[Expression of Netrin-1 in Patients with Acute Lymphoblastic Leukemia and Its Clinical Significance]

Abstract

Objective: To investigate the correlation of the Netrin-1 expression level with the clinical characteristics in children with acute lymphoblastic leukemia (ALL) and to explore its possible regulatory mechanism.

Methods: ELISA was used to detect the expression level of Netrin-1 in peripheral blood serum from 48 child ALL patients (newly diagnosed, recurrent), and its relevance with clinical indicators was statistically analyzed. The blood serum samples from 27 children with non malignant hematological diseases were choosen as controls. Leukemia cell lines of Jurkat,Molt-4,SUP-B15 and Raji were cultivated in vitro, after treated with different concentrations of recombinant human Netrin-1 protein， the invasive ability of the cells was detected by Transwell method； the effect of Netrin-1 to the proli feration of cells was detected by CCK-8 method； The expression and phosphorylation level of key molecules, such as FAK,Erk1/2,PI3K and Akt signaling pathway were detected by Western blot.

Results: The expression of Netrin-1 in child patients was significantly higher than that of the control group (P<0.05). With the increasing of Netrin-1 level, the level of Plt (r=0.483, P<0.05) increased, while the level of WBC (r=-0.290, P<0.05) decreased, and there were no significant correlation with age, Hb level and the proportion of immature cells in bone marrow. When the concentration of Netrin-1 was 25-50 ng/ml, the level of Netrin-1 positively correlated with WBC (r=0.886, P<0.05) ; the level of Netrin-1 significantly decreased when the patient's WBC was ＞50×10⁹/L and Plt ＞20×10⁹/L(P=0.042，P=0.001); The expression level of Netrin-1 was significantly different in the risk group(P=0.017), and level of Netrin-1 in high-risk group was significantly higher than that in low risk group and middle risk group, but there was no significant difference of Netrin-1 expression in sex, hepatosplenomegaly, MRD, recurrence and chromosome abnormality. Netrin-1 could promote the invasiveness of the four kinds of cells (P<0.05). With the increase of Netrin-1 concentration, the number of cells increased at first and then decreased, and the number of cells in the invading chamber was the highest when the concentration of Netrin-1 was 100 ng/ml; the survival rate of the four kinds of cells significantly increased when the concentration of Netrin-1 was 25 ng/ml(P<0.05), and SUP-B15 cells showed the highest cell survival rate at a concentration of 100 ng/ml; The survival rate of the four kinds of cells showed a tendency : survival of cells increased at low concentration of Netrin-1 and survival of cells decreased at high concentration of Netrin-1. The results of Western blot showed that Netrin-1 activated the phosphorylation level of key molecules such as FAK,Erk1/2,PI3K,Akt signaling pathway (P<0.05).

Conclusion: There is abnormal expression of Netrin-1 in serum of children with ALL. Netrin-1 may affect the occurrence and development of ALL by increasing the proliferation and invasiveness of leukemia cells, and may become a risk factor of ALL or a potential target in biotherapy.