g四链体抑制结肠癌细胞增殖和血管生成的研究

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1、山西医科大学硕士学位论文外血管生成。结论：结肠癌细胞HCT116和SW480成功摄取寡核苷酸PU22序列后可以抑制VEGF基因的表达，诱导细胞凋亡，抑制细胞增殖。其细胞分泌物也可抑制HUVEC体外血管生成能力。其作用机制可能与基因启动子区域G四链体的形成有关，本研究为结肠癌的治疗提供新的思路和方法。关键词：VEGF；G四链体；PU22；血管生成III山西医科大学硕士学位论文ResearchG-quadruplexinhibitcoloncancercellproliferationandangi

2、ogenesisAbstractObjective：WedesignandSynthesisofPU22fragments.weinvestigatedthatPU22influenceontheexpressionofVEGF,cellproliferation,cellapoptosisandangiogenesiscase,invitro.Methods：(1)WearedesignedandsynthesizedoligonucleotidefragmentsPU22bybioinfor

3、matics,whichanalysistheVEGFgenepromoterregion.MUTPU22sequenceformedbydisruptingPU22sequenceasacontrolsequence.Theycontrastwiththegenedatabaseinordertoensurethatspecific.(2)AftertheoligonucleotidePU22bringFITCfluorescencelabelingjoinHCT116cellsandSW48

4、0cells,weanalysisofcoloncancercelluptakeofoligonucleotidesPU22case.(3)ThetotalRNAextractedcoloncancerHCT116andSW480cellswhichweretreatedwithdifferentconcentrationsPU22orMUTPU22.ByReal-timePCRtechnologywedetectmRNAexpressionofVEGFgeneinordertoidentify

5、theoptimalconcentration.(4)Follow-upoftheexperimentweredividedintothreegroups:PU22treatedexperimentalgroup,MUTPU22treatedcontrolgroupanduntreatedcontrolgroup.(5)Thetotalproteinofcoloncancercellswasextracted.DetectVEGFgeneproteinwhichbyWesternblotting

6、technique.(6)AnalysisofapoptosisincoloncellsusingAnnexinV-FITCmethodbyflowcytometry.DetectcasesofcoloncancercellproliferationbyMTTassay.(7)ResearchPU22impactonangiogenesisbyhumanumbilicalveinendothelialcells(HUVEC)tubeformationinvitro.Results：IV山西医科大

7、学硕士学位论文(1)BybioinformaticsdatabasesuccessfullyscreenedanddesignoutPU22actingsequences,whichpossibleroleofthepromoterregionofVEGFandMUTPU22controlsequences.Theycontrastwiththegenedatabaseinordertoensurethatspecific.(2)WehavedetectedthatPU22successfull

8、yuptakeintocancercells,andtheuptakerategreaterthan90%,byfluorescentstaining.(3)qRT-PCRresultsshowedthathumancoloncancercelllineHCT116andSW480whicheffectwithdifferentconcentrationsPU22vegfgeneRNAexpressionlevelsarereduced.Amongthem,themostobviouseffec