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Loss-of-function variant in the gene encoding the KCNQ4 potassium channel causes autosomal dominant nonsyndromic hearing loss (DFNA2), and no effective pharmacotherapeutics have been developed to reverse channel activity impairment. Phosphatidylinositol 4,5-bisphosphate (PIP2), an obligatory phospholipid for maintaining KCNQ channel activity, confers differential pharmacological sensitivity of channels to KCNQ openers. Through whole-exome sequencing of DFNA2 families, we identified three novel KCNQ4 variants related to diverse auditory phenotypes in the proximal C-terminus (p.Arg331Gln), the C-terminus of the S6 segment (p.Gly319Asp), and the pore region (p.Ala271_Asp272del). Potassium currents in HEK293T cells expressing each KCNQ4 variant were recorded by patch-clamp, and functional recovery by PIP2 expression or KCNQ openers was examined. In the homomeric expression setting, the three novel KCNQ4 mutant proteins lost conductance and were unresponsive to KCNQ openers or PIP2 expression. Loss of p.Arg331Gln conductance was slightly restored by a tandem concatemer channel (WT-p.R331Q), and increased PIP2 expression further increased the concatemer current to the level of the WT channel. Strikingly, an impaired homomeric p.Gly319Asp channel exhibited hyperactivity when a concatemer (WT-p.G319D), with a negative shift in the voltage dependence of activation. Correspondingly, a KCNQ inhibitor and chelation of PIP2 effectively downregulated the hyperactive WT-p.G319D concatemer channel. Conversely, the pore-region variant (p.Ala271_Asp272del) was nonrescuable under any condition. Collectively, these novel KCNQ4 variants may constitute therapeutic targets that can be manipulated by the PIP2 level and KCNQ-regulating drugs under the physiological context of heterozygous expression. Our research contributes to the establishment of a genotype/mechanism-based therapeutic portfolio for DFNA2., Hearing loss: genetically tailored treatment strategy for congenital deafness People with a certain type of inherited hearing loss may stand to benefit from a personalized, genetically tailored treatment strategy. A research team from South Korea led by Byung Yoon Choi from Seoul National University College of Medicine identified three new mutations in KCNQ4, a gene that encodes a potassium channel protein found in the inner ear, that can cause congenital deafness. Strikingly, the authors identified the first hyperactive KCNQ4 variant. They inserted these gene variants into human cells in culture, and found that drugs known to affect the activity of this channel protein had different effects on different mutations. Impairments caused by two of the variants were prevented by drug treatment; the third variant proved resistant to the same therapeutic approach. The authors propose further validating the therapeutic responsiveness of different KCNQ4 variants in genetically engineered mice.

LMX1A, encoding the LIM homeobox transcription factor, is essential for inner ear development. Despite previous reports of three human LMX1A variants with nonsyndromic hearing loss (NSHL) in the literature, functional characterization of these variants has never been performed. Encouraged by identification of a de novo, heterozygous, missense variant (c.595A > G; p.Arg199Gly) located in the homeodomain of LMX1A in a subject with congenital severe-to-profound deafness through Exome sequencing, we performed luciferase assay to evaluate transcriptional activity of all LMX1A variants reported in the literature including p.Arg199Gly. Resultantly, p.Arg199Gly manifesting the most severe NSHL showed the biggest reduction of transcriptional activity in contrast with moderately reduced activity of p.Cys97Ser and p.Val241Leu associated with less severe progressive NSHL, proposing a genotype-phenotype correlation. Further, our dominant LMX1A variant exerted pathogenic effects via haploinsufficiency rather than dominant-negative effect. Collectively, we provide a potential genotype-phenotype correlation of LMX1A variants as well as the pathogenic mechanism of LMX1A-related NSHL.

Variant prioritization of exome sequencing (ES) data for molecular diagnosis of sensorineural hearing loss (SNHL) with extreme etiologic heterogeneity poses a significant challenge. This study used an automated variant prioritization system (“EVIDENCE”) to analyze SNHL patient data and assess its diagnostic accuracy. We performed ES of 263 probands manifesting mild to moderate or higher degrees of SNHL. Candidate variants were classified according to the 2015 American College of Medical Genetics guidelines, and we compared the accuracy, call rates, and efficiency of variant prioritizations performed manually by humans or using EVIDENCE. In our in silico panel, 21 synthetic cases were successfully analyzed by EVIDENCE. In our cohort, the ES diagnostic yield for SNHL by manual analysis was 50.19% (132/263) and 50.95% (134/263) by EVIDENCE. EVIDENCE processed ES data 24-fold faster than humans, and the concordant call rate between humans and EVIDENCE was 97.72% (257/263). Additionally, EVIDENCE outperformed human accuracy, especially at discovering causative variants of rare syndromic deafness, whereas flexible interpretations that required predefined specific genotype–phenotype correlations were possible only by manual prioritization. The automated variant prioritization system remarkably facilitated the molecular diagnosis of hearing loss with high accuracy and efficiency, fostering the popularization of molecular genetic diagnosis of SNHL.

Obesity and insulin resistance are primary risk factors for Non-Alcoholic Fatty Liver Disease (NAFLD). NAFLD is generally exhibited by non-progressive simple steatosis. However, a significant subset of patient's progress to nonalcoholic steatohepatitis (NASH) that is defined by the presence of steatosis, inflammation and hepatocyte injury with fibrosis. Unfortunately, there are no approved therapies for NAFLD or NASH and therefore therapeutic approaches are urgently needed. Niclosamide is an U.S. Food and Drug Administration (FDA)-approved anthelmintic drug that mediates its effect by uncoupling oxidative phosphorylation. Niclosamide and its salt forms, Niclosamide Ethanolamine (NEN), and Niclosamide Piperazine (NPP) have shown efficacy in murine models of diet induced obesity characterized by attenuation of the prominent fatty liver disease phenotype and improved glucose metabolism. While the exact mechanism(s) underlying these changes remains unclear, the ability to uncouple oxidative phosphorylation leading to increased energy expenditure and lipid metabolism or attenuation of PKA mediated glucagon signaling in the liver have been proposed. Unfortunately, niclosamide has very poor water solubility, leading to low oral bioavailability. This, in addition to mitochondrial uncoupling activity and potential genotoxicity have reduced enthusiasm for its clinical use. More recently, salt forms of niclosamide, NEN and NPP, have demonstrated improved oral bioavailability while retaining activity. This suggests that development of safer more effective niclosamide derivatives for the treatment of NAFLD and Type 2 Diabetes may be possible. Herein we explored the ability of a series of N-substituted phenylbenzamide derivatives of the niclosamide salicylanilide chemotype to attenuate hepatic steatosis using a novel phenotypic in vitro model of fatty liver and the high fat diet-fed mouse model of diet induced obesity. These studies identified novel compounds with improved pre-clinical properties that attenuate hepatic steatosis in vitro and in vivo. These compounds with improved drug properties may be useful in alleviating symptoms and protection against disease progression in patients with metabolic syndrome and NAFLD.

Objective To determine the incidence, etiology, and concurrent ophthalmic findings in small breed dogs that presented with cataracts. Animal studied Five hundred and sixty-one small breed dogs (942 eyes) were presented to the Veterinary Medical Teaching Hospital of Seoul National University between July 2002 and December 2007 with cataract. Procedure The medical records of small breed dogs with cataract were reviewed. The reason for presentation, cataract duration, bilaterality, breed, gender, age, vision, etiology, stage of cataract development, concurrent ophthalmic findings, IOP, ocular ultrasonography findings, and scotopic electroretinography (ERG) findings were investigated. Results The most frequently presented breeds were the Miniature/Toy Poodle (n = 112, 20.0%), Yorkshire Terrier (n = 110, 19.6%), and Shih Tzu (n = 95, 16.9%). The Miniature/Toy Poodle showed a significantly higher odds ratio for cataract formation (2.6). The proportion of female cataract patients was significantly higher than that of male cataract patients in the overall population (P

Objective To evaluate motility of silicone orbital implants and corneoscleral prostheses, with and without use of a motility coupling post (MCP) in dogs. Animals studied Eighteen mixed-breed dogs. Procedures The motility of an orbital silicone implant and corneoscleral prosthesis after enucleation (n = 6), evisceration (n = 6), or use of a MCP with evisceration (n = 6) in dogs were compared. One eye from each dog had surgery whereas the opposite eye was used as a control. Clinical evaluations were performed three times a week. Histopathology of the orbital tissues was performed 8 and 12 weeks after surgery. Results Implant motility in dogs with evisceration (vertical movement [VM] 8.04 ± 2.13; horizontal movement [HM] 11 ± 3.05) and evisceration with MCP (VM 9.61 ± 1.59); HM was significantly greater than the enucleation group (VM 0.51 ± 0.5; HM 1.22 ± 0.68) (P

The optimal dark adaptation time of electroretinograms (ERG's) performed on conscious dogs were determined using a commercially available ERG unit with a contact lens electrode and a built-in light source (LED-electrode). The ERG recordings were performed on nine healthy Miniature Schnauzer dogs. The bilateral ERG's at seven different dark adaptation tim es at an intensity of 2.5 cdㆍs/m² was performed. Signal averaging (4 flashes of light stimuli) was adopted to reduce electrophysiologic noise. As the dark adaptation time increased, a significant increase in the mean a-wave amplitudes was observed in comparison to base-line levels up to 10 min (p < 0.05). Thereafter, no significant differences in amplitude oc- cured over the dark adaptation time. Moreover, at this time the mean amplitude was 60.30 ± 18.47 µV. However, no significant changes were observed for the implicit times of the a-wave. The implicit times and amplitude of the b-wave increased significantly up to 20 min of dark adap- tation (p < 0.05). Beyond this time, the mean b-wave am- plitudes was 132.92 ± 17.79 µV. The results of the present study demonstrate that, the optimal dark adaptation time when performing ERG's, should be at least 20 min in con- scious Miniature Schnauzer dogs.

The medical records of 23 dogs that underwent medial canthoplasty for treatment of epiphora were reviewed. The most prevalent breed encountered was the shih tzu. Other affected breeds included the Pekingese, Maltese, toy poodle, and pug. All dogs had epiphora associated with medial canthal trichiasis and/or entropion. Other ocular abnormalities included conjunctivitis, keratitis, pigmentary keratitis, corneal ulceration, globe prolapse, and nasal fold trichiasis. After medial canthoplasty, the epiphora resolved in all dogs.

In pharmaceutical companies and research institutes, many toxicity tests are performed with laboratory animals. This study was performed to produce reference data for eye toxicity tests and to investigate the ophthalmic diseases of 408 ICR mice and 119 BALB/c mice, which are commonly used as subjects in toxicity tests. The experimental animals without clinical disorders were selected regardless of sex. The ophthalmic diseases were examined by using special ophthalmic instruments: direct ophthalmoscope, indirect ophthalmoscope, slit-lamp biomicroscope and focal illuminator. The most prevalent ocular variation within normal limits was hyaloid vessel remnant (ICR mice, 28.2%; BALB/c mice, 31.9%) and the incidence gradually decreased with age. The ocular diseases found in ICR mice were retinal degeneration (9.8%), corneal scar (4.2%), focal cataract (2.2%), anisocoria (1.2%), corneal ulcer (0.2%) and uveitis (0.2%). In BALB/c mice, corneal scar (9.2%), focal cataract (1.7%) and corneal ulcer (0.8%) were the ocular diseases found.

To compare the antiangiogenic effects of tetraarsenic oxide (As4O6) with those of diarsenic oxide (As2O3) in the rat cornea, rat cornea micropocket assay was conducted to induce angiogenesis by implantation of the pellet contained 1.0 ng of nerve growth factor (NGF). Ten of thirty eyes of Sprague-Dawley rats were randomly assigned to one of three groups, namely, control group (no medication), As2O3 group (50 mg/kg As2O3, PO, s.i.d.), and As4O6 group (50 mg/kg As4O6, PO, s.i.d.). After implantation, the number of new vessels, vessel length and clock hour of neovascularization were examined under the microscope from day 3 to day 7. The area of neovascularization was calculated using a mathematical formula. Although new vessels in control and As2O3 groups were first noticed at day 3, whereas those of As4O6 group were first observed on day 5. The number, length, clock hour of neovascularization and areas of the vessels in As4O6 group showed more significant inhibition than those of control and As2O3 groups from day 5 (P

The purpose of this study was to investigate spontaneous eye disease in New Zealand White (NZW) rabbits, which are commonly used for toxicity tests, and to provide reference materials for pharmaceutical companies and research centers. A total of 586 NZW rabbits were randomly chosen without sex preference and were examined using ocular equipment, including a direct ophthalmoscope, an indirect ophthalmoscope, a slit-lamp biomicroscope, a focal illuminator, and a fundus camera. This study showed that the incidence rate of temporary cataracts, regarded as a change within normal variation, was 0.5% in the NZW rabbits. Regarding abnormal ophthalmic disease, blepharitis was the most commonly observed ocular disease. Other findings included cataract, conjunctivitis, choroidal hypoplasia, keratitis, corneal scarring, eyelid laceration, posterior synechiae, uveitis, dacryocystitis, and persistent pupillary membrane. In total, the incidence rate of ophthalmic diseases was 9.6%. Based on sex and age distributions, females had more ocular diseases than males, and rabbits were less susceptible to eye diseases as they got older. In this study, photographs were taken to document findings, such as normal fundus, normal variations, ophthalmic disease, and histopathologic examination.

A mongrel male dog of three years old was referred to the Seoul National University Veterinary Teaching Hospital following a one month history of glaucoma. On ophthalmic examination, hyphema, glaucoma, uveitis, iridal mass, and loss of vision were noted in the right eye. Ultrasonography and computed tomography revealed a mass with involvement of the entire uvea. Radiographic evaluation did not reveal any evidence of distant metastasis. The right eye was surgically removed because of the high likelihood of neoplasia. A histologic diagnosis of malignant uveal melanoma was made.

This study evaluated the surgical outcome and complications of phacoemulsification and the implantation of an acryl foldable intraocular lens (IOL) with a squared edge in dogs with cataracts. Thirty-two eyes from 26 dogs were examined. The mean follow up period was 75.9 days ranging from 23 to 226 days. The complications after phacoemulsification were posterior capsular opacity (PCO) around the IOL (n = 11), ocular hypertension (n = 4), focal posterior synechia (n = 4), hyphema (n = 3) and corneal ulcer (n = 2). The complications associated with the IOL were decenteration of the optic (n = 2) and ventral haptic displacement (n = 1). Most cases of PCO were found only around the margin of the IOL, and all eyes had vision during the observation period. In conclusion, the implantation of an acryl-foldable lens with a squared edge at the time of phacoemulsification is an effective method for preserving the central visual field of dogs with cataract.

This study was performed to compare the effect of intratesticular (IT) injection of xylazine/ketamine combination for canine castration with those of intramuscular (IM) or intravenous (IV) injection. Xylazine and ketamine was administered simultaneously via intratesticularly (IT group), intramuscularly (IM group) or intravenously (IV group) at doses of 2 and 10 mg/kg, respectively. Pain response at the time of injection, mean induction time, mean arousal time, mean walking time and cardiopulmonary function during anesthesia were monitored after the xylazine and ketamine administration. In IV and IM groups, heart rates were significantly decreased 30 and 45 min after xylazine and ketamine administration, respectively (p < 0.05). Respiratory rates were significantly decreased in the IV group (p < 0.05). In the IT group, there was no significant changes in heart and respiratory rates. The occurrence of cardiac arrhythmias was less severe in IT group compared with those in IM and IV groups. The route of administration did not affect rectal temperature. Mean induction time was significantly (p < 0.05) longer in IT group than in IM and IV groups. On the contrary, mean arousal time and mean walking time were shortened in IT group. Clinical signs related to pain response at the time of injection and vomiting were less observed in IT group than in IM group, and head shaking was less shown in IT group than in IM and IV groups during recovery period. These results indicated that intratesticular injection of xylazine/ketamine for castration has several advantages such as less inhibition of cardiopulmonary function and fast recovery from anesthesia without severe complications, and would be an effective anesthetic method for castration in small animal practice.