Expression of Collagen XIII in Tissues of the Thyroid and Orbit With Relevance to Thyroid-Associated Ophthalmopathy.

Purpose: Antibodies against collagen XIII have previously been identified in patients with active thyroid-associated ophthalmopathy (TAO). Although collagen XIII expression has been described in extraocular muscles and orbital fat, its detailed localization in extraocular and thyroid tissues and the connection to autoimmunity for collagen XIII remain unclear. Our objective was to map the potential targets for these antibodies in the tissues of the orbit and thyroid. Methods: We evaluated the expression of collagen XIII in human patient and mouse orbital and thyroid tissues with immunostainings and RT-qPCR using Col13a1-/- mice as negative controls. COL13A1 expression in Graves' disease and goiter thyroid samples was compared with TGF-ß1 and TNF, and these were also studied in human thyroid epithelial cells and fibroblasts. Results: Collagen XIII expression was found in the neuromuscular and myotendinous junctions of extraocular muscles, blood vessels of orbital connective tissue and fat and the thyroid, and in the thyroid epithelium. Thyroid expression was also seen in germinal centers in Graves' disease and in neoplastic epithelium. The expression of COL13A1 in goiter samples correlated with levels of TGF-B1. Upregulation of COL13A1 was reproduced in thyroid epithelial cells treated with TGF-ß1. Conclusions: We mapped the expression of collagen XIII to various locations in the orbit, demonstrated its expression in the pathologies of the Graves' disease thyroid and confirmed the relationship between collagen XIII and TGF-ß1. Altogether, these data add to our understanding of the targets of anti-collagen XIII autoantibodies in TAO.

Morphology of the optic nerve head and factors affecting it in the Northern Finland birth cohort.

PURPOSE: To assess topographic characteristics of the optic nerve head (ONH) and retinal nerve fibre layer (RNFL) and study the effect of ocular and physiological factors on them in a middle-aged population. METHODS: A 1552-person randomised sample from Northern Finland population aged 45-49 was examined. Laser scanning tomography and optical coherency tomography were performed to obtain measurements for ONH and RNFL morphology. Measurements of the RNFL included global thickness and the six zones used in automated structure-function analysis (S-F analysis). Influence of central corneal thickness (CCT), refractive correction, intraocular pressure (IOP), anterior chamber angle, gender, blood pressure, height, weight and body mass index (BMI) on tomographic data was analysed. RESULTS: The optic disc area had a strong correlation with all other parameters of ONH morphology (R = 0.261 to 0.706) as did spherical equivalent (R = -0.280 to 0.280). The correlations between ONH and RNFL measurements were weaker (R = 0.057 to 0.180). Gender, CCT, anterior chamber angle, blood pressure, height and BMI had statistically significant, yet feeble, correlations with a number of ONH parameters. CONCLUSION: Other than spherical equivalent, the studied anatomical and physiological attributes had little predictive value on the ONH morphology. The optic disc area itself had a significant effect on other measurements of ONH tomography and should be taken into consideration when the thresholds for normal ONH morphology are calculated.

Antenatal and neonatal factors and morphology of the optic nerve head in the Northern Finland birth cohort.

PURPOSE: The optic nerve head (ONH) is a part of the brain that can be evaluated through the transparent medium of the eye. The purpose of this study was to explore the possible correlations among the properties of the optic nerve head, maternal factors during pregnancy and neonatal parameters in a randomized sample of a birth cohort. METHODS: The Northern Finland 1966 Birth Cohort has been prospectively monitored since their antenatal period. Data on pregnancy and neonatal period were collected during gestation and right after birth in 1966. A randomized sample of 3070 subjects underwent an ophthalmic assessment at the age of 46-48 years. The examination protocol included scanning laser ophthalmoscopy with the Heidelberg Retina Tomograph. The ophthalmological parameters assessed were the disc area and the neuroretinal rim volume of the ONH. RESULTS: We found that chronic pulmonary disease of the mother (p = 0.007), the number of gestational weeks (p = 0.030) and the mother's highest measured systolic blood pressure (p = 0.035) during pregnancy had a statistically significant effect on the disc area. Smaller disc size was associated with pulmonary disease and early gestation. There was a significant difference in rim volume between genders (p < 0.001). Women had larger neuroretinal rim volumes compared to men. CONCLUSION: In this population-based study, the vast majority of antenatal and neonatal factors showed no correlation with optic disc area or rim volume. Furthermore, even the factors with statistically significant correlation with ONH morphology had limited predictive value.

Optic Nerve Parameters and Cognitive Function in the Northern Finland Birth Cohort Eye Study.

PURPOSE: The optic nerve head (ONH) is a part of the brain that can be easily studied through the transparent medium of the eye. We explored the relationship between the properties of the optic nerve head, the retinal nerve fiber layer (RNFL) and cognitive function. METHODS: Participants of the Northern Finland Birth Cohort (NFBC) 1966 underwent an ophthalmic and cognitive assessment after randomization at age 46. The ophthalmological parameters obtained were the disc area and the neuroretinal rim volume of the ONH and the average RNFL thickness. The surrogates used for cognitive function were the paired associates learning test (PAL), level of education, grade point average (GPA) and Humphrey 24-2 perimetric test time (HFA). We did exploratory research between the ophthalmological parameters and the surrogates for cognition and the correlations between the surrogates for cognition. RESULTS: We found that a larger disc area was associated with a higher level of education, faster accomplishment of the HFA (R = -0.065) but a lower GPA (R = -0.084). An increase in neuroretinal rim volume was associated with fewer errors in the PAL test (R = -0.056), higher level of education, higher GPA (R = 0.072) and faster accomplishment of the HFA (R = -0.047). A thicker RNFL was associated with faster accomplishment of the HFA (R = -0.047). CONCLUSION: We were able to find statistically significant associations between the parameters of the optic nerve head, the RNFL and cognition in the NFBC Eye study. However, the correlations were negligible at best and of limited predictive value.

Comparison of imaging parameters between OCT, GDx and HRT in the Northern Finland birth cohort eye study.

PURPOSE: The purpose of the study was to assess and compare the optic nerve head (ONH) and retinal nerve fibre layer (RNFL) parameters and image quality parameters obtained by Cirrus HD-OCT, GDxECC and HRT3 in a population-based screening study. METHODS: This analysis examined 2566 subjects taking part in the Northern Finland Birth Cohort Eye study. Images with spectral domain OCT (Cirrus HD-OCT 4000), scanning laser ophthalmoscopy (HRT3) and scanning laser polarimetry (GDxECC) were obtained from each subject. The correlation of average and regional parameters of RNFL and ONH between devices was evaluated. RESULTS: The RNFL thickness was 90.9 µm when measured with OCT, 24.6 µm with HRT and 48.1 µm with GDx. There was a high correlation between the disc and cup measurements with the HRT and OCT and the RNFL thickness of the OCT and GDx (r > 0.5). A statistically significant correlation was found between RNFL measurements of the HRT and OCT in the superior, temporal and inferior quadrants. Optical coherence tomography (OCT) signal strength correlated with the image quality parameters of the HRT and GDx. The percentage of good quality images was the lowest with the GDx. CONCLUSION: The RNFL thickness in Northern Finland birth cohort was at a lower level compared to other studies. The study confirms the difference in measuring ONH parameters between the imaging devices. However, significant correlations between devices were found in the cup volume and cup disc area ratio parameters of the OCT and HRT. The correlations between image quality parameters and glaucoma detection parameters were relatively low.

Combined structure-function analysis in glaucoma screening.

AIM: To assess the applicability of a structure-function (S-F) analysis combining spectral-domain optical coherence tomography (SD-OCT) and standard automated perimetry (SAP) in glaucoma screening in a middle-aged population. METHODS: A randomised sample of 3001 Caucasian participants aged 45-49 years of the Northern Finland Birth Cohort Eye Study was examined. We performed an eye examination, including 24-2 SAP, optic nerve head (ONH) and retinal nerve fibre layer (RNFL) photography and SD-OCT of the peripapillary RNFL. The S-F report was generated by Forum Glaucoma Workplace software. OCT, SAP and the S-F analysis were evaluated against clinical glaucoma diagnosis, that is, the positive '2 out of 3' rule based on the clinician's evaluation of ONH and RNFL photographs and visual fields (VFs). RESULTS: At a specificity of 97.5%, the sensitivity for glaucomatous damage was 26% for abnormal OCT, 35% for SAP and 44% for S-F analysis. Estimated areas under the curve were 0.74, 0.85 and 0.76, and the corresponding positive predictive values were 8 %, 10% and 12%, respectively. By applying a classification tree approach combining OCT, SAP and defect localisation data, a sensitivity of 77% was achieved at 90% specificity. In a localisation analysis of glaucomatous structural and functional defects, the correlation with glaucoma increased significantly if the abnormal VF test points were located on borderline or abnormal OCT zones. CONCLUSION: SAP performs slightly better than OCT in glaucoma screening of middle-aged population. However, the diagnostic capability can be improved by S-F analysis.

Reduced evoked activity and cortical oscillations are correlated with anisometric amblyopia and impairment of visual acuity.

Amblyopia is a developmental disorder associated with abnormal visual experience during early childhood commonly arising from strabismus and/or anisometropia and leading to dysfunctions in visual cortex and to various visual deficits. The different forms of neuronal activity that are attenuated in amblyopia have been only partially characterized. In electrophysiological recordings of healthy human brain, the presentation of visual stimuli is associated with event-related activity and oscillatory responses. It has remained poorly understood whether these forms of activity are reduced in amblyopia and whether possible dysfunctions would arise from lower- or higher-order visual areas. We recorded neuronal activity with magnetoencephalography (MEG) from anisometropic amblyopic patients and control participants during two visual tasks presented separately for each eye and estimated neuronal activity from source-reconstructed MEG data. We investigated whether event-related and oscillatory responses would be reduced for amblyopia and localized their cortical sources. Oscillation amplitudes and evoked responses were reduced for stimuli presented to the amblyopic eye in higher-order visual areas and in parietal and prefrontal cortices. Importantly, the reduction of oscillation amplitudes but not that of evoked responses was correlated with decreased visual acuity in amblyopia. These results show that attenuated oscillatory responses are correlated with visual deficits in anisometric amblyopia.

Diagnostic performance of modern imaging instruments in glaucoma screening.

AIM: To evaluate the applicability of imaging devices (spectral-domain optical coherence tomography (Cirrus SD-OCT), scanning laser polarimetry (GDx) and scanning laser ophthalmoscopy (Heidelberg Retinal Tomograph, HRT3)) for glaucoma screening in a middle-aged unselected population. METHODS: Participants of the population-based Northern Finland Birth Cohort Eye Study, aged 45 to 49 years, underwent a comprehensive eye examination including modern imaging with five methods (retinal nerve fibre layer (RNFL) and macular ganglion cell layer +inner plexiform layer (GCIPL) analysis and their combination with SD-OCT, GDx and HRT). The performance of the automated classification of the imaging devices was assessed using a clinical glaucoma diagnosis as reference, that is, the '2 out of 3' rule based on the evaluation of optic nerve head and RNFL photographs and visual fields. RESULTS: We examined 6060 eyes of 3039 subjects; in the clinical evaluation, glaucomatous damage was found in 33 subjects (1.1%) in 43 eyes. The following sensitivities were obtained; RNFL analysis (53%), GCIPL analysis (50%), OCT combination analysis (61%), GDx (56%) and HRT (31%) with corresponding specificities of 95%, 92%, 90%, 88% and 96%. The area under the curve values were 0.76, 0.73, 0.75, 0.75 and 0.73, respectively. Post-test probabilities of glaucoma after positive imaging finding with each of these methods in this unselected population were 11%, 7%, 6%, 5% and 7%, respectively. CONCLUSION: Screening capabilities of the OCT, GDx and HRT were rather similar. The accuracy of all evaluated parameters was only moderate and thus screening with these parameters alone is not reliable.

Novel variants and phenotypes widen the phenotypic spectrum of GABRG2-related disorders.

PURPOSE: Next-generation sequencing (NGS) has made genetic testing of patients with epileptic encephalopathies easier - novel variants are discovered and new phenotypes described. Variants in the same gene - even the same variant - can cause different types of epilepsy and neurodevelopmental disorders. Our aim was to identify the genetic causes of epileptic encephalopathies in paediatric patients with complex phenotypes. METHODS: NGS was carried out for three patients with epileptic encephalopathies. Detailed clinical features, brain magnetic resonance imaging and electroencephalography were analysed. We searched the Human Gene Mutation Database for the published GABRG2 variants with clinical description of patients and composed a summary of the variants and their phenotypic features. RESULTS: We identified two novel de novo GABRG2 variants, p.P282T and p.S306F, with new phenotypes including neuroradiological evidence of neurodegeneration and epilepsy of infancy with migrating focal seizures (EIMFS). One patient carried previously reported p.P83S variant with autism spectrum disorder (ASD) phenotype that has not yet been described related to GABRG2 disorders and a more severe epilepsy phenotype than reported earlier. In all, the literature search yielded twenty-two articles describing 27 different variants that were divided into two categories: those with self-limiting epilepsies and febrile seizures and those with more severe drug-resistant epileptic encephalopathies. CONCLUSION: This study further expands the genotypic and phenotypic spectrum of epilepsies associated with GABRG2 variants. More knowledge is still needed about the influence of the environment, genetic background and other epilepsy susceptibility genes on the phenotype of the specific GABRG2 variants.

Fluoxetine does not enhance the effect of perceptual learning on visual function in adults with amblyopia.

Amblyopia is a common visual disorder that is treatable in childhood. However, therapies have limited efficacy in adult patients with amblyopia. Fluoxetine can reinstate early-life critical period-like neuronal plasticity and has been used to recover functional vision in adult rats with amblyopia. We conducted a Phase 2, randomized (fluoxetine vs. placebo), double-blind, multicenter clinical trial examined whether or not fluoxetine can improve visual acuity in amblyopic adults. This interventional trial included 42 participants diagnosed with moderate to severe amblyopia. Subjects were randomized to receive either 20 mg fluoxetine (n = 22) or placebo (n = 20). During the 10-week treatment period, all subjects performed daily computerized perceptual training and eye patching. At the primary endpoint, the mean treatment group difference in visual acuity improvement was only 0.027 logMAR units (95% CI: -0.057 to 0.110; p = 0.524). However, visual acuity had significantly improved from baseline to 10 weeks in both fluoxetine (-0.167 logMAR; 95% CI: -0.226 to -0.108; p < 0.001) and placebo (-0.194 logMAR; 95% CI: -0.254 to -0.133; p < 0.001) groups. While this study failed to provide evidence that fluoxetine enhances neuroplasticity, our data support other recent clinical studies suggesting that improvement of vision can be accomplished in adults with amblyopia.

Evaluating refraction and visual acuity with the Nidek autorefractometer AR-360A in a randomized population-based screening study.

PURPOSE: The evaluation of visual acuity (VA) and refraction in the Northern Finland Birth Cohort Eye study was performed using the Nidek AR-360A autorefractometer. The accuracy of the method for this population-based screening study was assessed. METHODS: Measurements of the refractive error were obtained from the right eyes of 1238 subjects (mean age 47), first objectively with the AR-360A and then subjectively by an optometrist. Agreement with the subjective refraction was calculated for sphere, cylinder, mean spherical equivalent (MSE), cylindrical vectors J45 and J0 and presbyopic correction (add). Visual acuity (VA) was measured using an ETDRS chart and the autorefractometer. RESULTS: The refractive error measured with the AR-360A was higher than the subjective refraction performed by the optometrist for sphere (0.007 D ± 0.24 D p = 0.30) and also for cylinder (-0.16 D ± 0.20 D p < 0.0005). The bias between the measurements of MSE, J45 and J0 was low: -0.07 D ± 0.22 D (p = 0.002), 0.01 D ± 0.43 D (p = 0.25) and -0.01 D ± 0.42 D (p = 0.43), respectively. The amount of add measured by the autorefractometer was higher than the subjective 0.35 D ± 0.29 D (p < 0.0005). There was a statistically significant correlation between VA (p < 0.0005) and the difference between the subjective and objective refraction. In 99.2% of the measurements, visual values were within one decimal line of each other. CONCLUSION: The Nidek AR-360A autorefractometer is an accurate tool for determining the refraction and VA in a clinical screening trial.

A 6-month study comparing efficacy, safety, and tolerability of the preservative-free fixed combination of tafluprost 0.0015% and timolol 0.5% versus each of its individual preservative-free components.

INTRODUCTION: The efficacy, safety and tolerability of the preservative-free (PF) fixed combination (FC) of tafluprost 0.0015% and timolol 0.5% (once daily) were compared to those of the individual components (PF tafluprost 0.0015% once daily and PF timolol 0.5% twice daily) in patients with open-angle glaucoma or ocular hypertension inadequately controlled on prior timolol or prostaglandin monotherapy for 6 months. METHODS: A stratified, double-masked, randomized, multicenter phase III study was conducted. A total of 189 prior timolol users were randomized within the timolol stratum (TS) to receive either FC (n = 95) or timolol 0.5% (TIM; n = 94). Furthermore, a total of 375 prior prostaglandin analog (PGA) users were randomized within the prostaglandin stratum (PS) to receive either FC (n = 188) or tafluprost 0.0015% (TAF; n = 187). To be eligible for participation in the study, the patients were required to have an intraocular pressure (IOP) of ≥22 mmHg when on timolol (TIM) or of ≥20 mmHg when on PGA in either treated eye at the screening and end-of-run-in visits. In addition to these, the study included visits at baseline, 2 and 6 weeks, 3 and 6 months and at a post-study visit. IOP was measured at 8 a.m., 10 a.m., 4 p.m., and 8 p.m. RESULTS: In the TS, a significant reduction from baseline IOP was seen with FC and TIM throughout the study. Average diurnal IOP change from baseline at month 3 was -8.55 mmHg (32%) for FC and -7.35 mmHg (28%) for TIM. The model-based treatment difference (FC-TIM) was -0.885 mmHg [95% confidence interval (CI) -1.745 to -0.024; p = 0.044] demonstrating the superiority of FC over TIM. In the PS, a significant reduction in IOP was seen with both FC and TAF throughout the study. The average diurnal IOP change from baseline at month 3 was -8.61 mmHg (33%) for FC and -7.23 mmHg (28%) for TAF. The model-based treatment difference (FC-TAF) was -1.516 mmHg (95% CI -2.044 to -0.988; p < 0.001) demonstrating the superiority of FC over TAF. In the TS, related ocular adverse events (AEs) were more frequent for patients treated with FC compared to TIM (16.8% versus 6.4%), whereas related non-ocular AEs were more frequent with TIM compared to FC (2.1% versus 0.0%). In the PS, AEs were similarly distributed between FC and TAF. The frequency of conjunctival hyperemia of FC was low (6.4%). CONCLUSION: The preservative-free fixed combination of tafluprost and timolol provided a substantial and significant IOP reduction in both strata. The IOP reduction was superior to both tafluprost 0.0015% and timolol 0.5% when given as monotherapies. Overall, the study treatments were safe and well tolerated. FUNDING: Santen Oy, Tampere, Finland.

Ang-2 upregulation correlates with increased levels of MMP-9, VEGF, EPO and TGFß1 in diabetic eyes undergoing vitrectomy.

PURPOSE: Angiogenesis in diabetic retinopathy (DR) is a multifactorial process regulated by hypoxia-induced growth factors and inflammatory cytokines. In addition to the angiogenic switch, the proteolytic processing and altered synthesis of the extracellular matrix are critical steps in this disease. This study was performed to evaluate the levels of matrix metalloproteinase-2 and matrix metalloproteinase-9 (MMP-2 and MMP-9), angiopoietin-1 and angiopoietin-2 (Ang-1 and Ang-2), vascular endothelial growth factor (VEGF), erythropoietin (EPO) and transforming growth factor-ß1 (totalTGFß1) in the vitreous of diabetic eyes undergoing vitrectomy compared with control eyes operated because of macular hole or pucker. METHODS: Prospective consecutive controlled observational study performed in the unit of vitreoretinal surgery in Finland during the years 2006-2008. Vitreous samples were collected before the start of the conventional 3-ppp vitrectomy. Vitreous MMP-2 and MMP-9, Ang-1 and Ang-2, VEGF, EPO and TGFß1 concentrations were measured from 69 patients with Type 1 or 2 diabetes and 40 controls. RESULTS: Comparison of eyes with DR with controls revealed that the mean vitreous concentrations of proMMP-2 (p = 0.0015), totalMMP-2 (p = 0.0011), proMMP-9 (p = 0.00001), totalMMP-9 (p < 0.00001), Ang-2 (p < 0.00001), VEGF (p < 0.00001), EPO (p < 0.00001) and totalTGFß1 (p = 0.000026) were significantly higher in the former group. A multivariate logistic regression analysis suggested intravitreal Ang-2 concentration being the key marker of PDR (p = 0.00025) (OR = 1507.9). CONCLUSION: The main new finding is that the intravitreal concentrations of Ang-2 correlated significantly with MMP-9, VEGF, EPO and TGFß1 levels in diabetic eyes undergoing vitrectomy. Thus, these factors could promote retinal angiogenesis synergistically.

Elevated messenger RNA expression and plasma protein levels of osteopontin and matrix metalloproteinase types 2 and 9 in patients with ascending aortic aneurysms.

OBJECTIVE: Ascending aortic aneurysms result from a degenerative process in the aortic wall, characterized by the loss of smooth muscle cells and elastic fibers. We hypothesized that there would be changes in plasma protein and aortic tissue messenger RNA levels of osteopontin, matrix metalloproteinase type 2, matrix metalloproteinase type 9, and tissue inhibitor of matrix metalloproteinases type 1 in ascending aortic aneurysm samples. METHODS: Plasma, aortic tissue, and aortic mRNA samples were collected from patients with an ascending aortic aneurysm or an abdominal aortic aneurysm and from control individuals. Plasma protein levels of osteopontin, matrix metalloproteinase (MMP) types 2 and 9, and tissue inhibitor of matrix metalloproteinases type 1 were determined by quantitative sandwich enzyme-linked immunosorbent assay. Aortic mRNA levels of these same proteins were analyzed with the quantitative real-time polymerase chain reaction (RT-PCR) method and protein levels from the aortic tissues were assayed by immunostaining. Quantitative RT-PCR results were estimated by the normalized expression method (ΔΔCt). RESULTS: Plasma protein levels were significantly elevated for osteopontin, MMP-2, and MMP-9 in the samples of ascending and abdominal aortic aneurysm group compared with controls. Plasma protein levels of MMP-9 were higher in the nonoperated compared with the operated ascending aortic aneurysm group. Aortic osteopontin, MMP-2, and MMP-9 mRNA levels were increased for ascending aortic aneurysm samples. CONCLUSIONS: This study reveals an important role of osteopontin, MMP-2 and MMP-9 in the development of ascending and abdominal aortic aneurysm.

Characterization of metabolic interrelationships and in silico phenotyping of lipoprotein particles using self-organizing maps.

Plasma lipid concentrations cannot properly account for the complex interactions prevailing in lipoprotein (patho)physiology. Sequential ultracentrifugation (UCF) is the gold standard for physical lipoprotein isolations allowing for subsequent analyses of the molecular composition of the particles. Due to labor and cost issues, however, the UCF-based isolations are usually done only for VLDL, LDL, and HDL fractions; sometimes with the addition of intermediate density lipoprotein (IDL) particles and the fractionation of HDL into HDL(2) and HDL(3) (as done here; n = 302). We demonstrate via these data, with the lipoprotein lipid concentration and composition information combined, that the self-organizing map (SOM) analysis reveals a novel data-driven in silico phenotyping of lipoprotein metabolism beyond the experimentally available classifications. The SOM-based findings are biologically consistent with several well-known metabolic characteristics and also explain some apparent contradictions. The novelty is the inherent emergence of complex lipoprotein associations; e.g., the metabolic subgrouping of the associations between plasma LDL cholesterol concentrations and the structural subtypes of LDL particles. Importantly, lipoprotein concentrations cannot pinpoint lipoprotein phenotypes. It would generally be beneficial to computationally enhance the UCF-based lipoprotein data as illustrated here. Particularly, the compositional variations within the lipoprotein particles appear to be a fundamental issue with metabolic and clinical corollaries.