MATERIALS AND METHODS: This study is a comparative, crosssectional and hospital-based study. We recruited 83 children with VKC and 83 healthy children as controls. All children underwent complete ocular examinations, CCT measurement using an ultrasound pachymeter and corneal topography using a Placido disc corneal analyser.
RESULTS: There was a statistically significant difference of means CCT and topographic indices in children with VKC compared to controls (p<0.05). The probability keratoconus reached 18% in children with VKC. The mean CCT was observed to be thinnest in the severe-to-very severe groups of VKC compared to the mild-to-moderate (p<0.05). The means simulated-K1 and -K2, apical keratometry, apical gradient curvature, superior-inferior index and keratoconus prediction index were significantly different in severe-tovery severe VKC compared to mild-to-moderate VKC and controls (p<0.05). However, there was no significant difference in mean cylinder value and percent probability keratoconus when comparing different groups of severity of VKC (p=0.912 and 0.070 respectively).
CONCLUSION: Children with VKC have thinner CCT and topographic indices changes compared to healthy children. Similar pattern was observed between groups with VKC. Degree of astigmatism and probability of keratoconus were similar in mild-to-moderate and severe-to-very severe groups.
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.
METHODS: A cross-sectional study was conducted to evaluate the corneal cell morphology of 47 keratoconus patients and 32 healthy eyes without any ocular disease. New keratoconus patients with different disease severities and without any other ocular co-morbidity were recruited from the ophthalmology department of a public hospital in Malaysia from June 2013 to May 2014. Corneal cell morphology was evaluated using an in vivo slit-scanning confocal microscope. Qualitative and quantitative data were analysed using a grading scale and the Nidek Advanced Visual Information System software, respectively.
RESULTS: The corneal cell morphology of patients with keratoconus was significantly different from that of healthy eyes except in endothelial cell density (P = 0.072). In the keratoconus group, increased level of stromal haze, alterations such as the elongation of keratocyte nuclei and clustering of cells at the anterior stroma, and dark bands in the posterior stroma were observed with increased severity of the disease. The mean anterior and posterior stromal keratocyte densities and cell areas among the different stages of keratoconus were significantly different (P < 0.001 and P = 0.044, respectively). However, the changes observed in the endothelium were not significantly different (P > 0.05) among the three stages of keratoconus.
CONCLUSION: Confocal microscopy observation showed significant changes in corneal cell morphology in keratoconic cornea from normal healthy cornea. Analysis also showed significant changes in different severities of keratoconus. Understanding the corneal cell morphology changes in keratoconus may help in the long-term monitoring and management of keratoconus.