[Establishment and identification of a H-2 completely mismatched microtransplantation model of leukemia mouse].

This study was purposed to establish and identify a H-2 completely mismatched microtransplantation model of leukemia mouse. The recipients were female BALB/c mice, while donors were male C57BL/6J mice. Recipients were inoculated intravenously with 1×10(6) of WEHI-3 cells, a cell line of myelomonocytic leukemia. Donors received 100 µg/kg G-CSF mobilization through hypodermic injection, every 12 hours, and it last 5 days. Chemotherapy regimens was MA (mitoxantrone+cytarabine), and it last 4 days. Recipients were given chemotherapy conditioning without GVHD prophylaxis after inoculation of leukemic cells for 2 days, and within 8 hours after last chemotherapy received donor mobilized spleen mononuclear cells (sMNC). The number of sMNC was (3, 6, 12) ×10(7), respectively. The early death rate, recovery level of WBC in peripheral blood and leukemia load were compared between chemotherapy and microtransplantation groups. The donor chimerism was detected by RT-PCR. From the clinical manifestation and pathological features, the GVHD in recipients was evaluated. The results showed that the early mortality in chemotherapy group was 25%, meanwhile those in the (3, 6, 12)×10(7) groups were 16.67%, 8.33%, 8.33%, respectively. The(3, 6)×10(7) groups has a stronger hematopoietic recovery capability than that in chemotherapy and 12×10(7) groups (P < 0.05) . There were more leukemic cells in chemotherapy mice than that in microtransplantation mice (P < 0.01) , and (12, 6)×10(7) groups had lower leukemia load than that in 3×10(7) group (P < 0.05) . No signs of GVHD were observed in microtransplantation mice. The donor microchimerism could be discovered at eraly 2 weeks after donor cell transfusion. It is concluded that a H-2 completely mismatched microtransplantation model of leukemia mouse has been successfully established, and it will provide a experimental base for studying microtransplantation in clinic.

HLA-mismatched stem-cell microtransplantation as postremission therapy for acute myeloid leukemia: long-term follow-up.

PURPOSE: Despite best current therapies, approximately half of patients with acute myeloid leukemia in first complete remission (AML-CR1) with no HLA-identical donors experience relapse. Whether HLA-mismatched stem-cell microtransplantation as a novel postremission therapy in these patients will improve survival and avoid graft-versus-host disease (GVHD) is still unknown. PATIENTS AND METHODS: One hundred one patients with AML-CR1 (9 to 65 years old) from four treatment centers received programmed infusions of G-CSF-mobilized HLA-mismatched donor peripheral-blood stem cells after each of three cycles of high-dose cytarabine conditioning without GVHD prophylaxis. Donor chimerism and microchimerism and WT1+CD8+ T cells were analyzed. RESULTS: The 6-year leukemia-free survival (LFS) and overall survival (OS) rates were 84.4% and 89.5%, respectively, in the low-risk group, which were similar to the rates in the intermediate-risk group (59.2% and 65.2%, respectively; P=.272 and P=.308). The 6-year LFS and OS were 76.4% and 82.1%, respectively, in patients who received a high dose of donor CD3+ T cells (≥1.1×10(8)/kg) in each infusion, which were significantly higher than the LFS and OS in patients who received a lower dose (<1.1×10(8)/kg) of donor CD3+ T cells (49.5% and 55.3%, respectively; P=.091 and P=.041). No GVHD was observed in any of the patients. Donor microchimerism (2 to 1,020 days) was detected in 20 of the 23 female patients who were available for Y chromosome analysis. A significant increase in WT1+CD8+ T cells (from 0.2% to 4.56%) was observed in 33 of 39 patients with positive HLA-A*02:01 antigen by a pentamer analysis. CONCLUSION: Microtransplantation as a postremission therapy may improve outcomes and avoid GVHD in patients with AML-CR1.

[Study on biological characteristics of cultured rhesus mesenchymal stem cells].

This study was aimed to isolate and culture rhesus mesenchymal stem cells (MSC), and to analyze its phenotype and biological characteristics. Bone marrow was aspirated from femur of rhesus, mononuclear cells were extracted, the nonadherent cells were removed after 24 hours, adherent cells were subcultured when they grew 80% confluence. After the fifth passage, the cells were used for examination. The phenotypes of MSC were analyzed by flow cytometry, differentiated cells were identified by relevant specific staining. Cytokines' mRNA expressed by MSC were detected by RT-PCR. The results showed that rhesus MSC gave rise to a population of adherent cells characterized by the presence of a predominant cell type with a typical fibroblast-like morphology. During the log phase of growth, MSC proliferated to a two-fold population at 31 - 40 hours. MSC could be ex vivo expanded by successive cycles of trypsinization, seeding, and culture. The phenotypes expressed on rhesus MSC were Flk-1, CD29, CD105, CD166 positive and CD34, CD45, CD33 nearly negative. In various induction differentiation conditions, rhesus MSC could differentiate into the osteoblast and lipocyte. IL-6, TGF-beta were highly expressed, TNF-alpha was lowly expressed and IL-2, Fas-L, IFN-gamma were not detected on rhesus MSC using RT-PCR method. It is concluded that rhesus MSC can be successfully isolated and culture-expanded and the biological characteristics of rhesus MSC are similar to those of human MSC.