Conditional knockout of hephaestin in the neural retina disrupts retinal iron homeostasis.

Iron accumulation has been implicated in degenerative retinal diseases. It can catalyze the production of damaging reactive oxygen species. Previous work has demonstrated iron accumulation in multiple retinal diseases, including age-related macular degeneration and diabetic retinopathy. In mice, systemic knockout of the ferroxidases ceruloplasmin (Cp) and hephaestin (Heph), which oxidize iron, results in retinal iron accumulation and iron-induced degeneration. To determine the role of Heph in the retina, we generated a neural retina-specific Heph knockout on a background of systemic Cp knockout. This resulted in elevated neural retina iron. Conversely, retinal ganglion cells had elevated transferrin receptor and decreased ferritin, suggesting diminished iron levels. The retinal degeneration observed in systemic Cp-/-, Heph-/- mice did not occur. These findings indicate that Heph has a local role in regulating neural retina iron homeostasis, but also suggest that preserved Heph function in either the RPE or systemically mitigates the degeneration phenotype observed in the systemic Cp-/-, Heph-/- mice.

Longitudinal Changes in Adult Bony Orbital Volume.

PURPOSE: To our knowledge, no prior study has measured bony orbital volume in a group of subjects over time. This study evaluates longitudinal changes in bony orbital volume with age. METHODS: A search was created for patients with digitized CT imaging studies of the orbit at least 8 years apart. Charts with a history of prior head trauma, head and neck tumors, sinus disease, head surgery, elevated intracranial pressure, thyroid eye disease, or conditions that could affect bone metabolism were excluded. Three outcome measures were used: orbital volume, medial orbital wall length, and lateral orbital wall length. Categorical data were summarized with frequency (%); normally distributed continuous data are summarized with mean (standard deviation), and non-normally distributed data were summarized with median. Normality was verified with Shapiro-Wilk Test. Paired t-tests were used to analyze the differences in the 3 outcome variables. RESULTS: One hundred and eighty-two subjects, 91 males and 91 females, were included. The median time between CT scans was 9.4 years. Orbital volume was significantly larger at the second scan than the first (p < 0.001). The average difference in orbital volume was 0.91 ml, an increase of 4.1%. Lateral orbital length was significantly shorter in the second scan than in the first (p = 0.046) though the difference was only 0.4 mm. There was no statistically significant change in medial orbital wall length. CONCLUSIONS: Bony orbital volume increases in individuals with age. This increase in orbital volume occurs in the context of soft tissue changes to contribute to the aging appearance of the periorbital region.

Comparison of Orbital Volume in Young Versus Senescent Human Skulls.

PURPOSE: To compare bony orbital volumes in young skulls with those of older skulls to elucidate aging-associated changes of the orbit. METHODS: One hundred Caucasian male skulls from the Hamann-Todd collection of the Cleveland Museum of Natural History were studied. There were 50 young skulls (age range, 19-33 years) and 50 senescent skulls (age range, 79-96 years). Volcanic sand was used to fill each orbit in an identical fashion and weighed as a proxy for volume. Digital calipers were used to perform linear measurements of the orbit. The relationship between orbit measures and skull size was assessed using Pearson's correlations and 95% CI, and statistical models to compare age groups adjusted for skull size. RESULTS: The volume of the orbits (P < 0.001), the horizontal diameter of the orbit (P = 0.015), and the orbital depth (P < 0.001) were significantly larger in the senescent group of skulls after adjusting for skull surface area. No significant differences were found in the vertical diameters of the orbit between the 2 groups. Skull size did not statistically differ between the age groups. CONCLUSIONS: Increases in the depth and horizontal dimensions of the orbit lead to increasing bony orbital volume with increasing age. These changes in size and shape of the orbit with age may contribute to phenotypic changes of aging and may affect disease processes and management.

Temporal Fat Pad Volume in Patients With Thyroid Eye Disease.

PURPOSE: To determine whether thyroid eye disease (TED) is associated with a change in temporal fossa soft tissue volume. METHODS: CT imaging studies were selected from patients with TED considering orbital decompression surgery and from an age-matched group of controls presenting to the Emergency Department for nontraumatic conditions requiring CT imaging. Measurements of the temporal fossa fat pad and soft tissue thickness were used as a proxy for volume and were performed using the zygomaticotemporal suture as a reference point. Categorical variables were described using frequencies and percentages, while continuous variables were described using medians and standard deviations. Chi-square tests were used to assess the relationship between gender and age group, while t-tests were used to examine the relationship between continuous variables and age group. RESULTS: A total of 56 CT scans were evaluated, including scans from 28 patients with TED and 28 controls. The following measurements were found to be significantly larger in the TED population: left fat pad measured 5 mm above the zygomaticotemporal suture (p = 0.012), right fat pad measured 15 mm above the suture (p = 0.005), right soft tissue measured 15 mm above the suture (p = 0.026), left fat pad measured 15 mm above the suture (p = 0.006), and the left soft tissue measure 15 mm above the suture (p = 0.032). CONCLUSIONS: Thyroid eye disease is associated with an increase in temporal fossa fat pad and soft tissue volume. These findings suggest that TED disease may produce pan-facial changes rather than changes confined to the peri-orbital region.

Anatomic Variations of the Infraorbital Foramen in Caucasian Versus African American Skulls.

PURPOSE: The infraorbital foramen (IOF) represents a highly conserved structure but demonstrates morphologic variability. The purpose of this study is to describe the IOF location, size, and supernumerary foramina in an African American population and compare it with a Caucasian population. METHODS: Sixty African American and 60 Caucasian skulls from the Hamann-Todd collection of the Cleveland Museum of Natural History were studied. The primary outcome was the number of accessory IOF and measurements of the location, size, shape, and direction of each foramen. Pearson chi-square, t tests, Fisher exact test, and Wilcoxon rank sum tests were used to analyze the data. RESULTS: The African American population had a smaller vertical IOF diameter (mean = 2.81 mm) compared with the Caucasian population (mean = 3.08 mm) on the right side (p < 0.01). The distance from the IOF to the anterior nasal spine on the left side was greater in the African American population (mean = 33.93 mm) compared with the Caucasian population (Caucasian mean = 32.84 mm, p = 0.03). The distance from the IOF to the zygomaticomaxillary suture was significantly shorter in the African American population (mean = 11.85 mm) compared with the Caucasian population (mean = 13.21 mm) on the left side (p = 0.01). Accessory foramina were found in 13 Caucasian skulls (21.7%) and 6 African American skulls (10%; p = 0.08). Two distinct types of IOF existed in each population, one close to the main foramen and one within the sutura notha. CONCLUSIONS: The IOF is smaller and exits more laterally, with a lower proportion of accessory foramina in an African American population compared with a Caucasian population. Both groups exhibit 2 distinct types of IOF. These morphologic differences should be noted during surgeries and anesthetic planning to avoid neurovascular complications.

Oxidative stress induces lysosomal membrane permeabilization and ceramide accumulation in retinal pigment epithelial cells.

Oxidative stress has been implicated in the pathogenesis of age-related macular degeneration, the leading cause of blindness in older adults, with retinal pigment epithelium (RPE) cells playing a key role. To better understand the cytotoxic mechanisms underlying oxidative stress, we used cell culture and mouse models of iron overload, as iron can catalyze reactive oxygen species formation in the RPE. Iron-loading of cultured induced pluripotent stem cell-derived RPE cells increased lysosomal abundance, impaired proteolysis and reduced the activity of a subset of lysosomal enzymes, including lysosomal acid lipase (LIPA) and acid sphingomyelinase (SMPD1). In a liver-specific Hepc (Hamp) knockout murine model of systemic iron overload, RPE cells accumulated lipid peroxidation adducts and lysosomes, developed progressive hypertrophy and underwent cell death. Proteomic and lipidomic analyses revealed accumulation of lysosomal proteins, ceramide biosynthetic enzymes and ceramides. The proteolytic enzyme cathepsin D (CTSD) had impaired maturation. A large proportion of lysosomes were galectin-3 (Lgals3) positive, suggesting cytotoxic lysosomal membrane permeabilization. Collectively, these results demonstrate that iron overload induces lysosomal accumulation and impairs lysosomal function, likely due to iron-induced lipid peroxides that can inhibit lysosomal enzymes.

Associations between arsenic (+3 oxidation state) methyltransferase (AS3MT) and N-6 adenine-specific DNA methyltransferase 1 (N6AMT1) polymorphisms, arsenic metabolism, and cancer risk in a chilean population.

Inter-individual differences in arsenic metabolism have been linked to arsenic-related disease risks. Arsenic (+3) methyltransferase (AS3MT) is the primary enzyme involved in arsenic metabolism, and we previously demonstrated in vitro that N-6 adenine-specific DNA methyltransferase 1 (N6AMT1) also methylates the toxic inorganic arsenic (iAs) metabolite, monomethylarsonous acid (MMA), to the less toxic dimethylarsonic acid (DMA). Here, we evaluated whether AS3MT and N6AMT1 gene polymorphisms alter arsenic methylation and impact iAs-related cancer risks. We assessed AS3MT and N6AMT1 polymorphisms and urinary arsenic metabolites (%iAs, %MMA, %DMA) in 722 subjects from an arsenic-cancer case-control study in a uniquely exposed area in northern Chile. Polymorphisms were genotyped using a custom designed multiplex, ligation-dependent probe amplification (MLPA) assay for 6 AS3MT SNPs and 14 tag SNPs in the N6AMT1 gene. We found several AS3MT polymorphisms associated with both urinary arsenic metabolite profiles and cancer risk. For example, compared to wildtypes, individuals carrying minor alleles in AS3MT rs3740393 had lower %MMA (mean difference = -1.9%, 95% CI: -3.3, -0.4), higher %DMA (mean difference = 4.0%, 95% CI: 1.5, 6.5), and lower odds ratios for bladder (OR = 0.3; 95% CI: 0.1-0.6) and lung cancer (OR = 0.6; 95% CI: 0.2-1.1). Evidence of interaction was also observed for both lung and bladder cancer between these polymorphisms and elevated historical arsenic exposures. Clear associations were not seen for N6AMT1. These results are the first to demonstrate a direct association between AS3MT polymorphisms and arsenic-related internal cancer risk. This research could help identify subpopulations that are particularly vulnerable to arsenic-related disease. Environ. Mol. Mutagen. 58:411-422, 2017. © 2017 Wiley Periodicals, Inc.