Objective To observe the molecular genetic characteristics of seven Chinese families with Leberprime;s hereditary optic neuropathy (LHON). Methods Ophthalmologic examinations were performed on seven probands, maternal members from seven Chinese families and 134 healthy controls. There were two LHON patients in seven Chinese families except probands. The entire mitochondrial genome was amplified using 24 pairs of oligonucleotide primers with overlapping fragments.The mutational site was analyzed through comparison of the Results and Cambridge reference sequence. The penetrance of mutation site was calculated and the haplotype was analyzed. Results Molecular analysis of mitochondrial DNA (mtDNA) in these pedigrees revealed the absence of three common LHON associated with ND4 G11778A, ND1 G3460A and ND6 T14484C mutations. The ND1 T3394C mutation in probands and other matrilineal relatives was present in four out of 134 Chinese healthy controls. Strikingly, these families exhibited very low penetrance of visual impairment. The penetrance was 12.50%, 22.22%, 16.76%, 6.25%, 9.09%, 11.11% and 28.57%. The Results of phylogenetic tree analysis of submitochondrial haplotype showed that these mtDNA polymorphism sites belong to the Asian haplogroups M9, M9, M, D4, M, M9 and M9. Conclusions T3394C mutation exists in seven Chinese LHON pedigrees, and the penetrance was ranged from 6.25% to 28.57%. The patients have different clinical manifestations.
Objective To find the new mutations of Leber's hereditary optic neuropathy (LHON). Methods Two LHON families were enrolled in this study. The probands and all maternal members in this two families were underwent ophthalmologic examinations. The ages of probands were seven and 14 years old respectively. A total of 358 healthy adults were enrolled in this study as control group. The genomic DNA from whole blood of participants were extracted. The entire mitochondrial genome of probands were PCR amplified and sequenced in 24 overlapping fragments using primers as designed. At the same time, the mtDNA of maternal relatives and 358 controls were also detected. Fourteen primate species were selected from GenBank to analyzed the phylogenetics of mitochondrial sequence. Results There was no ND4 G11778A, ND1 G3460A, ND6 T14484C mutational site in all maternal members. Molecular analysis of mtDNA in this two families identified the homoplasmic tRNAGluA14683G mutation and distinct set of variants belonging to the Asian haplogroup F1a1 and G2. The site was at theTpsi;C stem oftRNAGlu and extremely conserved among 14 primate species. It was anticipated that the A14683G increased the highly conserved C-G basepairing. Furthermore, the A14683G was absence in control group. Conclusion The tRNAGluA14683G mutation is likely a new mutation associated with LHON.
ObjectiveTo observe the effects of penetrance, different time of onset and mutation sites on retinal nerve fiber layer (RNFL) and macular thickness in patients with Leber's hereditary optic neuropathy (LHON).MethodsThis was a cross-sectional observational study. A total of 88 patients with LHON and 1492 relatives of the maternal relatives (gene carriers) who received treatment in People’s Liberation Army General Hospital from 2015 to 2017 were included in the study. Among the 1492 family members, there were 694 males and 798 females. Peripheral venous blood was extracted from all subjects for mitochondrial DNA testing, and penetrance was calculated. A total of 117 patients underwent BCVA and SD-OCT examinations, including 82 patients and 35 gene carriers. The BCVA examination was performed using the Snellen visual acuity chart, which was converted into logMAR visual acuity. The thickness of RNFL, ganglion cell complex (GCC) and inner limiting membrane (ILM)-RPE were measured with OCT instrument. The mean follow-up was 50.02±86.27 months. The disease course was divided into 6 stages including ≤3 months, 4-6 months, 7-12 months and >12 months. The thickness of RNFL, GCC and ILM-RPE in patients with different time of onset and mutation sites were comparatively analyzed by covariance analysis. Categorical variables were expressed as a percentage, and the χ2 test was used for comparison among multiple groups.ResultsAmong the 1492 family members, 285 were diagnosed with LHON and highly suspected clinical manifestations (19.10%), including 190 males (21.98%) and 95 females (11.90%). The total penetrance rates of 11778, 14484 and rare mutation sites were 19.84% (228/1149), 20.50% (33/161), and 13.19% (24/182) respectively; male penetrance rates were 28.87% (153/530), 27.28% (20/72), and 18.48% (17/92) and female penetrance rates were 12.12% (75/619),14.61% (13/89) and 7.78% (7/90). There was no significant difference in total (χ2=4.732), male (χ2=4.263) and female (χ2=4.263) penetrance between different mutation sites (P=0.094, 0.110, 0.349). Compared with non-pathogenic carriers, the thickness of the RNFL, GCC and ILM-RPE were all different in the four stages ( ≤3months, 4-6 months, 7-12 months and >12 months). The thickness of RNFL, GCC and ILM-RPE decreased with the time of onset (P=0.000). There were significant differences in the thickness of each of the GCC and ILM-RPE layers in the macular area of LHON patients with different mutation sites (P＜0.05). Among them, the site 11778 and 3460 had the most severe damage in all quadrants of macular GCC and ILM-RPE layer, followed by 14484 site, and the rare site had the least damage in all quadrants.ConclusionsThe penetrance of LHON patients is 19.10%. With the extension of the onset time (within 1 year), the RNFL layer of the optic disc and all quadrants of the macular GCC and ILM-RPE layer gradually thinned. Compared with 11778 and rare site, 14484 site, and the rare site had the lighter damage on the thickness of RNFL, GCC and ILM-RPE.
Leber hereditary optic neuropathy is an optic neuropathy associated with mitochondrial DNA. The disease affects young men mainly, which is considered to be due to denaturation of the retinal nerve ganglion cell and axonal loss of optic nerve, leading to optic atrophy. Nowadays, there are some development in studying Leber hereditary optic neuropathy by optical coherence tomography (OCT) and optical coherence tomography angiography (OCTA). It is great help to know the disease, forecast the progression of disease, and take action on intervention. In addition, there is a lack of in-depth study on OCT and OCTA characteristics among different mutation sites of LHON, different genders of the same site, different families of the same site or even different branches of the same family. It is expected to be improved in the future work.
ObjectiveTo analyze the characteristics and prognosis of visual field of Leber hereditary optic neuropathy (LHON) with G11778A mutation.MethodsA retrospective clinical study. Twenty-two (44 eyes) of LHON patients diagnosed with G11778A site mutation by mt-DNA examination from May 2008 to February 2018 in Ophthalmology Department of Dongfang Hospital of Beijing University of Chinese Medicine, were enrolled in this study. All patients underwent best corrected visual acuity (BCVA), visual field and optical coherence tomography (OCT). The BCVA examination was performed using the international standard visual acuity chart, which was converted into logarithm of the minimum angle of resolution (logMAR) BCVA for record. The thickness of the retinal nerve fiber layer (RNFL) in the 200μm×200μm annular region 1.73 mm outside the optic disc was measured by OCT. At least 7 visual field examinations were performed within one month before and after 2, 4, 8, 12, 18, 24 and 30 months of the course of disease by using Octopus 101 perimetry. Among 44 eyes, 27 eyes were detected with G2 procedure (G2 group) and 17 eyes were detected with LVC procedure (LVC group). The mean field defect (MD) and mean optical sensitivity (MS) were used as the main outcome indexes. According to the onset age, the patients were further divided into the ≤14 years old group and＞14 years old group. There was a significant difference in initial logMAR BCVA between the G2 group and LVC group (t=4.994, P=0.000), but there was no significant difference in gender (χ2=1.896, P=0.169) and age (t=0.337, P=0.708) between the two groups. Independent sample t test was used for comparison between groups, paired t test was used for comparison within groups, and one-way analysis of variance was used for comparison between groups. The statistical data were compared by χ2 test.ResultsIn the G2 group, the MD value of the subgroup of children (≤14 years old) decreased gradually during the follow-up period, and the MD value since 18 months after onset was significantly lower than the value of 2 months after onset (t=3.813, 4.590, 5.033; P=0.002, 0.001, 0.000). No obvious visual field index changes were seen in other subgroups (P＞0.05). The central scotoma was the most common type of visual field defect in the early stage, and the diffuse defect was the most common type of visual field defect in the late stage. There was a significant difference in the types of visual field distribution between the early and late stage in G2 group (χ2=17.414, P=0.015). There was no significant difference in the type of visual field distribution between the early and late stage in LVC group (χ2=4.541, P=0.474). The MD value in the G2 group remained stable within 8 months after onset, but significantly improved after 18 months after onset (t=2.100, 3.217, 3.566; P=0.046, 0.003, 0.001). The MS in the LVC group did not significantly improve during follow-up (P＞0.05). The average visual acuity of the G2 group was significantly improved from 12 months (t=3.039, 3.678, 4.264, 5.078; P=0.008, 0.002, 0.001, 0.000). The visual acuity of the eyes in the G2 group was better than that of the LVC group during all follow-up periods (P≤0.05). The RNFL thickness of all patients continued to decrease after onset, but the RNFL thickness was significantly higher at 4, 8, 18, 24, 30 months in the G2 group than those in the LVC group (t=2.471, 2.269, 2.474, 2.509, 2.782; P=0.018, 0.028, 0.017, 0.016, 0.008).ConclusionsThe main types of visual field defect of LHON with G11778A mutation are the central scotoma in the early stage, while the diffuse defect and central scotoma are both very common in the later stage. The visual field of LHON patients examined by G2 procedure is significantly improved during the follow-up, as well as the visual acuity improved significantly, and the visual field improvement in younger cases (≤14 years old) is better than that of older cases (＞14 years old), but the visual field of the LVC procedure cases did not improve during follow-up.
ObjectiveTo observe the changes of vision and visual evoked potentials (VEP) in patients with Leber hereditary optic neuropathy (LHON) before and after gene therapy.MethodsA retrospective cohort study. From December 2017 to October 2018, 35 cases of 70 eyes of m11778G.A/MT-ND4 mutant LHON patients who were diagnosed in the Tongji Hospital of Huazhong University of Science and Technology and received gene therapy were included in the study. There were 30 males (87.71%) and 5 females (12.29%), with the mean age of 23.31±6.72 years. The gene therapy method was intravitreal injection of rAAV2-ND4 (recombinant adeno-associated virus carrying NADH-ubiquinone oxidoreductase subunit 4 gene) into one eye. The eye with poor visual acuity was chosen as the injection eye. If both eyes had the same visual acuity, the right eye was designated as the injection eye. Seventy eyes were divided into the injected eye group and the non-injected eye group, in which were both 35 eyes. The best corrected visual acuity (BCVA) and pattern VEP (PVEP) examinations were performed in the injected eye group and the non-injected eye group before treatment (baseline), 1, 3, and 6 months after injection. Compare the changes of BCVA and PVEP between the injected eye group and the non-injected eye group at baseline, 1 month, 3 months, and 6 months after injection. Independent sample t test, paired sample t test or two independent sample nonparametric test were performed to compare the two groups.ResultsCompared with baseline, 1, 3, and 6 months after treatment, the BCVA of the injected eye group (t=3.530, 4.962, 5.281; P=0.001, 0.000, 0.000) and the non-injected eye group (t=3.288, 2.620, 2.252; P= 0.002, 0.013, 0.031) increased, and the difference was statistically significant; there was no statistically significant difference between VEP IT (tinjected eye group=−0.158, 1.046, −1.134; Pinjected eye group = 0.875, 0.303, 0.190; tnon-injected eye group=0.773, −0.607, −0.944; Pnon-injected eye group= 0.445, 0.548, 0.352) and VEP A (Zinjected eye group=−0.504, −0.934, −1.065; Pinjected eye group= 0.614, 0.351, 0.287; Znon-injected eye group=−0.521, −0.115, −0.491; Pnon-injected eye group = 0.602, 0.909, 0.623).ConclusionAfter gene therapy, the visual acuity of the injected and non-injected eyes of LHON patients improved; PVEP did not change significantly, and remained stable compared with baseline.
Leber hereditary optic neuropathy (LHON) is a maternally inherited mitochondrial disease. It is clinically recognizable by painless, bilateral loss of vision, and the prognosis of vision is generally poor. In recent years, the information provided by optical coherence tomography (OCT) and OCT angiography (OCTA) has greatly improved people's understanding of LHON, and new progress has been made in the intervention and treatment of LHON. A detailed understanding of the structural changes of retina and choroid under OCT and OCTA of the natural course and after treatment of LHON, may provide reference for revealing the pathogenesis, prediction of onset time, differential diagnosis, follow-up of treatment effect and prognosis of LHON.
Leber's hereditary optic neuropathy (LHON) is a rare hereditary optic nerve disease. At present, the understanding of its etiology and pathogenesis is relatively clear. With the emergence of new drugs such as idebenone and the possibility of gene therapy for LHON, it has brought hope for patients to recover. However, because genetic testing technology has not been widely developed in China, clinical misdiagnosis of LHON as optic neuritis still occurs from time to time. How to make timely identification and correct diagnosis of LHON still poses certain challenges for Chinese ophthalmologists. In addition, in terms of treatment, the choice of treatment methods and treatment costs in the pre-onset (gene mutation carriers) and different periods after the onset of LHON are also huge challenges for patients and their families.
ObjectiveTo investigate the feasibility of immoribund skin fibroblast cell line derived from Leber′s hereditary optic neuropathy (LHON) patients as a cell model. MethodsA basic research. Two LHON patients and 2 healthy volunteers were recruited from Department of Ophthalmology of Genetic Clinic of Henan Provincial Eye Hospital. The skin tissue of participants was obtained, and the 4 immortalized skin fibroblasts were constructed by SV40 virus infection, including 2 LHON patient cells (LHON-1 and LHON-2 cells) and 2 healthy volunteers cells (NC-1 and NC-2 cells). Mitochondrial morphology in cells was observed by electron microscope. The levels of reactive oxygen species (ROS), nicotinamide adenine dinucleotide-oxidation state (NAD+), nicotinamide adenine dinucleotide-reduction state (NADH) and adenosine triphosphate (ATP) in fibroblasts were detected. Cellular oxygen consumption was measured by seahorse mitochondrial pressure assay. Cell viability was detected using cell counting kit-8 (CCK8). One-way ANOVA was performed to compare the levels of ROS, NAD+, NADH and ATP in LHON and NC cells, as well as basal oxygen consumption, maximal oxygen consumption, ATP-coupled oxygen consumption, and cell viability. ResultsCompared with NC-1 and NC-2, the number of mitochondrial crest in LHON fibroblasts was significantly reduced, indicating abnormal mitochondrial morphology. Biochemical analysis showed that ROS levels in LHON cells increased, but NAD+/NADH and ATP levels decreased, and the oxygen consumption was significantly inhibited, indicating the presence of mitochondrial damage and respiratory dysfunction. The results of CCK-8 detection showed that the survival ability of LHON-1 and LHON-2 cells was worse under stress conditions. ConclusionImmortalized skin fibroblast cell lines from LHON patients presented mitochondrial dysfunction.