Indoleamine 2, 3-dioxygenase expression in cells of human acute monocyte leukemia (M(5)) and acute lymphocyte leukemia and therapeutic effect of its inhibitor 1-methyl tryptophan / 中国实验血液学杂志

The objective of this study was to investigate the expression and function of indoleamine 2, 3-dioxygenase (IDO) in leukemia. The IDO expressions in human acute monocyte leukemia (M(5)) and acute lymphocyte leukemia (ALL) were detected by immunofluorescence staining. Constructed leukemia mouse model was used to observe whether the IDO inhibitor, 1-methyl tryptophan (1-MT), has any effect in treating leukemia. The experimental group were fed with 1-MT solution every day while the mice in control group had no further treatment. The results showed that the average ratios of IDO expression were 29.4 +/- 11.2% in M(5) patients and 24.7 +/- 7.96% in ALL patients respectively. After statistical test, IDO expression level in leukemia cells was significantly higher than that of normal mononuclear cells. The tumor decreased gradually in mice treated with 1-MT. At the terminal point of the experiment (88 days after vaccination), the average survival time in the experimental group was 42.3 days while the mice in control group only lived 15.1 days in average, which difference was statistically significant (P < 0.05). Some of the leukemia mice in the experimental group long-term survived without tumor (more than three months after vaccination). It is concluded that human acute monocyte leukemia (M(5)) and acute lymphocyte leukemia (ALL) express IDO, and both can be treated by 1-MT in mice.

Immunological screening for multiple myeloma-associated antigens and their bioinformatics analysis / 中国实验血液学杂志

This study was aimed to screen the cell cDNA expression library of multiple myeloma HMy2 (MM HMy2) by using "serological analysis of cDNA expression library (SEREX)" technique. The obtained 30 positive clones were all sequenced, and analyzed by BLAST (basic local alignment search tool). The results indicated that 6 known genes and 12 new MM-associated genes were obtained, part of which sequences were spliced by EST (expressed sequence tag) splicing. 6 known genes such as for ring finger protein 167, KLF10, TPT1 protein, p02 protein, cDNA FLJ46859 fis, DNMT1 methyltrasferase etc. have been demonstrated a certain relationship with other tumor's formation, progress and prognosis. The structures and functions of the new genes preliminarily analyzed and predicted by means of bioinformatics showed that MMSA-3, MMSA-8 and MMSA-11 encoding 215, 160 and 122 amino acid residues respectively had the full open reading frames (ORF). All the new genes might be located at euchromosomes but MMSA-1 at sex chromosome. MMSA-4 was highly similar to the protein controlling the transcription of tumor antigen, MMSA-5 might take part in cell phagocytosis, MMSA-7 might inactivated NF-kappaB, and MMSA-12 might be a lymphocytic cytoplasmic protein. The specificity of new genes such as MMSA-3 and MMSA-7 were higher, by a preliminary analysis using CrELISA. It is concluded that tumor antigens screened by this study can be used for early immunological diagnosis, surveillance of minor residual foci, assessment of prognosis, and preparation of tumor vaccine and so on.

Construction of FAS-targeted RNAi-retroviral vector and its identification for biological activity / 中国实验血液学杂志

This study was aimed to construct mouse Fas-targeting si RNA-expressing recombinant retroviral vector in order to explore the therapeutic potential of Fas inhibition by siRNA in the treatment of aplastic anemia and to provide a basia for extensive development of RNA interference techninque. The U6(+) 27 promoter cassette and siFas sequence were obtained by PCR method. The U6-siFas fragment was cloned into the multiple restriction site of pLXSN-EGFP and directly downstream of EGFP gene. The resultant retroviral vector pLXSN/EGFP-siFas was packaged using PA317 cell line and tittered using NIH3T3 cell line. P815 cells were infected by the retroviral vector. EGFP expression in P815 was observed under fluorescent microscope and Fas inhibition effect was detected by immunohistochemistry. The results indicated that successfully constructed retrovirus vector pLXSN/EGFP-siFas was could not only deliver siRNA into mammalian cells efficiently and inhibit Fas expression in P815 cells, but also could express EGFP as marker and neomycine resistance gene to allow antibiotic selection. It is concluded that the successful construction of this retroviral vector would greatly facilitate the application of RNA interference and lay the foundation for therapeutic study of Fas inhibition in the treatment of aplastic anemia.

Construction and identification of Fas-targeting siRNA-expressing plasmid / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

<p><b>OBJECTIVE</b>To study the therapeutic potential of Fas inhibition in different diseases, a Fas-targeting siRNA (small interfering)-expressing plasmid was constructed.</p><p><b>METHODS</b>The U6 promoter cassette and siFas (small interfering RNA that inhibit Fas expression) template sequence were obtained by PCR method. They were cloned into modified pcDNA3.1. The resultant plasmid pU6-siFas was transfected into P815 cells with lipofectin2000 and selected under G-418-containing culture medium. Fas inhibition in stably transfected cells was detected by immunocytochemistry.</p><p><b>RESULTS</b>The plasmid pU6-siFas efficiently reduced the expression of Fas and conferred G-418 resistance in P815 cells.</p><p><b>CONCLUSION</b>The successful construction of the siRNA expressing plasmid will facilitate the application of RNA interference technique and lay the foundation for further study of Fas inhibition in the treatment of different diseases such as aplastic anemia and acute liver failure.</p>