Introduction

Inherited optic neuropathies (IONs) affect the optic nerve and the nervous system. Both dominant and recessive IONs are known. Many of the dominant IONs are caused by mutations of OPA1 (MIM: 605290).1, 2 Some patients have OPA3 (MIM: 606580) mutations.3 Both OPA1 and OPA3 encode inner mitochondrial proteins. Some patients with the OPA1 gene mutations show extraocular features, including hearing disorder, progressive external ophthalmoplegia and various neurological manifestations.4 Patients with the OPA3 gene mutations show dominant IONs and cataracts.

The autosomal-recessive IONs with known molecular defects are OPA7 (MIM#612989), OPA9 (MIM#616289), OPA10 (MIM#616732) and OPA11 (MIM#617302). They are caused by mutations of TMEM126A (MIM*612988),5 ACO2 (MIM*100850),6 RTN4IP1 (MIM*610502)7 and YME1L1 (MIM*607472),8 respectively. Angebault et al.7 reported four families with OPA10. They showed early-onset optic neuropathy and some neurological manifestations.7 RTN4IP1 mutations are designated as ‘optic atrophy 10 with or without ataxia, mental retardation and seizures.’ RTN4IP1 encodes a mitochondrial ubiquinol oxydo-reductase. Angebault et al.7 revealed that fibroblasts from patients with a RTN4IP1 mutation showed decreased activities of mitochondrial respiratory complex I and IV. The cells also showed susceptibility to ultraviolet light. Depletion of Rtn4ip1 by short hairpin RNA resulted in abnormal growth of mouse retinal ganglion cell dendrites.

We report brothers with optic neuropathy who had mutations of the RTN4IP1 gene. They showed extraocular manifestations resembling mitochondrial encephalopathy.

Clinical report

The 15-year-old male (II-1) was the first child of healthy and non-consanguineous Japanese parents (Figure 1). He had presented with poor vision since early childhood. He also showed color blindness and congenital nystagmus. His visual acuity was 0.02 in each eye. He presented with epileptic seizures and visual blurring. The electroencephalography showed spikes and high voltage slow waves from the left occipital region. Transient hyperlacticacidemia was noted on laboratory investigations (lactic acid 74.8 mg dl−1, pyruvic acid 3.22 mg dl−1). Leber’s hereditary optic neuropathy was suspected. However, mitochondrial DNA mutation was not found by the extensive screening system.9 A brain magnetic resonance imaging showed no significant abnormalities except for hypoplastic optic nerves. Magnetic resonance spectroscopy showed no lactate peak.

Figure 1
figure 1

Pedigree of the family.

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Physical growth was also within normal standards for a Japanese male. He had learning difficulty. Neurological examinations were normal expect for the ocular symptoms. He has been seizure-free for a number of years. Ataxia was not observed. Funduscopic examination revealed that bilateral optic disc was moderately pallor and there were depigmented areas around the disc (Figures 2a and b).

Figure 2
figure 2

Fundus examinations revealed pallor of the optic discs and depigmented areas around the disc. (a,b) Patient 1 (II-1) (c,d) Patient 2 (II-2) Depigmentation was more prominent in Patient 2 (II-2). LE, left eye; RE, right eye. A full color version of this figure is available at the Journal of Human Genetics journal online.

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The 10-year-old male (II-2) was the younger brother of Patient 1. He presented with poor vision, color blindness and congenital nystagmus. His visual acuity was 0.03 in each eye. He did not show hyperlacticacidemia or epileptic seizures. Physical growth was normal. His bilateral optic disc was pallor and depigmented area was found around the disc (Figures 2c and d). Depigmentation was more prominent in Patient 2 (II-2).

Psychomotor development was mildly delayed. His development quotient was 72 based on the Kyoto Scale of Psychological Development 2001 at 5 years old. Macular function was disturbed in both patients. The visual fields were severely constricted in both patients. The brothers did not show sensitivity to ultraviolet in daily activities. Their parents (I-1, I-2) with the heterozygous variant did not show ocular symptoms.

Materials and methods

The study was approved by the ethics committees of the respective institutions. All biological samples were obtained after written informed consent from the individuals of the family. The samples were analyzed using whole-exome sequencing. Three μg of DNA was sheared to 150–200 bp using the Covaris DNA Shearing System (Woburn, MA, USA). To capture the exonic DNA, we used the SureSelect XT Human All Exon V4/V5 capture library (Agilent, Santa Clara, CA, USA). The sequence library was constructed with the SureSelect XT Target Enrichment System for Illumina Paired-End Sequencing Library kit (Agilent) according to the manufacturer’s instructions. We performed DNA sequencing of 101 bp paired-end reads using the Illumina (San Diego, CA, USA) HiSeq 2000 sequencer. We analyzed exome data as described previously.10 The computer software ANNOVAR (http://annovar.openbioinformatics.org/en/latest/) was used to annotate of sequence data.

Results

We identified a c.308G>A NM_032730.5:exon2:c.G308A:p.R103H, substitution and a c.806+1G>A in the splice site substitution of RTN4IP1 (Figure 3). Patients 1 (II-1) and 2 (II-2) had compound heterozygous mutations. The parents were heterozygous for one of the each mutated alleles.

Figure 3
figure 3

RTN4IP1 mutations in the family of Patients 1 (II-1) and 2 (II-2) were compound heterozygous for the mutation. The parents were each heterozygous for one of the each mutated alleles. A full color version of this figure is available at the Journal of Human Genetics journal online.

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Discussion

We identified a c.308G>A (p.Arg103His) substitution and a c.806+1G>A splice site substitution of RTN4IP1. The c.308G>A (p.Arg103His) substitution has been previously reported by Angebault et al.7 The splice site variant is believed to have a deleterious effect. It is interesting that this mutation was repeatedly found in unrelated families. The splice site variant is a novel mutation. No other variants in optic atrophy-related genes including OPA1, OPA3, TMEM126A, ACO2 and YME1L1 were found.

Patient 1 (II-1) had been more severely affected by epileptic seizures from the age of 3 years. Transient hyperlacticacidemia was noted. These findings were suggestive of a mitochondrial encephalopathy. He is now free from seizures and there has been no further progression of disorder. Angebault et al.7 reported on a patient that showed myoclonic seizures and two patients who exhibited mild intellectual disability. Magnetic resonance imaging of the brothers showed only hypoplasia of optic nerve. No significant abnormalities including stroke-like lesions were observed.

Angebault et al.7 assessed the subcellular localization of RTN4IP1 with the outer membrane mitochondria. Rtn4ip1 plays important roles in controlling the Rtn4 function and disruption of Rtn4ip1 results in abnormal retinal ganglion cell neurite outgrowth.7 Ocular size of the morpholino-injected zebrafish was small. Looping swimming behavior suggesting visual impairment was observed in the zebrafish. Our patients showed congenital nystagmus and low visual acuity. Their ocular size was normal.

Our results further support the suggestion that RTN4IP1 abnormalities result in early-onset optic neuropathy and neurological features including mild intellectual disability and epilepsy. The clinical manifestations may resemble mitochondrial encephalopathy. We should consider RTN4IP1 mutations in similar patients.