Anti-angiogenic effect of interferon on JAK2V617F positive myeloproliferative neoplasms and its anti-angiogenic mechanisms

Abstract

To research the suppressive effect of interferon α2b (IFN-α2b) on angiogenesis of JAK2 tyrosine kinase gene point mutation (JAK2V617F) positive myeloproliferative neoplasm (MPN) and its anti-angiogenic mechanisms. (1) A total of 42 cases of JAK2V617F positive MPN patients in the No.1 Hospital of Baoding were selected between January 2012 and October 2014, including the newly diagnosed group before treatment (n=25) and the IFN-α2b treatment group (n=17). Ten cases of idiopathic immune thrombocytopenic purpura (ITP) patients were enrolled as controls. JAK2V617F/JAK2 ratio was detected by real-time fluorescent quantitative PCR to measure mutation; the expression levels of phosphorylated JAK2 (p-JAK2), vascular endothelial growth factor (VEGF), hypoxia-inducible factor-1α (HIF-1α) and CD105 marked microvascular density (MVD) in bone marrow were detected by immunohistochemistry. The correlations were analyzed among JAK2V617F mutation burden and VEGF, HIF-1α, and MVD. (2) Human erythroleukemia cell line HEL cells were treated with different concentrations of IFN-α2b. The apoptosis was detected by Hoechst staining method. The cell viability was detected by CCK-8 test. Cell migration ability was tested by transwell chambers. The protein expression levels of p-JAK2, VEGF, and HIF-1α were detected by Western blot. (1)The ratio of JAK2V617F/JAK2 in the IFN-α2b treatment group (22.69% ± 12.64%) was significantly lower than that in the newly diagnosed group (46.17% ± 19.32%) (P<0.01). The expression levels of p-JAK2, VEGF, and HIF-1α in the newly diagnosed group (82.41% ± 11.65%, 64.72% ± 25.01%, 45.12% ± 20.28%) were significantly higher than those in the IFN-α2b treatment group (60.93% ± 20.57%, 36.58% ± 15.95%, 32.15% ± 14.27%) and the control group (43.05% ± 12.59%, 25.69%± 1 3.75%, 25.07% ± 15.49%) (all P<0.01). MVD in the newly diagnosed group (26.58% ± 5.93%) was significantly higher than that in the treatment group (15.86%± 4.27%) and the control group (10.76% ± 4.01%) (P<0.01). Spearman correlation analysis showed positive correlation of JAK2V617F mutation with VEGF and MVD (r=0.589, P<0.05; r=0.577, P<0.05). (2)The apoptosis of HEL cells were increased after treated with 30 000 U/L of IFN-α2b in HEL cells after 24 h. The proliferation of HEL cell were inhibited by different concentrations of IFN-α2b in time- and dose-dependent manner. The number of membrane-permeating HEL cells after treated with 5 000 U/L of IFN-α2b in HEL cells after 24 h was significantly lower than that in the untreated cells (52.9 ± 7.5 vs 77.3 ± 6.1) (P<0.05). The expression levels of p-JAK2, VEGF, and HIF-1α were decreased in a dose-dependent manner. IFN-α2b may exert an anti-angiogenic effect by inhibiting the VEGF, HIF-1α, and MVD expression in MPN via JAK2 signal pathway.