反向寡脱氧核苷酸抑制细胞周期素d1表达及对黑素瘤细胞增殖的影响

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1、反向寡脱氧核苷酸抑制细胞周期素D1表达及对黑素瘤细胞增殖的影响反向寡脱氧核苷酸抑制细胞周期素D1表达及对黑素瘤细胞增殖的影响中华皮肤科杂志1999年第4期第32卷论著作者：陆前进　苏先狮　夏家辉　魏启幼　张其亮　王振明祝和成　施家琦　黄柏英　赵云　夏昆单位：410011长沙，湖南医科大学附属第二医院(陆前进　苏先狮　魏启幼　张其亮　王振明　赵云)，中国医学遗传学国家重点实验室(夏家辉　施家琦　夏昆)，癌变原理卫生部重点实验室(祝和成　黄柏英)关键词：黑色素瘤核酸；反义细胞周期蛋白类【摘要】目的　探讨细胞周期素D1反向寡脱氧核苷酸对恶性黑素瘤的潜在治疗作用。方法　合成针对细胞周期素D1mRNA翻

2、译起始部位序列的20bp硫代磷酸反向寡脱氧核苷酸(AS-ODN)。采用RT-PCR检测AS-ODN对黑素瘤(B16细胞)细胞周期素D1mRNA水平的影响；用流式细胞仪检测细胞周期素D1蛋白表达及细胞周期分布；固绿染色计数、集落形成抑制试验体外观察AS-ODN对B16细胞增殖的影响。结果　细胞周期素D1mRNA反向链明显抑制B16细胞细胞周期素D1mRNA和细胞周期素D1蛋白表达，降低S期细胞百分比，且明显抑制B16细胞生长和集落形成。结论　细胞周期素D1反向寡脱氧核苷酸能有效抑制黑素瘤细胞增殖。AntisenseOligodeoxynucleotideInhibitstheExpression

3、ofCyclinD1andItsInfluenceonGrowthofMelanomaCellsLUQianjin,SUXianshi,XIAJiahui,etal.SecondAffiliatedHospital,HumanMedicalUniversity,Changsha410011【Abstract】ObjectiveToexplorethetherapeuticpotentialofantisenseoligodeoxynucleotidetocyclinD1formalignantmelanoma.MethodsThree20-mersphosphorothioateoligode

4、oxynucleotides(ODNs)weresynthesized:(1)CyclinD1antisenseODN(AS-ODN)targetingtheinitiationsiteoftranslationincyclinD1mRNA;(2)CyclinD1senseODN(S-ODN)homologoustotheinitiationsiteoftranslationincyclinD1mRNA;(3)RandomODN(R-ODN).ThelevelofcyclinD1mRNAwasobservedbyRT-PCR.ThelevelofcyclinD1proteinandthedis

5、tributionofcellcycleprogressionwereexaminedbyflowcytometry.TheinhibitoryeffectofODNforthegrowthofmelanoma(B16)cellswasobservedbysolidgreenstainingandcolonyforminginhibitorytest.ResultsAS-ODNinhibitedobviouslythelevelofcyclinD1mRNAexpressionwhileS-ODNandR-ODNdidnot.ThelevelofcyclinD1proteinwassignifi

6、cantlylowerinAS-ODNtreatedcellsthanthatincontrols(P＞0.05).AS-ODNinhibitedB16cellsenteringSphasefromG1phase,however,bothS-ODNandR-ODNhadnosucheffect.CellstreatedwithAS-ODN,S-ODN,R-ODNdisplayedacolonyforminginhibitoryrateof63.64％,18.19％,and31.82％,andacellgrowthinhibitoryrateof64.73％,26.51％,and29.64％,r

7、espectively.ConclusionCyclinD1antisenseoligodeoxynucleotidehasstronginhibitoryeffectonthegrowthofmelanomacells.网址：www.jxdyf.com第7页，共7页【Keywords】MelanomaOligodeoxynucleotide,antisenseCyclins近几年对细胞周期调控研