Expression Of Notch Signaling Pathway In Patients With Acute Leukemia And The Relationship With VEGF Pathway

BackgroungLeukemia is a group of heterogenous hematopoietic malignant tumors due to differentiation block,apoptosis obstruction and malignant proliferation of hematopoietic stem cell or progenitor cell.It is the most commom hematological malignancy.Angiogenesis is defined as the formation of new capillaries from preexisting blood vessels,Vascular endothelial growth factor (VEGF) and its receptors are important factors in tumor angiogenesis,which have a major impact in the progress of the leukemia and poor prognosis. Recent study found that Notch signaling pathway not only plays an important role in the maintenance of normal hematopoietic function,but also in the incidence of leukemia.Notch signaling pathway was first identified in human acute T lymphoblastic leukemia(T-ALL)with chromosome translocaton t(7;9), however,some sholars recently noticed that Notch receptor was also ectopically expressed in B-ALL,AML,MM.But the role of Notch signaling pathway in the pathogenesis of leukemia is not clear.For the past few years,a series of evidence indicates the important roles of VEGF and DII4/Notch signaling pathways in modulation of angiogenesis,Aberrant activations of VEGF and DII4/Notch signalings have been found in bone marrow microenvironment of acute leukemia.Despite extensive studies on VEGF and its receptors in AL,the role of DII4/Notch and its relationship with VEGF in modulating angiogenesis are far from reached. ObjectivesIn the present study,we examine the protein and mRNA levels of Notch1, Notchl-IC,DII4 and Hes1 in peripheral blood mononuclear cells in patients with AL to investigate the role of Notch signaling pathway associated genes in the pathogenesis of acute leukemia,we examine the cellular levels of VEGF, VEGFR1,VEGFR2,Notch1 and DII4 in patients with AL to investigate the roles and clinical significances of the two pathways and discuss their relationship in angiogenesis to provide significant information for mechanisms and molecular target therapy.Methods1.Patients and controls:70 patients with acute leukemia treated in our institute between 2008 and 2009 were investigated,which divided into initial group and complete remission(CR) group.And 24 healthy volunteers were studied as control group.2.We applied immunocytochemistry and Western blot to detect the protein expression of Notch1-IC,DII4 and Hes1 in peripheral blood mononuclear cells of patients with AL.We established real-time quantitative RT-PCR methods to detect the expression of VEGF,VEGFR1,VEGFR2,Notch1,DII4 and Hes1 in patients with AL,andβ-actin was detected as internal control.3.The central tendency was responsed by the median M and variables were assessed by the Wilcoxon rank-sum test and Spearman rank correlation.Results1.The expression of Notch1-IC,DII4 and Hes1 protein had been detected in both normal controls and initial patients with acute leukemia in peripheral blood mononuclear cells,the expression in initial group was significantly higher than those in control group.2.The expression of Notch1-IC,DII4 and Hes1 protein in CR group was still higher than those in control group with no statistical significance.3.The expression of VEGFR2,Notch1 and DII4 mRNA in initial group was significantly higher than those in control group,whereas the up-regulation of VEGF was not statistically significant.4.The expression of VEGFR2,Notch1 and DII4 mRNA decreased in CR group but was still higher than those in control group,but the difference was not statistically significant.5.In control group Notch1 exhibited a positive correlation with VEGF and a negative correlation with VEGFR2 significantly.In initial group Notch1 and DII4 both exhibited significantly positive correlations with VEGFR2.In CR group neither Notch1 nor DII4 is positively correlated with VEGFR2.6.Both in the control group,initial group or CR group,Notch1 or DII4 showed a significantly positive correlations with VEGFR1:Notch1 exhibited a positive correlation with VEGFR1 in control group,DII4 had a positive correlation with VEGFR1 in initial group,and in CR group Notch1 and DII4 are both positively correlated with VEGFR1.Conclusions1.Aberrant activation of Notch signaling may exist in AL and the expression lever of Notch signaling associated gene was related with the disease state, suggesting that Notch signaling pathway may play an important role in the development of acute leukemia.In CR group Notch signalings were still activated.2.Aberrant activation of VEGF signaling may exist in AL mainly depended on the up-regulation of VEGFR2.Aberrant activation of DII4/Notch signaling in AL depended on the up-regulation of Notch1 and DII4.3.Notch1 was up-regulated by VEGF and could inhibit VEGFR2,which modulated the angiogenesis in normal.The interaction and correlation of VEGFR2/VEGF and DII4/Notch signaling pathways all modulated the angiogenesis in bone marrow microenvironment of AL.VEGF and DII4/Notch signalings were still activated in CR group.4.DII4/Notch signaling might up-regulate VEGFR1.