Levels and Significance of Serum Interleukin-35 and the New Regulatory T Cells -iTR35 Cells in Myelodysplastic Syndrome Patients / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the significance of serum interleukin-35 level and the new regulatory T cells -iTR35 cells in patients with myelodysplastic syndrome (MDS).</p><p><b>METHODS</b>Twenty three cases of newly diagnosed MDS were enrolled in this study from January 2014 to January 2016 in Department of Hematology of The First Hospital of Quanzhou in Fujian Province. According to MDS International Prognostic Scoring System (IPSS), the 23 patients were divided into 4 groups: high-risk (n=4), intermediate risk-2 (n=10), intermediate risk-1 (n=5) and low-risk group(n=4). Twenty healthy people of routine physical examination were used as control during the same period. Enzyme Linked Immunosorbent assay(ELISA) and flow cytometry(FCM) were used to detect the expression level of serum IL-35, the proportion of iTR35 cells, and the expression levels of associated molecules such as IL-12p35,IL-27EBl3 respectively.</p><p><b>RESULTS</b>The proportion of CD4CD25Foxp3Treg cells in peripheral blood of MDS patients was significantly higher than that in controls (P<0.01). The proportion of iTR35 cells was also higher than that in controls(P<0.01). However, the proportion of CD4+ CD25Foxp3T cells was not significantly different between 2 groups (P>0.05). The level of serum IL-35 in MDS patients was significantly higher than that in control group (P<0.05). The expression levels of IL-12p35 and IL-27EBl3 in the Treg cells were also significantly upregulated than those in control group(P<0.05), the expression levels of IL-35, IL-12p35 and IL-27EBl3 in MDS group positively correlated with the proportion of iTR35 cells(r=0.92, 0.99 and 0.52, P<0.05, respectively). IL-35 level and the proportion of iTR35 cells in 4 groups of MDS showed significantly difference in general term, no significant difference was found in IL-35 level between the high-risk group and intermediate risk-2 group (P>0.05), but the IL-35 levels in both groups were significantly higher than those in intermediate risk-1 group and the low-risk group (P<0.05), and the level in the intermediate risk-2 group was significantly higher than that in the intermediate risk-1 group and the low-risk group (P<0.05), while there was not different between the intermediate risk-1group and the low risk group (P>0.05). The proportion of iTR35 cells was not significantly different between the high-risk group and the intermediate risk-2 group. The proportions of iTR35 cells in the high-risk group and the intermediate risk-2 group were higher than those in the intermediate risk-1 group and the low-risk group respectively (P<0.05), but there was no differentce in population of iTR35 between the intermediate risk-1 group and the low-risk group (P>0.05).</p><p><b>CONCLUSION</b>The imbalance between IL-35 level and iTR35 cells propertion may play an important role in the development of MDS, which possibly to provides a new theoretical basis for the study of MDS immune targeting therapy.</p>

Expression Analysis and Epigenetics of MicroRNA let-7b in Acute Lymphoblastic Leukemia / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To study the expression and its mechamisms of microRNA let-7b in adult acute lymphoblastic leukemia (ALL), so as to provide the basis for searching a new targeted therapy.</p><p><b>METHODS</b>Firstly, methylation-specific polymerase chain reaction (MSP) was used to analyze the methylation status of CpG islands in microRNA let-7b promoter of bone marrow mononuclear cells in the patients with ALL and patients with non-hematologic malignancies as control, the real-time fluorescence quantitative polymerase chain reaction (qPCR) was used to detect the expression levels of microRNA let-7b in this 2 groups; and then 5-aza-2'-deoxycytidine (5-Aza-dC, DAC) was used to treat ALL cell line MOLT-4; after drug treatment, MSP was used to analyze the methylation status of the CpG islands in microRNA let-7b promoter; the qPCR was used to detect the expression levels of microRNA let-7b, and further explore the regulatory mechanism of microRNA let-7b expression.</p><p><b>RESULTS</b>Hypermethylation of CpG islands in microRNA let-7b promoter in ALL patients was significantly higher than that in patients with non-hematologic malignancies, and the relative expression level of microRNA let-7b was significantly reduced in ALL patients; 5-aza-dC could significantly inhibit the growth of MOLT-4 cells and arrest the cells in G1 phase, thus biosynthesis of RNA and protein was suppressed, and the apoptosis was promoted, meanwhile, 5-Aza-dC could increase the expression of microRNA let-7b.</p><p><b>CONCLUSION</b>In the patients with ALL, the expression of microRNA let-7b is regulated by methylation of CpG islands in the region of genomic promoter. The microRNA let-7b may act as a tumor suppressor, whose low expression is involved in ALL development, indicating the microRNA let-7b may become a new therapeutic target for ALL.</p>

Proliferation promotion and apoptotic inhibition effects of ribosomal protein RPL36A small interference RNA on U937 cells / 中国实验血液学杂志

This study was purposed to investigate the expression of RPL36A (ribosomal protein 36a) in the newly diagnosed acute myeloid leukemia (AML) cells and its mechanism at the molecular level. The RPL36A mRNA expression in the newly diagnosed AML cells, U937 cells and normal MNCs was determined by RT-PCR. Small interfering RNA (siRNA) targeting to RPL36A was transfected into U937 cells by Lipofectamine 2000 system. Proliferation, cell cycle, apoptosis of U937 were observed through MTT assay, flow cytometry, acridine orange/ethidium bromide (AO/EB) double staining, TUNEL and Annexin V/FITC respectively. RPL36A mRNA and protein expression levels were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analysis respectively. The results showed that RPL36A expression in the newly diagnosed AML cells and U937 cells was significantly upregulated. The average OD value of U937 cells transfected with RPL36A siRNA was significantly lower as compared with 3 control groups. The cell percentage in G2-and S-phase increased, which indicated the inhibition effect of RPL36A siRNA on cell proliferation. Remarkable cell apoptosis in U937 cells treated with RPL36A siRNA was observed by AO/EB, TUNEL analysis and Annexin V/FITC assay; RPL36A mRNA and protein expression level of U937 cells treated with siRNA were significantly declined in a time-dependent manner (r=0.9813 and 0.9537). It is concluded that the RPL36A expression in the AML cells is significantly enhanced and the RPL36A gene may be involved in regulation of cell cycle and cell apoptosis of AML, which promotes proliferation of AML cells and inhibits apoptosis of cells.