Christoph Kernstock

Artificial vision with wirelessly powered subretinal electronic implant alpha-IMS

This study aims at substituting the essential functions of photoreceptors in patients who are blind owing to untreatable forms of hereditary retinal degenerations. A microelectronic neuroprosthetic device, powered via transdermal inductive transmission, carrying 1500 independent microphotodiode-amplifier-electrode elements on a 9 mm2 chip, was subretinally implanted in nine blind patients. Light perception (8/9), light localization (7/9), motion detection (5/9, angular speed up to 35 deg s−1), grating acuity measurement (6/9, up to 3.3 cycles per degree) and visual acuity measurement with Landolt C-rings (2/9) up to Snellen visual acuity of 20/546 (corresponding to decimal 0.037 or corresponding to 1.43 logMAR (minimum angle of resolution)) were restored via the subretinal implant. Additionally, the identification, localization and discrimination of objects improved significantly (n = 8; p < 0.05 for each subtest) in repeated tests over a nine-month period. Three subjects were able to read letters spontaneously and one subject was able to read letters after training in an alternative-force choice test. Five subjects reported implant-mediated visual perceptions in daily life within a field of 15° of visual angle. Control tests were performed each time with the implants power source switched off. These data show that subretinal implants can restore visual functions that are useful for daily life.

PNPLA6 mutations cause Boucher-Neuhäuser and Gordon Holmes syndromes as part of a broad neurodegenerative spectrum

Boucher-Neuhäuser and Gordon Holmes syndromes are clinical syndromes defined by early-onset ataxia and hypogonadism plus chorioretinal dystrophy (Boucher-Neuhäuser syndrome) or brisk reflexes (Gordon Holmes syndrome). Here we uncover the genetic basis of these two syndromes, demonstrating that both clinically distinct entities are allelic for recessive mutations in the gene PNPLA6. In five of seven Boucher-Neuhäuser syndrome/Gordon Holmes syndrome families, we identified nine rare conserved and damaging mutations by applying whole exome sequencing. Further, by dissecting the complex clinical presentation of Boucher-Neuhäuser syndrome and Gordon Holmes syndrome into its neurological system components, we set out to analyse an additional 538 exomes from families with ataxia (with and without hypogonadism), pure and complex hereditary spastic paraplegia, and Charcot-Marie-Tooth disease type 2. We identified four additional PNPLA6 mutations in spastic ataxia and hereditary spastic paraplegia families, revealing that Boucher-Neuhäuser and Gordon Holmes syndromes in fact represent phenotypic clusters on a spectrum of neurodegenerative diseases caused by mutations in PNPLA6. Structural analysis indicates that the majority of mutations falls in the C-terminal phospholipid esterase domain and likely inhibits the catalytic activity of PNPLA6, which provides the precursor for biosynthesis of the neurotransmitter acetylcholine. Our findings show that PNPLA6 influences a manifold of neuronal systems, from the retina to the cerebellum, upper and lower motor neurons and the neuroendocrine system, with damage of this protein causing an extraordinarily broad continuous spectrum of associated neurodegenerative disease.

Decreased catalytic activity and altered activation properties of PDE6C mutants associated with autosomal recessive achromatopsia

Mutations in the gene encoding the catalytic subunit of the cone photoreceptor phosphodiesterase (PDE6C) have been recently reported in patients with autosomal recessive inherited achromatopsia (ACHM) and early-onset cone photoreceptor dysfunction. Here we present the results of a comprehensive study on PDE6C mutations including the mutation spectrum, its prevalence in a large cohort of ACHM/cone dysfunction patients, the clinical phenotype and the functional characterization of mutant PDE6C proteins. Twelve affected patients from seven independent families segregating PDE6C mutations were identified in our total patient cohort of 492 independent families. Eleven different PDE6C mutations were found including two nonsense mutations, three mutations affecting transcript splicing as shown by minigene assays, one 1 bp-insertion and five missense mutations. We also performed a detailed functional characterization of six missense mutations applying the baculovirus system to express recombinant mutant and wildtype chimeric PDE6C/PDE5 proteins in Sf9 insect cells. Purified proteins were analyzed using Western blotting, phosphodiesterase (PDE) activity measurements as well as inhibition assays by zaprinast and Pγ. Four of the six PDE6C missense mutations led to baseline PDE activities and most likely represent functional null alleles. For two mutations, p.E790K and p.Y323N, we observed reduction in PDE activity of approximately 60% and 80%, respectively. We also observed differences for Pγ inhibition. The p.E790K mutant, with an IC₅₀ value of 2.7 nm is 20.7-fold more sensitive for Pγ inhibition, whereas the p.Y323N mutant with an IC₅₀ of 158 nm is 3-fold less sensitive when compared with the wildtype control.

Retinal Nerve Fiber Layer Loss in Multiple System Atrophy

The retina can be regarded as a ‘‘window to the central nervous system’’ that might provide new insights into the multisystemic character of neurodegenerative diseases. 1 Retinal nerve fibers (RNFs) can in particular be studied with optical coherence tomography (OCT), which allows fast, noninvasive imaging of retinal tissue with resolution reaching that of low-power histological sections. 2 Latest generation OCT devices are able to substantially reduce test–retest variability by real time eye tracking and to automatically segment and quantify the RNF layer, providing immediate data on the distribution and structural integrity of RNF tissue. We investigated whether multiple system atrophy (MSA) is associated with decreased RNF layer thickness (RNFLT) and if so, whether such an effect correlates with indices of disease severity or disease duration.

Treatment of optic neuritis with erythropoietin (TONE): a randomised, double-blind, placebo-controlled trial—study protocol

Introduction Optic neuritis leads to degeneration of retinal ganglion cells whose axons form the optic nerve. The standard treatment is a methylprednisolone pulse therapy. This treatment slightly shortens the time of recovery but does not prevent neurodegeneration and persistent visual impairment. In a phase II trial performed in preparation of this study, we have shown that erythropoietin protects global retinal nerve fibre layer thickness (RNFLT-G) in acute optic neuritis; however, the preparatory trial was not powered to show effects on visual function. Methods and analysis Treatment of Optic Neuritis with Erythropoietin (TONE) is a national, randomised, double-blind, placebo-controlled, multicentre trial with two parallel arms. The primary objective is to determine the efficacy of erythropoietin compared to placebo given add-on to methylprednisolone as assessed by measurements of RNFLT-G and low-contrast visual acuity in the affected eye 6 months after randomisation. Inclusion criteria are a first episode of optic neuritis with decreased visual acuity to ≤0.5 (decimal system) and an onset of symptoms within 10 days prior to inclusion. The most important exclusion criteria are history of optic neuritis or multiple sclerosis or any ocular disease (affected or non-affected eye), significant hyperopia, myopia or astigmatism, elevated blood pressure, thrombotic events or malignancy. After randomisation, patients either receive 33 000 international units human recombinant erythropoietin intravenously for 3 consecutive days or placebo (0.9% saline) administered intravenously. With an estimated power of 80%, the calculated sample size is 100 patients. The trial started in September 2014 with a planned recruitment period of 30 months. Ethics and dissemination TONE has been approved by the Central Ethics Commission in Freiburg (194/14) and the German Federal Institute for Drugs and Medical Devices (61-3910-4039831). It complies with the Declaration of Helsinki, local laws and ICH-GCP. Trial registration number NCT01962571.

In vivo assessment of retinal vascular wall dimensions.

PURPOSE Retinal blood vessel diameter and arteriovenous ratio (AVR) are commonly used diagnostic parameters. Because vascular walls are typically not visible in funduscopy, clinical AVR estimation is based on the lumen rather than the entire vessel diameter. Here the authors used a transgenic mouse model to quantify AVR in vivo based on total vessel dimensions (wall and lumen). METHODS Confocal scanning laser ophthalmoscopy (cSLO) and indocyanine green angiography of the retinal vasculature were performed in wild-type and transgenic mice expressing green fluorescent protein (GFP) under the transcriptional control of the smooth muscle type α-actin (αSMA) promoter. Spectral-domain-OCT and ERG were performed to control for integrity of retinal structure and function in vivo and histology to demonstrate the location of GFP expression. RESULTS Native cSLO imaging and angiography yielded only inner vessel diameters similar to those observed through clinical funduscopy. In αSMA-GFP mice, autofluorescence imaging of the GFP-marked vascular walls also allowed the determination of outer vessel diameters. The mean AVR based on either inner diameter (AVR(id) = 0.72 ± 0.08) or outer diameter (AVR(od) = 0.97 ± 0.09) measurements were significantly different (P < 0.01). CONCLUSIONS Transgenic αSMA-GFP expression in murine vessel wall components allowed quantification of retinal vessel outer diameters in vivo. Although arterioles and venules differ in lumen and vessel wall width, they share a common outer diameter, leading to an AVR(od) close to unity. Because vessel walls are primary targets in common hypertensive and metabolic diseases, αSMA-GFP transgenic mice may prove valuable in the detailed assessment of such disorders in vivo.

Ataxia meets chorioretinal dystrophy and hypogonadism: Boucher-Neuhäuser syndrome due to PNPLA6 mutations

A 37-year-old woman presented with progressive visual loss and primary amenorrhoea due to hypogonadotropic hypogonadism since 14 years of age. These symptoms were followed by cerebellar ataxia and lower limb spasticity, leading to wheelchair dependency at the age of 30 years. At examination at the age of 37 years, her vision was reduced to perception of hand motion, thus meeting the criteria of legal blindness (defined as visual acuity <20/400). This progressive visual loss was caused by chorioretinal degeneration as demonstrated by fundoscopy and optical coherence tomography (figure 1). Whereas patellar tendon reflexes were increased, Achilles tendon reflexes were …

Phenotyping of Mouse Models with OCT

Optical coherence tomography (OCT) is an invaluable technique to perform noninvasive retinal imaging in small animal models such as mice. It provides virtual cross sections that correlate well with histomorphometric data with the advantage that multiple iterative measurements can be acquired in time line analyses to detect dynamic changes and reduce the amount of animals needed per study.

Electrophysiology and colour: a comparison of methods to evaluate inner retinal function

PurposeSeveral methods are routinely used in the clinic to diagnose and monitor diseases of inner retinal function. In this study, we compare four such methods in patients with diabetes and glaucoma, to determine correlations between their results and to determine which method is most sensitive for detecting disease.MethodsTwenty control subjects, 12 patients with early glaucoma and eight patients with diabetes mellitus, were enrolled in the study. All underwent four examinations: transient pattern electroretinogram (PERG), multifocal pattern electroretinogram (mfPERG), chromatic contrast threshold measurements (protan and tritan), and blue-on-yellow short-wavelength automated perimetry (SWAP).ResultsFor the total cohort of 40 subjects, the results show a significant correlation between the amplitudes of the PERG and those of the mfPERG, as well as between the tritan contrast thresholds and the SWAP MD. Furthermore, ROC analyses reveal that colour contrast thresholds could significantly distinguish between the patient and the control group. Glaucoma patients alone could also be distinguished.ConclusionsWe conclude that the methods compared in this study show correlations between their results if they are testing same pathway or underling cells, and that the colour contrast threshold is the most sensitive method to detect early functional deficits in diabetic and glaucoma patients.

Usher Syndrome and Color Vision

ABSTRACT Purpose: The aim of this study is to report on the results of color vision testing in a European cohort of patients with Usher syndrome (USH). We describe the results in relation to Usher type (USH1 and USH2), age and visual acuity. Methods and methods: The color vision of 220 genetically confirmed adult USH patients, aged 18–70 years, was analyzed with one of three methods: the Farnsworth D-15 Dichotomous test (D-15) along with the Lanthony desaturated 15 Hue tests (D-15d), the Roth 28-Hue test, or the Ishihara 14-plate test. Visual acuity was measured with either the ETDRS or the SNELLEN charts. The Confusion index, the Selectivity index and the Confusion angle were calculated for the panel tests and used for analysis. The numbers of plates that could not be read were analyzed for the Ishihara test. Results: For the panel tests, the degree of color loss (Confusion index) is similar in both subtypes of USH, but the polarization of error scores (Selectivity index) is significantly lower in USH1 than USH2, showing more diffuse errors than those found in USH2. There is no significant correlation between logMAR visual acuity and the Confusion or the Selectivity indices. Additionally, we find a significant correlation between patient age and the degree and the polarity of the loss only in USH2. There was no difference between USH1 and USH2 in the results of the Ishihara test. Conclusions: The examination of color vision in patients with USH shows a significant difference in the pattern of color vision loss in USH1 and USH2 patients, but not in the severity of the loss. In USH2, we find a correlation between patient age and the degree and the polarity of the loss. These results may be due to differences in the pathogenesis of retinal dystrophy in USH1 and USH2.