Exclusive homoplasmic 11778 mutation in mitochondrial DNA of Chinese patients with Leber's hereditary optic neuropathy.

PURPOSE: To investigate the degree of heteroplasmy of the 11778 mtDNA mutation in Chinese patients with Leber's hereditary optic neuropathy (LHON). METHODS: Seventeen Chinese Leber's pedigrees, including 24 patients, 17 unaffected maternal lineages, 4 internal controls, and 6 unrelated controls, were screened for the 11778 mtDNA mutation. This was carried out by analysis of the restriction fragment length polymorphism, single-strand conformation polymorphism, and DNA sequencing. RESULTS: All patients and unaffected maternal lineages, regardless of their symptoms, had homoplastic 11778 mtDNA mutation, which was revealed by restriction fragment length polymorphism analysis and single-strand conformation polymorphism analysis. CONCLUSION: Exclusive homoplasmy of the 11778 mtDNA mutation in Chinese LHON patients was found in this study. Homoplasmy of the 11778 mtDNA mutation cannot account for the variation in the clinical phenotype of Chinese Leber's patients.     

Leber遗传性视神经病变11778突变家系异质性的研究

目的研究Leber遗传性视神经病变（LHON）11778点突变家系的异质性,分析异质性对LHON的发病率和患者视力的影响。方法研究5个11778点突变LHON家系,其中母系成员68例,发病者27例;除2例携带者外,66例母系成员接受检查。应用聚合酶链式反应、限制性核酸内切酶酶切、变性高效液相色谱分析等技术,分析家系之间突变的异质性以及异质性与发病和视力预后的关系。结果在5个家系中,2个家系为同质突变（纯合突变）,包括24例母系成员和10例患者;3个家系为异质突变（杂合突变）,包括42例母系成员和17例患者。同质突变和异质突变母系成员之间LHON的发病率比较,差异无统计学意义;2种突变患者之间的视力比较,差异亦无统计学意义。结论LHON患者11778的突变存在异质性,但其异质性对发病和视力预后均无影响。     

Visual recovery in patients with Leber's hereditary optic neuropathy and the 11778 mutation.

Five patients with Leber's hereditary optic neuropathy (LHON) and the 11778 mitochondrial mutation spontaneously recovered 20/40 or better visual acuity in at least one eye after months to years of legal blindness. The patients ranged in age from 9 to 45 years, and the duration of visual loss before recovery ranged from several months to 5.9 years. These patients constitute only about 4% of the 136 affected LHON patients we have studied who also had the 11778 mutation in their mitochondrial DNA. Thus, even though the visual prognosis for most patients with LHON and the 11778 mutation is poor, a few individuals do recover near-normal vision in at least one eye even years after the initial visual loss.     

Phylogenetic assessment of the mitochondrial DNA displacement loop haplotype in Japanese patients with Leber's hereditary optic neuropathy harboring the mitochondrial DNA G11778A mutation

To investigate the anthropological background and the association of mitochondrial DNA (mtDNA) haplotype with the disease phenotype, the nucleotide sequence in the hypervariable segment of the displacement loop (D-loop) region of mtDNA was determined in Japanese patients with Leber's hereditary optic neuropathy (LHON) harboring the G11778A mutation. Genetic polymorphism of mtDNA was examined in 36 unrelated Japanese LHON patients who presented with bilateral optic nerve disease and had the mtDNA G11778A mutation. DNA was extracted from the peripheral blood after having obtained informed consent. The nucleotide sequence of the D-loop region (np 16,002-16,490) was directly determined. The intergenic deletion of the COII/tRNA(Lys) gene of mtDNA was also examined. From the data set of nucleotide alignments, the phylogeny of the mtDNA sequence and phenotypic diversity within the examined population were evaluated. One-base polymorphism was present at 37 different sites. The estimated value of nucleotide diversity was 0.69%. D-loop sequences were classified into 13 monophyletic clusters (CA to CM). There was not any definite ancestral haplotype of the D-loop sequence in the examined LHON population. Thus, the mutational event of G11778A appears to be independent of the evolutionary course in the D-loop haplotype. Patients with a CD plus CH cluster had a significantly older age at onset (p = 0.006), and had a family history being significantly lower as compared with patients with other clusters (p = 0.05). The mtDNA D-loop haplotype characterized by the presence of T16362C or C16290T, lacking G16129A and G16390A, may be a risk for older age at onset and other unusual clinical features in Japanese LHON patients with the G11778A mutation.     

Leber's hereditary optic neuropathy mitochondrial DNA mutations at nucleotides 11778 and 3460 in multiple sclerosis

OBJECTIVES: Leber's hereditary optic neuropathy (LHON) can be difficult to distinguish from optic neuritis seen in multiple sclerosis (MS). About half of the LHON patients harbor a mutation at nucleotide (nt) 11778 in the mitochondrial (mt) DNA. In addition, mutations at nt-3460 and nt-14484 have been associated with LHON. An association of LHON and MS has been suspected for decades, and, recently, the LHON nt-11778 and nt-3460 mtDNA mutations have been found in several patients with MS or MS-like disease. We attempted to determine which MS patients should be evaluated further for LHON mutations. METHODS: We screened 103 clinically definite MS patients (age range from 18 to 72 years, 27 men and 76 women) for the LHON nt-11778 and nt-3460 mtDNA mutations. RESULTS: Neither mutation was identified in the patients. CONCLUSIONS: Our findings confirm previous reports which found that both LHON mutations are rare in unselected MS patients. The reports to date suggest that MS patients with peripapillary teleangiectasia typical of LHON, with relatives harboring LHON or with early severe bilateral optic neuropathy, particularly if female, should be further evaluated for LHON mutations.     

G11778A突变型Leber遗传性视神经病变患者血清总超氧化物歧化酶活力和丙二醛含量的研究

目的 检测G11778A突变型Leber遗传性视神经病变(LHON)患者血清总超氧化物歧化酶(SOD)活力和丙二醛(MDA)含量并了解LHON患者体内氧化与抗氧化状态.方法 病例对照研究.纳入经眼科检查和线粒体基因测序确诊的G11778A基因突变型LHON患者19例,G11778A基因突变携带者(携带者)12例,正常对照者30例,分别应用黄嘌呤氧化酶法和硫代巴比妥酸法检测受试者血清总SOD活力和MDA含量.应用SPSS 12.0统计学软件包.采用单因素三水平设计定量资料方差分析,分别计算LHON患者、携带者及正常对照者血清总SOD活力和MDA含量的均数及标准差,比较3组受试者间各检测指标的均数差异;若有差异则进行多个均数两两比较的q检验.结果 LHON患者、携带者和正常对照者的血清总SOD活力分别为(22.01±7.21)×103 U/L、(43.44±14.70)×103U/L及(42.70 ±12.51)×103U/L,差异有统计学意义(F=20.875,P＜0.05).LHON患者血清总SOD活力较携带者和正常对照者均明显下降(q值分别为7.085,8.351;均P＜0.05),携带者与正常对照者的血清总SOD活力差异无统计学意义(q=0.269,P＞0.05).LHON患者、携带者及正常对照者的血清MDA含量分别为(10.39±4.38)mmol/L、(7.08±2.51)mmol/L及(2.97±1.81)mmol/L,差异有统计学意义(F=20.109,P＜0.05).LHON患者和携带者血清MDA含量均明显高于正常对照者(q值分别为9.069,4.748;均P＜0.05),LHON患者的血清MDA含量也高于携带者(q=3.618,P＜0.05).结论 LHON患者体内抗氧化能力明显下降,机体氧化与抗氧化系统失衡与LHON患者的发病有关.     

The clinical characteristics of pedigrees of Leber's hereditary optic neuropathy with the 11778 mutation

In a study of the phenotypic characteristics of pedigrees of Leber's hereditary optic neuropathy positive for the mitochondrial DNA mutation at position 11778, 28 of 49 pedigrees were represented by singleton cases. Seven families, including six singleton pedigrees, had maternal family members with a mixture of mutant and normal mitochondrial DNA (heteroplasmy). Seventy-two affected individuals from 43 families showed a male predominance of 81.9% (59/72) and ages of onset of visual loss ranging from 8 to 60 years. The time interval between affected eyes averaged 1.8 months; the duration of progression of visual loss in each eye averaged 3.7 months. Visual acuity was 20/200 or worse in 107 of 109 (98.2%) eyes. Telangiectatic microangiopathy, disk pseudoedema, or vascular tortuosity, ophthalmoscopic features believed to be classic of Leber's hereditary optic neuropathy, were noted in 30 of 52 patients. Visual-evoked responses were typically absent or abnormal. Electrocardiograms, fluorescein angiograms, cerebrospinal fluid analyses, brain computed tomography, and magnetic resonance imaging were usually normal. There were no consistent neurologic or systemic illnesses associated with these Leber's pedigrees. In many cases, the diagnosis would not have been suspected because of the absence of a compatible family history, typical clinical profile, or ophthalmoscopic appearance. Genetic analysis showed the mitochondrial DNA mutation at position 11778, which established the diagnosis of Leber's hereditary optic neuropathy and has allowed for a broader view of the clinical features of this disease.     

遗传性视神经病线粒体DNA原发性位点突变的研究

目的 探讨线粒体DNA(mtDNA)11778、3460、14484位点突变与遗传性视神经病(Leber′s hereditary optic neuropathy,Leber病)之间的关系。 方法 应用多聚酶链式反-单链构象多态性分析(PCR-SSCP)检测临床一对可疑Leber病同卵双生子患者及其亲属,有突变者,对突变序列进行序列测定。 结果 PCR-SSCP检测同卵双生子及母亲11778、3460位点所在的DNA区段存在突变,14484位点所在的DNA区段均未检出突变,将11778位点所在DNA区段克隆后测序显示11778位点仍为CGC,但发现一个新的突变位点(11915T→A)。 结论 Leber病发病机制为mtDNA点突变。多位点突变也被看作为Leber病的病因。11915位点可能也是多位点突变之一。可用PCR SSCP对临床有视神经萎缩和视神经炎可疑为Leber病患者(包括家系成员)做出遗传性视神经病的基因诊断。 (中华眼底病杂志, 1999, 15: 223-226)     

Mitochondrial DNA mutation in Leber's hereditary optic neuropathy.

Leber's hereditary optic neuropathy (LHON) causes acute or subacute central visual loss in healthy young males. Recently, it has been thought to be caused by a single nucleotide change in the ND4 gene in the mitochondrial genome. Mitochondrial DNA (mtDNA) of leukocytes and hair follicle cells from five patients in four families with LHON and nine relatives were analyzed by Sfa NI and Mae III enzyme digestion and DNA sequencing. Loss of Sfa NI site was found in all patients and maternal lineages but not in nonmaternal lineages and normal controls. Mae III digested all the mtDNAs that lost the Sfa NI site. The restriction fragment pattern of polymerase chain reaction (PCR) products exhibited mtDNA heteroplasmy in the hair follicle cells but not in blood cells of the proband in one family. Direct sequencing of PCR-amplified mtDNA fragments encompassing the ND4 gene of the patients disclosed a transition from guanine to adenine at nucleotide position 11778. These results confirm previous reports that a G to A point mutation is associated with LHON and that tissue variability and heteroplasmy of mtDNA exist in some, but not all, LHON patients.     

Secondary mutations of mitochondrial DNA in Japanese patients with Leber's hereditary optic neuropathy

Purpose: Based on studies on the pathogenesis of Leber's hereditary optic neuropathy (LHON), mitochondrial DNA (mtDNA) mutations have been divided into two types: primary and secondary. Primary mutations at nucleotide positions (nt) 11778, 3460, and 14484 can each cause LHON. Secondary mutations may be simultaneously found in LHON patients with a primary mutation, may occur at higher frequency in LHON patients than in normal controls, and may play an additional role in the pathogenesis of LHON. We examined the frequencies of secondary mutations of mtDNA at nt3394, 7444, 9438, 9804, 13708, and 15257 in 19 Japanese patients with LHON associated with primary mutations and 108 normal controls. Methods: Mutations were determined by restriction enzyme analysis or DNA sequencing using polymerase chain reaction (PCR) products. Results: One patient with an nt11778 mutation also had an nt13708 mutation. Another patient with an nt3460 mutation also had an nt7444 mutation. During DNA sequencing of the PCR fragment harboring nt3394, three novel mutations in the ND1 gene (nt3316, 3496, and 3497 mutations) were found in three patients with an nt11778 mutation. The frequency of these mutations in 108 control subjects was studied further: one (0.9%) had an nt3394 mutation, none (0%) had an nt9804 mutation, one (0.9%) had an nt13708 muation, two (1.9%) had nt3316 mutations, one (0.9%) had an nt3496 mutation, and two (1.9%) had nt3497 mutations. Conclusion: It is unlikely that the frequencies of secondary mutations in Japanese patients with LHON are higher than those in normal Japanese controls. It is possible that the mutations at nt3316, 3496, and 3497 are secondary mutations of LHON.     

7个中国Leber遗传性视神经病变家系线粒体DNA突变分析

目的 观察7个中国Leber遗传性视神经病变(LHON)家系的分子遗传学特征.方法 对7个家系先证者及其母系成员和134例正常健康者进行临床眼科检查.除7个先证者外,还确诊2例LHON患者.用24对有部分重叠的引物对受检者线粒体DNA全序列进行扩增,双向测序,结果与修正的剑桥参照序列进行比对,分析突变位点.计算突变位点的外显率,分析家系的单体型.结果 7个家系先证者及其母系成员均未携带ND4 G11778A、ND1 G3460A和ND6 T14484C这3个常见的原发突变位点,但均携有与LHON相关的ND1 T3394C突变位点.134位正常健康者中仅发现4例携带此突变位点.7个家系的ND1 T3394C外显率分别为12.50％、22.22％、16.67％、6.25％、9.09％、11.11％、28.57％.根据本亚线粒体单体型系统进化树分析结果,7个家系分别属于东亚线粒体单体型M9、M9、M、D4、M、M9、M9.结论 中国LHON家系中存在ND1 T3394C突变位点,该突变位点的外显率为6.25％～28.57％,表现度不一.     

Leber's hereditary optic neuropathy mitochondrial DNA mutations in normal-tension glaucoma

BACKGROUND: in Leber's hereditary optic neuropathy, increased optic nerve cupping has been reported by several authors. Recently, a mitochondrial DNA (mtDNA) mutation at nucleotide 11778 typically associated with Leber's hereditary optic neuropathy (LHON) was identified in a patient treated for glaucoma but lacking typical signs of LHON. The question arises: should all normal-tension glaucoma patients be further evaluated for LHON? METHODS: we screened 54 unselected patients with normal-tension glaucoma (age range 20-96 years, 16 men and 38 women) for the primary mtDNA LHON mutations at nucleotides 3460, 11778 and 14484. RESULTS: none of the patients harboured the mtDNA mutations at nucleotides 3460, 11778 or 14484 (95% confidence intervals for each mutation ranged from 0% to 5.3%). CONCLUSIONS: primary LHON mtDNA mutations are rare or absent in unselected normal-tension glaucoma patients. Therefore, unselected normal-tension glaucoma patients should not be screened for these mutations. It is probable that only normal-tension glaucoma patients with atypical features (rapid progression, early deep central scotoma, pallor of neuroretinal rim, elevated disc, peripapillary teleangiectasia) or a positive family history of visual loss compatible with a matrilinear transmission should be further evaluated.     

全血样等位基因特异性聚合酶链反应法筛查Leber遗传性视神经病变线粒体DNA G11778A位点突变

目的建立全血样等位基因特异性聚合酶链反应(PCR)法快速筛查Leber遗传性视神经病变(LHON)患者线粒体DNA G11778A位点突变。方法采用直接以全血样为模板的等位基因特异性PCR法检测14例阳性对照和10例阴性对照血样。同时对22例血样进行双盲检测。 结果24份对照血样检测结果表明该方法的准确率为100%,而且以全血样为模板的PCR特异性优于以纯化DNA为模板的。双盲检测22例血样发现7例阳性结果和15例阴性结果,与预期结果相一致。结论该方法简便、快速、准确、经济,非常适合临床基因诊断线粒体DNA G11778A位点突变的LHON患者,具有重要的临床推广应用价值。     

Clinical Analysis of Leber＇s Hereditary Optic Neuropathy Harboring mtDNA Mutation at nt11778

Purpose: To improve our diagnostic technique through the analysis of clinical features of Leber‘s hereditalw optic neuropathy (LHON) harboring mtDNA point mutation at nt11778. Methods: Detection of nt11778 mutation was performed on 38 patients clinically diagnosed as LHON in our ophthalmic center from year 1998 to 2000. Circumstances of onset and family history were obtained and ophthalmoscopy, fundus fluorescein angiography, visual field and visual evoked potential were performed on all 38 patients. Result: 30 In 38 patients (78.95 % ) harbor nt11778 mutation, including 28 male (93.33%) and 2 female (6.67%). The ratio of affected male to female is 14: 1. Patients harboring nt11778 mutation display typical clinical manifestations. Cenelusion: Identification of one of the three LHON specifically associated mtDNA mutations is essential to confirm the diagnosis. Eye Science 2001: 17: 31 - 34.     

DNA diagnosis of Leber's hereditary optic neuropathy

The point mutation at nt11778 in mitochondrial DNA is highly associated with Leber's hereditary optic neuropathy and eliminates a restriction enzyme SfaNI site in American blacks and Caucasians. DNA diagnosis was applied to a male Japanese patient with this disorder and his mother as a carrier. The mitochondrial DNA fragments (255bp) including this mutation were amplified by polymerase chain reaction. SfaNI digested the DNA fragments of normal Japanese subjects, but did not digest those of the patient or the carrier, The mutation within the SfaNI site is also associated with Japanese suffering from this disorder.     

Leber＇s遗传性视神经病变患者的线粒体DNA检测

目的：探讨Leber‘s遗传性视神经病变（Leber‘s hereditary optic neuropathy,LHON)相关的线粒体DNA原位点突变在视神经疾病中的诊意义。方法：79例各种,原因引起的双侧视神经疾病中,16例为临床诊断的LHPON患者,44例为可疑HLON患者,2例为酒精性弱视患者,4例为多发性硬化症患者,5例为常染色体显性遗传的视神经萎缩患者,4例为原发性开角型 青光眼患者,3例为脊髓小脑退行性变和1例乙胺丁醇引起的视神经萎缩患者,用聚合酶链反应（Polymerase chain reatction,PCR)及限制性片段长度多态性技术。检测外周血DNA中提取的线粒体DNA的3460位点、11778位点及14484位点,分析3个原发位点的突变。结果：31例（39．2％）呈11778位点突变阳性,其中包括16例临床诊断为LHON的患者、13例（29．5％）可疑LHON患者及2例酒精性弱视患者,余48例均未检出上述3个原发位点突变。结论：线粒体DNA的检测分析为确立或排除LHON提供了诊断依据,尤其是对无家族史或原因不明的双侧性视神经炎的患者更具有诊断价值。     

Leber's hereditary optic neuropathy

Leber's hereditary optic neuropathy (LHON) is a rare disease primarily affecting the retinal ganglion cells. In most cases patients with LHON develop permanent visual loss with a large central scotoma in the visual field of both eyes. The optic disc becomes partially or completely pale. At the onset of the disease many patients are considered to suffer from an optic neuritis and are treated under the diagnostic and therapeutic regimen of optic neuritis. LHON is mostly only considered when high dose cortisone therapy fails to be effective or the second eye is affected. Thereafter, molecular genetic analysis will prove LHON in these cases. Detailed anamnesis including pedigree analysis in combination with observance of the peripapillary microangiopathic alterations at the fundus will help to speed up the diagnosis of LHON, but even after exact clinical and molecular genetic diagnosis of LHON some aspects of the disease still remain a mystery today.