Roadmap Out of COVID-19.

The recent coronavirus disease (COVID-19) was declared as a public health emergency by the World Health Organisation on 30th January 2020, and has now affected more than 100 countries. Healthcare institutions and governments worldwide have raced to contain the disease, albeit to varying degrees of success. Containment strategies adopted range from complete lockdowns to remaining open with public advisories regarding social distancing. However, general principles adopted by most countries remain the same, mainly to avoid gatherings in large numbers and limit social interactions to curb the spread of disease. In Singapore, this disease had a very different progression. The first wave of the disease started with the confirmation of the first COVID-19 positive patient in Singapore on 23rd January 2020. Initially, the daily number of confirmed cases were low and manageable. With a rise in unlinked cases, the Disease Outbreak Response System Condition (DORSCON) status was raised from yellow to orange. New cluster outbreaks in foreign worker dormitories led to the rampant spread of disease, with daily spikes of COVID-19 cases. As of 7th June 2020, we have a total of 37,910 confirmed cases of COVID-19 infections, the highest in Southeast Asia, 12,999 active cases and a manageable mortality count of 25 deaths. This details our unique method for dealing with a pandemic, including a brief demographic of trauma patients during this period. We were able to conserve sufficient resources to ensure that our essential services can still continue. Moving on, we have to ensure the continued protection of our population, especially the vulnerable groups such as the elderly and the immunocompromised, as we reopen.

Mutants of PC12 cells with altered cyclic AMP responses.

PC12 cells, derived from a rat pheochromocytoma, were mutagenized and selected in media containing agents known to elevate intracellular concentrations of cyclic AMP (cAMP). More than 40 clones were isolated by selection with cholera toxin or 2-chloroadenosine or both. The variants that were deficient in accumulating cAMP were obtained by using a protocol in which 1 microM 8-bromo-cAMP was included in addition to the agonist. Certain of these variants were partially characterized with respect to the site of altered cAMP metabolism. The profiles of adenylate cyclase activity responsiveness of certain variants to guanosine-5'-(beta, gamma-imido) triphosphate and to forskolin resembled those of UNC and cyc phenotypes of S49 lymphoma cells, which are functionally deficient in the GTP-sensitive coupling protein, Ns. Other variants were characterized by increased cyclic nucleotide phosphodiesterase activity at low substrate concentration. Diverse morphological traits were observed among the variants, but it was not possible to assign them to a particular cAMP phenotype. Two revertants of a PC12 mutant were isolated and observed to have regained a cellular cAMP response to 2-chloroadenosine and to forskolin. It is hoped that these PC12 mutants will have utility for defining cAMP-mediated functions, including any links to the action of nerve growth factor, in cells derived from the neural crest.

Roadmap Out of COVID-19

@#The recent coronavirus disease (COVID-19) was declared as a public health emergency by the World Health Organisation on 30th January 2020, and has now affected more than 100 countries. Healthcare institutions and governments worldwide have raced to contain the disease, albeit to varying degrees of success. Containment strategies adopted range from complete lockdowns to remaining open with public advisories regarding social distancing. However, general principles adopted by most countries remain the same, mainly to avoid gatherings in large numbers and limit social interactions to curb the spread of disease. In Singapore, this disease had a very different progression. The first wave of the disease started with the confirmation of the first COVID-19 positive patient in Singapore on 23rd January 2020. Initially, the daily number of confirmed cases were low and manageable. With a rise in unlinked cases, the Disease Outbreak Response System Condition (DORSCON) status was raised from yellow to orange. New cluster outbreaks in foreign worker dormitories led to the rampant spread of disease, with daily spikes of COVID-19 cases. As of 7th June 2020, we have a total of 37,910 confirmed cases of COVID-19 infections, the highest in Southeast Asia, 12,999 active cases and a manageable mortality count of 25 deaths. This details our unique method for dealing with a pandemic, including a brief demographic of trauma patients during this period. We were able to conserve sufficient resources to ensure that our essential services can still continue. Moving on, we have to ensure the continued protection of our population, especially the vulnerable groups such as the elderly and the immunocompromised, as we reopen.

Expression of vitreous cytokines in proliferative vitreoretinopathy: a prospective study.

PURPOSE: Proliferative vitreoretinopathy (PVR) is a major cause of failure of retinal detachment surgery. It is believed to be a wound-healing process in the retina. Many of the cellular functions are influenced by cytokines and growth factors such as interleukins (ILs). The present study was conducted to investigate the presence of transforming growth factor-beta 2 (TGF-beta2), basic fibroblast growth factor (bFGF), IL-1beta, IL-6, and protein in the vitreous of patients with retinal detachment and to determine the value of these mediators in predicting the future development of PVR. METHODS: A prospective study was conducted in 140 consecutive patients with rhegmatogenous retinal detachment in whom vitrectomy was considered necessary. Vitreous samples were analyzed for the presence of TGF-beta2, bFGF, IL-1beta, IL-6, and protein. Patients were then followed up for 3 months for the development of postoperative PVR. RESULTS: The mean levels of TGF-beta2, bFGF, IL-1beta, and protein in the vitreous were significantly higher (P < 0.05) in patients with preoperative PVR compared with those without. The mean levels of TGF-beta2, bFGF, IL-6, and protein in the vitreous were significantly higher (P < 0.05) in patients who had postoperative PVR compared with those who did not. Multivariate logistic regression analysis showed IL-6 and protein to be significant (P < 0.05), independent, predictive risk factors for the development of PVR. CONCLUSIONS: The various cytokines may play a role in the pathobiology of PVR. High vitreous levels of IL-6 and protein were identified as significant risk factors for PVR. A model was developed to predict the probability of development of postoperative PVR in these patients, and it may be used to indicate intravitreal pharmacologic treatment for those at risk.

A prospective study of matrix metalloproteinases in proliferative vitreoretinopathy.

PURPOSE: The migration, proliferation, differentiation, and adhesion of cells and other cellular functions are influenced by the surrounding extracellular matrix, in normal and wound-healing conditions. The matrix metalloproteinases (MMPs) are a family of enzymes that degrade and remodel the extracellular matrix and, thus, play a central role in the wound-healing process. Proliferative vitreoretinopathy (PVR), a wound-healing process in the retina, is a major cause of the failure of retinal detachment surgery. The role of MMPs in the pathobiology of PVR is unknown. We have investigated the presence of MMPs in the vitreous of patients with retinal detachment and the predictive value of MMPs for the future development of PVR. METHODS: A prospective study was conducted on 140 consecutive patients with a rhegmatogenous retinal detachment in whom vitrectomy was considered necessary because of a giant retinal tear and the presence of preoperative PVR, among other reasons. Vitreous samples were obtained and analyzed by zymography for the presence of MMPs. The patients were then followed up for the development of postoperative PVR (mild and severe). RESULTS: Two species of MMPs were detected in the vitreous: MMP-2 and MMP-9. MMP-2 was detected in all of the vitreous samples obtained, whereas MMP-9 was found in only 64 (47%) of 136 samples. The levels of MMPs detected were not significantly associated with the presence of preoperative PVR (P > 0.05), but they were significantly associated (P < 0.05) with the development of postoperative PVR (mild and severe). CONCLUSIONS: The results from this prospective study suggest that MMPs may be an important predictor and may also play a role in the development of postoperative PVR.

A new way to use the Ishihara test.

The Ishihara plates are widely used as a test for colour vision. Originally designed for the purpose of detecting congenital red-green colour blindness, the test also has some value in demonstrating acquired colour vision defects. There are, however, several disadvantages in the present arrangement of the plates. A modification of the test, involving the rearrangement of the order of the plates, is presented which, together with a new recording chart, simplifies both the administration and the interpretation of the test.

Risk factors for proliferative vitreoretinopathy after primary vitrectomy: a prospective study.

AIM: To assess clinical variables and vitreous protein as risk factors for the development of postoperative proliferative vitreoretinopathy (PVR). METHODS: A prospective study was conducted on 140 patients with a rhegmatogenous retinal detachment in whom a primary vitrectomy was performed. 12 clinical variables were recorded and vitreous samples obtained for measurement of protein concentration. Univariate and multivariate logistic regression analysis was used to determine the risk factors for PVR. RESULTS: Complete data were available for 136 of 140 patients. 40 of the 136 patients (29.4%) developed postoperative PVR. Univariate regression revealed that significant (p<0.05) risk factors included aphakia, presence of preoperative PVR, size of detachment, the use of silicone oil, and high vitreous protein level. Multivariate regression analysis revealed only aphakia (odds ratio 2.72), the presence of preoperative PVR (odds ratio 3.01), and high vitreous protein concentration (odds ratio 1.11) to be significant (p<0.05) independent, predictive risk factors for the development of PVR. CONCLUSIONS: This study has shown that the significant risk factors for PVR are preoperative PVR, aphakia, and high vitreous protein levels. Two models (clinical factors only and clinical factors and vitreous protein) were constructed to predict the probability of developing postoperative PVR and may be used to identify those at risk for possible intravitreal pharmacological treatment.

Effects of single, short-term exposures of human retinal pigment epithelial cells to thiotepa or 5-fluorouracil: implications for the treatment of proliferative vitreoretinopathy.

AIM: To investigate the effects of single, short-term (5 or 30 minutes) exposures to thiotepa or 5-fluorouracil (5-FU) on collagen lattice contraction and retinal pigment epithelial (RPE) cell proliferation. METHODS: For collagen contraction studies, RPE cells seeded into free floating type I collagen lattices were exposed to single 5 or 30 minute treatments with thiotepa (0.06-4 mg/ml), or 5-FU (0.25-25 mg/ml), or phosphate buffered saline alone as a control. For proliferation studies, RPE cell monolayers were similarly exposed to these agents. The degree of contraction, effects on cell number, and viability were determined up to 14 days after treatment. RESULTS: Contraction of collagen lattices containing RPE cells and proliferation of RPE cells were significantly inhibited (p < 0.05) by thiotepa and 5-FU at concentrations above 0.06 mg/ml and 0.25 mg/ml respectively (for both 5 and 30 minute treatments), compared with controls. Cell death did not occur except for exposure of the RPE cells in collagen lattices to the highest concentration of thiotepa (4 mg/ml). CONCLUSION: It was concluded that single 5 or 30 minute exposures to thiotepa or 5-FU significantly inhibited collagen contraction and the proliferation of RPE cells. These findings suggest that short, single, non-toxic exposures to thiotepa or 5-FU which can be reproduced clinically may be useful in the modulation of proliferative vitreoretinopathy.

Silicone oil concentrates fibrogenic growth factors in the retro-oil fluid.

AIM: To determine whether silicone oil concentrates protein and growth factors in the retro-oil fluid. METHODS: A laboratory analysis of intraocular fluid and vitreous specimens obtained from patients undergoing removal of silicone oil, revision vitrectomy, or primary vitrectomy for macular hole, proliferative vitreoretinopathy (PVR), or retinal detachment. Patients were prospectively recruited from routine vitreoretinal operating lists. Vitreous cavity fluid and vitreous samples were analysed for the presence of transforming growth factor beta (TGF-beta2), basic fibroblast growth factor (bFGF), interleukin 6 (IL-6), and total protein using either commercially available enzyme linked immunosorbent assays (ELISA) or protein assay kits. RESULTS: The median levels of bFGF, IL-6, and protein in the retro-oil fluid were raised (p<0.05) compared to all the other vitreous and vitreous cavity fluid samples. bFGF, IL-6, and protein levels were raised in PVR vitreous compared to non-PVR vitreous. TGF-beta2 levels were not significantly raised in retro-oil fluid or in PVR vitreous. CONCLUSIONS: The concentration of fibrogenic (bFGF) and inflammatory (IL-6) growth factors and protein is raised in retro-silicone oil fluid. This may contribute to the process of retro-oil perisilicone proliferation and subsequent fibrocellular membrane formation.

Systemic conditions in the Emergency Eye Clinic.

PURPOSE: A systemic disease may be diagnosed for the first time in patients who present to the eye department with ophthalmological manifestations. This study documents the pattern of systemic disorders which were encountered in an emergency eye clinic for the first time. SUBJECTS AND METHOD: Over a period of six months details of all patients who had a previously unknown systemic condition diagnosed in the Emergency Eye Clinic were recorded. Patients were seen in this clinic upon referral from other practitioners. All patients, with the exception of those with minor ophthalmic problems, were screened for hypertension and glycosuria. The patients were subsequently referred to their general practitioner or the appropriate hospital specialist for further management. RESULTS: During this period 1025 new patients visited the emergency eye clinic. Previously undiagnosed systemic disease was present in 86 patients (8.4%). The commonest disorders were neurological 35 (41%), cardiovascular 17 (20%), glycosuria 13 (15%) and respiratory 5 (6%). Visual disturbance and pain were the most frequent ocular presentation. In 54 patients (62.8%) the ocular condition was a direct manifestation of the systemic condition and in 32 patients (37.2%) the systemic disease was a coincidental finding. CONCLUSIONS: Awareness of ophthalmological manifestations of systemic disorders helps in the early diagnosis of these conditions and reduces the resulting morbidity and possibly mortality. This study illustrates the hidden role of the emergency eye clinic in identifying patients with an underlying systemic condition.

Macular hole surgery without prone positioning.

PURPOSE: To investigate the role of vitrectomy without prone posturing in the anatomic and functional outcome of macular hole surgery (MHS). METHODS: Forty-one consecutive eyes of 41 patients with stage II-IV full-thickness macular holes underwent pars plana vitrectomy and 16% C3F8 tamponade. In 25 cases posturing group (P), subjects were instructed to assume prone positioning for 10 days postoperatively, whereas in 16 cases non-posturing group (NP) patients were advised to avoid lying supine but no other posturing instructions were given. Preoperative, intraoperative and postoperative clinical data were collected, with macular hole closure rate and change in LogMAR visual acuity, contrast sensitivity, metamorphopsia, and 25-Visual Function Questionnaire (VFQ-25) being the primary outcome measures. RESULTS: Over a mean follow-up of 4.21.2 months, anatomical hole closure was noted in 22/25 (88%) and 14/16 (87.5%) in groups P and NP respectively. Visual acuity improved by a mean of eight letters and there was no significant difference in the two groups (P=0.724). Similarly, postoperative prone posturing did not have an effect on the final contrast sensitivity, metamorphopsia, and VFQ-25 composite scores (P=0.238, P=0.472, and P=0.87, respectively). However, eyes in group NP developed significantly more severe cataract in the early postoperative period (P=0.02). CONCLUSIONS: Prone posturing following MHS provides no functional or anatomic benefit but it is associated with slower progression of cataract. Combined phacovitrectomy without face down positioning may be considered for all phakic patients undergoing MHS.

Glutamine and glutamate metabolism in normal and heat shock conditions in Drosophila Kc cells: conditions supporting glutamine synthesis maximize heat shock polypeptide expression.

We have previously reported that Drosophila Kc cells require glutamine for maximal expression of heat shock proteins in stressed conditions (Sanders and Kon: J. Cell. Physiol. 146:180-190, 1991). The mechanism of this effect has been investigated by comparing the metabolic utilization of glutamine in conditions which support hsp expression with that of glutamate in conditions where up to 100-fold less hsp is synthesized. This comparison showed that free ammonia was generated by cells incubated in the presence of glutamine in 37 degrees C (heat shock) conditions, but not at 25 degrees C, and not in the presence of glutamate in either normal or heat shock conditions. There was no difference in the amount of [14C]O2 generated from either [14C]-labeled amino acid in the tricarboxylic acid cycle, but three- to four-fold more alanine was synthesized in cells incubated in glutamine than in glutamate. Treating the cells with aminotransferase inhibitors to artificially increase NH3 release raised hsp expression in the presence of glutamate to maximal levels characteristic of glutamine. This potentiation correlated with inhibition of alanine aminotransferase. Since only NH3 production correlated with hsp expression in heat shock conditions in the presence of glutamine, and NH3 addition to glutamate also resulted in maximal hsp expression, we measured glutamine production in glutamate plus NH3 and observed net glutamine synthesis. The supposition that glutamine itself is responsible for the regulatory changes supporting maximal hsp expression was supported by the finding that the glutamine analog, 6-diazo-5-oxo-L-norleucine (DON), mimicked the effects of glutamine. We conclude that glutamine imposes regulatory changes which alter nitrogen metabolism and support hsp expression in Kc cells.

A new colour vision test for clinical use.

Several tests are available for assessing colour vision but they can be expensive, complicated or too time consuming to perform. We have produced a new plate test based on pseudoisochromatic principles. The test, using an error score, examines both the red-green and blue-yellow axes, with four levels of difficulty for each axis. Results from a pilot study show that error scores from congenital red-green blind subjects are significantly higher than those of age-matched controls (p < 0.01) only when using the red-green plates and not the blue-yellow plates. In optic neuritis patients, error scores using both the red-green and blue-yellow plates were significantly higher than those of controls throughout the 6 month follow-up. The test, including scoring, takes 6 minutes to complete. These preliminary results suggest that the new test is effective for screening congenital red-green blindness and monitoring colour vision defects in acquired diseases such as optic neuritis.

Potentiation of guanosine 3':5'-monophosphate accumulation in C-6 glial tumor cells.

C-6 glial tumor cells treated with norepinephrine and sodium azide accumulated cyclic GMP to concentrations approximately 10-fold greater than the sum of the separate responses. Isoproterenol, but not phenylephrine, was an effective substitute for norepinephrine, and the response was blocked by propranolol and sotalol. Nitroprusside, but neither cyanide nor isobutyl-methylxanthine, replaced azide. The potentiation was not affected by removal of CA2+ OR Na+ from the extracellular medium and was not blocked by cocaine. The potentiative accumulation of cyclic GMP in C-6 cells differs from the recently described stimulation by catecholamines of soluble guanylate cyclase of renal cortex. The potentiative phenomenon is compared with the few known instances in which cyclic AMP augments cyclic GMP formation and may be associated with synergistic modifications of cellular functions.

Glutamine is a powerful effector of heat shock protein expression in Drosophila Kc cells.

We have investigated the effects of extracellular anions on the regulation of expression of the heat shock response in Drosophila Kc cells incubated in defined balanced salt solutions. Widely varying chloride concentrations had no effect on normal or heat shock protein (hsp) expression. Increasing glutamate concentrations from zero to 15 mM increased hsp expression more than 100-fold while affecting expression of non-heat-shock proteins minimally. Glutamine was 20-100-fold more potent than glutamate in supporting hsp expression, while other amino acids were less effective or supported no detectable hsp synthesis in heat shock. Inhibition of glutamine synthetase with methionine-sulfoximine resulted in very low hsp expression with glutamate and normal high level expression with glutamine, confirming the importance of glutamine. The absence of glucose and treatment with 2-deoxyglucose did not change the requirement for adequate glutamine for hsp expression. Cells heat shocked under conditions which gave very low hsp expression resumed growth when returned to normal medium as well as cells which expressed normal levels of hsps. Measurements of free amino acid levels in cells heat shocked in the presence and absence of glutamine showed a correlation between glutamine levels and amount of hsp expression. We conclude that a physiological process regulated by glutamine or a glutamine metabolite is important for normal hsp expression in heat shock conditions in Drosophila.

Phenylephrine and pilocarpine in the treatment of post-operative irido-corneal adhesion.

Following cataract surgery, entrapment of the iris within the surgical wound is often managed by intensive use of miotics. As the radial fibres stretch, only a small amount of fraction is exerted upon the entrapped iris. Application of a combination of phenylephrine and pilocarpine drops causes simultaneous contraction of the pupil sphincter and the radial muscle fibres. This study investigated the relative magnitude of forces induced in the iris periphery by pilocarpine and phenylephrine and the effectiveness of adding g. phenylephrine 10% to g. pilocarpine 4% drops in the treatment of postoperative irido-corneal adhesions. The investigation was divided into two parts. First, the forces induced in the iris periphery upon exposure to pilocarpine and phenylephrine were measured in 6 cadaver irises. The mean force was 27.5 +/- 5.7 x 10(-3) N for pilocarpine and 23.3 +/- 4.0 x 10(-3) N for phenylephrine. The combination of the two drugs produced a force of 54.2 +/- 6.6 x 10(-3) N (p < 0.05). In the second part of the study intensive pilocarpine 4% drops were administered to 17 patients who had iris-wound entrapment on the first post-operative day. Patients with persistent adhesion were commenced on intensive g. phenylephrine 10% and assessed after 90 minutes. Of the 17 patients, 6 responded to pilocarpine drops alone; in a further 7 the irido-corneal adhesion was released only by the addition of phenylephrine drops, and in 4 patients drops were ineffective in relieving the adhesion. This study indicates that addition of phenylephrine 10% to pilocarpine 4% drops enhances the effectiveness of pharmacological treatment of post-operative irido-corneal adhesion.

Transpupillary thermotherapy for the treatment of subfoveal choroidal neovascularization associated with age-related macular degeneration.

OBJECTIVE: To evaluate the efficacy of transpupillary thermotherapy (TTT) for the treatment of choroidal neovascularization (CNV) secondary to age-related macular degeneration (AMD). DESIGN: Prospective, non-randomized, non-masked, case-selected series. METHODS: All patients with subfoveal CNV due to AMD and initial visual acuity (VA) between 6/9 and 6/60 were offered the opportunity to undergo TTT. Recruited subjects were treated using a diode laser (810 nm) with a beam size of 1.2-3.0 mm and power settings of 460-1200 mW. Treatment was applied for 60 seconds in a subthreshold manner. MAIN OUTCOME MEASURES: Differences in VA and changes in the angiographic appearance of CNV. RESULTS: Thirty-one occult/predominantly occult and five classic/predominantly classic membranes were treated with TTT and were followed-up for a mean of 6.0 +/- 1.2 months. Following an average of 1.5 +/- 0.7 (range 1-4) laser sessions, VA remained stable ( -1 to +1 Snellen line) in 24 (66.7%) eyes, improved by > 1 line in two (5.6%) eyes and decreased significantly (> or = 2 Snellen lines) in 10 (27.8%) eyes. Angiographically confirmed closure of CNV was detected in 22 (61.1%) patients. Membranes persisted in 11 (30.6%) eyes and recurred in three (8.3%) eyes. There was no association between reduction, elimination or persistence of angiographic leakage of CNV and change in VA after treatment (p = 0.347). CONCLUSIONS: Transpupillary thermotherapy may be effective at preserving vision and reducing CNV leakage in a number of patients with exudative AMD. Further studies are required to compare TTT with the natural course of subfoveal CNV and alternative treatment options.