The Mechanism Research Of Differentiation Inducer And Differentiation Inhibition Factor In Neural Tube Defects And Tumor Cells Growth

PART ONE RESEARCH ON THE EXPRESSION OF DIFFERENTIATION INDUCER AND DIFFERENTIATION INHIBITION FACTOR IN NEURAL TUBE DEFECTSSECTION ONE THE EXPRESSION OF DIFFERENTIATION INDUCER AND DIFFERENTIATION INHIBITION FACTOR IN MENINGOENCEPHALOCELEObjective: To research the relative expression level of differentiation resistance gene HA117 and differentiation inducer gene in meningoencephalocele and normal brain tissue.Methods: Collected 15 cases surgical removal organization of meningoencephalocele,5 cases of brain trauma resection tissue as normal controls.1.qRT-PCR was utilized to examine the change of mRNA level for HA117 、 DPF3 、 RGS6 、 STRA6 、 RAR α 、 CYP26-A1 between meningoencephalocele and normal brain tissue.2.Western blotting was utilized to examine the change of protein level for DPF3、RGS6、STRA6、RAR α 、 CYP26-A1 between meningoencephalocele and normal brain tissue.Results: 1.The relative expression levels of HA117 mRNA between normal andmeningoencephalocele groups has no difference(P > 0.05),the relative expression levels of DPF3、RGS6、STRA6、RXRα、Cyp26a1m RNA were significant increased in meningoencephalocele(P﹤0.01).2.The relative expression levels of DPF3、RGS6、STRA6、RXRα、Cyp26a1protein were significant increased in meningoencephalocele(P﹤0.01).Conclusion: The cause of the meningoencephalocele can be high expression of differentiation inducer gene DPF3、RGS6、STRA6、RXRα、Cyp26a1.SECTION TWO THE EXPRESSION OF DIFFERENTIATION INDUCER AND DIFFERENTIATION INHIBITION FACTOR IN NTDS MODEL OF C57 MOUSEObjective: To research the relative expression level of differentiation resistance gene HA117 and differentiation inducer gene in NTDs model of C57 mouse brain and normal mouse brain tissue.Methods: 1.High-dose ARTA lavage to stimulate pregnant mouse,and screen the fetal mouse with NTDs.2.q RT-PCR was utilized to examine the change of m RNA level for HA117、DPF3、RGS6、STRA6、RARα、CYP26-A1 between NTDs model of C57 mouse brain and normal mouse brain tissue.Results: 1.Successful build NTDs mouse model,compared with normal fetal mouse,the afterbrain of NTDs was abnormal,visible hemangioma appearance change,spine neural tube recently developed interruption,neural tube on the surface of the skin was weak,and the tail was dysplasia.2.The relative expression levels of HA117 m RNA in NTDs mouse model groups were significant increased(P ﹤ 0.01),the relative expression levels of STRA6、RXRα、Cyp26a1 m RNA were significant higher expression(P﹤0.01),and the relative expression levels of DPF3、RGS6 had no difference(P > 0.05).Conclusion: The mechanism of High-dose ARTA stimulating pregnant mouse lead to NTDs can be: ARTA stimulate the expression of non-coding RNA HA117 that inhibite the differentiation of Neural tube,stimulate STRA6,RARA,CYP26A1 gene expression that causes abnormal differentiation.PART TWO RESEARCH ON THE EXPRESSION OF DIFFERENTIATION INDUCER AND DIFFERENTIATION INHIBITION FACTOR IN GLIOBLASTOMA MULTIFORMESECTION ONE THE EXPRESSION OF DIFFERENTIATION INDUCER AND DIFFERENTIATION INHIBITION FACTOR IN GLIOBLASTOMA MULTIFORMEObjective: To research the relative expression level of differentiation resistance gene HA117 and differentiation inducer gene in GBM and normal brain tissue.Methods: Collected 25 cases surgical removal organization of GMB,5 cases of brain trauma resection tissue as normal controls.1.q RT-PCR was utilized to examine the change of m RNA level for HA117、DPF3、RGS6、STRA6、RARα、CYP26-A1 between GMB and normal brain tissue.2.Western blotting was utilized to examine the change of protein level for DPF3、RGS6、STRA6、RARα、CYP26-A1 between GMB and normal brain tissue.Results: 1.The relative expression levels of HA117 m RNA were significant increased in GMB(P﹤0.01),the relative expression levels of DPF3、STRA6、RXRα、Cyp26a1 m RNA were significant decreased in GMB(P ﹤ 0.01),and the relative expression level of RGS6 has no difference(P > 0.05).2.The relative expression levels of DPF3、RGS6、STRA6、RXRα、Cyp26a1 protein were significant decreased in GMB(P﹤0.05).Conclusion: The expression of differentiation inducer gene DPF3、RGS6、STRA6、RXRα、Cyp26a1 were increased and differentiation inhibition factor HA117 was inhabited in GMB.SECTION TWO THE MECHANISM OF TARGETED INTERFERENCE DIFFERENTIATION INHIBITION GENE HA117 REVERSE THE DRUG RESISTANCE OF U251 CELLSObjective: To probe the mechanism of targeted interference differentiation inhibition gene HA117 reverse the drug resistance of U251cellsMethods: 1.Establish HA117-RNAi U251 cell lines,verify the interference effect and detect the effect of virus on cell apoptosis;2.q RT-PCR was utilized to examine the change of m RNA level for HA117、DPF3、RGS6、STRA6、RARα、CYP26-A1 in normal U251、LV-vector U251、HA117-RNAi U251;3.MTT was used to examine the effect of different concentrations of TMZ and TMZ+0.5μm ARTA on three groups cells.4.Western blotting was utilized to examine the change of protein level for DPF3、RGS6、STRA6、RARα、CYP26-A1 in three groups cells.Results: 1.Successful build HA117-RNAi U251 cell lines,and the relative expression levels of HA117 m RNA was significant decreased in HA117-RNAi U251 cell lines(P﹤0.01),and FCM analysis the effect of virus on cell apoptosis infected by Lv-HA117 had no difference in three groups cells(P > 0.05);2.The relative expression levels of DPF3、STRA6Cyp26a1 m RNA were significant increased in HA117-RNAi U251 cell lines group(P﹤0.01),and the relative expression level of RGS6、RARαm RNA had no difference(P > 0.05).3.Drug testing showed that: there wno difference between normal U251 and LV-vector U251 groups,compared with U251 and LV-vector U251 groups the sensitivity of TMZ was increased three times on HA117-RNAi U251 cell lines group(P﹤0.01),after addition 0.5u M ARTA,the sensitivity of TMZ was increased four times on HA117-RNAi U251 cell lines group(P﹤0.01).4.The relative expression levels of DPF3、Cyp26a1 protein were significant increased in HA117-RNAi U251 cell lines group(P﹤0.01),and the relative expression level of RGS6、STRA6、RARαprotein had no difference(P > 0.05).Conclusion: 1.High expression of differentiation inhibition gene HA117 lead to the chemotherapy drug resistance in U251 cell lines,the mechanism was suppressed the expression of differentiation inducer gene DPF3 and Cyp26a1.2.Targeted interference differentiation inhibition gene HA117 can reverse the drug resistance of U251 cells.PART THREE THE MECHANISM OF ARTESUNATE IN TREATMENT OF INFANTILE HEMANGIOMA BY SUPPRESS THE EXPRESSION OF ANGIOGENESIS INDUCER GENE HIF-1Α-VEGF-AObjective: To determine the mechanism of inhibition of proliferation and invasion of mouse hemangioendothelioma endothelial cells(EOMA)in vitro and tumor growth in vivo by artesunate.Methods: The effects of artesunate on EOMA cell viability were determined by MTT assay;apoptosis was evaluated by Annexin V/Propidium Iodide(PI)staining and flow cytometric analysis,and cell invasion ability was determined using a transwell invasion assay.q RT-PCR and Western blotting were utilized to examine the change of Vascular Endothelial Growth Factor–A(VEGF-A),VEGFR-1,VEGFR-2,Hypoxia Inducible Factor-1 α(HIF-1α),caspase-3 after the effect of artesunate on EOMA cell.Inoculated EOMA cells were injected into the subcutaneous tissues of nude mice toobserve the effect of artesunate therapy on the vascular tumor,and compared with pingyangmycin.Results: Artesunate treatment(0?600 μg/mL)inhibited cell growth in a time-and dose?dependent manner.Artesunate at 300 μg/ml significantly reduced the proliferation and invasion of EOMA cells,and decreased the expression of VEGF-A,VEGFR-1,VEGFR-2,HIF-1 α on the level of m RNA and protein(p<0.05),and up-regulation the protein of caspase-3(p<0.05)in vitro.Artesunate significantly inhibited the growth of the tumor(p<0.05),and the curative effect was similar to that seen with pingyangmycin(p>0.05).Conclusion: Artesunate effectively inhibited HIF-1 α and VEGF-A expression in vascular tumors,and the effect was similar to that seen with pingyangmycin.