A deep intronic TCTN2 variant activating a cryptic exon predicted by SpliceRover in a patient with Joubert syndrome.

The recent introduction of genome sequencing in genetic analysis has led to the identification of pathogenic variants located in deep introns. Recently, several new tools have emerged to predict the impact of variants on splicing. Here, we present a Japanese boy of Joubert syndrome with biallelic TCTN2 variants. Exome sequencing identified only a heterozygous maternal nonsense TCTN2 variant (NM_024809.5:c.916C >T, p.(Gln306Ter)). Subsequent genome sequencing identified a deep intronic variant (c.1033+423G>A) inherited from his father. The machine learning algorithms SpliceAI, Squirls, and Pangolin were unable to predict alterations in splicing by the c.1033+423G>A variant. SpliceRover, a tool for splice site prediction using FASTA sequence, was able to detect a cryptic exon which was 85-bp away from the variant and within the inverted Alu sequence while SpliceRover scores for these splice sites showed slight increase (donor) or decrease (acceptor) between the reference and mutant sequences. RNA sequencing and RT-PCR using urinary cells confirmed inclusion of the cryptic exon. The patient showed major symptoms of TCTN2-related disorders such as developmental delay, dysmorphic facial features and polydactyly. He also showed uncommon features such as retinal dystrophy, exotropia, abnormal pattern of respiration, and periventricular heterotopia, confirming these as one of features of TCTN2-related disorders. Our study highlights usefulness of genome sequencing and RNA sequencing using urinary cells for molecular diagnosis of genetic disorders and suggests that database of cryptic splice sites predicted in introns by SpliceRover using the reference sequences can be helpful in extracting candidate variants from large numbers of intronic variants in genome sequencing.

[A case of acute encephalitis with refractory repetitive partial seizures successfully controlled by very-high-dose phenobarbital therapy found in a boy].

We experienced a case of acute encephalitis with refractory, repetitive partial seizures (AERRPS) found in an 8-year-old boy. Convulsive status epilepticus developed at the onset, which was intractable to the treatment with intravenous thiopental sodium even at the maximum dose of 9 mg/kg/hr. Since the adverse effect developed, thiopental sodium was discontinued. Phenobarbital (PB) was administrated at a very high daily dose up to 80 mg/kg, reaching serum trough level of 250 µg/ml, which was markedly effective to the treatment. Because seizures reappeared during tapering the dosage of PB, potassium bromide (KBr) at a daily dose of 80 mg/kg was additionally administrated. PB was successfully tapered into a daily dose of 20 mg/kg with a trough serum level around 80 µg/ml. He recovered in motor functions, but had disturbance of memory and apneic seizures. A very-high-dose PB therapy in an early period may be helpful for the treatment of intractable convulsive status epilepticus.

Rehabilitation Approach for Children With Joubert Syndrome and Related Disorders.

Joubert syndrome and related disorders (JSRD) are rare and intractable diseases characterized by delayed psychomotor development, hypotonia and/or ataxia, and abnormal respiratory and eye movements. Cerebellar vermis agenesis and molar tooth signs are distinct on cerebral magnetic resonance imaging (MRI). Children with JSRD present with delayed psychomotor development, including intellectual disability and emotional or behavioral problems. Rehabilitation treatments are provided to promote psychomotor development. However, limited reports and evidence exist on rehabilitation treatments for children with JSRD. Three children with JSRD received rehabilitation treatment. The children received rehabilitation treatment once a week to once every one to two months at our hospital and/or other facilities. All patients received physical, occupational, and speech-language-hearing therapy, depending on their symptoms and conditions. In children with tracheostomies due to abnormal respiration, respiratory physical therapy and speech-language-hearing therapy, including augmentative and alternative communication, were needed. For hypotonia and ataxia, an orthotic intervention was considered in all three cases, and foot or ankle-foot orthoses were used in two cases. Although there is no specific or established rehabilitation method for children with JSRD, appropriate rehabilitation approaches, including physical, occupational, speech-language-hearing therapies and orthotic intervention, should be considered and provided to improve their function and expand their activity and participation. Orthotic intervention for hypotonia seems reasonable for improving gross motor development and function in children with JSRD.

[Studies on bone mineral status with bone quantitative ultrasonography in severely handicapped school children--correlations with gross motor function and dietary status].

OBJECTIVE: Severely handicapped children and adolescents have reduced bone mineral density and high prevalence of pathological fractures. Bone quantitative ultrasonography (QUS) is a radiation-free method for assessing bone density. It is portable and easy to use in subjects with severe bodily deformities. METHODS: We evaluated 166 students (age 6-20 years) at a school for disabled children for bone mineral density using the osteo-sono-assessment index (OSI) calculated by measuring the velocity of ultrasound waves, the speed of sound (SOS) and the transmission index (TI), at the calcaneus. All examinations were performed using an AOS-100 analyzer (ALOKA Ltd., Tokyo, Japan). The Gross Motor Function Classification System (GMFCS) for cerebral palsy was also applied. We assessed OSI for dietary texture modifications and methods of feeding. RESULTS: Those with pathological fractures tended to have lower OSI than other students. Such fractures were individually unrelated to age, sex and GMFCS. OSI was significantly higher at GMFCS level I than level II. OSI in levels I to III was equally significantly higher than that in levels IV and V. As to feeding methods, the tube feeding group tended to have lower OSI than the oral ingestion group. In the oral ingestion group, those receiving a regular diet had significantly higher OSI than the mixed-minced diet group. However, students with a gastrostomy tended to have higher OSI than those receiving gastro-nasal tube feeding. CONCLUSIONS: Gross motor function (applied GMFCS) is a major factor affecting bone mineral density. Tube feeding reduces bone mineral density. However, forced oral intake may also reduce it. In the tube feeding group, a modified diet of appropriate texture delivered via gastrostomy may be the key to improving bone mineral density.

[Case of infantile autism with pediatric Wernicke's encephalopathy due to severe eating disorder].

Wernicke's encephalopathy (WE) or thiamine deficiency is fatal if left untreated. We report a case of a 3-year-old boy with infantile autism and a severe eating disorder who developed WE after 3 weeks of starvation without thiamine supplementation. The eating disorder started when he entered preschool. He presented with unconsciousness and a cluster of seizures. Cranial magnetic resonance imaging (MRI) showed high-intensity signal changes in the basal ganglia on T2-weighted images and fluid-attenuated inversion recovery (FLAIR). Treatment with high-dose intravenous thiamine was effective. Pediatric patients with WE tends to show no typical symptoms or brain lesions on MRI as seen in adult WE patients typically along alcoholics. Brain lesions similar to those in hypoxia or mitochondrial diseases such as Leigh's encephalopathy, are observed in patients with pediatric WE, and this makes diagnosis difficult. WE should be considered when patients with severe eating disorders present with unconsciousness and/or frequent seizures, and show basal ganglia lesions on MRI, differential diagnosis should include WE.

[Effect of tacrolimus in a case of autoimmune encephalitis].

We report a 17-year-old boy who was diagnosed as autoimmune encephalitis with various neurological complications such as hemiplegia, aphasia and seizures. An autoimmune process was considered to be responsible for the repeated episodes of encephalitis because the symptoms were highly responsive to steroids and anti-glutamate receptor antibodies were detected in the CSF. After administration of the immunosuppressant tacrolimus, we could taper the steroid dosage. He has had no relapse for three years to date. We demonstrated the possibility of steroid-sparing treatment with tacrolimus for a patient with steroid-responsive encephalitis. There were few reports describing tacrolimus therapy for encephalitis. Tacrolimus may be effective for selected patients with recurrent encephalitis in which an autoimmune mechanism is considered as the pathogenesis.