ABSTRACT
The purpose of this study was to explore the effect of NOD2 signalling pathway activated by muramyl dipeptide (MDP) on the immunomodulation effect of human monocyte-derived dendritic cells (DC) loaded with leukemia cell lysates. Peripheral blood mononuclear cells (PBMNC) were isolated by density gradient centrifugation, These cells were cultured with three cytokines for 7 days to induce their maturation. On the 5th day, cells were loaded with leukemia cell HL-60 lysates. NOD2 expression was detected by RT-PCR and Western blot. The phenotype of DC were analyzed by flow cytometry, and ELISA was used to assay levels of IL-12 (p40) . The results showed that MDP could trigger NOD2 mRNA and protein expression in different groups of DC, especially in sensitized DC+MDP group, which was significantly higher than that in the DC+MDP group and sensitized DC without MDP stimulation, the difference was statistically significant (P < 0.05). Besides, the expression of surface molecules (HLA-DR, CD80, CD83, CD86, CD40) in the group of DC loaded with leukemia cell lysate and stimulated by MDP (sensitized DC+MDP) reached the highest level, followed by the group of DC loaded with leukemia cell lysate without MDP and DC only stimulated by MDP, non-treated DC were the lowest (P < 0.05). Similarly, compared with untreated unstimulated DC, after loading with HL-60 lysates or only stimulating with MDP, the secretion of IL-12p40 increased, but IL-12p40 level (573.86 ± 32.09 pg/ml) in DC+MDP group was higher than that in group of sensitized DC (365.03 ± 28.86 pg/ml) (P < 0.05), and it in sensitized DC+MDP group reached the highest (898.30 ± 61.08) pg/ml, compared to other groups (P < 0.05). It is concluded that MDP can significantly enhance the NOD2 mRNA and protein expression in sensitized DC, promote the expression of HLA-DR, synergistic costimulatory molecules and adhesion molecules of DC, at the same time, MDP can increase secretion of inflammatory factors IL-12p40. This study will provide a new ideas for DC application in leukemia immunotherapy.
Subject(s)
Humans , Acetylmuramyl-Alanyl-Isoglutamine , Pharmacology , Cells, Cultured , Dendritic Cells , Allergy and Immunology , Metabolism , HL-60 Cells , Interleukin-12 Subunit p40 , Bodily Secretions , Leukemia , Allergy and Immunology , Metabolism , Leukocytes, Mononuclear , Metabolism , Membrane Proteins , Metabolism , Nod2 Signaling Adaptor Protein , MetabolismABSTRACT
The study was purposed to investigate the effect of phosphorylated-chk1 on cell cycle and apoptosis of human erythroleukemic cell line K562 and K562/A02, and to explore the mechanism of chk1 in regulation of drug-resistance of leukemia cells. After treatment with adrimycin for six hours, the cell cycle distribution was detected by flow cytometry; the Chk1mRNA expression was detected by RT-PCR and the Chk1 phosphorylation level was detected by Western blot. Under the condition of down-regulation of Chk1mRNA expression in cells transfected with Chk1 short hairpin RNA, the cell apoptosis rates were detected by flow-cytometry following adrimycin. The results indicated that the proportion of K562/A02 cell line in G2/M phase was (54.12 +/- 0.57)% at 6 hours after drug treatment, significantly higher than that of K562 cell line (36.99 +/- 1.28)%. No evident difference of the Chk1mRNA expression was observed between K562 and K562/A02 cell lines, while elevated Chk1 phosphorylation following DNA damage induced by adriamycin was observed in the K562/A02 cell line (0.79 +/- 0.56), significantly higher than that in K562 cell line (0.27 +/- 1.47). The cell apoptosis rate of the Chk1 shRNA group in K562/A02 cell line was 3.84-fold of blank vector group, but that in K562 cell line was 1.30-fold of blank vector group. It is concluded that the increased chk1 activity that delay the progress of cell cycle are associated with cellular resistance to adrimycin in the K562/A02 cell line.
Subject(s)
Humans , Apoptosis , Physiology , Checkpoint Kinase 1 , Doxorubicin , Pharmacology , Drug Resistance, Neoplasm , Genetics , G2 Phase , K562 Cells , Mitosis , Phosphorylation , Protein Kinases , Genetics , Metabolism , RNA Interference , RNA, Messenger , Genetics , Signal TransductionABSTRACT
To evaluate the clinical efficacy of recombinant human Interleukin 11 in the treatment of pre-aplastic anemia, six patients with pre-aplastic anemia were injected with rhIL-11 of 6 million units once a day during 7-14 days. Blood platelet counts were taken on day 8, 15, 30 and 60 after the treatment, and bone marrow examination was performed on day 15 as compared with those before treatment. The results showed that platelet counts in 3 out of 6 patients increased remarkably (50%), one of the six increased moderately (16.7%), another case of the six increased slightly (16.7%), platelet in one out of six did not significantly increase (16.7%), the total efficacy rate is 83.3%, the amount of megakaryocyte in bone marrow of all six patients increased, the side effect of the rhIL-11 treatment was light. In conclusion, the efficacy of recombinant human Interleukin-11 in the treatment of thrombocytopenia patients with pre-aplastic anemia is satisfactory. As the number of the cases is too small to conclude, further exploration needs accumulation of more applications.