PF242 LONG NON‐CODING RNA MANTIS INHIBITS THE PROLIFERATION OF ACUTE MYELOID LEUKEMIA CELL LINE VIA NOTCH SIGNALING PATHWAY

Abstract

Background:Chemotherapy and stem cell transplantation are effective to many acute myeloid leukemia (AML) patients, while resistance and recurrence still affect patient's prognosis. It is important to study the pathogenesis and molecular mechanism of AML.Long non‐coding RNA (lncRNA) plays an essential role in the process of proliferation, invasion and metastasis of many kinds of tumors by regulating signal pathways and biological functions. The role of lncRNA is similar to oncogenes or tumor suppressor genes. lncRNA can regulate the maturation and differentiation of blood cells, and participate the pathogenesis of malignant hematological diseases, such as leukemia. lncRNA MANTIS is a new type of differentially regulated lncRNA with the function of promoting endothelial angiogenesis. Whether lncRNA MANTIS can affect the pathogenesis of AML need more study.Aims:To investigate the expression of MANTIS in AML. To study the role of MANTIS in the proliferation and apoptosis of U937 cells. To explore the relationship between MANTIS and Notch signaling pathway.Methods:Thirty‐six patients with acute myeloid leukemia (non M3 type) and eleven health donors were enrolled to collect and isolate bone marrow mononuclear cells to detect the expression of lncRNA MANTIS by Real‐time PCR. Culture the U937 cell line. Overexpression pcDNA‐ MANTIS was transfected into U937 cells using Lipofectamine 3000 (Invitrogen). The expression of MANTIS in overexpression pcDNA‐ MANTIS U937 cells was detected by Real‐time PCR. Cell proliferation was measured by CCK‐8 Cell Counting Kit. Apoptosis was detected by Annexin ‐FITC/PI double staining and flow cytometry analysis. Expression of Notch1, Dll4 and Hes1 protein were detected by western blot. The data were presented as means  ±  standard deviation, and the difference between two groups was evaluated using Student's test. A probability level of 0.05 is used to establish significance and indicated as P < 0.05.Results:The expression of MANTIS in AML patients was significantly lower than that in normal controls, and there was statistical significance between the two groups (P < 0.01). In U937 cell line, the expression of MANTIS was also significantly lower than normal controls (P < 0.05).The expression of MANTIS in the over‐expression pcDNA‐MANTIS U937 cells (U937/MANTIS) was 441 times higher than the pcDNA plasmid U937 cells (U937/vector) (P < 0.05). The proliferation inhibition rate in U937/MANTIS group was higher than the U937/vector group (P < 0.05). In U937/MANTIS group, the inhibition rate at 12 h, 24 h, 48 h and 72 h were (11.16 ± 2.13)%, (15.47 ± 0.31)%, (21.86 ± 1.71)% and (42.97 ± 2.61)%, respectively. Over‐expression pcDNA‐MANTIS induced U937 cells apoptosis. The apoptotic rates of U937/untreated, U937/vector and U937/MANTIS group were (31.71+0.32)%, (35.42+1.94)% and (59.06+2.37)%, respectively, and the difference was statistically significant (P < 0.05). Notch1, Dll4 and Hes1, which were key proteins in Notch signaling pathway, were increased in U937/MANTIS group. Protein level was higher in the U937/MANTIS group than the U937/vector group.Summary/Conclusion:Expression of lncRNA MANTIS was low in the bone marrow of AML and U937 cell line. Increased expression of lncRNA MANTIS can inhibit the proliferation and induce apoptosis of AML cell line, and increase the level of Notch1, Dll4 and Hes1. The lncRNA MANTIS affected the AML cell line via Notch signaling pathway.image