Robert J. Casson

Genome-wide association study identifies susceptibility loci for open angle glaucoma at TMCO1 and CDKN2B-AS1

We report a genome-wide association study for open-angle glaucoma (OAG) blindness using a discovery cohort of 590 individuals with severe visual field loss (cases) and 3,956 controls. We identified associated loci at TMCO1 (rs4656461[G] odds ratio (OR) = 1.68, P = 6.1 × 10−10) and CDKN2B-AS1 (rs4977756[A] OR = 1.50, P = 4.7 × 10−9). We replicated these associations in an independent cohort of cases with advanced OAG (rs4656461 P = 0.010; rs4977756 P = 0.042) and two additional cohorts of less severe OAG (rs4656461 combined discovery and replication P = 6.00 × 10−14, OR = 1.51, 95% CI 1.35–1.68; rs4977756 combined P = 1.35 × 10−14, OR = 1.39, 95% CI 1.28–1.51). We show retinal expression of genes at both loci in human ocular tissues. We also show that CDKN2A and CDKN2B are upregulated in the retina of a rat model of glaucoma.

Definition of glaucoma: clinical and experimental concepts

Glaucoma is a term describing a group of ocular disorders with multi‐factorial etiology united by a clinically characteristic intraocular pressure‐associated optic neuropathy. It is not a single entity and is sometimes referred to in the plural as the glaucomas. All forms are potentially progressive and can lead to blindness. The diverse conditions that comprise glaucoma are united by a clinically characteristic optic neuropathy: glaucomatous optic neuropathy (GON). Evidence suggests that the primary site of neurological injury is at the optic nerve head. This fact enables the conditions to be grouped, irrespective of the causal mechanism(s). The term experimental glaucoma implies model resemblance to the human condition. We propose that ‘experimental glaucoma’ be restricted to animal models with demonstrable features of GON and/or evidence of a primary axonopathy at the optic nerve head. A fundamental inadequacy in this framework is any reference to the pathogenesis of GON, which remains unclear.

Recharacterization of the RGC-5 Retinal Ganglion Cell Line

PURPOSE The transformed RGC-5 retinal ganglion cell line is used widely in glaucoma research. Increased resistance to glutamate was noted in published literature and led to the recharacterization of the RGC-5 cell line. METHODS Characterization of the RGC-5 cell line was performed by sequencing of a region of the nuclear Thy1 gene and mitochondrial DNA sequencing of a region of the d-loop and tRNA(Phe) gene. Marker expression was examined in undifferentiated cells, and cells differentiated with 50 microg/mL succinyl concanavalin A (S Con A) for 3 days. Glutamate sensitivity was examined in undifferentiated and S Con A differentiated cells by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay after 24-hours of glutamate treatment. RESULTS RGC-5 cells were found to be of mouse (Mus musculus), not rat (Rattus norvegicus), origin by mitochondrial and nuclear DNA analyses. RGC-5 DNA sequenced in a second laboratory was subsequently found to be of M. musculus origin. Cells stained positively for the neuronal markers beta-tubulin and PGP9.5 and for the microtubule-associated protein tau, but not for known markers of ganglion cells such as neurofilaments or Thy1.2, suggesting that they likely represented a lineage of mouse neuronal precursor cells. Differentiation with S Con A did not increase RGC-5 sensitivity to glutamate excitotoxicity or increase the expression of retinal or ganglion cell marker proteins. CONCLUSIONS Investigators using cells designated as RGC-5 should confirm the species to be of rat origin and retinal-specific marker expression before considering their use as retinal ganglion-like cells.

Microglial Activation in the Visual Pathway in Experimental Glaucoma: Spatiotemporal Characterization and Correlation with Axonal Injury

PURPOSE Glia are the main cellular CNS elements initiating defense mechanisms against destructive influences and promoting regenerative processes. The aim of the current work was to characterize the microglial response within the visual pathway in a rat model of experimental glaucoma and to correlate the microglial response with the severity of axonal degeneration. METHODS Experimental glaucoma was induced in each right eye of adult Sprague-Dawley rats by translimbal laser photocoagulation of the trabecular meshwork. Rats were subsequently killed at various times from 3 days to 6 weeks. Tissue sections were obtained from globes, optic nerves, chiasmata, and optic tracts for immunohistochemistry and toluidine blue staining. RESULTS This model of experimental glaucoma led to a marked activation of microglia in the retina, optic nerve, and tract. Indeed, microglial activity remained elevated, even after intraocular pressure returned to basal levels. It is postulated that this process accompanies ongoing axonal degeneration. The degree of activation in the optic nerve correlated with axonal damage. Activation was characterized by increased density and morphologic changes. Both major histocompatibility complex (MHC) class I and MHC class II surface proteins were persistently upregulated in optic nerves and localized to microglial cells; however, this did not correlate with any significant T-cell infiltration. Interestingly, MHC class II expression was not detected in the retina. CONCLUSIONS The present data may have implications for the study of the pathology associated with the visual pathway in diseases such as glaucoma.

Possible role of excitotoxicity in the pathogenesis of glaucoma

Excitotoxicity describes the process of neuronal injury by excess stimulation of amino acid receptors. This form of insult was first described in the retina, and subsequently has been shown to be an important component of the pathogenesis of ischaemic and traumatic injury in the central nervous system. Furthermore, there is increasing evidence that excitotoxicity is involved in several chronic neurological conditions, and anti‐excitotoxic treatment has already been approved for some of these conditions. A large‐scale trial is currently underway that will determine the efficacy of an anti‐excitotoxic drug (memantine) in the management of glaucoma. This review provides an overview of neurotransmission and the mechanisms of excitotoxicity. The evidence for excitotoxicity as a component of certain neurological diseases, including glaucoma, is discussed.

Mitomycin : Clinical applications in ophthalmic practice

Mitomycin (mitomycin C; MMC) is an antibiotic isolated from Streptomyces caespitosus. The drug is a bioreductive alkylating agent that undergoes metabolic reductive activation, and has various oxygen tension-dependent cytotoxic effects on cells, including the cross-linking of DNA. It is widely used systemically for the treatment of malignancies, and has gained popularity as topical adjunctive therapy in ocular and adnexal surgery over the past 2 decades. In ophthalmic medicine, it is principally used to inhibit the wound healing response and reduce scarring of surgically fashioned ostia. Hence, it has been used as adjunctive therapy in various ocular surgeries, such as glaucoma filtering surgeries, dacryocystorhinostomy, corneal refractive surgery and surgeries for ocular cicatrisation. In addition, it has been used as an adjunct in the surgical management of pterygia, ocular surface squamous neoplasia, primary acquired melanosis with atypia and conjunctival melanoma. In many of these surgeries and ophthalmic pathologies, MMC showed a significant beneficial effect.

Optic nerve and neuroprotection strategies

AbstractBackground Experimental studies have yielded a wealth of information related to the mechanism of ganglion cell death following injury either to the mylinated ganglion cell axon or to the ganglion cell body. However, no suitable animal models exist where injury can be directed to the optic nerve head region, particularly the unmylinated ganglion cell axons. The process of relating the data from the various animal models to many different types of optic neuropathies in man must, therefore, be cautious.Results Extensive studies on the isolated optic nerve have yielded valuable information on the way white matter is affected by ischaemia and how certain types of compounds can attenuate the process. Moreover, there are now persuasive data on how ganglion cell survival is affected when the ocular blood flow is reduced in various animal models. As a consequence, the molecular mechanisms involved in ganglion cell death are fairly well understood and various pharmacological agents have been shown to blunt the process when delivered before or shortly after the insult.Conclusions A battery of agents now exist that can blunt animal ganglion cell death irrespective of whether the insult was to the ganglion cell body or the mylinated axon. Whether this information can be applied for use in patients remains a matter of debate, and major obstacles need to be overcome before the laboratory studies may be applied clinically. These include the delivery of the pharmacological agents to the site of ganglion cell injury and side effects to the patients. Moreover, it is necessary to establish whether effective neuroprotection is only possible when the drug is administered at a defined time after injury to the ganglion cells. This information is essential in order to pursue the idea that a neuroprotective strategy can be applied to a disease like glaucoma, where ganglion cell death appears to occur at different times during the lifetime of the patient.

Long term effect on intraocular pressure of phacotrabeculectomy compared to trabeculectomy

Aim: To compare the long term mean intraocular pressure (IOP) reduction after non-augmented single site phacotrabeculectomy with that after trabeculectomy and to determine the relation between preoperative IOP and IOP reduction. Methods: A group of 44 consecutive patients with chronic open angle glaucoma who underwent phacotrabeculectomy were matched to a trabeculectomy control group and the results of surgery were compared. Linear regression analysis of preoperative IOP and IOP reduction was undertaken. Results: The mean IOP reduction was significantly less in the phacotrabeculectomy group (6.7 (SD 2.1) mm Hg) than in the trabeculectomy group (11.0 (1.4) mm Hg) (p=0.0017). There was a significant difference in surgical success between the groups. The preoperative IOP was significantly related to the postoperative reduction in IOP in both groups (p<0.001). Conclusions: In elderly white patients with chronic open angle glaucoma, phacotrabeculectomy is not as effective as trabeculectomy in reducing IOP. In both procedures the magnitude of IOP reduction is proportional to the preoperative IOP.

Comparison of the ICare rebound tonometer with the Goldmann applanation tonometer by experienced and inexperienced tonometrists.

PurposeTo assess the agreement between ICare® rebound tonometer and Goldmann applanation tonometer in the hands of experienced and inexperienced tonometrists.Patients and methodsTwo tonometrists, experienced with both Goldmann applanation tonometry (GAT) and ICare® Tonometry (ICT) measured intraocular pressure (IOP), in a masked fashion, in 100 patients. In another series of 58 patients, ICT was performed by an inexperienced tonometrist and GAT by an experienced tonometrist.ResultsIn approximately 80% of patients, the difference in IOP between GAT and ICT was ⩽2 mmHg in group 1 and ⩽3 mmHg in group 2. The 95% limits of agreement were −4.0–4.4 mmHg in group 1 and −6.0–5.0 mmHg in group 2.ConclusionICT compares reasonably with GAT, in both experienced and inexperienced hands. Its ease of use, portability, and sterility make it an attractive tonometer. Its degree of accuracy in inexperienced hands would make it a useful instrument for health care workers with limited ophthalmic experience.

Common variants near ABCA1 , AFAP1 and GMDS confer risk of primary open-angle glaucoma

Primary open-angle glaucoma (POAG) is a major cause of irreversible blindness worldwide. We performed a genome-wide association study in an Australian discovery cohort comprising 1,155 cases with advanced POAG and 1,992 controls. We investigated the association of the top SNPs from the discovery stage in two Australian replication cohorts (932 cases and 6,862 controls total) and two US replication cohorts (2,616 cases and 2,634 controls total). Meta-analysis of all cohorts identified three loci newly associated with development of POAG. These loci are located upstream of ABCA1 (rs2472493[G], odds ratio (OR) = 1.31, P = 2.1 × 10−19), within AFAP1 (rs4619890[G], OR = 1.20, P = 7.0 × 10−10) and within GMDS (rs11969985[G], OR = 1.31, P = 7.7 × 10−10). Using RT-PCR and immunolabeling, we show that these genes are expressed within human retina, optic nerve and trabecular meshwork and that ABCA1 and AFAP1 are also expressed in retinal ganglion cells.