Publications by authors named "Holly Chinnery"

51 Publications

Corneal immune cell morphometry as an indicator of local and systemic pathology: A review.

Clin Exp Ophthalmol 2021 Jul 9. Epub 2021 Jul 9.

Department of Optometry and Vision Sciences, University of Melbourne, Parkville, Victoria, Australia.

The corneal epithelium contains a population of resident immune cells commonly referred to as dendritic cells (DCs), or Langerhans cells. A unique advantage of the transparent cornea being situated at the surface of the eye is that these cells can be readily visualised using in vivo confocal microscopy. Over the past decade, interest in the involvement of corneal DCs in a range of ocular and systemic diseases has surged. For most studies, the number of corneal DCs has been the main outcome of interest. However, more recently attention has shifted towards understanding how DC morphology may provide insights into the inflammatory status of the cornea, and in some cases, the health of the peripheral nervous system. In this review, we provide examples of recent methodologies that have been used to classify and measure corneal DC morphology and discuss how this relates to local and systemic inflammatory conditions in humans and rodents.
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http://dx.doi.org/10.1111/ceo.13972DOI Listing
July 2021

Distribution of Corneal TRPV1 and Its Association With Immune Cells During Homeostasis and Injury.

Invest Ophthalmol Vis Sci 2021 Jul;62(9)

Department of Optometry and Vision Sciences, University of Melbourne, Parkville, Australia.

Purpose: Given the role of corneal sensory nerves during epithelial wound repair, we sought to examine the relationship between immune cells and polymodal nociceptors following corneal injury.

Methods: Young C57BL/6J mice received a 2 mm corneal epithelial injury. One week later, corneal wholemounts were immunostained using β-tubulin-488, TRPV1 (transient receptor potential ion channel subfamily V member-1, a nonselective cation channel) and immune cell (MHC-II, CD45 and CD68) antibodies. The sum length of TRPV1+ and TRPV1- nerve fibers, and their spatial association with immune cells, was quantified in intact and injured corneas.

Results: TRPV1+ nerves account for ∼40% of the nerve fiber length in the intact corneal epithelium and ∼80% in the stroma. In the superficial epithelial layers, TRPV1+ nerve terminal length was similar in injured and intact corneas. In intact corneas, the density (sum length) of basal epithelial TRPV1+ and TRPV1- nerve fibers was similar, however, in injured corneas, TRPV1+ nerve density was higher compared to TRPV1- nerves. The degree of physical association between TRPV1+ nerves and intraepithelial CD45+ MHC-II+ CD11c+ cells was similar in intact and injured corneas. Stromal leukocytes co-expressed TRPV1, which was partially localized to CD68+ lysosomes, and this expression pattern was lower in injured corneas.

Conclusions: TRPV1+ nerves accounted for a higher proportion of corneal nerves after injury, which may provide insights into the pathophysiology of neuropathic pain following corneal trauma. The close interactions of TRPV1+ nerves with intraepithelial immune cells and expression of TRPV1 by stromal macrophages provide evidence of neuroimmune interactions in the cornea.
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http://dx.doi.org/10.1167/iovs.62.9.6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8267209PMC
July 2021

Altered Corneal Epithelial Dendritic Cell Morphology and Phenotype Following Acute Exposure to Hyperosmolar Saline.

Invest Ophthalmol Vis Sci 2021 02;62(2):38

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, Australia.

Purpose: The purpose of this study was to assess the morphological and phenotypic responses of corneal epithelial dendritic cells (DCs) to acute topical hyperosmolar stress, given a pathogenic role for tear hyperosmolarity in dry eye disease (DED).

Methods: C57BL/6J mice were anesthetized and received 350 mOsm/L (physiological; n = 5 mice), 450 mOsm/L (n = 6), or 600 mOsm/L (n = 6) saline on a randomly assigned eye. Corneas were harvested 2 hours later. Immunofluorescent staining was performed using CD45, CD86, and CD68 antibodies to investigate DC morphology (density, viability, field area, circularity, and dendritic complexity) and immunological phenotype. Flow cytometry was used to confirm CD86 and CD68 expression in CD11c+ DCs, using C57BL/6J mice that received topical applications of 350 mOsm/L, 450 mOsm/L, or 600 mOsm/L (n = 5 per group) bilaterally for 2 hours.

Results: Following exposure to 450 mOsm/L topical saline for 2 hours, DCs in the central and peripheral cornea were larger (field area: Pcentral = 0.005, Pperipheral = 0.037; circularity: Pcentral = 0.026, and Pperipheral = 0.013) and had higher expression of CD86 compared with 350 mOsm/L controls (immunofluorescence: P < 0.0001; flow cytometry: P = 0.0058). After application of 600 mOsm/L saline, DC morphology was unchanged, although the percentage of fragmented DCs, and phenotypic expression of CD86 (immunofluorescence: P < 0.0001; and flow cytometry: P = 0.003) and CD68 (immunofluorescence: P = 0.024) were higher compared to 350 mOsm/L controls.

Conclusions: Short-term exposure to mild hyperosmolar saline (450 mOsm/L) induced morphological and phenotypic maturation in corneal epithelial DCs. More severe hyperosmolar insult (600 mOsm/L) for 2 hours appeared toxic to these cells. These data suggest that hyperosmolar conditions activate corneal DCs, which may have implications for understanding DC activation in DED.
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http://dx.doi.org/10.1167/iovs.62.2.38DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7910639PMC
February 2021

Identification of presumed corneal neuromas and microneuromas using laser-scanning in vivo confocal microscopy: a systematic review.

Br J Ophthalmol 2021 Feb 10. Epub 2021 Feb 10.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, Australia

Background/aims: This systematic review critically evaluated peer-reviewed publications describing morphological features consistent with, or using terms related to, a 'neuroma' or 'microneuroma' in the human cornea using laser-scanning in vivo confocal microscopy (IVCM).

Methods: The review was prospectively registered on PROSPERO (CRD42020160038). Comprehensive literature searches were performed in Ovid MEDLINE, Ovid Embase and the Cochrane Library in November 2019. The review included primary research studies and reviews that described laser-scanning IVCM for examining human corneal nerves. Papers had to include at least one of a pre-specified set of keyword stems, broadly related to neuromas and microneuromas, to describe a corneal nerve feature.

Results: Twenty-five papers (20 original studies; 5 reviews) were eligible. Three original studies evaluated corneal nerve features in healthy eyes. Most papers assessed corneal nerves in ocular and systemic conditions; seven studies did not include a control/comparator group. There was overlap in terminology used to describe nerve features in healthy and diseased corneas (eg, bulb-like/bulbous, penetration, end/s/ing). Inspection of IVCM images within the papers revealed that features termed 'neuromas' and 'microneuromas' could potentially be physiological corneal stromal-epithelial nerve penetration sites. We identified inconsistent definitions for terms, and limitations in IVCM image acquisition, sampling and/or reporting that may introduce bias and lead to inaccurate representation of physiological nerve characteristics as pathological.

Conclusion: These findings identify a need for consistent nomenclature and definitions, and rigorous IVCM scanning and analysis protocols to clarify the prevalence of physiological, as opposed to pathological, corneal nerve features.
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http://dx.doi.org/10.1136/bjophthalmol-2020-318156DOI Listing
February 2021

Morphometric Changes to Corneal Dendritic Cells in Individuals With Mild Cognitive Impairment.

Front Neurosci 2020 9;14:556137. Epub 2020 Dec 9.

Department of Optometry and Vision Sciences, University of Melbourne, Parkville, VIC, Australia.

Purpose: There has been increasing interest in identifying non-invasive, imaging biomarkers for neurodegenerative disorders of the central nervous system (CNS). The aim of this proof-of-concept study was to investigate whether corneal sensory nerve and dendritic cell (DC) parameters, captured using confocal microscopy (IVCM), are altered in individuals with mild cognitive impairment (MCI) and Alzheimer's disease (AD).

Methods: Fifteen participants were recruited from the Australian Imaging Biomarkers and Lifestyle (AIBL) study in Melbourne, VIC, Australia. The cohort consisted of cognitively normal (CN) individuals ( = 5), and those with MCI ( = 5) and AD ( = 5). Participants underwent a slit lamp examination of the anterior segment, followed by corneal imaging using laser-scanning confocal microscopy (IVCM) of the central and inferior whorl regions. Corneal DC density, field area, perimeter, circularity index, aspect ratio, and roundness were quantified using Image J. Quantitative data were derived for corneal nerve parameters, including nerve fiber length (CNFL), fiber density (CNFD), branch density (CNBD), and diameter.

Results: Corneal DC field area and perimeter were greater in individuals with MCI, relative to CN controls, in both the central and inferior whorl regions ( < 0.05 for all comparisons). In addition, corneal DCs in the whorl region of MCI eyes had lower circularity and roundness indices and a higher aspect ratio relative to CNs ( < 0.05 for all comparisons). DC density was similar across participant groups in both corneal regions. There was a trend toward lower quantitative parameters for corneal nerve architecture in the AD and MCI groups compared with CN participants, however, the inter-group differences did not reach statistical significance. Central corneal nerve diameters were similar between groups.

Conclusion: This study is the first to report morphological differences in corneal DCs in humans with MCI. These differences were evident in both the central and mid-peripheral cornea, and in the absence of significant nerve abnormalities or a difference in DC density. These findings justify future large-scale studies to assess the utility of corneal IVCM and DC analysis for identifying early stage pathology in neurodegenerative disorders of the CNS.
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http://dx.doi.org/10.3389/fnins.2020.556137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7755610PMC
December 2020

Processing of positive newborn screening results: a qualitative exploration of current practice in England.

BMJ Open 2020 12 12;10(12):e044755. Epub 2020 Dec 12.

Florence Nightingale Faculty of Nursing, Midwifery & Palliative Care, King's College London, London, UK.

Objective: To explore current communication practices for positive newborn screening results from the newborn bloodspot screening (NBS) laboratory to clinicians to highlight differences, understand how the pathways are implemented in practice, identify barriers and facilitators and make recommendations for future practice and research.

Design: A qualitative exploratory design was employed using semi-structured interviews.

Setting: Thirteen NBS laboratories in England.

Participants: Seventy-one clinicians; 22 NBS laboratory staff across 13 laboratories and 49 members of relevant clinical teams were interviewed.

Results: Assurance of quality and consistency was a priority for all NBS laboratories. Findings indicated variation in approaches to communicating positive NBS results from laboratories to clinical teams. This was particularly evident for congenital hypothyroidism and was largely influenced by local arrangements, resources and the fact individual laboratories had detailed standard operating procedures for how they work. Obtaining feedback from clinical teams to the laboratory after the child had been seen could be challenging and time-consuming for those involved. Pathways for communicating carrier results for cystic fibrosis and sickle cell disease could be ambiguous and inconsistent which in turn could hamper the laboratories efforts to obtain timely feedback regarding whether or not the result had been communicated to the family. Communication pathways for positive NBS results between laboratories and clinical teams could therefore be time-consuming and resource-intensive.

Conclusion: The importance placed on ensuring positive NBS results were communicated effectively and in a timely fashion from the laboratory to the clinical team was evident from all participants. However, variation existed in terms of the processes used to report positive NBS results to clinical teams and the people involved. Variant practice identified may reflect local needs, but more often reflected local resources and a more consistent 'best practice' approach is required, not just in the UK but perhaps globally.

Trial Registration Number: ISRCTN15330120.
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http://dx.doi.org/10.1136/bmjopen-2020-044755DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7735110PMC
December 2020

Psychological Impact of NBS for CF.

Int J Neonatal Screen 2020 06 30;6(2):27. Epub 2020 Mar 30.

Faculty of Sports, Health and Applied Science, St Mary's University, London TW1 4SX, UK;

Newborn screening for cystic fibrosis has resulted in diagnosis often before symptoms are recognised, leading to benefits including reduced disease severity, decreased burden of care, and lower costs. The psychological impact of this often unsought diagnosis on the parents of seemingly well children is less well understood. The time during which the screening result is communicated to families but before the confirmatory test results are available is recognised as a period of uncertainty and it is this uncertainty that can impact most on parents. Evidence suggests this may be mitigated against by ensuring the time between communication and confirmatory testing is minimized and health professionals involved in communicating positive newborn screening results and diagnostic results for cystic fibrosis to families are knowledgeable and able to provide appropriate reassurance. This is particularly important in the case of false positive results or when the child is given a Cystic Fibrosis Screen Positive, Inconclusive Diagnosis designation. However, to date, there are no formal mechanisms in place to support health professionals undertaking this challenging role, which would enable them to meet the expectations set out in specific guidance.
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http://dx.doi.org/10.3390/ijns6020027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7422999PMC
June 2020

The effect of high-fat diet-induced metabolic disturbance on corneal neuroimmune features.

Exp Eye Res 2020 12 15;201:108298. Epub 2020 Oct 15.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, Australia. Electronic address:

Purpose: The highly innervated cornea is susceptible to nerve loss secondary to systemic diseases such as diabetes and metabolic disturbances caused by high-fat diet. In this study, we characterize the effect of high-fat diet on the mouse corneal neuroimmune phenotype, including changes to corneal nerve density and resident immune cells, alongside the clinical assessment of corneal thickness and endothelial cell density.

Methods: Male C57Bl6/J mice, aged 10 weeks, were fed a high-fat diet (60 kcal% fat, 5.2 kcal/g) or control diet (10 kcal%, 3.8 kcal/g) for 16 weeks. At the study endpoint, metabolic parameters (HbA1c, weight, fasting glucose, body fat) were measured to confirm metabolic disturbance. Clinical imaging of the anterior segment was performed using optical coherence tomography to measure the corneal epithelial and stromal thickness. Corneal sensory nerves were visualized using flatmount immunostaining and confocal microscopy. The topographical distribution and density of sensory nerves (BIII-tubulin), intraepithelial CD45 and MHC- II cells, stromal macrophages (IBA1CD206) and endothelial cells (ZO-1) were analysed using FIJI.

Results: High-fat diet mice had significantly higher blood HbA1c, higher body weight, a higher percentage of body fat and elevated fasting glucose compared to the control diet mice. Corneal epithelial and stromal thickness was similar in both groups. The sum length of the basal nerve plexus was lower in the central and peripheral cornea of mice fed a high-fat diet. In contrast, the sum length of superficial nerve terminals was similar between groups. Epithelial immune cell density was two-fold higher in the central corneas of high-fat diet mice compared to control diet mice. IBA1CD206 macrophage density was similar in the anterior stroma of both groups but was significantly higher in the posterior stroma of the peripheral cornea in the high-fat diet mice compared to controls. The percentage of nerve-associated MHC-II cells in the epithelium and stroma was higher in HFD mice compared to controls. Endothelial cell density was similar in the corneas of high-fat diet mice compared to controls.

Conclusion: Together with corneal neuropathy, corneal immune cells in mice fed a high-fat diet were differentially affected depending on their topographical distribution and location within cornea, and appeared in closer proximity to epithelial and stromal nerves, suggesting a local neuroimmune disruption induced by systemic metabolic disturbance.
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http://dx.doi.org/10.1016/j.exer.2020.108298DOI Listing
December 2020

Qualitative exploration of health professionals' experiences of communicating positive newborn bloodspot screening results for nine conditions in England.

BMJ Open 2020 10 1;10(10):e037081. Epub 2020 Oct 1.

Paediatrics, Institute of Child Health, Merseyside, UK.

Objective: To explore health professionals' experiences of communicating positive newborn bloodspot screening (NBS) results, highlight differences, share good practice and make recommendations for future research.

Design: Qualitative exploratory design was employed using semi-structured interviews SETTING: Three National Health Service provider organisations in England PARTICIPANTS: Seventeen health professionals involved in communicating positive newborn bloodspot screening results to parents for all nine conditions currently included in the newborn bloodspot screening programme in England.

Results: Findings indicated variation in approaches to communicating positive newborn bloodspot screening results to parents, largely influenced by resources available and the lack of clear guidance. Health professionals emphasised the importance of communicating results to families in a way that is sensitive to their needs. However, many challenges hindered communication including logistical considerations; difficulty contacting the family and other health professionals; language barriers; parental reactions; resource considerations; lack of training; and insufficient time.

Conclusion: Health professionals invest a lot of time and energy trying to ensure communication of positive newborn bloodspot screening results to families is done well. However, there continues to be great variation in the way these results are communicated to parents and this is largely influenced by resources available but also the lack of concrete guidance. How best to support health professionals undertaking this challenging and emotive task requires further exploration. We recommend evaluation of a more cohesive approach that meets the needs of parents and staff while being sensitive to the subtleties of each condition.

Trial Registration Number: ISRCTN15330120.
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http://dx.doi.org/10.1136/bmjopen-2020-037081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7534703PMC
October 2020

Retinal Functional and Structural Changes in the 5xFAD Mouse Model of Alzheimer's Disease.

Front Neurosci 2020 13;14:862. Epub 2020 Aug 13.

Department of Optometry and Vision Sciences, University of Melbourne, Parkville, VIC, Australia.

Alzheimer's disease is characterized by the aberrant deposition of protein in the brain and is the leading cause of dementia worldwide. Increasingly, there have been reports of the presence of these protein hallmarks in the retina. In this study, we assayed the retina of 5xFAD mice, a transgenic model of amyloid deposition known to exhibit dementia-like symptoms with age. Using OCT, we found that the retinal nerve fiber layer was thinner in 5xFAD at 6, 12, and 17 months of age compared with wild-type littermates, but the inner plexiform layer was thicker at 6 months old. Retinal function showed reduced ganglion cell responses to light in 5xFAD at 6, 12, and 17 months of age. This functional loss was observed in the outer retina at 17 months of age but not in younger mice. We showed using immunohistochemistry and ELISA that soluble and insoluble amyloid was present in the retina and brain at all ages. In conclusion, we report that amyloid is present in brain and retina of 5xFAD mice and that the pattern of neuronal dysfunction occurs in the inner retina at the early ages and progresses to encompass the outer retina with age. This implies that the inner retina is more sensitive to amyloid changes in early disease and that the outer retina is also affected with disease progression.
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http://dx.doi.org/10.3389/fnins.2020.00862DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7438734PMC
August 2020

In vivo immune cell dynamics in the human cornea.

Exp Eye Res 2020 10 23;199:108168. Epub 2020 Aug 23.

Department of Ophthalmology and Neuroscience Center of Excellence, School of Medicine, Louisiana State University Health New Orleans, 2020 Gravier St., Suite D, New Orleans, LA, 70112, USA.

In vivo confocal microscopy (IVCM) allows the evaluation of the living human cornea at the cellular level. The non-invasive nature of this technique longitudinal, repeated examinations of the same tissue over time. Image analysis of two-dimensional time-lapse sequences of presumed immune cells with and without visible dendrites at the corneal sub-basal nerve plexus in the eyes of healthy individuals was performed. We demonstrated evidence that cells without visible dendrites are highly dynamic and move rapidly in the axial directions. A number of dynamic cells were observed and measured from three eyes of different individuals. The total average displacement and trajectory speeds of three cells without visible dendrites (N = 9) was calculated to be 1.12 ± 0.21 and 1.35 ± 0.17 μm per minute, respectively. One cell with visible dendrites per cornea was also analysed. Tracking dendritic cell dynamics in vivo has the potential to significantly advance the understanding of the human immune adaptive and innate systems. The ability to observe and quantify migration rates of immune cells in vivo is likely to reveal previously unknown insights into corneal and general pathophysiology and may serve as an effective indicator of cellular responses to intervention therapies.
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http://dx.doi.org/10.1016/j.exer.2020.108168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7898771PMC
October 2020

The neuroregenerative effects of topical decorin on the injured mouse cornea.

J Neuroinflammation 2020 May 4;17(1):142. Epub 2020 May 4.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, 3053, Australia.

Background: The cornea is innervated with a rich supply of sensory nerves that play important roles in ocular surface health. Any injury or pathology of the corneal nerves increases the risk of dry eye disease and infection. This study aims to evaluate the therapeutic potential of topical decorin to improve corneal nerve regeneration in a mouse model of sterile epithelial abrasion injury.

Methods: Bilateral central corneal epithelial abrasions (2-mm, Alger Brush) were performed on young C57BL/6 J mice to remove the corneal sensory nerves. Decorin, or vehicle, was applied topically, three times per day for 1 week or every 2 h for 6 h. Spectral-domain optical coherence tomography was performed to measure the abrasion area and corneal thickness. Wholemount immunofluorescence staining was used to assess sensory nerve regeneration (β-tubulin III) and immune cell density (CD45, Iba1, CD11c). To investigate the specific role of dendritic cells (DCs), Cx3cr1 mice, which spontaneously lack resident corneal epithelial DCs, were also investigated. The effect of prophylactic topical administration of recombinant human decorin (applied prior to the abrasion) was also investigated. Nerve tracing (NeuronJ software) was performed to compare recovery of basal nerve axons and superficial nerve terminals in the central and peripheral cornea.

Results: At 6 h after injury, topical decorin application was associated with greater intraepithelial DC recruitment but no change in re-epithelialisation or corneal thickness, compared to the vehicle control. One week after injury, sub-basal nerve plexus and superficial nerve terminal density were significantly higher in the central cornea in the decorin-treated eyes. The density of corneal stromal macrophages in the decorin-treated eyes and their contralateral eyes was significantly lower compared to saline-treated corneas. No significant improvement in corneal nerve regeneration was observed in Cx3cr1 mice treated with decorin.

Conclusions: Decorin promotes corneal epithelial nerve regeneration after injury. The neuroregenerative effect of topical decorin was associated with a higher corneal DC density during the acute phase, and fewer macrophages at the study endpoint. The corneal neuroregenerative effects of decorin were absent in mice lacking intraepithelial DCs. Together, these findings support a role for decorin in DC-mediated neuroregeneration following corneal abrasion injury.
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http://dx.doi.org/10.1186/s12974-020-01812-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199348PMC
May 2020

Tunneling Nanotubes and the Eye: Intercellular Communication and Implications for Ocular Health and Disease.

Biomed Res Int 2020 8;2020:7246785. Epub 2020 Apr 8.

Department of Ophthalmology, Casey Eye Institute, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, OR, USA 97239.

Cellular communication is an essential process for the development and maintenance of all tissues including the eye. Recently, a new method of cellular communication has been described, which relies on formation of tubules, called tunneling nanotubes (TNTs). These structures connect the cytoplasm of adjacent cells and allow the direct transport of cellular cargo between cells without the need for secretion into the extracellular milieu. TNTs may be an important mechanism for signaling between cells that reside long distances from each other or for cells in aqueous environments, where diffusion-based signaling is challenging. Given the wide range of cargoes transported, such as lysosomes, endosomes, mitochondria, viruses, and miRNAs, TNTs may play a role in normal homeostatic processes in the eye as well as function in ocular disease. This review will describe TNT cellular communication in ocular cell cultures and the mammalian eye , the role of TNTs in mitochondrial transport with an emphasis on mitochondrial eye diseases, and molecules involved in TNT biogenesis and their function in eyes, and finally, we will describe TNT formation in inflammation, cancer, and stem cells, focusing on pathological processes of particular interest to vision scientists.
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http://dx.doi.org/10.1155/2020/7246785DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7171654PMC
February 2021

Novel alterations in corneal neuroimmune phenotypes in mice with central nervous system tauopathy.

J Neuroinflammation 2020 Apr 28;17(1):136. Epub 2020 Apr 28.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Australia.

Background: Tauopathy in the central nervous system (CNS) is a histopathological hallmark of frontotemporal dementia (FTD) and Alzheimer's disease (AD). Although AD is accompanied by various ocular changes, the effects of tauopathy on the integrity of the cornea, which is densely innervated by the peripheral nervous system and is populated by resident dendritic cells, is still unknown. The aim of this study was to investigate if neuroimmune interactions in the cornea are affected by CNS tauopathy.

Methods: Corneas from wild type (WT) and transgenic rTg4510 mice that express the P301L tau mutation were examined at 2, 6, 8, and 11 months. Clinical assessment of the anterior segment of the eye was performed using spectral domain optical coherence tomography. The density of the corneal epithelial sensory nerves and the number and field area of resident epithelial dendritic cells were assessed using immunofluorescence. The immunological activation state of corneal and splenic dendritic cells was examined using flow cytometry and compared between the two genotypes at 9 months of age.

Results: Compared to age-matched WT mice, rTg4510 mice had a significantly lower density of corneal nerve axons at both 8 and 11 months of age. Corneal nerves in rTg4510 mice also displayed a higher percentage of beaded nerve axons and a lower density of epithelial dendritic cells compared to WT mice. From 6 months of age, the size of the corneal dendritic cells was significantly smaller in rTg4510 compared to WT mice. Phenotypic characterization by flow cytometry demonstrated an activated state of dendritic cells (CD86 and CD45 CD11bCD11c) in the corneas of rTg4510 compared to WT mice, with no distinct changes in the spleen monocytes/dendritic cells. At 2 months of age, there were no significant differences in the neural or immune structures between the two genotypes.

Conclusions: Corneal sensory nerves and epithelial dendritic cells were altered in the rTg4510 mouse model of tauopathy, with temporal changes observed with aging. The activation of corneal dendritic cells prior to the gradual loss of neighboring sensory nerves suggests an early involvement of corneal immune cells in tau-associated pathology originating in the CNS.
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http://dx.doi.org/10.1186/s12974-020-01803-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7189727PMC
April 2020

Ocular Phenotype of Relaxin Gene Knockout (Rln) Mice.

Curr Eye Res 2020 10 16;45(10):1211-1221. Epub 2020 Mar 16.

School of BioSciences, The University of Melbourne , Parkville, Australia.

: To test if relaxin deficiency affects ocular structure and function we investigated expression of relaxin () and RXFP receptors (), and compared ocular phenotypes in relaxin gene knockout ( ) and wild type ( ) mice. : and mRNA expression was detected in ocular tissues of Rln+/+ mice using RT-PCR. The eyes of 11 and 5 male mice were investigated. Corneal and retinal thickness was assessed using optical coherence tomography. Intraocular pressure was measured using a rebound tonometer. Retinal, choroidal and sclera morphology and thickness were evaluated histologically. Eyes were collected and fixed for immunofluorescence staining or used for RNA extraction to evaluate mRNA expression using real-time PCR. : mRNA was expressed only in the retina, whereas transcripts were detected in the retina, cornea and sclera/choroid. was only present in the cornea. None of these genes were expressed in the lacrimal gland, eyelid or lens. Intraocular pressure was higher and central cornea of mice was significantly thicker and had significantly larger endothelial cells and a lower endothelial cell density than mice. Immunohistochemistry demonstrated no significant difference in AQP3 and AQP5 staining in the cornea or other regions between wildtype and mice. mRNA expression of was significantly higher in than in corneas, whereas and were significantly decreased. Expression of and was significantly decreased in compared to uvea. No significant differences in these genes were detected in the retina. Retinal, choroidal and scleral thicknesses were not different and morphology appeared normal. : The findings indicate that loss of affects expression of several genes in the uvea and cornea and results in thicker corneas with altered endothelial cells. Many of the gene changes suggest alterations in extracellular matrix and fluid transport between cells.
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http://dx.doi.org/10.1080/02713683.2020.1737714DOI Listing
October 2020

Corneal Epithelial "Neuromas": A Case of Mistaken Identity?

Cornea 2020 Jul;39(7):930-934

Department of Ophthalmology, School of Medical Sciences, Faculty of Medicine, University of New South Wales, Sydney, New South Wales, Australia.

Laser scanning in vivo confocal microscopy is a useful clinical tool to assess the corneal nerves in human and laboratory animals. With this new technology, the use of terms such as "neuromas" and "microneuromas" is becoming popular to describe nerve structures seen in humans. Here, we point out that the sites where stromal nerves enter the corneal epithelium are often hyperreflective and can appear dysmorphic when imaged using in vivo confocal microscopy. Furthermore, we clarify what is known anatomically about how the nerves enter the corneal epithelium from the stroma, and we urge colleagues to differentiate between hyperreflective foci at the corneal stromal-epithelial nerve penetration sites and alterations in nerve morphology secondary to injury or disease.
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http://dx.doi.org/10.1097/ICO.0000000000002294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7279054PMC
July 2020

Topographical and Morphological Differences of Corneal Dendritic Cells during Steady State and Inflammation.

Ocul Immunol Inflamm 2020 Aug 20;28(6):898-907. Epub 2019 Aug 20.

Department of Optometry and Vision Sciences, The University of Melbourne , Melbourne, Australia.

Purpose: We report novel differences in mouse corneal DC morphology and density during local and systemic inflammation.

Methods: Local inflammation was induced by topical application of saline or TLR9 agonist CpG-ODN on abraded C57BL6J mouse corneas. Systemic inflammation was induced by intraperitoneal injection of lipopolysaccharide (LPS) in CD11c-YFP mice. Corneal epithelial DCs from uninjured, injured and contralateral eyes were analysed by confocal microscopy.

Results: Following local CpG delivery on the injured cornea, the DC density and size increased in both central and peripheral regions. Contralateral uninjured eyes displayed enlarged DC morphology in the central cornea compared to naïve cohorts. After systemic LPS, the size of DCs in the central cornea was lower at 2 hours, returning to baseline after 24 hours.

Conclusions: Corneal DCs respond differently in terms of shape and distribution during local and systemic inflammation. These features can serve as indicators in ocular and systemic diseases.
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http://dx.doi.org/10.1080/09273948.2019.1646775DOI Listing
August 2020

The Effects of Aging on Corneal and Ocular Surface Homeostasis in Mice.

Invest Ophthalmol Vis Sci 2019 06;60(7):2705-2715

Department of Optometry and Vision Sciences, University of Melbourne, Parkville, Victoria, Australia.

Purpose: Aging is a risk factor for dry eye disease. The aim of this study was to investigate if aging is associated with a range of signs of dry eye disease, including tear hyperosmolarity, reduced nerve density, and increased dendritic cell number, in mice.

Method: Healthy C57BL/6 female mice, aged 2 months (young, n = 10) and 22 months (aged, n = 11) were used. Clinical assessments included corneal sensitivity (Cochet-Bonnet esthesiometry), tear secretion (Phenol red thread test), tear film osmolarity (TearLab osmometer), and corneal thickness (optical coherence tomography). The sum length of the corneal superficial terminals and sub-basal nerves, density of vertical nerve projections, and density and tree area of resident epithelial dendritic cells, were assessed using immunofluorescence and confocal microscopy.

Results: Aged mice had significantly higher tear secretion, lower corneal sensitivity, and a thicker corneal stroma but thinner epithelium. There was no significant intergroup difference for tear osmolarity. Aged mice showed a significantly lower sum length of nerves in the superficial terminals and sub-basal plexus, relative to young mice. Dendritic cell density and morphology were similar in both groups.

Conclusions: In mice, aging is associated with higher tear secretion and corneal epithelial thinning, together with lower corneal nerve density and sensitivity. However, aging was not significantly associated with changes to tear osmolarity or dendritic cell density or size, despite a significant reduction in total nerve length. These findings demonstrate that aged mice exhibit some changes to ocular surface parameters that parallel the anomalies evident in dry eye disease.
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http://dx.doi.org/10.1167/iovs.19-26631DOI Listing
June 2019

Regional and functional heterogeneity of antigen presenting cells in the mouse brain and meninges.

Glia 2019 05 26;67(5):935-949. Epub 2018 Dec 26.

Department of Anatomy and Developmental Biology, Monash Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.

The central nervous system (CNS) is considered to be immune privileged, owing in part to the absence of major histocompatibility (MHC) class II cells in the healthy brain parenchyma. However, systemic inflammation can activate microglia to express MHC class II, suggesting that systemic inflammation may be sufficient to mature microglia into functional antigen presenting cells (APCs). We examined the effects of systemic lipopolysaccharide (LPS)-induced inflammation on the phenotype and function of putative APCs within the mouse brain parenchyma, as well as its supporting tissues-the choroid plexus and meninges. Microglia isolated from different regions of the brain demonstrated significant heterogeneity in their ability to present antigen to naïve OT-II CD4 T cells following exposure to systemic LPS. Olfactory bulb microglia (but not cortical microglia) intimately interacted with T cells in vivo and stimulated T cell proliferation in vitro, albeit in the absence of co-stimulation. In contrast, myeloid cells within the choroid plexus and meninges were immunogenic and upregulated the co-stimulatory molecule CD80 following systemic inflammation. Dural APCs, which clustered around LYVE-1 lymphatics, were more efficient at stimulating naïve T cell proliferation than choroid plexus APCs, suggesting that the dura may be an under-appreciated site for immune interactions. This study has highlighted the functional diversity of myeloid cells within the sub-compartments of the CNS and its supporting tissues. Furthermore, these findings demonstrate that systemic inflammation can mature selected microglia populations and choroid plexus/meningeal myeloid cells into functional APCs, which may contribute to the pathogenesis of neuroinflammation and neurodegenerative diseases.
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http://dx.doi.org/10.1002/glia.23581DOI Listing
May 2019

Anterior segment optical coherence tomography: its application in clinical practice and experimental models of disease.

Clin Exp Optom 2019 05 1;102(3):208-217. Epub 2018 Oct 1.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, Australia.

Optical coherence tomography (OCT) provides non-invasive, high-resolution in vivo imaging of the ocular surface and anterior segment. Over the years, it has become an essential tool for evaluating the anterior segment of the eye to monitor ocular development and ocular pathologies in both the clinical and research fields of ophthalmology and optometry. In this review, the clinical applications relating to the use of anterior segment OCT for imaging and quantifying normal and pathological features of the ocular surface, cornea, anterior chamber, and aqueous outflow system are summarised in a range of human ocular diseases. Applications of anterior segment OCT technology that have improved imaging and quantitation of ocular inflammation in experimental animal models of ocular diseases, such as anterior uveitis, microbial keratitis and glaucoma, are also described. The capacity to longitudinally evaluate anterior segment anatomical changes during development, and inflammation facilitates the understanding of the dynamics of tissue responses, and further enhances the intra-operative in vivo imaging during procedures, such as corneal transplantation and drug delivery. Future developments including in vivo ultrahigh-resolution anterior segment OCT, automated analyses of anterior segment OCT images and functional extensions of the technique, may revolutionise the clinical evaluation of anterior segment, corneal and ocular surface diseases.
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http://dx.doi.org/10.1111/cxo.12835DOI Listing
May 2019

Recovery of the sub-basal nerve plexus and superficial nerve terminals after corneal epithelial injury in mice.

Exp Eye Res 2018 06 14;171:92-100. Epub 2018 Mar 14.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria 3010, Australia. Electronic address:

Our aim was to compare regeneration of the sub-basal nerve plexus (SBNP) and superficial nerve terminals (SNT) following corneal epithelial injury. We also sought to compare agreement when quantifying nerve parameters using different image analysis techniques. Anesthetized, female C57BL/6 mice received central 1-mm corneal epithelial abrasions. Four-weeks post-injury, eyes were enucleated and processed for PGP9.5 to visualize the corneal nerves using wholemount immunofluorescence staining and confocal microscopy. The percentage area of the SBNP and SNT were quantified using: ImageJ automated thresholds, ImageJ manual thresholds and manual tracings in NeuronJ. Nerve sum length was quantified using NeuronJ and Imaris. Agreement between methods was considered with Bland-Altman analyses. Four-weeks post-injury, the sum length of nerve fibers in the SBNP, but not the SNT, was reduced compared with naïve eyes. In the periphery, but not central cornea, of both naïve and injured eyes, nerve fiber lengths in the SBNP and SNT were strongly correlated. For quantifying SBNP nerve axon area, all image analysis methods were highly correlated. In the SNT, there was poor correlation between manual methods and auto-thresholding, with a trend towards underestimating nerve fiber area using auto-thresholding when higher proportions of nerve fibers were present. In conclusion, four weeks after superficial corneal injury, there is differential recovery of epithelial nerve axons; SBNP sum length is reduced, however the sum length of SNTs is similar to naïve eyes. Care should be taken when selecting image analysis methods to compare nerve parameters in different depths of the corneal epithelium due to differences in background autofluorescence.
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http://dx.doi.org/10.1016/j.exer.2018.03.012DOI Listing
June 2018

Laser scanning in vivo confocal microscopy (IVCM) for evaluating human corneal sub-basal nerve plexus parameters: protocol for a systematic review.

BMJ Open 2017 Nov 3;7(11):e018646. Epub 2017 Nov 3.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Victoria, Australia.

Introduction: Laser scanning in vivo confocal microscopy (IVCM) enables non-invasive, high-resolution imaging of the cornea. In recent years, there has been a vast increase in researchers using laser scanning IVCM to image and quantify corneal nerve parameters. However, a range of methodological approaches have been adopted. The primary aim of this systematic review is to critically appraise the reported method(s) of primary research studies that have used laser scanning IVCM to quantify corneal sub-basal nerve plexus (SBNP) parameters in humans, and to examine corneal nerve parameters in healthy individuals.

Methods And Analysis: A systematic review of primary studies that have used laser scanning IVCM to quantify SBNP parameters in humans will be conducted. Comprehensive electronic searches will be performed in Ovid MedLine, Embase and the Cochrane Library. Two reviewers will independently assess titles and abstracts, and exclude studies not meeting the inclusion criteria. For studies judged eligible or potentially eligible, full texts will be independently assessed by two reviewers to determine eligibility. A third reviewer will resolve any discrepancies in judgement. Risk of bias will be assessed using a custom tool, covering five methodological domains: participant selection, method of image capture, method of image analysis, data reporting and other sources of bias. A systematic narrative synthesis of findings will be provided. A multilevel random-effects meta-analysis will be performed for corneal nerve parameters derived from healthy participants. This review will be reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement.

Ethics And Dissemination: As this review considers published data, ethical approval is not required. We foresee that this synthesis will serve as a reference for future studies, and can be used to inform best practice standards for using IVCM in clinical research. A manuscript reporting the results of the review will be published and may also be presented at scientific conferences.
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http://dx.doi.org/10.1136/bmjopen-2017-018646DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5695406PMC
November 2017

Optical Coherence Tomography Reveals Changes to Corneal Reflectivity and Thickness in Individuals with Tear Hyperosmolarity.

Transl Vis Sci Technol 2017 May 22;6(3). Epub 2017 May 22.

Department of Optometry and Vision Sciences, Faculty of Medicine, Dentistry & Health Sciences, The University of Melbourne, Parkville, Victoria, Australia 3010.

Purpose: To investigate whether tear hyperosmolarity, a feature of dry eye disease (DED), affects central corneal thickness (CCT), corneal light reflectivity, and/or tear film reflectivity.

Methods: This prospective, cross-sectional study involved 48 participants (38 with hyperosmolar tears and 10 controls with normo-osmolar tears). Symptoms and signs of DED (tear osmolarity, sodium fluorescein tear break-up time, ocular surface staining, Schirmer test) were assessed. CCT, and the reflectivity of the cornea and the tear-epithelial interface were quantified relative to background noise using Fourier-domain optical coherence tomography (FD-OCT).

Results: CCT of eyes with severe tear hyperosmolarity, defined as eyes in the upper quartile of the hyperosmolar group, was less than control eyes (539.1 ± 7.4 vs. 583.1 ± 15.0 μm, = 0.02) and eyes with less severe tear hyperosmolarity, defined as hyperosmolar eyes in the lower quartile (622.7 ± 5.8 μm, < 0.0001). CCT showed a negative linear relationship with tear osmolarity for values above 316 mOsmol/L ( = 0.17, = 0.01). Central corneal reflectivity was lower in hyperosmolar eyes than normo-osmolar eyes (45.1 ± 0.3 vs. 48.1 ± 0.6 pixels, = 0.02); the greatest relative difference was in the anterior stroma, where corneal reflectivity was 4.7 ± 1.9% less in hyperosmolar eyes ( < 0.01). Peak reflectivity of the tear-epithelial interface was 4.8% ± 3.5% higher in the hyperosmolar group than the normo-osmolar tear group ( = 0.04).

Conclusion: Individuals with significant tear hyperosmolarity and clinical signs of symptoms of DED show reduced CCT and altered corneal reflectivity.

Translational Relevance: Anterior segment FD-OCT provides novel insight into corneal microstructural differences in individuals with DED.
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http://dx.doi.org/10.1167/tvst.6.3.6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5444496PMC
May 2017

Omega-3 supplementation is neuroprotective to corneal nerves in dry eye disease: a pilot study.

Ophthalmic Physiol Opt 2017 07 12;37(4):473-481. Epub 2017 Mar 12.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Australia.

Purpose: To investigate whether oral, long-chain omega-3 (ω-3) essential fatty acid (EFA) supplementation, for 3 months, induces changes to the central corneal sub-basal nerve plexus in dry eye disease and whether nerve alterations correlate with clinical findings.

Methods: This prospective, comparative study involved the final 12 participants enrolled in a randomised, double-masked, placebo-controlled clinical trial of 60 participants with moderate dry eye disease. Participants received either placebo (olive oil 1500 mg/day; n = 4) or ω-3 EFA supplements (~1000 mg/day eicosapentaenoic acid + ~500 mg/day docosahexaenoic acid; n = 8) for 90 days. The main outcome measure was the mean change in central corneal sub-basal plexus nerve parameters between days one and 90, quantified using in vivo confocal microscopy. Secondary outcomes included mean change in tear osmolarity, corneal dendritic cell density and basal epithelial cell density.

Results: Compared with baseline, the reduction in OSDI score and tear osmolarity at day 90 were greater in the ω-3 EFA group than the placebo group (OSDI: ω-3 EFA, mean ± SEM: -15.6 ± 2.8 vs placebo: -2.8 ± 4.1 units, t = 2.6, p = 0.04; tearosmolarity: ω-3 EFA: -22.63 ± 5.7 vs placebo: -8 ± 2.7 mOsmol/L, t = 2.3, p = 0.04). At day 90, corneal total nerve branch density (CTBD: 91.1 ± 8.6 vs 45.1 ± 13.4 branches/mm , F = 14, p = 0.004) and corneal nerve branch density on the main fibre (CNBD: 63.4 ± 6.5 vs 27.9 ± 11.5 branches/mm , F = 6, p = 0.03) were higher in the ω-3 EFA group compared with placebo. Relative to day 1, CNBD (branches/mm ) increased at day 90 in the ω-3 EFA group (+20.0 ± 9.2, t = 3.2 p = 0.01) compared with placebo (-10.8 ± 3.2). Similar changes were evident for corneal nerve fibre length (CNFL, mm/mm ), which increased from baseline at day 90 in the omega-3 EFA group (+2.9 ± 1.6, t = 3.4 p = 0.01) compared with placebo (-2.7 ± 0.5). There was a negative correlation between CTBD and tear osmolarity (r = -0.70, p = 0.01). No significant changes were observed for basal epithelial cell or corneal dendritic cell density.

Conclusion: These pilot study findings suggest that ω-3 EFA supplementation imparts neuroprotective effects in the corneal sub-basal plexus that correlate with the extent of tear osmolarity normalisation.
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http://dx.doi.org/10.1111/opo.12365DOI Listing
July 2017

Characterization of the Circumlimbal Suture Model of Chronic IOP Elevation in Mice and Assessment of Changes in Gene Expression of Stretch Sensitive Channels.

Front Neurosci 2017 10;11:41. Epub 2017 Feb 10.

Department of Optometry and Vision Sciences, University of Melbourne Parkville, VIC, Australia.

To consider whether a circumlimbal suture can be used to chronically elevate intraocular pressure (IOP) in mice and to assess its effect on retinal structure, function and gene expression of stretch sensitive channels. Anesthetized adult C57BL6/J mice had a circumlimbal suture (10/0) applied around the equator of one eye. In treated eyes ( = 23) the suture was left in place for 12 weeks whilst in sham control eyes the suture was removed at day two ( = 17). Contralateral eyes served as untreated controls. IOP was measured after surgery and once a week thereafter. After 12 weeks, electroretinography (ERG) was performed to assess photoreceptor, bipolar cell and retinal ganglion cell (RGC) function. Retinal structure was evaluated using optical coherence tomography. Retinae were processed for counts of ganglion cell density or for quantitative RT-PCR to quantify purinergic () or stretch sensitive channel () gene expression. Immediately after suture application, IOP spiked to 33 ± 3 mmHg. After 1 day, IOP had recovered to 27 ± 3 mmHg. Between weeks 2 and 12, IOP remained elevated above baseline (control 14 ± 1 mmHg, ocular hypertensive 19 ± 1 mmHg). Suture removal at day 2 (Sham) restored IOP to baseline levels, where it remained through to week 12. ERG analysis showed that 12 weeks of IOP elevation reduced photoreceptor (-15 ± 4%), bipolar cell (-15 ± 4%) and ganglion cell responses (-19 ± 6%) compared to sham controls and respective contralateral eyes (untreated). The retinal nerve fiber layer was thinned in the presence of normal total retinal thickness. Ganglion cell density was reduced across all quadrants (superior -12 ± 5%; temporal, -7% ± 2%; inferior -9 ± 4%; nasal -8 ± 5%). Quantitative RT-PCR revealed a significant increase in gene expression (+11 ± 4%), whilst other genes were not significantly altered (). Our results show that circumlimbal ligation produces mild chronic ocular hypertension and retinal dysfunction in mice. Consistent with a sustained change to purinergic signaling we found an up-regulation of Entpd1.
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http://dx.doi.org/10.3389/fnins.2017.00041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5301305PMC
February 2017

Macrophage physiology in the eye.

Pflugers Arch 2017 04 23;469(3-4):501-515. Epub 2017 Feb 23.

Department of Anatomy and Developmental Biology and Monash Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.

The eye is a complex sensory organ composed of a range of tissue types including epithelia, connective tissue, smooth muscle, vascular and neural tissue. While some components of the eye require a high level of transparency to allow light to pass through unobstructed, other tissues are characterized by their dense pigmentation, which functions to absorb light and thus control its passage through the ocular structures. Macrophages are present in all ocular tissues, from the cornea at the anterior surface through to the choroid/sclera at the posterior pole. This review will describe the current understanding of the distribution, phenotype, and physiological role of ocular macrophages, and provide a summary of evidence pertaining to their proposed role during pathological conditions.
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http://dx.doi.org/10.1007/s00424-017-1947-5DOI Listing
April 2017

A Randomized, Double-Masked, Placebo-Controlled Clinical Trial of Two Forms of Omega-3 Supplements for Treating Dry Eye Disease.

Ophthalmology 2017 01 3;124(1):43-52. Epub 2016 Nov 3.

Department of Optometry and Vision Sciences, The University of Melbourne, Parkville, Australia. Electronic address:

Purpose: To assess the efficacy of 2 forms of oral long-chain omega-3 (ω-3) essential fatty acid (EFA) supplements, phospholipid (krill oil) and triacylglyceride (fish oil), for treating dry eye disease (DED).

Design: Randomized, double-masked, placebo-controlled clinical trial.

Participants: This study was conducted at a single site and involved 60 participants with mild to moderate DED who were randomized (1:1:1) to 1 of 3 groups: placebo (olive oil), krill oil, or fish oil supplements.

Methods: Participants received 1 of the 3 interventions: placebo (olive oil 1500 mg/day), krill oil (945 mg/day eicosapentaenoic acid [EPA], + 510 mg/day docosahexaenoic acid [DHA]), or fish oil (1000 mg/day EPA + 500 mg/day DHA) for 90 days, with monthly study visits.

Main Outcome Measures: Primary outcome measures were mean change in (1) tear osmolarity and (2) DED symptoms (Ocular Surface Disease Index [OSDI] score) between days 1 and 90. Secondary outcomes included mean change in key clinical signs (tear stability, tear production, ocular surface staining, bulbar and limbal redness, tear volume, anterior blepharitis, meibomian gland capping) and tear inflammatory cytokine levels.

Results: In total, 54 participants completed the study. At day 90, tear osmolarity was reduced from baseline with both krill oil (mean ± standard error of the mean: -18.6±4.5 mOsmol/l; n = 18; P < 0.001) and fish oil (-19.8±3.9 mOsmol/l; n = 19; P < 0.001) supplements, compared with placebo (-1.5±4.4 mOsmol/l; n = 17). OSDI score was significantly reduced at day 90 relative to baseline in the krill oil group only, compared with placebo (-18.6±2.4 vs. -10.5±3.3; P = 0.02). At day 90, there were also relative improvements in tear breakup time and ocular bulbar redness, compared with placebo, for both forms of ω-3 EFAs. Basal tear levels of the proinflammatory cytokine interleukin 17A were significantly reduced in the krill oil group, compared with placebo, at day 90 (-27.1±10.9 vs. 46.5±30.4 pg/ml; P = 0.02).

Conclusions: A moderate daily dose of both forms of long-chain ω-3 EFAs, for 3 months, resulted in reduced tear osmolarity and increased tear stability in people with DED. Omega-3 EFAs in a predominantly phospholipid form (krill oil) may confer additional therapeutic benefit, with improvements in DED symptoms and lower basal tear levels of interleukin 17A, relative to placebo.
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http://dx.doi.org/10.1016/j.ophtha.2016.09.023DOI Listing
January 2017

Questionnaire study to gain an insight into the manufacturing and fitting process of artificial eyes in children: an ocularist perspective.

Int Ophthalmol 2017 Oct 28;37(5):1175-1183. Epub 2016 Oct 28.

Public Health/Health Visiting, Faculty of Health and Social Science, Department of Nursing and Clinical Science, Bournemouth University, Poole, UK.

Purpose: To gain an insight into the manufacturing and fitting of artificial eyes in children and potential improvements to the process.

Method: An online qualitative survey was distributed to 39 ocularists/prosthetists in Europe and Canada. Participants were recruited through purposive sampling, specifically maximum variation sampling from the researcher's contacts and an online search.

Results: The findings highlighted the current impression technique as being the most difficult yet most important part of the current process for both the ocularist and child patient. Negatively affecting obtaining a good impression, the child patients distress can be reduced by their parents by providing encouragement, reassurance, practicing the insertion and removal of the artificial eye and being matter of fact. Whilst improvements to the current process provided mixed views, the incorporation of current technology was perceived as not being able to meet the requirements to produce aesthetically pleasing artificial eyes.

Conclusion: The current artificial eye process can be seen as an interaction with its success being dependent on the child patient's acceptance and adjustment which is dependent on the factors associated to the process. Investigation into the needs of the patient and whether technology can improve the process are the next steps in its advancement.
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http://dx.doi.org/10.1007/s10792-016-0383-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5633635PMC
October 2017

Longitudinal Changes to Tight Junction Expression and Endothelial Cell Integrity in a Mouse Model of Sterile Corneal Inflammation.

Invest Ophthalmol Vis Sci 2016 06;57(7):3477-84

Purpose: We previously reported that applying toll-like receptor (TLR) ligands to an injured cornea induces corneal edema at 24 hours, which subsides by 1 week. We tested the hypotheses that endothelial expression of the tight-junction protein, zonula occludens-1 (ZO-1), would be altered during experimental sterile corneal inflammation and that endothelial cell density (ECD) would remain unaffected.

Methods: Anesthetized C57BL/6J mice received central 1-mm corneal abrasions followed by topical application of saline or cytosine-phosphate-guanosine oligodeoxynucleotide (CpG-ODN, TLR-9 agonist). At 24 hours, 1 week and 4 weeks post treatment, spectral-domain optical coherence tomography images were captured. Eyes were enucleated and processed for zonula occludens-1 (ZO-1) immunofluorescent staining. Corneal flatmounts were analyzed for endothelial ZO-1 expression, cell density, polymegethism, and polymorphism. Corneal stromal inflammatory cell infiltration was evaluated at 4 weeks by immunostaining for CD45.

Results: Central corneal thickness (CCT) was increased in CpG-ODN treated eyes at 24 hours, had normalized by 1 week, but was again thickened by 4 weeks. In eyes with CpG-ODN, endothelial cell ZO-1 expression was reduced at 24 hours but returned to normal levels by 1 week. Endothelial cell density was not altered at 24 hours or 1 week. By 4 weeks, only CpG-ODN eyes showed relatively reduced ECD, as well as large numbers of CD45+ cells in the stroma. Changes to ECD correlated with CCT (r = -0.53, P < 0.01). Compared with naïve controls, more saline- and CpG-ODN-treated eyes exhibited polymegethism.

Conclusions: This study provides novel insights into the interplay between endothelial cell integrity, corneal edema, and chronic stromal leukocyte activation during sterile corneal inflammation in mice.
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http://dx.doi.org/10.1167/iovs.15-19005DOI Listing
June 2016

A case of mistaken identity: CD11c-eYFP(+) cells in the normal mouse brain parenchyma and neural retina display the phenotype of microglia, not dendritic cells.

Glia 2016 08 18;64(8):1331-49. Epub 2016 May 18.

Department of Anatomy and Developmental Biology and Monash Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.

Under steady-state conditions the central nervous system (CNS) is traditionally thought to be devoid of antigen presenting cells; however, putative dendritic cells (DCs) expressing enhanced yellow fluorescent protein (eYFP) are present in the retina and brain parenchyma of CD11c-eYFP mice. We previously showed that these mice carry the Crb1(rd8) mutation, which causes retinal dystrophic lesions; therefore we hypothesized that the presence of CD11c-eYFP(+) cells within the CNS may be due to pathology associated with the Crb1(rd8) mutation. We generated CD11c-eYFP Crb1(wt/wt) mice and compared the distribution and immunophenotype of CD11c-eYFP(+) cells in CD11c-eYFP mice with and without the Crb1(rd8) mutation. The number and distribution of CD11c-eYFP(+) cells in the CNS was similar between CD11c-eYFP Crb1(wt/wt) and CD11c-eYFP Crb1(rd8/rd8) mice. CD11c-eYFP(+) cells were distributed throughout the inner retina, and clustered in brain regions that receive input from the external environment or lack a blood-brain barrier. CD11c-eYFP(+) cells within the retina and cerebral cortex of CD11c-eYFP Crb1(wt/wt) mice expressed CD11b, F4/80, CD115 and Iba-1, but not DC or antigen presentation markers, whereas CD11c-eYFP(+) cells within the choroid plexus and pia mater expressed CD11c, I-A/I-E, CD80, CD86, CD103, DEC205, CD8α and CD135. The immunophenotype of CD11c-eYFP(+) cells and microglia within the CNS was similar between CD11c-eYFP Crb1(wt/wt) and CD11c-eYFP Crb1(rd8/rd8) mice; however, CD11c and I-A/I-E expression was significantly increased in CD11c-eYFP Crb1(rd8/rd8) mice. This study demonstrates that the overwhelming majority of CNS CD11c-eYFP(+) cells do not display the phenotype of DCs or their precursors and are most likely a subpopulation of microglia. GLIA 2016. GLIA 2016;64:1331-1349.
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http://dx.doi.org/10.1002/glia.23005DOI Listing
August 2016