【作者】 劳万生；

【导师】 方美云；

【作者基本信息】 大连医科大学 ， 内科学， 2013， 博士

【摘要】 目的：探讨初治免疫性血小板减少症（ITP）患者巨噬细胞移动抑制因子（Macrophagemigration inhibitory factor,MIF）表达与糖皮质激素（Glucocorticoid，GC）抵抗的关系。方法：收集2009年2月—2012年11月大连医科大学附属第一医院，大连210医院血液科住院初治ITP患者90例（女性56例，男性34例），所有患者均采用GC治疗,根据GC疗效分为敏感组60例（女性36例，男性24例），抵抗组30例（女性20例，男性10例），通过实时荧光定量RT-PCR SYBR Green染料法检测敏感组和抵抗组患者外周血单个核细胞（Peripheral blood mononuclear cells,PBMC）MIF的mRNA表达，采用ELISA方法检测敏感组和抵抗组患者血浆中MIF的蛋白表达，比较组间差异。所有数据采用平均数±标准差表示，应用SPSS13.0统计软件分析试验结果。组间比较采用t检验，P＜0.05认为具有统计学意义。结果：1.激素抵抗组ITP患者外周血单个核细胞MIF的mRNA表达（28.71±5.97）高于激素敏感组ITP患者（27.19±6.30），但两组无统计学意义（P＞0.05）。激素抵抗组ITP患者血浆中MIF的蛋白表达（13.96±6.15）也高于激素敏感组ITP患者（12.82±5.54），两组相比无统计学意义（P＞0.05）。2.根据性别，年龄，病情，病程分组比较，激素抵抗组ITP患者外周血单个核细胞MIF的mRNA表达与激素敏感组相比较有升高趋势，但均无显著性差异（P＞0.05）。3.根据性别，年龄，病情，病程分组比较，激素抵抗组ITP患者外周血单个核细胞MIF的蛋白表达与激素敏感组相比较有升高趋势，但均无显著性差异（P＞0.05）。结论：与激素敏感组ITP患者相比，激素抵抗组ITP患者MIF基因mRNA水平和蛋白水平均有升高趋势，但无显著性差异，推测MIF的表达可能与ITP患者GC抵抗无密切相关。背景及目的：糖皮质激素（GC）是治疗免疫性血小板减少症（ITP）的主要药物。然而，经过3个月的标准治疗后，临床上少数患者对GC治疗反应很差或者无反应，称为GC抵抗。GC抵抗是临床治疗上的难题，而GC抵抗的分子机制仍不明确。FKBP5蛋白，作为HSP90的分子伴侣，具有调节GC受体敏感性的作用。研究发现FKBP5的基因多态性（rs1360780）在与应激有关的精神病中能调节激素的敏感性。本研究目的调查FKBP5基因多态性与ITP患者GC疗效的关系，探讨ITP患者GC抵抗的可能机制。方法:搜集212个ITP患者和110个健康志愿者，其中激素抵抗患者55名,激素敏感ITP患者157名。测定FKBP5单核甘酸多态位点rs1360780，应用实时荧光定量PCR和cycling探针技术进行基因分型，分析FKBP5基因多态位点等位基因及单倍体在ITP患者和健康对照组中的表达，探讨该基因与ITP患者激素疗效的关系。数据分析使用SPSS13.0软件，组间比较使用卡方检验。结果：1. FKBP5rs1360780位点各基因型在ITP患者组和对照组中的分布符合遗传平衡规律。2.激素敏感ITP患者的CC,CT,和TT的基因型频率分别是：53%,43%,4%, C和T等位基因频率分别是：75%,25%。；正常对照组的CC, CT,和TT的基因型频率分别是：59%,36%,5%, C和T等位基因频率分别是：77%,23%；二组间各基因型(P=0.40)及等位基因（P=0.62，OR=1.13,95%CI=0.76-1.70）相比较无显著性差异。3.激素抵抗ITP患者的CC, CT,和TT的基因型频率分别是：59%,39%,2%, C和T等位基因频率分别是：78%,22%；正常对照组相比，二组间各基因型(P=0.51)及等位基因频率(P=0.89，OR=0.92,95%CI=0.53-1.60)无显著性差异。4.与激素敏感ITP患者相比较，激素抵抗ITP患者的基因型频率(P=0.67)和等位基因频率(P=0.52，OR=1.23,95%CI=0.73-2.06)无显著性差异。结论：1. FKBP5rs1360780位点基因各基因型在ITP组和正常对照组中的分布符合哈迪-温伯格遗传平衡定律；2. FKBP5rs1360780位点多态性可能与ITP患者GC疗效无关联。背景及目的：糖皮质激素（GC）是治疗免疫性血小板减少症（ITP）的主要药物。然而，经过标准治疗后，临床上大约10-30%的患者对激素治疗反应很差或者无反应，称为糖皮质激素抵抗。研究发现FKBP51的过度表达和FKBP52的低表达能影响GC的免疫抑制作用。本实验目的是探讨初治免疫性血小板减少症（ITP）患者FKBP51和FKBP52的mRNA表达与糖皮质激素抵抗的关系。方法：收集于2009年2月—2012年11月大连医科大学附属第一医院，大连210医院血液科住院初治ITP患者90例（女性56例，男性34例），所有患者均首选GC治疗，根据GC疗效分为敏感组60例（女性36例，男性24例），抵抗组30例（女性20例，男性10例），通过实时荧光定量RT-PCR SYBR Green染料法检测敏感组和抵抗组患者外周血单个核细胞FKBP51和FKBP52的mRNA表达，比较组间差异。结果：1.激素抵抗组ITP患者外周血单个核细胞FKBP51的mRNA表达（26.20±5.18）与激素敏感组ITP患者（27.80±5.99）相比，无统计学意义（P＞0.05）。激素抵抗组ITP患者血浆中FKBP52的mRNA表达（17.35±4.96）与激素敏感组ITP患者（18.82±5.54）相比，无统计学意义（P＞0.05）。2.根据性别，年龄，病情，病程分组比较，激素抵抗组ITP患者外周血单个核细胞FKBP51的mRNA表达与激素敏感组相比较有升高趋势，但均无显著性差异（P＞0.05）。3.根据性别，年龄，病情，病程分组比较，激素抵抗组ITP患者外周血单个核细胞FKBP52的mRNA表达相与激素敏感组比较有降低趋势，但均无显著性差异（P＞0.05）。结论：1.与激素敏感组ITP患者相比，激素抵抗组ITP患者FKBP51的mRNA表达与激素敏感组相比较虽有升高趋势，但均无显著性差异，推测FKBP51的mRNA表达可能与ITP患者GC抵抗无关。2.与激素敏感组ITP患者相比，激素抵抗组ITP患者FKBP52的mRNA表达与激素敏感组相比较虽有降低趋势，但均无显著性差异，推测FKBP52的mRNA表达可能与ITP患者GC抵抗无关。更多还原

【Abstract】 Objective: To investigate the expression of macrophage migration inhibitoryfactor (MIF) in patients with primary immune thrombocytopenia(ITP) and itscorrelation with glucocorticoid resistance.Methods: The MIF mRNA expression was analyzed by semiquantitative real-timeRT-PCR in90newly diagnosed ITP patients. All the patients with ITP were dividedinto several subgroups according to glucocorticoid response, gengder, age, stages andseverity of desease, and difference between each pair of subgroups were analyzed. MIFprotein expression in serum was performed by ELISA.Results: We found no association of GC resistance and MIF mRNA and proteinexpression in ITP patients. According to the gengder, age, stages and severity ofdesease, the patients were further subdivided into8groups, no statistical difference wasfound.Conclusion: The present study suggested that the MIF expression does notcontribute to GC resistance in ITP. Objective: The FKBP5gene codes for the FK506-binding protein51(FKBP5), aco-chaperone of hsp90, which regulates glucocorticoid (GC) receptor sensitivity. TheFKBP5gene single nucleotide polymorphisms (SNP), rs1360780, has been found tomodulate GC sensitivity in stress-related psychiatric disorders. The aim of the presentstudy was to examine the effects of rs1360780on the treatment outcome of patientssuffering from idiopathic thrombocytopenic purpura (ITP) administered with GC.Methods: The polymorphism of FKBP5, rs1360780, was genotyped in55GC-resistant ITP patients,157GC-sensitive ITP patients and110unrelated healthyindividuals using real-time PCR and cycling probe technology with DNA extractedfrom peripheral blood.Results: No significant differences in FKBP5rs1360780genotypes (P=0.51) andalleles (P=0.89) were observed between the GC-resistant ITP patients and the healthycontrols. There were no significant differences observed between the GC-sensitive ITPpatients and the healthy controls (P=0.40for genotypes and P=0.62for T allele), as wellas between the GC-sensitive ITP patients and the GC-resistant patients (P=0.67forgenotypes and for T allele).Conclusion: The present study demonstrates that the FKBP5polymorphism maynot affect the response of ITP patients to GC treatment. Objective: Glucocorticoids (GCs) are considered the important drugs used intreatment of immune thrombocytopenia (ITP). However, about only10-30%patientswith ITP develop GC resistance after standard treatment with GC. Many studies hasbeen shown that the FKBP51overexpression and low expression of FKBP52affectedimmunosuppressive effects glucocorticoids. Here, we investigated the association of GCresistance with FKBP51and FKBP52mRNA expression in this case-control study.Methods:The FKBP51and FKBP52mRNA expression was analyzed bysemiquantitative real-time RT-PCR in90newly diagnosed ITP patients. All the patientswith ITP were divided into several subgroups according to glucocorticoid response,gengder, age, stages and severity of desease, and difference between each pair ofsubgroups were analyzed.Results: We found no association of GC resistance and FKBP51mRNAexpression in GC-resistant ITP patiences （26.20±5.18） compared with theGC-sensitive（27.80±5.99）. We found no association of GC resistance and FKBP52mRNA expression in GC-resistant ITP patiences （17.35±4.96）compared with theGC-sensitive（18.82±5.54）. According to gengder, age, stages and severity of desease,the patients were further subdivided into8groups, no statistical difference was found inGC-resistant ITP patiences compared with the GC-sensitive.Conclusion:The present study suggested that the FKBP51and FKBP52mRNAexpression does not contribute to GC resistance in ITP.更多还原