【作者】 李岩；

【导师】 高丽；

【作者基本信息】 郑州大学 ， 儿科学， 2013， 硕士

【摘要】 背景与目的癫痫是常见的慢性神经系统疾病,约有近60%的患者于儿童时期起病,直接影响全球约5千万人的健康。虽然大多数癫痫患者对抗癫痫药物(antiepileptic drugs, AEDs)反应良好,但仍有近三分之一的患者治疗无效,属难治性癫痫(refractory epilepsy, RE)。尽管近年来有一些新的抗癫痫药物上市,但并未使难治性癫痫患者的发病率降低。反复的癫痫发作导致患者身体上的残疾和社会适应性降低,是一个严重的全球性的公共卫生问题。是什么原因导致难治性癫痫的产生呢?是否因机体对AEDs产生耐药、其机制又是什么呢?为此,医学界进行了不断的研究。目前研究发现,癫痫药物的耐药机制是一种很复杂的现象,可能涉及多种机制,但具体机制目前并不十分清楚。从国内外对抗肿瘤药物耐药机制的各种研究发现,多药耐药现象与多药耐药(multi-drug resistance, MDR)基因1及由其编码的P-糖蛋白(permeability glycoprotein, P-gp)有关。P-gp是典型的药物转运蛋白,可将进入大脑或神经元内的药物“泵”出,调控细胞内的药物含量。因此,由于各种原因,使P-gp在血脑屏障上过量表达,可能限制AEDs渗透入大脑,进而导致耐药的发生。2003年Siddiqui等首次发现MDR1C3435T位点的单核苷酸多态性(Single Nucleotide Polymorphism, SNP)与抗癫痫药物的耐药有关,认为SNP使P-gp的表达增加、功能增强,从而减少AED在脑内的分布水平自此研究首次提出抗癫痫药物的耐药是由基因决定的观点后,众多学者对此进行了大量研究，但结果差异较大，一些研究甚至得出相反的结论。考虑到以上研究结果的差异可能与种族地区差异有关,本文对MDR1基因C3435T、T129C两位点的多态性进行分析,探讨其与我国河南汉族儿童难治性癫痫之间的相关性及儿童难治性癫痫耐药机制形成的分子遗传学机制。方法收集2012年01月-08月于河南省人民医院就诊的癫痫患儿及健康体检患儿的外周血样,共260例,包括难治性癫痫患儿60例,疗效良好的癫痫患儿及健康儿童各100例。采用多聚合酶链反应扩增后继以限制性内切酶片段长度多态性分析方法,分别测定难治性癫痫组、疗效良好组及健康对照组的基因型分布和等位基因频率,应用SPSS17.0软件进行统计学分析,以P<0.05认为有统计学差异。结果C3435T位点：难治性癫痫组的基因型以TC为主,其分布频率与疗效良好组之间无相关性,但明显高于健康对照组,且差异有统计学意义(P=0.031),但二者等位基因间无统计学差异。T129C位点：疗效良好组与对照组基因型数目相同,三组均未发现CC基因型,难治组TC基因型分布频率、T等位基因频率高于疗效良好组及健康对照组,差异有统计学意义(P=0.029,0.034)。结论MDR1基因C3435T及T129C的单核苷酸多态性可能与河南汉族儿童难治性癫痫存在相关性,并参与了难治性癫痫耐药机制的形成。更多还原

【Abstract】 Backgrounds and ObjectiveEpilepsy is the most prevalent chronic neurological disorder, approximately60%of epileptics had their seizure initially from childhood. Although a majority of patients with epilepsy are responsive to the presently available antiepileptic drugs, nearly one-third of them continue to exhibit recurrent seizures, which is called refractory epilepsy. The introduction of several new AEDs in the recent years has not improved the outcome of these patients. Patients with drug-resistant epilepsies are physically and socially disabled, which has become a serious global public health problem.What are the causes of the forming to refractory epilepsy? is it because the body resistant to AEDs? Then, what is the mechanisms? To this end, the medical profession has made unremitting efforts.Recent studies showed that resistance to AED is considered as a complex phenomenon that may involve many mechanisms, none of which is well understood. Through the research of the mechanisms for resistance anticancer drugs, we found that the resistance is associated with the P-glycoprotein which is encoded by Multi-drug resistance gene1. P-gp is the archetypal example of multidrug transport proteins.Which can pump drugs that entered into the brain and neurons,and regulation of intracellular drug content. So, some rensons that made P-gp over expression on the blood-brain barrier may limit AEDs penetration into the brain,which leading to the occurrence of resistance. In2003, Siddiqui et al. firstly reported the C3435T polymorphism in the MDR1gene as being associated with resistance to AEDs,the Single Nucleotide Polymorphism cound increased the expression of P-gp and enhanced its function, thereby reduced the level of AED distributed in the brain. Since this study thought that drug resistance in epilepsy might be genetically determined for the first time, many scholars have conducted a lot of research of it. while, the result is quite different, Some studies have even come to the opposite conclusions.Taking into account the above findings may be related to racial differences in regional disparities, this study will analysis the relationship between two single nucleotide polymorphisms of multidrug resistance gene1and Henan Han children with refractory epilepsy, and detect their mechanisms of molecular genetics.MethodsDNA samples were obtained from Henan Provincial People’s Hospital, A total of260cases were enrolled, including60cases of refractory epilepsy,100cases of drug-responsive epilepsy and100cases of healthy children. The genotypes and allele frequencies for MDR1polymorphisms were determined by polymerase chain reaction-restriction fragment length polymorphism analysis. SPSS17.0software was applied for statistical treatment. P<0.05was considered statistically significant.ResultsC3435T:The genotype of refractory epilepsy group was given priority to with TC, the frequency distribution has no correlation between it and drug responsive group,but significantly higher than healthy control group, and the difference was statistically significant (P=0.031), but there was no statistical difference between the two alleles.T129C:The drug-responsive group and healthy control group have the same number of genotypes, the CC genotype was not found in all of the three groups.The distribution of TC genotype and T allele frequencies of refractory epilepsy group differed significantly from that in drug-responsive group and healthy control group (P=0.029,0.034).ConclusionC3435T and T129C polymorphisms of the MDR1gene were associated with refractory epilepsy in Chinese Henan Han children and may play some role in the cause of resistance mechanisms for refractory epilepsy.更多还原