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Ntentakis DP, Correa VSMC, Ntentaki AM, Delavogia E, Narimatsu T, Efstathiou NE, Vavvas DG. Effects of newer-generation anti-diabetics on diabetic retinopathy: a critical review. Graefes Arch Clin Exp Ophthalmol 2023;Abstract
Diabetic retinopathy (DR) is the leading etiology of blindness in the working population of the USA. Its long-term management relies on effective glycemic control. Seven anti-diabetic classes have been introduced for patients with type 2 diabetes (T2D) in the past two decades, with different glucose-lowering and cardiovascular benefits. Yet, their effects specifically on DR have not been studied in detail. A systematic review of the literature was conducted to investigate this topic, focusing on the available clinical data for T2D. Published studies were evaluated based on their level of statistical evidence, as long as they incorporated at least one endpoint or adverse event pertaining to retinal health. Fifty nine articles met our inclusion criteria and were grouped per anti-diabetic class as follows: alpha-glucosidase inhibitors (1), peroxisome proliferator-activated receptor gamma (PPAR-γ) agonists (8), amylin analogs (1), glucagon-like peptide-1 (GLP-1) receptor agonists (28), dipeptidyl peptidase 4 (DPP-4) inhibitors (9), and sodium glucose co-transporter-2 (SGLT-2) inhibitors (9), plus one retrospective study and two meta-analyses evaluating more than one of the aforementioned anti-diabetic categories. We also reviewed publicly-announced results of trials for the recently-introduced class of twincretins. The available data indicates that most drugs in the newer anti-diabetic classes are neutral to DR progression; however, there are subclasses differences in specific drugs and T2D populations. In particular, there is evidence suggesting there may be worse diabetic macular edema with PPAR-gamma agonists, potential slight DR worsening with semaglutide (GLP-1 receptor agonist), and potential slight increase in the incidence of retinal vein occlusion in elderly and patients with advanced kidney disease receiving SGLT-2 inhibitors. All these warrant further investigation. Longer follow-up and systematic assessment of at least one DR-related endpoint are highly recommended for all future trials in the T2D field, to ultimately address this topic.
Chang Y-H, Staffa SJ, Yavuz Saricay L, Zurakowski D, Gise R, Dagi LR. Sensitivity, specificity and cutoff identifying optic atrophy by macular ganglion cell layer volume in syndromic craniosynostosis. Ophthalmology 2023;Abstract
PURPOSE: Determine sensitivity, specificity, and cut-off of macular ganglion cell layer (GCL) volume consistent with optic atrophy in children with syndromic craniosynostosis (CS). Investigate whether obstructive sleep apnea (OSA), Chiari malformation, history of elevated intracranial pressure (ICP), CS diagnosis, age, or sex independently alter GCL volume with CS. DESIGN: Retrospective cross-sectional study. SUBJECTS: Patients with syndromic CS evaluated at Boston Children's Hospital (2010 - 2022) with reliable macular optical coherence tomography (OCT) scans. METHODS: Latest ophthalmic examination that included OCT macula scans was identified. Age at examination, sex, ethnicity, best-corrected logMAR visual acuity, cycloplegic refraction, and funduscopic optic nerve appearance were recorded in addition to history of primary or recurrent elevated ICP, Chiari malformation and OSA. Spectral domain-OCT software quantified segmentation of macula retinal layers, and was manually checked. MAIN OUTCOME MEASURES: Primary outcome was determining sensitivity, specificity and optimal cutoff of GCL volume consistent with optic atrophy. Secondary outcome was determining possible independent association of previously elevated ICP, OSA, Chiari, CS diagnosis, logMAR acuity, age, or sex with altered GCL volume. RESULTS: Median age at examination was 11.9 years (interquartile range (IQR), 8.5-14.8 years). Fifty-eight of 61 patients had reliable macula scans, 74% were female, and diagnoses were Apert (n=14); Crouzon (n=17); Muenke (n=6); Pfeiffer (n= 6); and Saethre-Chotzen (n=15). Optimal cutoff identifying optic atrophy was GCL volume <1.02mm3 with a sensitivity of 83% and specificity of 77%. Univariate analysis demonstrated significantly lower macular GCL volume with optic atrophy on fundus exam (P<0.001), Apert syndrome (P<0.001), history of elevated ICP (P=0.015), Chiari malformation (P=0.001), OSA (P<0.001), in males (P=0.027) and with worse logMAR acuity (-0.36, P<0.001). Multivariable median regression analysis confirmed that only OSA (P=0.005), optic atrophy on fundus exam (P=0.003), and worse logMAR acuity (P=0.042) independently associated with lower GCL volume. CONCLUSIONS: Macular GCL volume <1.02mm3 predicted optic atrophy in patients with CS with sensitive of 83% and specificity of 77%. OSA, a treatable often concomitant disorder, was independently associated with lower GCL volume. Surveillance for optic neuropathy by GCL volume proved effective in a population where cognitive skills can limit acquisition of other key ophthalmic measures.
Mazumder AG, Julé AM, Sun D. Astrocytes of the optic nerve exhibit a region-specific and temporally distinct response to elevated intraocular pressure. Mol Neurodegener 2023;18(1):68.Abstract
BACKGROUND: The optic nerve is an important tissue in glaucoma and the unmyelinated nerve head region remains an important site of many early neurodegenerative changes. In both humans and mice, astrocytes constitute the major glial cell type in the region, and in glaucoma they become reactive, influencing the optic nerve head (ONH) microenvironment and disease outcome. Despite recognizing their importance in the progression of the disease, the reactive response of optic nerve head astrocytes remains poorly understood. METHODS: To determine the global reactive response of ONH astrocytes in glaucoma we studied their transcriptional response to an elevation in IOP induced by the microbead occlusion model. To specifically isolate astrocyte mRNA in vivo from complex tissues, we used the ribotag method to genetically tag ribosomes in astrocytes, restricting analysis to astrocytes and enabling purification of astrocyte-associated mRNA throughout the entire cell, including the fine processes, for bulk RNA-sequencing. We also assessed the response of astrocytes in the more distal myelinated optic nerve proper (ONP) as glaucomatous changes manifest differently between the two regions. RESULTS: Astrocytes of the optic nerve exhibited a region-specific and temporally distinct response. Surprisingly, ONH astrocytes showed very few early transcriptional changes and ONP astrocytes demonstrated substantially larger changes over the course of the experimental period. Energy metabolism, particularly oxidative phosphorylation and mitochondrial protein translation emerged as highly upregulated processes in both ONH and ONP astrocytes, with the former showing additional upregulation in antioxidative capacity and proteolysis. Interestingly, optic nerve astrocytes demonstrated a limited neuroinflammatory response, even when challenged with a more severe elevation in IOP. Lastly, there were a greater number of downregulated processes in both astrocyte populations compared to upregulated processes. CONCLUSION: Our findings demonstrate an essential role for energy metabolism in the response of optic nerve astrocytes to elevated IOP, and contrary to expectations, neuroinflammation had a limited overall role. The transcriptional response profile is supportive of the notion that optic nerve astrocytes have a beneficial role in glaucoma. These previously uncharacterized transcriptional response of optic nerve astrocytes to injury reveal their functional diversity and a greater heterogeneity than previously appreciated.
Seresirikachorn K, Thiamthat W, Annopawong K, Wanichwecharungruang B, Friedman DS, Vu DM. Treatment Outcomes for Juvenile Open Angle Glaucoma in Thailand. J Glaucoma 2023;32(11):976-982.Abstract
PRCIS: Juvenile open angle glaucoma (JOAG) patients with thick central corneas and negative family history were more likely to undergo surgery, mainly trabeculectomy with half requiring additional surgery within 10 years. PURPOSE: To assess the characteristics and treatment outcomes of patients with JOAG in Thailand. PATIENTS AND METHODS: This retrospective, multicenter study included all patients diagnosed with JOAG over 12 years from 2 tertiary hospitals in Bangkok, Thailand. RESULTS: A total of 200 eyes from 104 patients were included in this study. The mean age of onset was 24.0±10.1 years (range: 5-40 y), with male predominance (60.5%). Over 90% of patients had bilateral JOAG and 25% had a positive family history. Negative family history (adjusted odds ratio=4.59, P =0.02) and thick central corneal thickness were surgical predictors (every 10 µm adjusted odds ratio=1.29, P =0.01). Over 70% of cases needed glaucoma surgery. Trabeculectomy with Mitomycin-C was performed on 131 eyes (65.5%) with a cumulative probability of complete success of 71.0%, 57.8%, 39.2%, and 26.9% and qualified success of 86.3%, 73.6%, 64.8%, and 45.7% at 1, 3, 5, and 10 years, respectively. The mean follow-up after surgery was 94.9 ± 69.8 months (range: 13-153 mo). There were no serious postoperative complications. Myopia and the number of baseline glaucoma medications were significantly associated with surgical failure. CONCLUSIONS: Trabeculectomy with mitomycin C was the most common primary surgery performed in Thai patients with JOAG, and successfully reduced intraocular pressure without significant complications. Patients with thicker corneas were more likely to undergo surgery. By 10 years, half of the patients required additional surgery and risk factors for failure included myopia and the number of medications.
Xu CL, Adu-Brimpong J, Moshfeghi HP, Rosenblatt TR, Yu MD, Ji MH, Wang SK, Zaidi M, Ghoraba H, Michalak S, Callaway NF, Kumm J, Nudleman E, Wood EH, Patel NA, Stahl A, Lepore D, Moshfeghi DM. Telemedicine retinopathy of prematurity severity score (TeleROP-SS) versus modified activity score (mROP-ActS) retrospective comparison in SUNDROP cohort. Sci Rep 2023;13(1):15219.Abstract
Identifying and planning treatment for retinopathy of prematurity (ROP) using telemedicine is becoming increasingly ubiquitous, necessitating a grading system to help caretakers of at-risk infants gauge disease severity. The modified ROP Activity Scale (mROP-ActS) factors zone, stage, and plus disease into its scoring system, addressing the need for assessing ROP's totality of binocular burden via indirect ophthalmoscopy. However, there is an unmet need for an alternative score which could facilitate ROP identification and gauge disease improvement or deterioration specifically on photographic telemedicine exams. Here, we propose such a system (Telemedicine ROP Severity Score [TeleROP-SS]), which we have compared against the mROP-ActS. In our statistical analysis of 1568 exams, we saw that TeleROP-SS was able to return a score in all instances based on the gradings available from the retrospective SUNDROP cohort, while mROP-ActS obtained a score of 80.8% in right eyes and 81.1% in left eyes. For treatment-warranted ROP (TW-ROP), TeleROP-SS obtained a score of 100% and 95% in the right and left eyes respectively, while mROP-ActS obtained a score of 70% and 63% respectively. The TeleROP-SS score can identify disease improvement or deterioration on telemedicine exams, distinguish timepoints at which treatments can be given, and it has the adaptability to be modified as needed.
Raparia E, Ballios BG, Place EM, Husain D, Huckfeldt RM. RP2 X-LINKED RETINITIS PIGMENTOSA CARRIER STATE PRESENTING WITH VASCULAR LEAKAGE AND UNILATERAL MACULAR ATROPHY. Retin Cases Brief Rep 2023;17(5):533-537.Abstract
PURPOSE: We describe the unusual clinical presentation of a 33-year-old woman subsequently identified as a carrier of RP2-associated X-linked retinitis pigmentosa. METHODS: Case report. RESULTS: A 33-year-old woman without a known family history of retinal disease presented with unilateral reduced visual acuity and central scotoma in the left eye. Examination showed underlying macular atrophy in the left eye and a bilateral tapetal-like reflex. Full-field electroretinogram was abnormal in the left eye but normal in the right eye. Notable findings on wide-field imaging included bilateral peripheral vascular leakage on fluorescein angiography and a bilaterally symmetric radial pattern of hyperfluorescence on fundus autofluorescence. Genetic testing demonstrated a pathogenic variant in the gene RP2 confirming that she was a carrier of X-linked retinitis pigmentosa. CONCLUSION: We describe clinical features of the carrier state of RP2-XLRP and expand potential findings to include peripheral vascular leakage. This case highlights the importance of awareness of the carrier state, particularly if a family history cannot be provided.
Singla E, Jha UP, Muralidharan S, Singh RB, Ichhpujani P. Management of multi-surface ocular burns caused by molten iron. Trauma Case Rep 2023;48:100925.Abstract
Ocular thermal burns are medical emergencies that require immediate intervention before the standard management protocol, which involves obtaining a detailed history and performing an ophthalmic examination. In this case report, we report the clinical manifestations of ocular burns caused by molten iron and the steps taken for good clinical outcomes. The patient presented with an inferior epithelial defect and limbal and lower lid ischemia at four hours post-injury. Over the course of treatment, due to non-resolving epithelial defect and increased superior lid notching, amniotic membrane transplantation (AMT) and lid repair by pentagon wedge excision were performed. Following AMT, the corneal surface completely healed with residual opacity and neovascularization. Additionally, limbal ischemia was significantly reduced with the restoration of normal lid anatomy. Corneal burns initiate a cascade of inflammatory reactions disrupting the balance between pro- and anti-angiogenic factors, leading to corneal neovascularization. The eyelid damage can lead to necrosis of tissues with eschar formation and eventually quantitative tissue loss. Therefore, timely intervention is the key to the successful management of ocular burns.
Vu DM, Elze T, Miller JW, Lorch AC, VanderVeen DK, Oke I, Oke I. Risk Factors for Glaucoma Diagnosis and Surgical Intervention following Pediatric Cataract Surgery in the IRIS® Registry. Ophthalmol Glaucoma 2023;Abstract
PURPOSE: To compare demographic and clinical factors associated with glaucoma following cataract surgery (GFCS) and glaucoma surgery rates between infants, toddlers, and older children using a large ophthalmic registry. DESIGN: Retrospective cohort study. PARTICIPANTS: Patients in the IRIS® Registry (Intelligent Research in Sight) who underwent cataract surgery at ≤17 years old and between January 1, 2013 and December 31, 2020. METHODS: Glaucoma diagnosis and procedural codes were extracted from the electronic health records of practices participating in the IRIS Registry. Children with glaucoma diagnosis or surgery before cataract removal were excluded. The Kaplan-Meier estimator was used to determine the cumulative probability of GFCS diagnosis and glaucoma surgery after cataract surgery. Multivariable Cox regression was used to identify factors associated with GFCS and glaucoma surgery. MAIN OUTCOME MEASURES: Cumulative probability of glaucoma diagnosis and surgical intervention within five years after cataract surgery. RESULTS: The study included 6,658 children (median age 10.0 years; 46.2% female). The five-year cumulative probability of GFCS was 7.1% (95% CI 6.1%-8.1%) and glaucoma surgery was 2.6% (95% CI 1.9%-3.2%). The five-year cumulative probability of GFCS for children <1 year old was 22.3% (95% CI 15.7%-28.4%). Risk factors for GFCS included aphakia (HR 2.63; 95% CI 1.96-3.57), unilateral cataract (HR 1.48; 95% CI 1.12-1.96), and Black race (HR 1.61; 95% CI 1.12-2.32). The most common surgery was glaucoma drainage device (GDD) insertion (32.6%), followed by angle surgery (23.3%), cyclophotocoagulation (15.1%), and trabeculectomy (5.8%). CONCLUSIONS: GFCS diagnosis in children in the IRIS Registry was associated with young age, aphakia, unilateral cataract, and Black race. GDD surgery was the preferred surgical treatment, consistent with the World Glaucoma Association 2013 consensus recommendations for GFCS management.
Jayaram H, Kolko M, Friedman DS, Gazzard G. Glaucoma: now and beyond. Lancet 2023;402(10414):1788-1801.Abstract
The glaucomas are a group of conditions leading to irreversible sight loss and characterised by progressive loss of retinal ganglion cells. Although not always elevated, intraocular pressure is the only modifiable risk factor demonstrated by large clinical trials. It remains the leading cause of irreversible blindness, but timely treatment to lower intraocular pressure is effective at slowing the rate of vision loss from glaucoma. Methods for lowering intraocular pressure include laser treatments, topical medications, and surgery. Although modern surgical innovations aim to be less invasive, many have been introduced with little supporting evidence from randomised controlled trials. Many cases remain undiagnosed until the advanced stages of disease due to the limitations of screening and poor access to opportunistic case finding. Future research aims to generate evidence for intraocular pressure-independent neuroprotective treatments, personalised treatment through genetic risk profiling, and exploration of potential advanced cellular and gene therapies.
Azad AD, Yuan M, Weinert M, Rosenblatt TR, Miller JW, Lorch A. The Transition to Ophthalmology Residency: A National Survey of the Combined Ophthalmology PGY-1 Program. J Acad Ophthalmol (2017) 2023;15(2):e188-e196.Abstract
Background  In 2017, the Accreditation Council for Graduate Medical Education announced all ophthalmology residency programs would provide a combined transitional or joint preliminary program for first postgraduate year (PGY-1) residents, with mandatory implementation by 2023. Purpose  This study aimed to survey ophthalmology residency program directors, postgraduate year 2 (PGY-2) ophthalmology residents who were a part of the first, official combined ophthalmology PGY-1 year, and postgraduate year 3 (PGY-3) residents who were a PGY-1 resident the year prior to integration to evaluate characteristics and perspectives on the combined ophthalmology PGY-1 year. Methods  A national, internet survey-based study approved by the Association of University Professors of Ophthalmology (AUPO) was disseminated to the AUPO listserv of program directors (PDs) and PGY-2 and PGY-3 ophthalmology residents from July to August 2022 and then again April to June 2023. Results  Twenty-six PDs completed the survey (response rate 20.3% out of 128 PDs). Forty-one PGY-2 ophthalmology residents who underwent the combined ophthalmology PGY-1 year and 33 PGY-3 ophthalmology residents also completed the survey. Most PGY-1 curricula focused on exposure to comprehensive ophthalmology and provided indirect ophthalmoscope, slit lamp, and refraction skills training to residents. Early exposure to fundamentals and clinical workflows were commonly cited benefits to the integration. When PDs were surveyed about how well-prepared PGY-1 residents who went through the combined year are for the PGY-2 relative to the prior year's class, 16 (61.5%) responded "better prepared." PGY-2 residents also reported a relatively higher level of clinical preparedness and familiarity with ophthalmology co-residents than PGY-3 residents. Several areas of improvement cited by both PDs and residents were identified including a dedicated didactic curriculum and more time in ophthalmology during the PGY-1 year. Conclusions  We found an overall net benefit from the integration of the combined ophthalmology PGY-1 year. Benefits include early exposure to clinical skills and knowledge specific to ophthalmology, leading to increased confidence and preparedness for the rigorous transition to ophthalmology residency. We also identified many areas for improvement to optimize the PGY-1 year including a formal curriculum and additional time in ophthalmology. Programs should work closely with their residents, faculty, and non-ophthalmology PDs to refine the PGY-1 for the benefit of future ophthalmologists.
Dissing-Olesen L, Walker AJ, Feng Q, Barr HJ, Walker AC, Xie L, Wilton DK, Das I, Benowitz LI, Stevens B. FEAST: A flow cytometry-based toolkit for interrogating microglial engulfment of synaptic and myelin proteins. Nat Commun 2023;14(1):6015.Abstract
Although engulfment is a hallmark of microglia function, fully validated platforms that facilitate high-throughput quantification of this process are lacking. Here, we present FEAST (Flow cytometric Engulfment Assay for Specific Target proteins), which enables interrogation of in vivo engulfment of synaptic material by brain resident macrophages at single-cell resolution. We optimize FEAST for two different analyses: quantification of fluorescent material inside live cells and of engulfed endogenous proteins within fixed cells. To overcome false-positive engulfment signals, we introduce an approach suitable for interrogating engulfment in microglia from perfusion-fixed tissue. As a proof-of-concept for the specificity and versatility of FEAST, we examine the engulfment of synaptic proteins after optic nerve crush and of myelin in two mouse models of demyelination (treatment with cuprizone and injections of lysolecithin). We find that microglia, but not brain-border associated macrophages, engulf in these contexts. Our work underscores how FEAST can be utilized to gain critical insight into functional neuro-immune interactions that shape development, homeostasis, and disease.
Yin Z, Rosenzweig N, Kleemann KL, Zhang X, Brandão W, Margeta MA, Schroeder C, Sivanathan KN, Silveira S, Gauthier C, Mallah D, Pitts KM, Durao A, Herron S, Shorey H, Cheng Y, Barry J-L, Krishnan RK, Wakelin S, Rhee J, Yung A, Aronchik M, Wang C, Jain N, Bao X, Gerrits E, Brouwer N, Deik A, Tenen DG, Ikezu T, Santander NG, McKinsey GL, Baufeld C, Sheppard D, Krasemann S, Nowarski R, Eggen BJL, Clish C, Tanzi RE, Madore C, Arnold TD, Holtzman DM, Butovsky O. APOE4 impairs the microglial response in Alzheimer's disease by inducing TGFβ-mediated checkpoints. Nat Immunol 2023;24(11):1839-1853.Abstract
The APOE4 allele is the strongest genetic risk factor for late-onset Alzheimer's disease (AD). The contribution of microglial APOE4 to AD pathogenesis is unknown, although APOE has the most enriched gene expression in neurodegenerative microglia (MGnD). Here, we show in mice and humans a negative role of microglial APOE4 in the induction of the MGnD response to neurodegeneration. Deletion of microglial APOE4 restores the MGnD phenotype associated with neuroprotection in P301S tau transgenic mice and decreases pathology in APP/PS1 mice. MGnD-astrocyte cross-talk associated with β-amyloid (Aβ) plaque encapsulation and clearance are mediated via LGALS3 signaling following microglial APOE4 deletion. In the brains of AD donors carrying the APOE4 allele, we found a sex-dependent reciprocal induction of AD risk factors associated with suppression of MGnD genes in females, including LGALS3, compared to individuals homozygous for the APOE3 allele. Mechanistically, APOE4-mediated induction of ITGB8-transforming growth factor-β (TGFβ) signaling impairs the MGnD response via upregulation of microglial homeostatic checkpoints, including Inpp5d, in mice. Deletion of Inpp5d in microglia restores MGnD-astrocyte cross-talk and facilitates plaque clearance in APP/PS1 mice. We identify the microglial APOE4-ITGB8-TGFβ pathway as a negative regulator of microglial response to AD pathology, and restoring the MGnD phenotype via blocking ITGB8-TGFβ signaling provides a promising therapeutic intervention for AD.
Chinn RN, Wilkinson CL, Staffa SJ, Michalak SM, Shoshany TN, Bishop K, Hunter DG, Gaier ED. Amblyopia treatment outcomes in patients with neurodevelopmental disorders. J AAPOS 2023;27(5):276.e1-276.e8.Abstract
PURPOSE: To compare amblyopia treatment outcomes between patients with neurodevelopmental disorders and their typically developing peers. METHODS: Of 2,311 patients diagnosed with amblyopia between 2010 and 2014 at Boston Children's Hospital, 460 met inclusion criteria (age 2-12 with anisometropic, strabismic, or mixed amblyopia [interocular difference (IOD) ≥2 lines]). Treatment and visual outcomes were analyzed according to neurodevelopmental status: neurodevelopmental delay (DD) versus typical development (TD). RESULTS: The DD group (n = 54) and TD group (n = 406) were similar in demographics, amblyogenic risk factors, baseline visual measures, prescribed therapy, and adherence (P ≥ 0.10). Between-visit follow-up time was longer for the DD group (0.65 [0.42- 0.97] years) than for the TD group (0.5 [0.36-0.82] years; P = 0.023). IOD improved similarly in each group by the last visit (DD, -0.15 logMAR [-0.31 to -0.02]; TD, -0.2 logMAR [-0.38 to -0.1]; P = 0.09). Each group reached amblyopia resolution by the last visit at similar frequencies (DD, 23/54 [43%]; TD, 211/406 [52%]; P > 0.2). DD diagnosis did not independently influence amblyopia resolution (HR, 0.77; 95% CI, 0.53-1.12; P = 0.17), but each additional month of interval time between follow-up visits reduced the likelihood of resolution by 2.7% (HR, 0.67; 95% CI, 0.51-0.87; P = 0.003). CONCLUSIONS: Patients with DD and those with TD responded similarly to amblyopia therapy; however, follow-up intervals were longer in patients with DD and correlated with the likelihood of persistent amblyopia, suggesting that greater efforts at assuring follow-up may benefit patients with DD.
Cullen PF, Mazumder AG, Sun D, Flanagan JG. Rapid isolation of intact retinal astrocytes: a novel approach. Acta Neuropathol Commun 2023;11(1):154.Abstract
Astrocytes are a major category of glial support cell in the central nervous system and play a variety of essential roles in both health and disease. As our understanding of the diverse functions of these cells improves, the extent of heterogeneity between astrocyte populations has emerged as a key area of research. Retinal astrocytes, which form the direct cellular environment of retinal ganglion cells somas and axons, undergo a reactive response in both human glaucoma and animal models of the disease, yet their contributions to its pathology and progression remain relatively unknown. This gap in knowledge is largely a function of inadequate isolation techniques, driven in part by the sparseness of these cells and their similarities with the more abundant retinal Müller cells. Here, we present a novel method of isolating retinal astrocytes and enriching their RNA, tested in both normal and ocular hypertensive mice, a common model of experimental glaucoma. Our approach combines a novel enzyme assisted microdissection of retinal astrocytes with selective ribosome immunoprecipitation using the Ribotag method. Our microdissection method is rapid and preserves astrocyte morphology, resulting in a brief post-mortem interval and minimizing loss of RNA from distal regions of these cells. Both microdissection and Ribotag immunoprecipitation require a minimum of specialized equipment or reagents, and by using them in conjunction we are able to achieve > 100-fold enrichment of astrocyte RNA.

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