The Role And Mechanism Of Id1Invovled The Proliferation And Angiogenesis Of Colon Cancer

Objective: To explore the role and the possible mechanism ofId1(inhibitor of differentiation-1)invovled the proliferation andangiogenesis of colon cancer.Method:(1) The expressions of Id1mRNA and protein were detectedby fluorescence quantitative polymerase chain reaction(real-time PCR)and Western blot assay in HT-29cells under hypoxia0h、6h、12h、24h、36h environment in vitro respectively.(2) Two human colon cancerHT-29sub-cell lines of forced expression of Id1-siRNA and Id1wereestablished. After transfection with Id1-siRNA and Id1expression plasmid,HT-29cells were cultured with the analysis of real-time PCR and Westernblot assay to detect expressions of HIF-1alpha (hypoxia-induciblefactor-1alpha, HIF-1alpha) and VEGF (vascular endothelial growthfactor,VEGF) on mRNA and protein. As conditioned medium cultivationof HUVECs, each group culture supernatant were collected. Cellproliferation of HT-29cell lines and the HUVECs cultured withsupernatant collected were determined by MTT experiment. The abilities of tube formation of HUVECs in the conditoned medium ofId1-overexpression HT-29cells were observed by tuble formationexperiment. Tumorigenesis and MVD(microvessel density,MVD) in HT-29cells forced expression of Id1cell-transplanted tumor tissues in nude micewere observed. By SP immunohistochemical method, the expression of Id1was detected.(3)HT-29cells were divided into four groups:Id1-overexpression group, Id1-overexpression+LY294002(inhibitor ofPI3K/Art pathway) group, the empty carrier group and empty carrier+LY294002group, respectively. Expressions of both mRNA and protein ofHIF-1alpha and VEGF were examined by real-time PCR and Western blot.Results:(1) Hypoxia can promote the expressions of Id1on mRNAand protein in colon cancer HT-29cells.(2) Establish two human coloncancer HT-29sub-cell lines of forced expression of Id1-siRNA and Id1expression plasmid. Western blot identify to establish successfully.(3)Significantly increase expressions of HIF-1alpha and VEGF on protein andmRNA levels in HT-29cells were observed in Id1-overexpression HT-29cells and significantly decreased expressions of protein and mRNA levelsin HT-29cells Id1-siRNA transfectants were found.(4)The abilities ofproliferation of HUVECs in the conditoned medium ofId1-overexpression HT-29cells were significantly increased, and theabilities of proliferation of HUVECs in the conditoned mediumof Id1- siRNA colon cancer cells were significantly decreased.(5)The tubularstructure and the abilities of tubular formation of HUVECs in theconditoned medium of Id1-overexpression colon cancer cells weresignificantly increased.(6)Tumor formation in nude mice experimentshows that Id1-overexpression can promote the abilities of tmorigenesisand the structure of MVD in HT-29cells and the expression of Id1.(7)LY294002can significantly down-regulate the expressions of bothHIF-1alpha and VEGF on both protein and mRNA levels in HT-29cells.Conclusion:(1) Hypoxia can promote the expressions of Id1onmRNA and protein in colon cancer HT-29cells.(2) Id1-overexpression canup-regulate the expressions of HIF-1alpha and VEGF in HT-29cells andplay an important role in colon cancer proliferation and angiogenesis.(3)Through PI3K/Art signaling pathway, Id1can promote colon cancerproliferation and angiogenesis by regulate the expressions of HIF-1alphaand VEGF.