bone marrow stroma and it is worthy of further study to explore their roles in pathogenesis and development of leukemia.

[Expression of SCL gene in bone marrow stromal cells from patients with leukemia].

In order to investigate expression of SCL (stem cell leukemia) gene in bone marrow stromal cells (BMSC) and bone marrow cells from patients with leukemia and normal individuals, bone marrow mononuclear cells from AML (18 cases), CML (17 cases), ALL (7 cases) and normal individuals (33 cases) were cultured long-term in vitro. Nonadherent cells (hematopoietic cells) and amplified adherent cells (BMSC) were collected respectively. RT-PCR-ELISA assay was then performed to detect expression of SCL gene. The expression ratio of SCL gene were analyzed and its expression level was normalized by beta(2)M gene acting as an internal reference for the purpose of semi-quantitative analysis. The results indicated that the expression ratio of SCL gene was lower in BMSC from AML (27.8%) and CML (11.8%) than that in normal controls (69.7%, P < 0.05), while was higher in the nonadherent cells from CML (64.3%) than that in its corresponding BMSC (P < 0.05). Semi-quantitative analysis showed that SCL gene expression level in nonadherent cells from AML was higher than that in its corresponding BMSC (P < 0.05). In conclusion, the low-level expression state of SCL gene in BMSC from patients with AML and CML may be involved in the abnormal regulation of hematopoiesis in myelocytic leukemia.

Pure red cell aplasia following major ABO-incompatible allogeneic hematopoietic stem cell transplantation.

Six out of 20 patients undergoing a major ABO-incompatible allogeneic stem cell transplantation (allo-HSCT) developed pure red cell aplasia (PRCA), which did not show any effects on granulocyte and platelet engraftment, and incidence of grade II-IV aGVHD. All the 6 cases of PRCA were in blood group O recipients of grafts from blood group A donors (n = 5) or blood group B donor (n = 1), suggesting that donor/recipient pair (A/O) is associated with a high risk of PRCA after major ABO-incompatible allo-HSCT. Erythroid engraftment occurred spontaneously in four cases without specific intervention other than the RBC transfusion, which coincided with the decrease of isoagglutinin titers below 8, and the remaining 2 patients with prolonged erythroid aplasia( > 300 days) despite therapy with erythropoietin (EPO) were successfully treated by plasma exchange with donor-type plasma replacement. Cyclosporine did not appear to have played any role in causing PRCA in our patients, however, the occurrence of GVHD may facilitate the recovery of erythropoiesis.