Genetic characteristics of 234 Italian patients with macular and cone/cone-rod dystrophy.

Two-hundred and thirty-four Italian patients with a clinical diagnosis of macular, cone and cone-rod dystrophies (MD, CD, and CRD) were examined using next-generation sequencing (NGS) and gene sequencing panels targeting a specific set of genes, Sanger sequencing and-when necessary-multiplex ligation-dependent probe amplification (MLPA) to diagnose the molecular cause of the aforementioned diseases. When possible, segregation analysis was performed in order to confirm unsolved cases. Each patient's retinal phenotypic characteristics were determined using focal and full-field ERGs, perimetry, spectral domain optical coherence tomography and fundus autofluorescence. We identified 236 potentially causative variants in 136 patients representing the 58.1% of the total cohort, 43 of which were unpublished. After stratifying the patients according to their clinical suspicion, the diagnostic yield was 62.5% and 53.8% for patients with MD and for those with CD/CRD, respectively. The mode of inheritance of all cases confirmed by genetic analysis was 70% autosomal recessive, 26% dominant, and 4% X-linked. The main cause (59%) of both MD and CD/CRD cases was the presence of variants in the ABCA4 gene, followed by variants in PRPH2 (9%) and BEST1 (6%). A careful morpho-functional evaluation of the phenotype, together with genetic counselling, resulted in an acceptable diagnostic yield in a large cohort of Italian patients. Our study emphasizes the role of targeted NGS to diagnose MDs, CDs, and CRDs, as well as the clinical usefulness of segregation analysis for patients with unsolved diagnosis.

Retinal Morpho-Functional Changes Following 0.19 mg Fluocinolone Acetonide Intravitreal Implant for Chronic Diabetic Macular Edema.

PURPOSE: To evaluate morpho-functional outcomes of the intravitreal fluocinolone acetonide (FAc) implant. METHODS: Retrospective, observational, single-center study. Primary endpoint was the mean change in central macular thickness (CMT) from baseline to month 1-3. Secondary endpoints included mean CMT change from baseline to month 4-8 and 9-14 and mean best corrected visual acuity (BCVA), photopic negative response (PhNR) and b-wave of flash full-field electroretinogram (ERG) changes from baseline to month 1-3, 4-8, and 9-14. RESULTS: Fourteen patients (18 eyes) were included. Mean (standard deviation) CMT decreased from 473 (196) µm at baseline to 371 (163) µm at month 1-3 (mean difference - 102.3 ± 98.35 µm, 95% CI ± 46.4 µm; p < 0.0001) and this decrease tended to endure up to month 9-14. BCVA did not change significantly. There was an improvement in mean PhNR amplitude from 2.76 (1.65) µV at baseline to 3.73 (2.32) µV at month 1-3 (mean difference 0.91 (1.14) µV, 95% CI ± 0.54 µV, p = 0.003); b-wave amplitude improved from 8.83 (4.52) µV at baseline versus 10.05 (5.04) µV at month 1-3 (mean difference 1.22 (2.23) µV, 95% CI ± 1.08 µV, p = 0.0384). These ERG positive changes tended to endure up to month 9-14, although they did not reach statistical significance after month 3. CONCLUSIONS: Intravitreal FAc implant significantly improved anatomic as well as functional outcomes related to middle and inner retinal layers, known to be altered in diabetic retinopathy. Our findings support the hypothesis that intravitreal FAc implant may exert a protective effect in diabetic retinas with diabetic macular edema.

Impaired Ca2+ Sensitivity of a Novel GCAP1 Variant Causes Cone Dystrophy and Leads to Abnormal Synaptic Transmission Between Photoreceptors and Bipolar Cells.

Guanylate cyclase-activating protein 1 (GCAP1) is involved in the shutdown of the phototransduction cascade by regulating the enzymatic activity of retinal guanylate cyclase via a Ca2+/cGMP negative feedback. While the phototransduction-associated role of GCAP1 in the photoreceptor outer segment is widely established, its implication in synaptic transmission to downstream neurons remains to be clarified. Here, we present clinical and biochemical data on a novel isolate GCAP1 variant leading to a double amino acid substitution (p.N104K and p.G105R) and associated with cone dystrophy (COD) with an unusual phenotype. Severe alterations of the electroretinogram were observed under both scotopic and photopic conditions, with a negative pattern and abnormally attenuated b-wave component. The biochemical and biophysical analysis of the heterologously expressed N104K-G105R variant corroborated by molecular dynamics simulations highlighted a severely compromised Ca2+-sensitivity, accompanied by minor structural and stability alterations. Such differences reflected on the dysregulation of both guanylate cyclase isoforms (RetGC1 and RetGC2), resulting in the constitutive activation of both enzymes at physiological levels of Ca2+. As observed with other GCAP1-associated COD, perturbation of the homeostasis of Ca2+ and cGMP may lead to the toxic accumulation of second messengers, ultimately triggering cell death. However, the abnormal electroretinogram recorded in this patient also suggested that the dysregulation of the GCAP1-cyclase complex further propagates to the synaptic terminal, thereby altering the ON-pathway related to the b-wave generation. In conclusion, the pathological phenotype may rise from a combination of second messengers' accumulation and dysfunctional synaptic communication with bipolar cells, whose molecular mechanisms remain to be clarified.

USH2A-Related Retinitis Pigmentosa: Staging of Disease Severity and Morpho-Functional Studies.

Usher syndrome type 2A (USH2A) is a genetic disease characterized by bilateral neuro-sensory hypoacusia and retinitis pigmentosa (RP). While several methods, including electroretinogram (ERG), describe retinal function in USH2A patients, structural alterations can be assessed by optical coherence tomography (OCT). According to a recent collaborative study, RP can be staged considering visual acuity, visual field area and ellipsoid zone (EZ) width. The aim of this study was to retrospectively determine RP stage in a cohort of patients with USH2A gene variants and to correlate the results with age, as well as additional functional and morphological parameters. In 26 patients with established USH2A genotype, RP was staged according to recent international standards. The cumulative staging score was correlated with patients' age, amplitude of full-field and focal flicker ERGs, and the OCT-measured area of sub-Retinal Pigment Epithelium (RPE) illumination (SRI). RP cumulative score (CS) was positively correlated (r = 0.6) with age. CS was also negatively correlated (rho = -0.7) with log10 ERG amplitudes and positively correlated (r = 0.5) with SRI. In USH2A patients, RP severity score is correlated with age and additional morpho-functional parameters not included in the international staging system and can reliably predict their abnormality at different stages of disease.

Combined Intravitreal Dexamethasone Implant and Cataract Surgery in Patients with Diabetic Retinopathy: Effect on Retinal Morphology and Function.

INTRODUCTION: Cataract surgery can be associated with vision-threatening complications in patients with diabetes. This study aimed to assess the functional and anatomic outcomes of the intravitreal dexamethasone (DEX) implant, administered at the time as cataract surgery, in patients with diabetic retinopathy and diabetic macular edema (DME). METHODS: This was a retrospective, observational, and single-center study. The primary endpoint was the mean change in central macular thickness (CMT) from baseline to month 1. Secondary endpoints included mean change in best corrected visual acuity (BCVA) from baseline to month 1 and 3, mean change in CMT from baseline to month 3, the photopic negative response (PhNR) and the b wave of flash full-field electroretinogram from baseline to month 1, and the incidence of adverse events. RESULTS: Twenty-four eyes of 21 patients were included in the study. The mean (range) age of patients was 69 (63-87) years and 13 (61.9%) were men. Mean (standard deviation) CMT significantly decreased from 447 (134) µm at baseline to 341 (134) µm at month 1 (mean difference - 106 ± 134 µm, 95% CI - 183.9 to - 28.1 µm; p = 0.0087). BCVA significantly improved from 46 (20) ETDRS letters at baseline to 59 (22) ETDRS letters at month 1 (mean difference 13 ± 21 letters, 95% CI 0.8-25.2 letters; p = 0.0375). Regarding electrophysiology, there was a statistically significant reduction in mean PhNR from 5.24 (1.67) µV at baseline to 3.73 (1.19) µV at month 1 (mean difference - 1.51 ± 0.42 µV, 95% CI - 2.4 to - 0.7 µV, p = 0.0008); whereas b wave amplitude did not change (12.69 ± 6.89 µV at baseline versus 12.29 ± 6.30 µV at month 1; p = 0.8347). Four (16.7%) eyes developed ocular hypertension over the course of follow-up, which was successfully controlled with topical hypotensive medication. CONCLUSION: Perioperative DEX implant significantly improved both anatomic and functional outcomes in patients with DME who underwent cataract surgery.