METHODS: Keratoconic (n = 74) and control subjects (n = 96) were recruited based on clinical diagnostic tests and selection criteria. DNA extracted from the blood samples was used to genotype VSX1 polymorphisms. In-house designed primers and optimization of PCR conditions were carried out to amplify exons 1 and 3 of the VSX1 gene. PCR conditions including percentage GC content, melting temperatures, and differences in melting temperatures of primers were evaluated to produce sensitive and specific DNA amplifications.
RESULTS: Genotyping was successfully carried out in 4 exons of the VSX1 gene. Primer annealing temperatures were observed to be crucial in enhancing PCR sensitivity and specificity. Annealing temperatures were carefully evaluated to produce increased specificity, yet not allowing sensitivity to be compromised. In addition, exon 1 of the VSX1 gene was amplified using 2 different sets of primers to produce 2 smaller amplified products with absence of non-specific bands. DNA amplification of exons 1 and 3 consistently showed single band products which were successfully sequenced to yield reproducible data.
CONCLUSIONS: The use of in-house designed primers and optimized PCR conditions allowed sensitive and specific DNA amplifications that produced distinct single bands. The in-house designed primers and DNA amplification protocols established in this study provide an addition to the current repertoire of primers for accurate molecular characterization of VSX1 gene polymorphisms in keratoconus research.
MATERIALS AND METHODS: The tenon tissue was harvested from a patient undergoing strabismus surgery. The human tenon fibroblast cell culture and isolation were performed according to the standard laboratory cell culturing protocol. The cells were divided into three groups: control, treatment with irradiated and non-irradiated riboflavin. There were five different concentrations (0.00156%, 0.003125%, 0.00625%, 0.0125%, 0.025%) in each group of riboflavin. The fibroblasts were treated with riboflavin and the cellular viability was assessed at 24-hour and 48-hour post treatment with MTT 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyl tetrazolium bromide colorimetric assay. The absorbance values were analysed using Magellan microplate reader data analysis. A triplicate of readings was taken. The data were presented as mean ± standard deviation of the triplicates. Statistical analysis was performed with Statistical Package for Social Sciences (SPSS) analysis version 23.
RESULTS: Irradiated riboflavin caused a concentration-dependent cell death in human tenon fibroblast cell culture (p
DESIGN: Prospective analysis of all ocular trauma injuries presenting to the Department of Ophthalmology in UMMC from 1 January 2008 to 31 December 2008.
PARTICIPANTS: A total of 603 eyes of 546 patients were recruited for the study.
METHODS: All patients presenting to the department with ocular trauma injuries were assessed by an ophthalmologist. Data on the type and source of injury, demographic profile of the patients, and clinical presentation were documented using a uniform and validated datasheet.
RESULTS: Among eye injury cases, 481 patients (88.1%) were male, with a male-to-female ratio of 7.4:1. Of the patients, 412 (75.5%) were Malaysian while the remaining 134 (24.5%) were of non-Malaysian nationality. The average age was 31.5 years (range 1-81 years). A total of 238 injured eyes (43.6%) were work-related. The common sources of eye trauma include the use of high-powered tools (30.8%), motor vehicle accident (23.1%), and domestic accidents (17.7%). Only six patients (2.5%) reported to having used eye protective device (EPD) at time of their work-related injuries.
CONCLUSIONS: A major cause of preventable ocular injuries in Malaysia was work-related trauma. Ocular injuries can be reduced by the use of eye protection devices and the implementation of appropriate preventive strategies to address each risk factor. Effective training is an integral part of occupational safety and health, which should be made mandatory at the workplace. In addition, there should be a continual assessment of safety and health issues at the workplace. A long-term database of all ocular injuries in Malaysia is recommended, to aid research on a larger scale and the development of new preventive strategies for ocular injuries.