The T-large Granular Lymphocyte Leukemia Clinical Features And Efficacy Of Ifn-¦Ã In The Regulation Of The Hematopoietic Role

Objective: To improve the recognization of characteristics of T-LGLL.Methods: Retrospectively analyze the documents of 59 patients with T-LGLL diagnosed between 2000 and 2009 in our hospital.Results: The median age at T-LGLL diagnosis was 50 years without a male or female predilection. The patients mainly complained of fatigue. Of those 59 patients, 40.7% had splenomegaly, and 13.6% heptomegaly. Rheumatoid arthritis was present in 1 patient. The most frequent hematological abnormalities were anemia in 81.4% of patients with median Hgb level of 65g/L. Pure red cell aplasia (PRCA) was more common in this study. Neutropenia was present in 66.1% of patients and 13.6% of patients were agranulocytosis. The median count of LGL in peripheral blood was 1.64×109/L and most of them were less than 2.0×109/L. Fourty-eight patients (81.3%) had the CD3+CD8+CD57+CD56- LGL phenotype. Fourty-eight patients accepted treatment and 87.5% of patients responded. With CsA therapy in 33 patients, 60.6% achieved HCR. The total relapse rate was 35.7%. The median follow-up time was 20 months.Conclusion: T-LGLL was an indolent disease, mainly presented with anemic symptoms. Nearly half of the patients had splenomegaly. Pure red cell aplasia was commonly associated with T-LGLL in this group while rheumatoid arthritis was found only in 1 patient. LGL count in peripheral blood was mostly≤2.0×109/L. The patients had a good response to immunosuppressive therapy. Purpose:To explore the role of murine IFN-γ-(mIFN-γ) in the regulation of hemopoiesis through lentivirus transduction.Methods:The mRNA was extracted from the splenic cells of C57 mouse and mIFN-y ORF was amplificated using RT-PCR and then cloned into lentiviral vector pCDHl-MCS1-EF1-copGFP (pCDH) to construct vector pCDH1-mIFN-y-EF1-copGFP (pCDH-mIFN-y-GFP). The lentiviruses were prepared following transfecting 293T cells by calcium phosphate precipitation. The murine bone marrow mononuclear cells (BMMNCs) were transduced with those two kinds of lentiviruses, and cultured in methylcellulose semisolid medium for colony-forming assay 48 hours later. The transduced BMMNCs were also transplanted into lethally-irradiated mice by tail vein injection and the peripheral blood cell counts 2 weeks later and green fluorescent protein (GFP) were monitored regularly. Bone marrow stromal cells (BMSCs) were transduced with lentivirus and transplanted into normal C57 mice, and the peripheral blood cell counts were also monitored regularly.Result:The titer of lentivirus was measured to be about 4.5×105/ml and 3.4×105/ml for pCDH-mIFN-γ-GFP or pCDH-GFP respectively using a bioassay with the 293T cell line. The expression of mIFN-y in the supernant of 3T3 was confirmed with ELISA kit and the concentration was 188pg/ml. The colony-forming assay showed that the number of colonies in the mIFN-y group was significantly decreased compared to the control. It indicated that mIFN-y may injure the hematopoietic progenitors directly and make its colony-forming capacity decreased. The circulating white blood cell count, platelet count and hemoglobin in mIFN-y group was significantly lower than control recipient mice at 2 weeks, while no difference was observed at 8 weeks. This indicated that the hematopoiesis rebuilding in the mIFN-y group was delayed than control group. It may be caused by the injury of the hematopoietic progenitors which overexpressed mIFN-y. GFP positive cells were detectable in the peripheral blood 8 weeks after transplantation and this indicated that the transduced cells could survive in vivo for at least 8 weeks. BMSCs trandsduced with lentivirus were transplanted into normal C57 mice. When the peripheral blood cell count decreased, the myeloid tissue was examined for pathologic analysis. This part was just ongoing and there was little results for analysis.In conclusion, we constructed mIFN-y expressing vector pCDH-mIFN-γ-GFP. It showed that not only the colony-forming capacity of transduced BMMNCs was inhibited, but also the hematopoiesis rebuilding was delayed. This result indicates that mIFN-y may damage the hematologic progenitors, supporting the hypothesis that IFN-y plays an important role in the mechanism of bone marrow failure.