线粒体脑肌病患者的基因突变研究

目的 探讨线粒体脑肌病患者骨骼肌细胞线粒体DNA基因突变情况及发病机制。方法 观察总结5例线粒体脑肌病患者的临床表现,影像学变化特点,并应用PCR、限制性内切酶BglⅠ、ApaⅠ酶切,PAGE电泳鉴定DNA片段长度的方法,检测5例患者骨骼肌细胞中mtDNA是否发生nt3243和8344位点A→G突变。结果 5例患者（3例MELAS和2例MERRF）在临床表现和影像学改变等方面均与国外学者的研究结果相符。1例MELAS患者仅存在3243A→G点突变,1例MERRF患者存在8344A→G点突变,1例MERRF上述2个位点均存在突变。另2例呈家系起病的MELAS患者这2个位点都无突变。结论 3243及8344位点突变分别与MELAS和MERRF的发病有关,MERRF患者可以同时存在上述2个位点的突变。临床表现仍是确诊和分类的主要依据。Ⅰ     

线粒体DNA突变与线粒体脑肌病

本文简述了线粒体DNA的基本结构、功能及特性,并着重讨论线粒体DNA点突变与几种临床常见的线粒体疾病如Leber's病,MELAS,MERRF之间的联系及特异性线粒体DNA点突变的基因诊断意义。     

线粒体肌病和脑肌病患者骨骼肌细胞线粒体DNA缺失分析

目的为了检测线粒体肌病和脑肌病患者的骨骼肌细胞的线粒体ＤＮＡ的缺失情况。方法从６例原发性线粒体肌病和１例脑肌病患者的骨骼肌活检标本中，提取总ＤＮＡ，以线粒体ＤＮＡ全长为探针进行分子杂交。结果发现１例ＭＥＲＲＦ患者有５ｋｂ的线粒体ＤＮＡ基因缺失，另１例线粒体肌病患者有１５ｋｂ的线粒体ＤＮＡ基因缺失，剂量分析表明缺失型线粒体ＤＮＡ分别占总线粒体ＤＮＡ的１９．３％和１０．７％。结论线粒体ＤＮＡ基因缺失是线粒体疾病的重要病因之一     

浙江温岭帕金森病线粒体DNA基因突变与多态性研究

目的 研究温岭散发性帕金森病(PD)线粒体DNA(mtNDA)基因突变与国人散发性PD相关性.方法 对88例散发性PD患者和60例正常健康人的基因位点G1719A、G4580A、C7028T进行扩增,将其异常结果进行基因测序,确定基因突变发生位点.结果 在PD患者G1719A附近有4例1711(G→A)1738(A→G),3例1738(A→G),1例1664(G→A)突变;在G4580A附近有1例4476(A→G),1例4638(A→G),1例4651(→T)突变;在C7028T附近有1例6984(C→T),1例6979( G→C) 6984(C→T),3例7083(C→),4例6963(G→A),1例6910(C→A)突变,共发现11种突变类型,对照组无发现突变位点.结论 散发性PD患者存在线粒体基因位点突变,预示线粒体基因突变参与了PD的发病过程.     

线粒体基因G13513A突变导致的线粒体脑肌病六例临床表型分析

目的 报告6例mtDNA G13513A点突变引起的线粒体脑肌病患者的临床、影像学特点,总结mtDNA G13513A突变所致的线粒体病的临床表型.方法 对35例mtDNA常见突变(包括大片段缺失及A3243G、T3271C、A8344G、T8993G/C点突变)检查为阴性的线粒体脑肌病患者,用线粒体DNA全长测序和(或)聚合酶链反应-限制性片段长度多态法检测mtDNA G13513A点突变,分析阳性患者的临床特点,复习文献报道的mtDNA G13513A所致线粒体病的病例.结果 35例患者中有6例存在mtDNA G13513A突变.该6例患者均出现偏盲、轻偏瘫或偏身感觉障碍等卒中样发作表现,其中3例成人发病者以卒中样发作为主要症状,伴随癫痫、头痛、身材矮小、神经性耳聋等,头颅MRI显示以顶-枕-颢叶受累为主的大片病灶,符合成人型线粒体脑肌病伴高乳酸血症和卒中样发作(MELAS)的临床和影像学特点;3例青少年发病者除卒中样发作外,还有构音障碍、共济失调、眼外肌瘫痪等脑干受累的症状,MRI检查可见枕-颞叶大脑皮质非对称性病灶,以及双侧基底节和脑干的对称性病灶,符合青少年型MELAS-Leigh叠加综合征的临床和影像学特点.肌肉病理检查在5例患者发现不整红边纤维.经复习文献,发现mtDNA G13513A突变患者还存在婴幼儿型Leigh或Leigh样综合征表型.结论 mtDNA G13513A点突变是线粒体脑肌病较常见的致病性突变,主要导致Leigh综合征、MELAS-Leigh叠加综合征或MELAS综合征,其临床表型具有年龄依赖性.

Abstract：

Objective To report 6 Chinese patients with mitochondrial encephalomyopathy caused by mitochondrial DNA(mtDNA)G13513A mutation and discuss the mitochondrial phenotype associated with this mutation based on the data of our patient series as well as the reports by others.Methods Direct sequencing of polymerase chain reaction(PCR)products or PCR-RFLP analysis Was performed to screen mtDNA G13513A mutation in 35 cases with mitoehondrial encephalomyopathy.who carried no mtDNA common mutations(1arge 8eale deletion,A3243G,T3271 C,A8344G,or T8993G/C).The clinical features,MRI changes were retrospectively collected and analyzed.Published studies of all patients with mtDNA G13513A mutation were also reviewed.Results Six patients were identified carrying mtDNA G13513A mutation.All patients presented stroke-like episodes with hemianopsia.hemiparesis or hemiparesthesia.Three adult patients presented clinical and radiological features of adult-onset mitochondrial myopathy,encephalopathy,lactic acidosis,and stroke-like episodes(MELAS),including stroke-like episodes,epilepsy,headache,short stature,sensorineural deafness,multifocal lesions on parietal,occipital and temporal lobes on cranial MRI scans.Three iuvenile.onset patients presented the clinical and brain MRI features of MELAS-Leigh syndrome(LS)overlap syndrome.In addition to the stroke-like episodes,they also showed brain stem lesions with dysarthria,ataxia,and ophthalmopJegia. Brain MRI revealed asymmetrical lesions in the cortex of the oecipital and temporal lobes,as well as symmetrical lesions in the bilateral basal ganglia and brainstem.Muslce biopsy showed ragged redfibem in 5 patients.The infant-onset LS or Leigh-like syndrome with mtDNA G135 13A was described in the English literature.Conclusions mtDNA G13513A mutation is a common pathogenic mutmion for mitochondrial encephalomyopathy,which can result in Leigh syndrome,MELAS-LS overlap syndrome and adult MELAS.The onset of various phenotypes is relatively age-dependent.     

Leber遗传性视神经病的线粒体基因突变及临床特征

目的　了解Leber遗传性视神经病 (LHON)线粒体基因突变及其临床特征。方法　采用PCR法扩增96例视神经病变患者mtDNA中ND1、ND4、ND6上的 3个片段后 ,用 377测序仪对mtDNA上 11778、346 0、14 4 84位点进行序列分析确定有无碱基突变。结果　 96例患者中 4 0例 (41 7% )线粒体基因突变 ,其中男性 34例 (85 % ) ,女性 6例 (15 % ) ;34例 (85 % )为 11778位点突变 ,2例 (5 % ) 346 0位点突变 ,4例 (10 % ) 14 4 84位点突变 ;346 0及 14 4 84突变均为散发病例 ;有家族史者突变表达占 87 0 % (2 0 /2 3) ,无家族史者突变占 2 7 4 % (2 0 /73) ;头颅MR显示 11778突变者中 3例有脑白质区病灶 ,1例LHON合并垂体瘤 ;12例突变阳性者检查脑脊液 ,11例有炎性脱髓鞘改变。结论　LHON以 11778突变常见 ;起病及病程多样化、散发病例比例较高、眼底微血管病改变不易发现、脑脊液改变与炎性脱髓鞘雷同导致LHON在临床上易漏诊误诊 ,应尽早行突变检测明确诊断。     

线粒体脑肌病伴高乳酸血症和卒中样发作综合征与一种新的线粒体DNA基因突变

目的 检测1例线粒体脑肌病伴高乳酸血症和卒中样发作综合征(MELAS)患者脑组织和外周血线粒体DNA(mtDNA)的基因突变类型。方法 应用聚合酶链反应-限制性片段长度多态性(PCR—RFLP)方法对这例MELAS患者脑组织和外周血mtDNA的A3243G点突变进行检测，对检测过程中发现的异常扩增产物进行DNA测序分析。结果 该例MELAS患者脑组织和外周血白细胞PCR扩增产物行聚丙烯酰胺凝胶电泳时产生了两条带，一条为494bp，另一条为218bp。494bp的扩增产物是目的片段，而218bp的片段是一种异常扩增产物。我们对218bp的PCR扩增产物进行测序，发现在mtDNA3314—3589之间有276bp的碱基缺失。结论 mtDNA3314-3589位点之间276bp的碱基缺失可能是导致MELAs的一种新的基因突变类型。     

脑皮质畸形和线粒体病性癫痫的基因机制

症状性癫痫中 ,脑皮质畸形伴发癫痫和线粒体DNA突变致进展型肌阵挛是其重要的类型。神经元移行和分化发育异常是脑皮质畸形最主要的原因之一 ,多种人类及动物的基因突变与此有关 ,并发现了脑皮质畸形引起癫痫可能机制的相关证据 ;多个位点的线粒体DNA突变引起症状性癫痫的病理生理机制还不完全清楚 ,但线粒体内Ca2 + 内环境的改变导致呼吸链功能障碍及ATP合成的降低是其可能的原因     

全基因组检测线粒体脑肌病的基因突变研究

目的探讨全基因组检测线粒体脑肌病(ME)基因突变的临床意义。方法分析8例ME的临床特征、24h视频脑电图(VEEG)、肌电图(EMG)、头颅MRI、全基因组检测基因突变。结果 8例全基因组检测基因突变表明,存在核基因突变8例、有氨基酸改变7例、线粒体基因突变8例;其中t RNA基因突变5例、TRNL1基因突变4例、ATP6基因突变3例、ND5和TRNS2基因突变各1例。核酸3243AG改变4例(50%),其他有8860AG,11719GA,14766CT,8993TG等4例(50%),氨基酸改变4例。结论ME患者大多存在核基因的突变,线粒体的5个mt DNA均可发生突变。本组患者核酸改变仅50%发生在3243位点,检测核基因和线粒体基因是诊断ME的依据之一。     

神经干细胞与脑胶质瘤干细胞☆

所有的肿瘤组织并不是由均一的肿瘤细胞所组成的,不同的细胞具有不同的增殖、浸润和转移能力,亦即肿瘤的异质性。其中存在少数担当着干细胞角色的肿瘤细胞,具有干细胞的基本特性,包括自我更新能力、无限的增殖能力和多向分化潜能,为肿瘤干细胞。神经干细胞具有很强的自我更新机制,获得较少突变即有可能恶性转化,而且干细胞存活时间较长,这意味着干细胞比成熟细胞发生细胞复制的错误几率更大,因外界环境的刺激而发生突变的机会更多,最终形成脑胶质瘤干细胞,同时调节神经干细胞增殖和自我更新的基因在脑胶质瘤的脑胶质瘤干细胞中也表达,这也是支持神经干细胞是脑胶质瘤干细胞来源的;也有推测认为它可能起源于已分化的细胞,由这些细胞突变发生去分化得来,并通过基因突变而获得了干细胞自我更新的特性,从而形成脑胶质瘤干细胞。通过探讨神经干细胞与脑胶质瘤干细胞,为脑胶质瘤的治疗提供依据。     

胶质瘤IDH1基因突变与胶质瘤干细胞相关基因表达的相关性研究

目的 研究原发性、原位复发性胶质瘤中异柠檬酸脱氢酶1(IDHI)基因突变情况,分析胶质瘤干细胞相关基因的变化情况与IDH1基因突变的相关性. 方法 选取新疆自治区人民医院神经外科自2010年8月至2011年8月间手术切除并经病理证实的人脑胶质瘤标本43例,其中原发30例,复发13例,应用PCR检测标本中IDH1基因多态性,流式细胞术检测肿瘤组织中CD133、nestin的表达. 结果 基因测序显示原发胶质瘤中17例(56.67％)发生IDH1突变,复发胶质瘤中13例(53.85％)发生IDHl突变,突变类型均为R132H型.流式细胞仪检测显示原发、复发胶质瘤标本中nestin、CD133的阳性表达率与肿瘤的病理分级均成正相关关系(P＜0.05).伴IDHl突变的胶质瘤标本nestin、CD133阳性表达率较IDH1野生型胶质瘤增多,差异有统计学意义(P＜0.05).与原发肿瘤比较,复发肿瘤中nestin、CD133阳性细胞表达率明显增高,差异有统计学意义(P＜0.05). 结论 IDH1突变在原发、复发性胶质瘤中发生频率均很高,可上调干细胞相关基因Nestin和CD133的表达.复发肿瘤中升高的干细胞相关基因与肿瘤发生及复发密切相关.     

神经胶质瘤中缓激肽B2受体的表达水平与病理级别的相关性

目的　分析神经胶质瘤中缓激肽B2受体表达水平与病理分级的相关关系,为缓激肽及其类似物的临床应用提供实验依据。方法　采用神经胶质瘤患者术后标本,用H&E染色进行神经胶质瘤的病理诊断及分级,用免疫组化法和WesternBlot法测定不同病理分级的神经胶质瘤中缓激肽B2受体的表达水平。结果　H&E染色显示26例神经胶质瘤中I级为9例,II级为9例,III级为8例,IV级为0例。胶质瘤边缘水肿带不表达缓激肽B2受体,B2受体位于胶质瘤细胞。WesternBlot结果显示在三组不同病理分级的神经胶质瘤中,B2受体表达水平为Ⅰ级与Ⅱ级( 39480. 88±5119. 96对51354. 25±6168. 77, n=8, t=2. 447, P<0. 05 ),Ⅰ级与Ⅲ级( 39480. 88±5119. 96对73032. 13±8802. 71, n=8, t=7. 710, P<0. 001 ),Ⅱ级与Ⅲ级( 51354. 25±6168. 77对73032. 13±8802. 71, n=8, t=5. 704, P<0. 001);神经胶质瘤中缓激肽B2受体表达水平与其病理分级I级~III级呈显著正相关(r=0. 895,P<0. 001),呈I级相似文献   

胶质瘤组织UCH37表达水平与病人预后的相关性

目的 探讨胶质瘤组织泛素化羧基末端水解酶37（UCH37）表达水平与病人预后的相关性。方法 选取2017年6月至2019年6月手术切除的脑胶质瘤组织104例和瘤旁脑组织64例,采用免疫组化法检测组织UCH37表达水平,根据免疫组化染色评分分成高表达组、低表达组;采用PCR法检测组织UCH37 mRNA水平。术后随访18个月,记录死亡、生存情况。结果 104例中,死亡20例,存活84例;UCH37高表达62例,低表达42例。脑胶质瘤组织UCH37高表达率（59.62%）明显高于瘤旁组织（17.19%;P<0.05）,而且脑胶质瘤组织UCH37 mRNA表达量明显高于瘤旁组织（P<0.05）。多因素logistic回归分析显示UCH37高表达是脑胶质瘤术后死亡的独立危险因素（P<0.05）。UCH37高表达组中位生存期（9个月）较低表达组（13个月）明显缩短（P<0.05）。ROC曲线分析显示,UCH37 mRNA评估病人术后死亡的最佳界值为2.725,曲线下面积为0.797（95%置信区间0.695~0.899）,敏感度为70.00%,特异度为70.10%。结论 脑胶质瘤UCH37呈高表达,表达水平越高,预后越差。肿瘤组织UCH37 mRNA水平检测对病人预后有一定评估价值。     

lncRNA-FTX与脑胶质瘤病人预后的相关性分析

目的 探讨长链非编码RNA（lncRNA）FTX表达水平与脑胶质瘤病人预后的相关性。方法 前瞻性收集2016年1月~2018年1月手术切除的脑胶质瘤88例（46例获得瘤旁组织），采用实时逆转录PCR检测lncRNA FTX表达水平，以脑胶质瘤组织lncRNA FTX表达水平均值为界分为高表达组（n=57）和低表达组（n=31）。术后随访3年，主要观察指标为无进展生存期、总生存期。结果 脑胶质瘤组织lncRNA FTX表达水平[（7.54±2.15）]明显高于瘤旁组织[（2.65±0.69）；P<0.001]。多因素Cox比例回归风险模型分析显示，lncRNA FTX高表达是脑胶质瘤生存预后不良的独立危险因素（RR=1.589；95% CI 1.004~2.515；P=0.048）。生存曲线分析显示，高表达组无进展生存期（15.10个月）和总生存期（20.24个月）较低表达组（分别为18.96和25.53个月）明显缩短（P<0.05）。结论 脑胶质瘤lncRNA FTX呈高表达，与病人不良生存预后有关。     

X-linked NDUFA1 gene mutations associated with mitochondrial encephalomyopathy

OBJECTIVE: Mitochondrial complex I deficiency is the commonest diagnosed respiratory chain defect, being genetically heterogeneous. The male preponderance of previous patient cohorts suggested an X-linked underlying genetic defect. We investigated mutations in the X-chromosomal complex I structural genes, NDUFA1 and NDUFB11, as a novel cause of mitochondrial encephalomyopathy. METHODS: We sequenced 12 nuclear genes and the mitochondrial DNA-encoded complex I genes in 26 patients with respiratory chain complex I defect. Novel mutations were confirmed by polymerase chain reaction restriction length polymorphism. Assembly/stability studies in fibroblasts were performed using two-dimensional blue native gel electrophoresis. RESULTS: Two novel p.Gly8Arg and p.Arg37Ser hemizygous mutations in NDUFA1 were identified in two unrelated male patients presenting with Leigh's syndrome and with myoclonic epilepsy and developmental delay, respectively. Two-dimensional blue native gel electrophoresis showed decreased levels of intact complex I with no accumulation of lower molecular weight subcomplexes, indicating that assembly, stability, or both are compromised. INTERPRETATION: Mutations in the X-linked NDUFA1 gene result in complex I defect and encephalomyopathy. Assembly/stability analysis might give an explanation for the different clinical phenotypes and become useful for future diagnostic purposes.     

TRIM38非CpG岛DNA甲基化与胶质瘤异柠檬酸脱氢酶突变的关系研究

目的 探讨TRIM38基因非CpG岛DNA甲基化与胶质瘤异柠檬酸脱氢酶（IDH）基因突变之间的关系。方法 利用中国胶质瘤基因组图谱计划（CGGA）数据库的多组学数据和临床资料，比较在IDH野生型或突变型的胶质瘤中，TRIM38非CpG岛DNA甲基化的改变模式以及与基因表达和临床预后的关系。结果 共纳入CGGA胶质瘤325例及非肿瘤对照脑组织（NTB组）11例，分析发现IDH野生型胶质瘤TRIM38非CpG岛DNA甲基化和基因表达，相对NTB组分别发生低甲基化（P =0.000）和高表达（P=0.007），且两者之间呈负相关（P=0.017）。生存分析显示，TRIM38非CpG岛DNA甲基化水平与IDH野生型肿瘤的预后有关（P=0.061）。结论 IDH突变可能通过限制TRIM38基因非CpG岛DNA低甲基化介导的肿瘤促癌基因表达上调，为IDH突变相关的胶质瘤提供“保护作用”。     

Correlation between magnetic resonance imaging and histopathology of intracranial glioma.

Postmortem histopathology of eight gliomas was studied in correlation with magnetic resonance imaging (MRI) and computed tomography (CT) findings. MRI demonstrated the lesions more clearly and widely than CT. Also, T2-weighted images (T2WI) had a greater ability to depict the lesion than T1-weighted images (T1WI). The areas in which neoplastic cells had invaded corresponded to the high intensity areas on T2WI in four cases of glioblastoma multiforme. In the case of a grade II astrocytoma, neoplastic cells were scattered beyond the region corresponding to the high intensity area on T2WI. In the case of a grade III astrocytoma, neoplastic cells did not come up to the line corresponding to the margin of the high intensity area on T2WI. In the remaining two cases, although the high intensity areas on T2WI were depicted as being larger than the areas in which neoplastic cells were seen histopathologically, the high intensity regions corresponding to the outside zones of the tumour-infiltrated area were thought to be a radiation necrosis in one case and a 'periventricular high intensity' in the other. The high cellularity of the glioma was seen mainly as a low intensity area on T1WI and as an isointensity or a slightly high intensity area on T2WI. However, the signal intensities of glioma on MRI, reflecting T1 or T2 values of the tumour tissues, did not correlate with the malignancy of the tumour.     

New mutations in TK2 gene associated with mitochondrial DNA depletion

Mitochondrial deoxyribonucleic acid depletion syndromes are autosomal recessive disorders characterized by a reduction of the amount of mitochondrial deoxyribonucleic acid, which impairs the synthesis of respiratory chain complexes. Mutations in the deoxyguanosine kinase and polymerase gamma genes have been identified in hepatocerebral forms, whereas thymidine kinase 2 gene mutations have been found in patients with isolated myopathy, encephalomyopathy, or spinal muscular atrophy. Mutations in the gene encoding the beta subunit of the adenosine diphosphate-forming succinyl-coenzyme A synthetase have also been reported in a family. In this report, the clinical, molecular, morphologic, and biochemical features of five children from two independent families with an infantile encephalomyopathy are characterized. The affected children manifested muscle mitochondrial deoxyribonucleic acid depletion and three novel thymidine kinase 2 gene mutations. They consist of a homozygous substitution resulting in Ala to Val change at the highly conserved position 181 of thymidine kinase in the first family, and two heterozygous substitutions in the second family: a Cys to Trp change at residue 108 and a Leu to Pro change at residue 257 of the enzyme. Common clinical features associated with these TK2 mutations are a normal early developmental phase followed by psychomotor regression, encephalopathy often with epileptic seizures, and myopathy with features of a progressive dystrophic process.     

NAD(P)H氧化酶与胶质瘤恶性程度相关性研究

目的 检测不同恶性程度胶质瘤组织NAD(P)H氧化酶(NOX)的表达及其产物ROS的含量,探讨NOX及细胞内ROS与胶质瘤增殖及恶性程度的关系.方法 收集南方医科大学珠江医院神经外科白2007年8月至2010年8月手术切除的人脑胶质瘤标本30例,其中低级别胶质瘤(Ⅰ和Ⅱ级)10例、Ⅲ级胶质瘤10例和Ⅳ级胶质瘤10例,另取10例行内减压术的脑损伤患者的正常脑组织作为对照组.RT-PCR检测4组脑组织NOX1～5 mRNA的表达,流式细胞仪测定脑组织ROS的含量,Western blotting和免疫荧光染色分别检测脑组织NOX蛋白的表达和分布.结果 4组脑标本NOX1～5 mRNA、ROS含量均不同,差异均有统计学意义(P＜0.05),且NOX4mRNA 、ROS含量在正常对照组、低级别胶质瘤(Ⅰ～Ⅱ级)、Ⅲ级胶质瘤和Ⅳ级胶质瘤中的表达逐渐增加,差异均有统计学意义(P＜0.05);Westenm blotting和免疫荧光染色结果显示NOX在对照组标本中呈低表达,在胶质瘤中呈高表达,且胶质瘤恶性程度越高,表达水平越高.结论 NOX的表达及ROS含量与胶质瘤的恶性程度有关,可能对胶质瘤的发生和恶性增殖起重要作用.