Vitamin A is an essential fat-soluble vitamin that occurs in various chemical forms. It is essent... more Vitamin A is an essential fat-soluble vitamin that occurs in various chemical forms. It is essential for several physiological processes. Either hyper- or hypovitaminosis can be harmful. One of the most important vitamin A functions is its involvement in visual phototransduction, where it serves as the crucial part of photopigment, the first molecule in the process of transforming photons of light into electrical signals. In this process, large quantities of vitamin A in the form of 11-cis-retinal are being isomerized to all-trans-retinal and then quickly recycled back to 11-cis-retinal. Complex machinery of transporters and enzymes is involved in this process (i.e., the visual cycle). Any fault in the machinery may not only reduce the efficiency of visual detection but also cause the accumulation of toxic chemicals in the retina. This review provides a comprehensive overview of diseases that are directly or indirectly connected with vitamin A pathways in the retina. It includes the...
USH2A mutation is the most common cause of retinitis pigmentosa, with or without hearing impairme... more USH2A mutation is the most common cause of retinitis pigmentosa, with or without hearing impairment. Patients most commonly exhibit hyperautofluorescent ring on fundus autofluorescence imaging (FAF) and rod-cone dystrophy on electrophysiology. A detailed study of three USH2A patients with a rare pattern of double hyperautofluorescent rings was performed. Twenty-four patients with typical single hyperautofluorescent rings were used for comparison of the ages of onset, visual fields, optical coherence tomography, electrophysiology, and audiograms. Double rings delineated the area of pericentral retinal degeneration in all cases. Two patients exhibited rod-cone dystrophy, whereas the third had a cone-rod dystrophy type of dysfunction on electrophysiology. There was minimal progression on follow-up in all three. Patients with double rings had significantly better visual acuity, cone function, and auditory performance than the single ring group. Double rings were associated with combinat...
Gelsolin amyloidosis typically presents with corneal lattice dystrophy and is most frequently ass... more Gelsolin amyloidosis typically presents with corneal lattice dystrophy and is most frequently associated with pathogenic GSN variant p.Asp214Asn. Here we report clinical and histopathological features of gelsolin amyloidosis associated with a novel GSN variant p.Glu580Lys. We studied DNA samples of seven members of a two-generation family. Exome sequencing was performed in the proband, and targeted Sanger sequencing in the others. The heterozygous GSN variant p.Glu580Lys was identified in six patients. The patients exhibited corneal dystrophy (5/6), loose skin (5/6) and/or heart arrhythmia (3/6) and one presented with bilateral optic neuropathy. The impact of the mutation on the protein structure was evaluated in silico. The substitution is located in the fifth domain of gelsolin protein, homologous to the second domain harboring the most common pathogenic variant p.Asp214Asn. Structural investigation revealed that the mutation might affect protein folding. Histopathological analysi...
Mutations in RPGRORF15 are associated with rod-cone or cone/cone-rod dystrophy, the latter associ... more Mutations in RPGRORF15 are associated with rod-cone or cone/cone-rod dystrophy, the latter associated with mutations at the distal end. We describe the phenotype associated with a novel variant in the terminal codon of the RPGRORF15 c.3457T>A (Ter1153Lysext*38), which results in a C-terminal extension. Three male patients from two families were recruited, aged 31, 35, and 38 years. Genetic testing was performed by whole exome sequencing. Filtered variants were analysed according to the population frequency, ClinVar database, the variant’s putative impact, and predicted pathogenicity; and were classified according to the ACMG guidelines. Examination included visual acuity (Snellen), colour vision (Ishihara), visual field, fundus autofluorescence (FAF), optical coherence tomography (OCT), and electrophysiology. All patients were myopic, and had central scotoma and reduced colour vision. Visual acuities on better eyes were counting fingers, 0.3 and 0.05. Electrophysiology showed sev...
In contrast to USH2A, variants in ADGRV1 are a minor cause of Usher syndrome type 2, and the asso... more In contrast to USH2A, variants in ADGRV1 are a minor cause of Usher syndrome type 2, and the associated phenotype is less known. The purpose of the study was to characterize the retinal phenotype of 18 ADGRV1 patients (9 male, 9 female; median age 52 years) and compare it with that of 204 USH2A patients (111 male, 93 female; median age 43 years) in terms of nyctalopia onset, best corrected visual acuity (BCVA), fundus autofluorescence (FAF), and optical coherence tomography (OCT) features. There was no statistical difference in the median age at onset (30 and 18 years; Mann–Whitney U test, p = 0.13); the mean age when 50% of the patients reached legal blindness (≥1.0 log MAR) based on visual acuity (64 years for both groups; log-rank, p = 0.3); the risk of developing advanced retinal degeneration (patch or atrophy) with age (multiple logistic regression, p = 0.8); or the frequency of cystoid macular edema (31% vs. 26%, Fisher’s exact test, p = 0.4). ADGRV1 and USH2A retinopathy were...
Mutations in rhodopsin gene (RHO) are a frequent cause of retinitis pigmentosa (RP) and less ofte... more Mutations in rhodopsin gene (RHO) are a frequent cause of retinitis pigmentosa (RP) and less often, congenital stationary night blindness (CSNB). Mutation p.G90D has previously been associated with CSNB based on the examination of one family. This study screened 60 patients. Out of these 60 patients, 32 were affected and a full characterization was conducted in 15 patients. We described the clinical characteristics of these 15 patients (12 male, median age 42 years, range 8–71) from three families including visual field (Campus Goldmann), fundus autofluorescence (FAF), optical coherence tomography (OCT) and electrophysiology. Phenotypes were classified into four categories: CSNB (N = 3, 20%) sector RP (N = 3, 20%), pericentral RP (N = 1, 6.7%) and classic RP (N = 8, 53.3% (8/15)). The phenotypes were not associated with family, sex or age (Kruskal–Wallis, p > 0.05), however, cystoid macular edema (CME) was observed only in one family. Among the subjects reporting nyctalopia, 69% ...
Purpose: to determine a detailed clinical and haplotypic variability of the Slovenian USH2A patie... more Purpose: to determine a detailed clinical and haplotypic variability of the Slovenian USH2A patients with homozygous c.11864G>A (p.Trp3955Ter) nonsense mutation and to develop sensitive, accurate and rapid screening test. Methods: Ten unrelated homozygous patients with detailed ophthalmological exam were included in our study. The High-Resolution Melting (HRM) method was developed for fast and reliable detection of the c.11864G>A mutation. Results: The c.11864G>A mutation represents the vast majority of pathogenic alleles in Slovenian USH2A-Usher syndrome population (84%). The median age of onset of nyctalopia was 16 years and all patients younger than 40 years had hyperautofluorescent rings on fundus autofluorescence imaging. The Kaplan Meier survival analysis showed a decline of central vision after the age of 40, with 50% patients reaching visual acuity (VA) ≤ 0.05 at the average age of 66 years visual field diameter less than 20° at the average age of 59 years. There wa...
Rejection is the leading cause of failure of limbal allogafts. Resident dendritic cell (DC) matur... more Rejection is the leading cause of failure of limbal allogafts. Resident dendritic cell (DC) maturation plays a critical role in host allosensitization. There are two lineages: myeloid (mDC) and lymphoid (pDC), with different biological properties. The aim was to analyse the distribution of DC subtypes in limbal explant cultures on amniotic membrane (AM), cultivated on either the epithelial or stromal side and to compare the results with directly isolated cells from cadaveric whole corneoscleral tissue divided into specific areas. The expression of CD11c (mDC), CD303/CD123 (pDC) and costimulatory molecules CD80, CD86 and activation markers HLA-DR, CD83 was investigated by flow cytometry. Additionally, the corneal epithelium marker CK12 and ABCB5, a new epithelial stem cell marker, were investigated. Cells positive for pDC and mDC markers were found in all examined areas, with a nonsignificant prevalence of pDC. In limbal explant cultures on AM, the percentage of pDC and mDC was simil...
The aim of this study is to report on the results of color vision testing in a European cohort of... more 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. 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. 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 di...
To determine the spectrum of BEST1 mutations and to study the phenotype in Slovenian families wit... more To determine the spectrum of BEST1 mutations and to study the phenotype in Slovenian families with Best vitelliform macular dystrophy (BVMD) to identify genotype-phenotype correlations. Twenty patients from five families underwent the ophthalmological examination including electrooculogram (EOG; N = 17), fundus autofluorescence imaging (N = 16) and optical coherence tomography (N = 14). Mutational screening was performed by direct DNA sequencing of the BEST1 gene. Mutation c.43G>C (p.Gly15Arg) was detected in three patients from family M presenting with different clinical stages of Best disease. Mutation c.313G>C (p.Arg105Gly) was found in families K, ST, S, B and was associated with incomplete clinical penetrance and variable retinal changes, including extramacular and multifocal lesions. In three patients from family K, an atypical form of BVMD was observed; there were additional peripheral lesions outside of the vascular arcades in addition to the typical macular lesions. Multiple alterations between the vitelliruptive and pseudohypopyon stages over a period of 11 years were seen in one patient. Two previously unreported disease-associated variants in the BEST1 gene (p.Gly15Arg and p.Arg105Gly) were found in Slovenian patients with Best disease. Our data expand the mutation spectrum of the BEST1 gene and further support the broad phenotypic variability observed clinically and with optical coherence tomography (OCT) and AF imaging.
Background: Pars plana vitrectomy (PPV) is preferred surgical procedure for the management of com... more Background: Pars plana vitrectomy (PPV) is preferred surgical procedure for the management of complex rhegmatogenous retinal detachment (RRD). The purpose of this study was to evaluate the anatomical results of primary PPV for the treatment of primary complex RRD and to determine the influence of lens status, tamponading agent, preoperative proliferative vitreoretinopathy (PVR) and axial length (AL) of the eye upon the anatomical outcome. Methods: A retrospective consecutive chart analysis was performed on 117 eyes from 117 patients with complex RRD managed with PPV. Fifty-nine eyes were phakic and 58 pseudophakic eyes. All patients had a minimum follow-up period of 12 months. Eyes were classified into groups using independent variables (first classification based upon lens status and tamponade used, second classification based upon lens and PVR status and third classification based upon AL of the eye). The groups were compared for anatomical outcomes (dependent variables) using nonparametric-or, in case of normally distributed data, parametricstatistical tests. Results: Retinal reattachment rate in phakic eyes was 94.9% compared to 93.1% in pseudophakic, with no statistically significant difference between the two. The overall retinal reattachment rate with single surgery was 94.0%. Final reattachment rate was 97.4%. In case of established PVR ≥ C1, the reattachment rate was not statistically different (92.6%) from eyes with no PVR (91.1%) irrespective of lens status. A statistically significant difference was found between redetachment rates only between phakic eyes with gas tamponade compared to silicon oil (SO) (p = 0.001). Reattachment rate proved to be similar in both AL groups (≤24 mm and > 24 mm). Conclusions: High anatomical success rate of primary vitrectomy for complex RRD with either gas or SO tamponade was achieved in phakic as well as pseudophakic eyes irrespective of AL of the eye.
Inducing selective or targeted cell apoptosis without affecting large number of neighbouring cell... more Inducing selective or targeted cell apoptosis without affecting large number of neighbouring cells remains a challenge. A plausible method for treatment of posterior capsular opacification (PCO) due to remaining lens epithelial cells (LECs) by reactive chemistry induced by localized single electrode microplasma discharge at top of a needle-like glass electrode with spot size~3 μm is hereby presented. The focused and highly-localized atmospheric pressure microplasma jet with electrode discharge could induce a dose-dependent apoptosis in selected and targeted individual LECs, which could be confirmed by real-time monitoring of the morphological and structural changes at cellular level. Direct cell treatment with microplasma inside the medium appeared more effective in inducing apoptosis (caspase 8 positivity and DNA fragmentation) at a highly targeted cell level compared to treatment on top of the medium (indirect treatment). Our results show that single cell specific micropipette plasma can be used to selectively induce demise in LECs which remain in the capsular bag after cataract surgery and thus prevent their migration (CXCR4 positivity) to the posterior lens capsule and PCO formation.
To evaluate the effect of prolonged limbal explants cultured without any scaffolds or on amniotic... more To evaluate the effect of prolonged limbal explants cultured without any scaffolds or on amniotic membrane (AM) on the viability, proliferation and differentiation potential of putative phenotypically defined cultured limbal mesenchymal (LMSC) and epithelial stem cells (LESC). Limbal explants were cultivated on cryopreserved intact AM or plastic plates using medium supplemented with only human serum. AM was positioned with either the epithelial or stromal side up. The outgrowing cells were immunophenotyped for the co-expression of mesenchymal stem cell markers (CD73/CD90/CD105 positive and CD45 negative), proliferation and putative progenitor markers (CXCR4, CD117), epithelial markers and antigen presenting cell markers (CD80, CD83, CD86) by flow cytometry. Immunohistochemistry on limbal cultures cultivated on AM was carried out with antibodies against pan-cytokeratin, p63, Ki67. Morphological and immunostaining analyses revealed two distinct stem cell population types, which could ...
The 12th Congress of the European Neuro-Ophthalmology Society (EUNOS) met in Ljubljana, Slovenia,... more The 12th Congress of the European Neuro-Ophthalmology Society (EUNOS) met in Ljubljana, Slovenia, 21–24 June 2015 at the Cankarjev Dom Cultural and Congress Centre.
Usher syndrome (USH), the most prevalent cause of hereditary deafness-blindness, is an autosomal ... more Usher syndrome (USH), the most prevalent cause of hereditary deafness-blindness, is an autosomal recessive and genetically heterogeneous disorder. Three clinical subtypes (USH1-3) are distinguishable based on the severity of the sensorineural hearing impairment, the presence or absence of vestibular dysfunction, and the age of onset of the retinitis pigmentosa. A total of 10 causal genes, 6 for USH1, 3 for USH2, and 1 for USH3, and an USH2 modifier gene, have been identified. A robust molecular diagnosis is required not only to improve genetic counseling, but also to advance gene therapy in USH patients. Here, we present an improved diagnostic strategy that is both cost-and time-effective. It relies on the sequential use of three different techniques to analyze selected genomic regions: targeted exome sequencing, comparative genome hybridization, and quantitative exon amplification. We screened a large cohort of 427 patients (139 USH1, 282 USH2, and six of undefined clinical subtype) from various European medical centers for mutations in all USH genes and the modifier gene. We identified a total of 421 different sequence variants predicted to be pathogenic, about half of which had not been previously reported. Remarkably, we detected large genomic rearrangements, most of which were novel and unique, in 9% of the patients. Thus, our strategy led to the identification of biallelic and monoallelic mutations in 92.7% and 5.8% of the USH patients, respectively. With an overall 98.5% mutation characterization rate, the diagnosis efficiency was substantially improved compared with previously reported methods.
Platelets are thought to be involved in the pathogenesis of diabetic retinopathy. The BglII gene ... more Platelets are thought to be involved in the pathogenesis of diabetic retinopathy. The BglII gene polymorphism of the α2β1 integrin, which is a platelet collagen receptor, has been suggested as a genetic risk factor for diabetic retinopathy in Japanese subjects. The aim of this study was to look for a relationship between the BglII gene polymorphism of the α2β1 integrin
Vitamin A is an essential fat-soluble vitamin that occurs in various chemical forms. It is essent... more Vitamin A is an essential fat-soluble vitamin that occurs in various chemical forms. It is essential for several physiological processes. Either hyper- or hypovitaminosis can be harmful. One of the most important vitamin A functions is its involvement in visual phototransduction, where it serves as the crucial part of photopigment, the first molecule in the process of transforming photons of light into electrical signals. In this process, large quantities of vitamin A in the form of 11-cis-retinal are being isomerized to all-trans-retinal and then quickly recycled back to 11-cis-retinal. Complex machinery of transporters and enzymes is involved in this process (i.e., the visual cycle). Any fault in the machinery may not only reduce the efficiency of visual detection but also cause the accumulation of toxic chemicals in the retina. This review provides a comprehensive overview of diseases that are directly or indirectly connected with vitamin A pathways in the retina. It includes the...
USH2A mutation is the most common cause of retinitis pigmentosa, with or without hearing impairme... more USH2A mutation is the most common cause of retinitis pigmentosa, with or without hearing impairment. Patients most commonly exhibit hyperautofluorescent ring on fundus autofluorescence imaging (FAF) and rod-cone dystrophy on electrophysiology. A detailed study of three USH2A patients with a rare pattern of double hyperautofluorescent rings was performed. Twenty-four patients with typical single hyperautofluorescent rings were used for comparison of the ages of onset, visual fields, optical coherence tomography, electrophysiology, and audiograms. Double rings delineated the area of pericentral retinal degeneration in all cases. Two patients exhibited rod-cone dystrophy, whereas the third had a cone-rod dystrophy type of dysfunction on electrophysiology. There was minimal progression on follow-up in all three. Patients with double rings had significantly better visual acuity, cone function, and auditory performance than the single ring group. Double rings were associated with combinat...
Gelsolin amyloidosis typically presents with corneal lattice dystrophy and is most frequently ass... more Gelsolin amyloidosis typically presents with corneal lattice dystrophy and is most frequently associated with pathogenic GSN variant p.Asp214Asn. Here we report clinical and histopathological features of gelsolin amyloidosis associated with a novel GSN variant p.Glu580Lys. We studied DNA samples of seven members of a two-generation family. Exome sequencing was performed in the proband, and targeted Sanger sequencing in the others. The heterozygous GSN variant p.Glu580Lys was identified in six patients. The patients exhibited corneal dystrophy (5/6), loose skin (5/6) and/or heart arrhythmia (3/6) and one presented with bilateral optic neuropathy. The impact of the mutation on the protein structure was evaluated in silico. The substitution is located in the fifth domain of gelsolin protein, homologous to the second domain harboring the most common pathogenic variant p.Asp214Asn. Structural investigation revealed that the mutation might affect protein folding. Histopathological analysi...
Mutations in RPGRORF15 are associated with rod-cone or cone/cone-rod dystrophy, the latter associ... more Mutations in RPGRORF15 are associated with rod-cone or cone/cone-rod dystrophy, the latter associated with mutations at the distal end. We describe the phenotype associated with a novel variant in the terminal codon of the RPGRORF15 c.3457T>A (Ter1153Lysext*38), which results in a C-terminal extension. Three male patients from two families were recruited, aged 31, 35, and 38 years. Genetic testing was performed by whole exome sequencing. Filtered variants were analysed according to the population frequency, ClinVar database, the variant’s putative impact, and predicted pathogenicity; and were classified according to the ACMG guidelines. Examination included visual acuity (Snellen), colour vision (Ishihara), visual field, fundus autofluorescence (FAF), optical coherence tomography (OCT), and electrophysiology. All patients were myopic, and had central scotoma and reduced colour vision. Visual acuities on better eyes were counting fingers, 0.3 and 0.05. Electrophysiology showed sev...
In contrast to USH2A, variants in ADGRV1 are a minor cause of Usher syndrome type 2, and the asso... more In contrast to USH2A, variants in ADGRV1 are a minor cause of Usher syndrome type 2, and the associated phenotype is less known. The purpose of the study was to characterize the retinal phenotype of 18 ADGRV1 patients (9 male, 9 female; median age 52 years) and compare it with that of 204 USH2A patients (111 male, 93 female; median age 43 years) in terms of nyctalopia onset, best corrected visual acuity (BCVA), fundus autofluorescence (FAF), and optical coherence tomography (OCT) features. There was no statistical difference in the median age at onset (30 and 18 years; Mann–Whitney U test, p = 0.13); the mean age when 50% of the patients reached legal blindness (≥1.0 log MAR) based on visual acuity (64 years for both groups; log-rank, p = 0.3); the risk of developing advanced retinal degeneration (patch or atrophy) with age (multiple logistic regression, p = 0.8); or the frequency of cystoid macular edema (31% vs. 26%, Fisher’s exact test, p = 0.4). ADGRV1 and USH2A retinopathy were...
Mutations in rhodopsin gene (RHO) are a frequent cause of retinitis pigmentosa (RP) and less ofte... more Mutations in rhodopsin gene (RHO) are a frequent cause of retinitis pigmentosa (RP) and less often, congenital stationary night blindness (CSNB). Mutation p.G90D has previously been associated with CSNB based on the examination of one family. This study screened 60 patients. Out of these 60 patients, 32 were affected and a full characterization was conducted in 15 patients. We described the clinical characteristics of these 15 patients (12 male, median age 42 years, range 8–71) from three families including visual field (Campus Goldmann), fundus autofluorescence (FAF), optical coherence tomography (OCT) and electrophysiology. Phenotypes were classified into four categories: CSNB (N = 3, 20%) sector RP (N = 3, 20%), pericentral RP (N = 1, 6.7%) and classic RP (N = 8, 53.3% (8/15)). The phenotypes were not associated with family, sex or age (Kruskal–Wallis, p > 0.05), however, cystoid macular edema (CME) was observed only in one family. Among the subjects reporting nyctalopia, 69% ...
Purpose: to determine a detailed clinical and haplotypic variability of the Slovenian USH2A patie... more Purpose: to determine a detailed clinical and haplotypic variability of the Slovenian USH2A patients with homozygous c.11864G>A (p.Trp3955Ter) nonsense mutation and to develop sensitive, accurate and rapid screening test. Methods: Ten unrelated homozygous patients with detailed ophthalmological exam were included in our study. The High-Resolution Melting (HRM) method was developed for fast and reliable detection of the c.11864G>A mutation. Results: The c.11864G>A mutation represents the vast majority of pathogenic alleles in Slovenian USH2A-Usher syndrome population (84%). The median age of onset of nyctalopia was 16 years and all patients younger than 40 years had hyperautofluorescent rings on fundus autofluorescence imaging. The Kaplan Meier survival analysis showed a decline of central vision after the age of 40, with 50% patients reaching visual acuity (VA) ≤ 0.05 at the average age of 66 years visual field diameter less than 20° at the average age of 59 years. There wa...
Rejection is the leading cause of failure of limbal allogafts. Resident dendritic cell (DC) matur... more Rejection is the leading cause of failure of limbal allogafts. Resident dendritic cell (DC) maturation plays a critical role in host allosensitization. There are two lineages: myeloid (mDC) and lymphoid (pDC), with different biological properties. The aim was to analyse the distribution of DC subtypes in limbal explant cultures on amniotic membrane (AM), cultivated on either the epithelial or stromal side and to compare the results with directly isolated cells from cadaveric whole corneoscleral tissue divided into specific areas. The expression of CD11c (mDC), CD303/CD123 (pDC) and costimulatory molecules CD80, CD86 and activation markers HLA-DR, CD83 was investigated by flow cytometry. Additionally, the corneal epithelium marker CK12 and ABCB5, a new epithelial stem cell marker, were investigated. Cells positive for pDC and mDC markers were found in all examined areas, with a nonsignificant prevalence of pDC. In limbal explant cultures on AM, the percentage of pDC and mDC was simil...
The aim of this study is to report on the results of color vision testing in a European cohort of... more 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. 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. 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 di...
To determine the spectrum of BEST1 mutations and to study the phenotype in Slovenian families wit... more To determine the spectrum of BEST1 mutations and to study the phenotype in Slovenian families with Best vitelliform macular dystrophy (BVMD) to identify genotype-phenotype correlations. Twenty patients from five families underwent the ophthalmological examination including electrooculogram (EOG; N = 17), fundus autofluorescence imaging (N = 16) and optical coherence tomography (N = 14). Mutational screening was performed by direct DNA sequencing of the BEST1 gene. Mutation c.43G>C (p.Gly15Arg) was detected in three patients from family M presenting with different clinical stages of Best disease. Mutation c.313G>C (p.Arg105Gly) was found in families K, ST, S, B and was associated with incomplete clinical penetrance and variable retinal changes, including extramacular and multifocal lesions. In three patients from family K, an atypical form of BVMD was observed; there were additional peripheral lesions outside of the vascular arcades in addition to the typical macular lesions. Multiple alterations between the vitelliruptive and pseudohypopyon stages over a period of 11 years were seen in one patient. Two previously unreported disease-associated variants in the BEST1 gene (p.Gly15Arg and p.Arg105Gly) were found in Slovenian patients with Best disease. Our data expand the mutation spectrum of the BEST1 gene and further support the broad phenotypic variability observed clinically and with optical coherence tomography (OCT) and AF imaging.
Background: Pars plana vitrectomy (PPV) is preferred surgical procedure for the management of com... more Background: Pars plana vitrectomy (PPV) is preferred surgical procedure for the management of complex rhegmatogenous retinal detachment (RRD). The purpose of this study was to evaluate the anatomical results of primary PPV for the treatment of primary complex RRD and to determine the influence of lens status, tamponading agent, preoperative proliferative vitreoretinopathy (PVR) and axial length (AL) of the eye upon the anatomical outcome. Methods: A retrospective consecutive chart analysis was performed on 117 eyes from 117 patients with complex RRD managed with PPV. Fifty-nine eyes were phakic and 58 pseudophakic eyes. All patients had a minimum follow-up period of 12 months. Eyes were classified into groups using independent variables (first classification based upon lens status and tamponade used, second classification based upon lens and PVR status and third classification based upon AL of the eye). The groups were compared for anatomical outcomes (dependent variables) using nonparametric-or, in case of normally distributed data, parametricstatistical tests. Results: Retinal reattachment rate in phakic eyes was 94.9% compared to 93.1% in pseudophakic, with no statistically significant difference between the two. The overall retinal reattachment rate with single surgery was 94.0%. Final reattachment rate was 97.4%. In case of established PVR ≥ C1, the reattachment rate was not statistically different (92.6%) from eyes with no PVR (91.1%) irrespective of lens status. A statistically significant difference was found between redetachment rates only between phakic eyes with gas tamponade compared to silicon oil (SO) (p = 0.001). Reattachment rate proved to be similar in both AL groups (≤24 mm and > 24 mm). Conclusions: High anatomical success rate of primary vitrectomy for complex RRD with either gas or SO tamponade was achieved in phakic as well as pseudophakic eyes irrespective of AL of the eye.
Inducing selective or targeted cell apoptosis without affecting large number of neighbouring cell... more Inducing selective or targeted cell apoptosis without affecting large number of neighbouring cells remains a challenge. A plausible method for treatment of posterior capsular opacification (PCO) due to remaining lens epithelial cells (LECs) by reactive chemistry induced by localized single electrode microplasma discharge at top of a needle-like glass electrode with spot size~3 μm is hereby presented. The focused and highly-localized atmospheric pressure microplasma jet with electrode discharge could induce a dose-dependent apoptosis in selected and targeted individual LECs, which could be confirmed by real-time monitoring of the morphological and structural changes at cellular level. Direct cell treatment with microplasma inside the medium appeared more effective in inducing apoptosis (caspase 8 positivity and DNA fragmentation) at a highly targeted cell level compared to treatment on top of the medium (indirect treatment). Our results show that single cell specific micropipette plasma can be used to selectively induce demise in LECs which remain in the capsular bag after cataract surgery and thus prevent their migration (CXCR4 positivity) to the posterior lens capsule and PCO formation.
To evaluate the effect of prolonged limbal explants cultured without any scaffolds or on amniotic... more To evaluate the effect of prolonged limbal explants cultured without any scaffolds or on amniotic membrane (AM) on the viability, proliferation and differentiation potential of putative phenotypically defined cultured limbal mesenchymal (LMSC) and epithelial stem cells (LESC). Limbal explants were cultivated on cryopreserved intact AM or plastic plates using medium supplemented with only human serum. AM was positioned with either the epithelial or stromal side up. The outgrowing cells were immunophenotyped for the co-expression of mesenchymal stem cell markers (CD73/CD90/CD105 positive and CD45 negative), proliferation and putative progenitor markers (CXCR4, CD117), epithelial markers and antigen presenting cell markers (CD80, CD83, CD86) by flow cytometry. Immunohistochemistry on limbal cultures cultivated on AM was carried out with antibodies against pan-cytokeratin, p63, Ki67. Morphological and immunostaining analyses revealed two distinct stem cell population types, which could ...
The 12th Congress of the European Neuro-Ophthalmology Society (EUNOS) met in Ljubljana, Slovenia,... more The 12th Congress of the European Neuro-Ophthalmology Society (EUNOS) met in Ljubljana, Slovenia, 21–24 June 2015 at the Cankarjev Dom Cultural and Congress Centre.
Usher syndrome (USH), the most prevalent cause of hereditary deafness-blindness, is an autosomal ... more Usher syndrome (USH), the most prevalent cause of hereditary deafness-blindness, is an autosomal recessive and genetically heterogeneous disorder. Three clinical subtypes (USH1-3) are distinguishable based on the severity of the sensorineural hearing impairment, the presence or absence of vestibular dysfunction, and the age of onset of the retinitis pigmentosa. A total of 10 causal genes, 6 for USH1, 3 for USH2, and 1 for USH3, and an USH2 modifier gene, have been identified. A robust molecular diagnosis is required not only to improve genetic counseling, but also to advance gene therapy in USH patients. Here, we present an improved diagnostic strategy that is both cost-and time-effective. It relies on the sequential use of three different techniques to analyze selected genomic regions: targeted exome sequencing, comparative genome hybridization, and quantitative exon amplification. We screened a large cohort of 427 patients (139 USH1, 282 USH2, and six of undefined clinical subtype) from various European medical centers for mutations in all USH genes and the modifier gene. We identified a total of 421 different sequence variants predicted to be pathogenic, about half of which had not been previously reported. Remarkably, we detected large genomic rearrangements, most of which were novel and unique, in 9% of the patients. Thus, our strategy led to the identification of biallelic and monoallelic mutations in 92.7% and 5.8% of the USH patients, respectively. With an overall 98.5% mutation characterization rate, the diagnosis efficiency was substantially improved compared with previously reported methods.
Platelets are thought to be involved in the pathogenesis of diabetic retinopathy. The BglII gene ... more Platelets are thought to be involved in the pathogenesis of diabetic retinopathy. The BglII gene polymorphism of the α2β1 integrin, which is a platelet collagen receptor, has been suggested as a genetic risk factor for diabetic retinopathy in Japanese subjects. The aim of this study was to look for a relationship between the BglII gene polymorphism of the α2β1 integrin
Uploads
Papers by Marko Hawlina