Novel method to rescue a lethal phenotype through integration of target gene onto the X-chromosome.

The loss-of-function mutations of serine protease inhibitor, Kazal type 1 (SPINK1) gene are associated with human chronic pancreatitis, but the underlying mechanisms remain unknown. We previously reported that mice lacking Spink3, the murine homologue of human SPINK1, die perinatally due to massive pancreatic acinar cell death, precluding investigation of the effects of SPINK1 deficiency. To circumvent perinatal lethality, we have developed a novel method to integrate human SPINK1 gene on the X chromosome using Cre-loxP technology and thus generated transgenic mice termed "X-SPINK1". Consistent with the fact that one of the two X chromosomes is randomly inactivated, X-SPINK1 mice exhibit mosaic pattern of SPINK1 expression. Crossing of X-SPINK1 mice with Spink3+/- mice rescued perinatal lethality, but the resulting Spink3-/-;XXSPINK1 mice developed spontaneous pancreatitis characterized by chronic inflammation and fibrosis. The results show that mice lacking a gene essential for cell survival can be rescued by expressing this gene on the X chromosome. The Spink3-/-;XXSPINK1 mice, in which this method has been applied to partially restore SPINK1 function, present a novel genetic model of chronic pancreatitis

Two Novel Mutations in the EYS Gene Are Possible Major Causes of Autosomal Recessive Retinitis Pigmentosa in the Japanese Population

Retinitis pigmentosa (RP) is a highly heterogeneous genetic disease including autosomal recessive (ar), autosomal dominant (ad), and X-linked inheritance. Recently, arRP has been associated with mutations in EYS (Eyes shut homolog), which is a major causative gene for this disease. This study was conducted to determine the spectrum and frequency of EYS mutations in 100 Japanese arRP patients. To determine the prevalence of EYS mutations, all EYS exons were screened for mutations by polymerase chain reaction amplification, and sequence analysis was performed. We detected 67 sequence alterations in EYS, of which 21 were novel. Of these, 7 were very likely pathogenic mutations, 6 were possible pathogenic mutations, and 54 were predicted non-pathogenic sequence alterations. The minimum observed prevalence of distinct EYS mutations in our study was 18% (18/100, comprising 9 patients with 2 very likely pathogenic mutations and the remaining 9 with only one such mutation). Among these mutations, 2 novel truncating mutations, c.4957_4958insA (p.S1653KfsX2) and c.8868C>A (p.Y2956X), were identified in 16 patients and accounted for 57.1% (20/35 alleles) of the mutated alleles. Although these 2 truncating mutations were not detected in Japanese patients with adRP or Leber's congenital amaurosis, we detected them in Korean arRP patients. Similar to Japanese arRP results, the c.4957_4958insA mutation was more frequently detected than the c.8868C>A mutation. The 18% estimated prevalence of very likely pathogenic mutations in our study suggests a major involvement of EYS in the pathogenesis of arRP in the Japanese population. Mutation spectrum of EYS in 100 Japanese patients, including 13 distinct very likely and possible pathogenic mutations, was largely different from the previously reported spectrum in patients from non-Asian populations. Screening for c.4957_4958insA and c.8868C>A mutations in the EYS gene may therefore be very effective for the genetic testing and counseling of RP patients in Japan

Optical coherence tomography and video recording of a case of bilateral contractile peripapillary staphyloma

Purpose: To report a case of bilateral contractile peripapillary staphyloma (CPS) examined by optical coherence tomography (OCT) and video recording. Observation: We report the clinical history of a male infant with bilateral CPS. Two ophthalmic examinations under general anesthesia were performed during follow-up examinations. During the first fundus examination when the infant was 4 months old, we observed staphylomatous excavation around the optic disc, an almost normal disc appearance at the bottom of the excavation, and irregular contraction bilaterally. OCT images showed deep excavation around the optic disc bilaterally and hyperreflective tissue beneath the sclera in the right eye. During the second examination when the child was 4 years and 1 month, in addition to the previous fundus findings, the fundus examination showed white fibrous tissue on the optic disc and pigmentation around the optic disc in the right eye and a retinal cyst in the left eye. Bilateral simultaneous video recordings showed that the contractions were not synchronized between the eyes. Conclusions: In a patient with bilateral CPS, we observed unsynchronized contraction between the eyes and the presence of sequential hyperreflective tissue under the sclera using bilateral ophthalmic approaches. Keywords: Contractile peripapillary staphyloma, Swept-source optical coherence tomography, Video recordin

Atypical intravitreal growth of retinoblastoma with a multi-branching configuration

Purpose: To report the clinical and histopathological findings of atypical intravitreal growth of a retinoblastoma with a multi-branching configuration. Observations: A 7-month-old boy was referred to our hospital due to leukocoria in the right eye. Ophthalmic examinations identified multi-branching vessels surrounded by diaphanous tissue behind the lens in the right eye. Imaging modalities showed microphthalmos, band-shaped calcification, and cystic lesions in that eye. Because it was difficult to rule out congenital anomalies such as persistent fetal vasculature due to the atypical clinical features of retinoblastoma, we performed a biopsy using a limbal approach. An intraoperative rapid pathological examination led to the definitive diagnosis of retinoblastoma. The right eye was enucleated and postoperative adjuvant chemotherapy was administered. Immunohistochemical staining of the enucleated eyeball showed that the tumoral cells and diaphanous tumoral tissue around the vessels were positive for neuron-specific enolase and Ki-67 and partially positive for glial fibrillary acidic protein (GFAP). The vessels of the diaphanous tissues near the tumoral mass were stained by GFAP and those behind the lens were stained faintly. Conclusions and importance: We described an atypical retinoblastoma of pseudo-persistent fetal vasculature with a multi-branching configuration, which expanded the clinical spectrum of retinoblastoma. Such a specific growth pattern of the embryonic tumor might occur with a combination of retinal development, retinal vasculature, and hyaloid vascular system

X-linked Retinitis Pigmentosa in Japan: Clinical and Genetic Findings in Male Patients and Female Carriers

X-linked retinitis pigmentosa (XLRP) is a type of severe retinal dystrophy, and female carriers of XLRP demonstrate markedly variable clinical severity. In this study, we aimed to elucidate the clinical findings of male patients with and female carriers of XLRP in a Japanese cohort and demonstrate the genetic contribution. Twelve unrelated families (13 male patients, 15 female carriers) harboring pathogenic mutations in RPGR or RP2 were included, and comprehensive ophthalmic examinations were performed. To identify potential pathogenic mutations, targeted next-generation sequencing was employed. Consequently, we identified 11 pathogenic mutations, of which five were novel. Six and five mutations were detected in RPGR and RP2, respectively. Only one mutation was detected in ORF15. Affected male patients with RP2 mutations tended to have lower visual function than those with RPGR mutations. Female carriers demonstrated varying visual acuities and visual fields. Among the female carriers, 92% had electroretinographical abnormalities and 63% had a radial autofluorescent pattern, and the carriers who had higher myopia showed worse visual acuity and more severe retinal degeneration. Our results expand the knowledge of the clinical phenotypes of male patients with and female carriers of XLRP and suggest the possibility that RP2 mutations are relatively highly prevalent in Japan

Clinical features of a toddler with bilateral bullous retinoschisis with a novel RS1 mutation

Purpose: To report the clinical and genetic findings of a male toddler who presented bilateral bullous retinoschisis with a novel RS1 mutation. Observations: This is an observational case report of a patient referred to our hospital with esotropia. A comprehensive ophthalmic examination was performed with the boy (age, 1 year 4 months) under general anesthesia that included fundus examinations, fluorescein angiography (FA), swept-source optical coherence tomography (SS-OCT), and full-field electroretinography (FF-ERG). Genetic analysis of the coding region in the RS1 gene was performed by Sanger sequencing for the patient and mother. There was a family history of X-linked retinoschisis (XLRS). Fundus examinations and FA showed bullous retinoschisis bilaterally in the inferior retina. The SS-OCT images showed two kinds of schisis in the inner nuclear layer (INL) and more proximally. In general, the inner plexiform layer, ganglion cell layer, and retinal nerve fiber layer are in the proximal INL; however, in this case there was hyperreflective tissue with a rough surface instead of normal retinal layers. In addition, in the schisis cavity between the hyperreflective tissue and separated retina, a number of hyperreflective fiber-like strands arose from the hyperreflective tissue and extended to the schisis cavity. During the follow-up period, the bullous retinoschisis collapsed spontaneously in the right eye. FF-ERG showed a reduced b-wave and relatively preserved a-wave in all components. Genetic analysis showed a novel RS1 mutation (c.185_186insT, p.E62DfsX24 in exon 4) in the patient and mother. Conclusions and importance: We report the detailed retinal structure in a genetically identified case of bullous retinoschisis. The notable finding was that the cavity of bullous retinoschisis contained a number of fiber-like strands as observed in the cavity of typical retinoschisis

Maternal Uniparental Isodisomy of Chromosome 4 and 8 in Patients with Retinal Dystrophy: SRD5A3-Congenital Disorders of Glycosylation and RP1-Related Retinitis Pigmentosa

Purpose: Uniparental disomy (UPD) is a rare chromosomal abnormality. We performed whole-exosome sequencing (WES) in cases of early-onset retinal dystrophy and identified two cases likely caused by UPD. Herein, we report these two cases and attempt to clarify the clinical picture of retinal dystrophies caused by UPD. Methods: WES analysis was performed for two patients and their parents, who were not consanguineous. Functional analysis was performed in cases suspected of congenital disorders of glycosylation (CDG). We obtained clinical case data and reviewed the literature. Results: In case 1, a novel c.57G>C, p.(Trp19Cys) variant in SRD5A3 was detected homozygously. Genetic analysis suggested a maternal UPD on chromosome 4, and functional analysis confirmed CDG. Clinical findings showed early-onset retinal dystrophy, intellectual disability, and epilepsy. In case 2, an Alu insertion (c.4052_4053ins328, p.[Tyr1352Alafs]) in RP1 was detected homozygously. Maternal UPD on chromosome 8 was suspected. The clinical picture was consistent with RP1-related retinitis pigmentosa. Although the clinical features of retinal dystrophy by UPD may vary, most cases present with childhood onset. Conclusions: There have been limited reports of retinal dystrophy caused by UPD, suggesting that it is rare. Genetic counseling may be encouraged in pediatric cases of retinal dystrophy