银屑病患者骨髓CFUHPP集落形成及集落细胞 p16基因启动子甲基化的研究.doc

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1、银屑病患者骨髓CFUHPP集落形成及集落细胞p16基因启动子甲基化的研究作者：张瑞丽,牛旭平,李新华,张开明,尹国华【摘要】本研究检测银屑病患者骨髓高增殖潜能集落形成单位（CFUHPP）的集落形成能力及集落细胞p16基因启动子甲基化状态，并探讨两者之间的关系。收集了24例银屑病患者及正常对照者骨髓，采用密度梯度离心法分离单个核细胞，并于含SCF/GMCSF/IL3/IL6细胞因子组合的甲基纤维素半固体培养液中，培养14天计数CFUHPP集落，然后收集集落。提取纯化集落细胞的DNA，经亚硫酸盐修饰后，采用甲基化特异PCR（

2、MSP）检测CFUHPP集落细胞的p16基因启动子甲基化状态。结果发现：在甲基纤维素半固体培养基中，银屑病患者骨髓CFUHPP集落数显著低于正常对照（t=5.91，p＜0.01），且集落形态较小；正常对照骨髓CFUHPP集落细胞的p16基因启动子甲基化阳性率较高（66.7%），而银屑病患者骨髓CFUHPP集落细胞p16基因启动子甲基化阳性率（37.5%）低于正常人。结论：银屑病患者骨髓CFUHPP细胞集落形成能力降低；银屑病患者骨髓CFUHPP集落细胞的p16基因甲基化降低可能与其相对较低的CFUHPP集落形成能力密切

3、相关。【关键词】高增殖潜能集落形成单位CFUHPPColonyFormationofBoneMarrowHematopoieticProginitorCellsinPsoriaticPatientsandMethylationofp16GenePromotorinCFUHPPColonyCellsAbstractThisstudywaspurposedtoinvestigatethecolonyformationofhighproliferativepotentialcolonyformingunits(CFUHPP)fr

4、ombonemarrowderivedhematopoieticcellsofpsoriaticpatientsandp16genepromotormethylationinCFUHPPcells，andtoexploretherelationshipbetweenthecolonyformationandthemethylationstatusofp16genepromoter.Bonemarrowderivedmononuclearcellsfrompsoriaticpatientsandnormalcontrolswer

5、eseparatedbydensitygradientcentrifugation,andwereculturedinmethycellulosesemisolidculturemediumwithSCF，GMCSF，IL3andIL6for14daystomeasurethecoloniesofCFUHPP.TheCFUHPPcolonycellswerecollectedandmethylationstatusofp16genepromoterofCFUHPPcellDNAmodifiedwithsodiumbis

6、ulfitewasdetectedbythemethylationspecificpolymerasechainreaction(MSP).Theresultsshowedthatinmethycellulosesemisolidculturesystem,thenumberandthesizeofCFUHPPcoloniesofbonemarrowofpsoriaticpatientswereallsignificantlylessthanthatofnormalcontrols,thepositivefrequencyof

7、p16genepromotermethylationinCFUHPPcellswaslowerthanthatinCFUHPPcolonycells7ofnormalcontrols.ItisconcludedthatthecolonyformationcapabilityofCFUHPPfrombonemarrowhematopoieticprogenitorcellsinpsoriaticpatientsislowerthanthatinnormalcontrols,andthelowerpositivefrequency

8、ofP16genepromotermethylationinCFUHPPcellsperhapscloselycorrelatedwithlowerCFUHPPcolonyformingcapability.Keywordsps