Yu Meng Wang

TBX6 Null Variants and a Common Hypomorphic Allele in Congenital Scoliosis

BACKGROUNDnCongenital scoliosis is a common type of vertebral malformation. Genetic susceptibility has been implicated in congenital scoliosis.nnnMETHODSnWe evaluated 161 Han Chinese persons with sporadic congenital scoliosis, 166 Han Chinese controls, and 2 pedigrees, family members of which had a 16p11.2 deletion, using comparative genomic hybridization, quantitative polymerase-chain-reaction analysis, and DNA sequencing. We carried out tests of replication using an additional series of 76 Han Chinese persons with congenital scoliosis and a multicenter series of 42 persons with 16p11.2 deletions.nnnRESULTSnWe identified a total of 17 heterozygous TBX6 null mutations in the 161 persons with sporadic congenital scoliosis (11%); we did not observe any null mutations in TBX6 in 166 controls (P<3.8×10(-6)). These null alleles include copy-number variants (12 instances of a 16p11.2 deletion affecting TBX6) and single-nucleotide variants (1 nonsense and 4 frame-shift mutations). However, the discordant intrafamilial phenotypes of 16p11.2 deletion carriers suggest that heterozygous TBX6 null mutation is insufficient to cause congenital scoliosis. We went on to identify a common TBX6 haplotype as the second risk allele in all 17 carriers of TBX6 null mutations (P<1.1×10(-6)). Replication studies involving additional persons with congenital scoliosis who carried a deletion affecting TBX6 confirmed this compound inheritance model. In vitro functional assays suggested that the risk haplotype is a hypomorphic allele. Hemivertebrae are characteristic of TBX6-associated congenital scoliosis.nnnCONCLUSIONSnCompound inheritance of a rare null mutation and a hypomorphic allele of TBX6 accounted for up to 11% of congenital scoliosis cases in the series that we analyzed. (Funded by the National Basic Research Program of China and others.).

Comparison of corneal dynamic parameters and tomographic measurements using Scheimpflug imaging in keratoconus

Aim To compare the diagnostic ability of corneal tomography and corneal dynamic response measurements in normal and keratoconus eyes. Methods Consecutive patients with grade II–III keratoconus and age-matched normal subjects were recruited. Corneal imaging was performed using Pentacam (Oculus Optikgeräte, Wetzlar, Germany) and Corvis (Oculus Optikgeräte). A beta version of Corvis software was used with three additional parameters: maximal change of arc length, deformation amplitude (DA) ratio 1 and DA ratio 2. Diagnostic ability of both devices to differentiate normal and keratoconus eyes was evaluated using receiver-operating characteristic (ROC) curves. The areas under the ROC curve (AUC) and partial AUC (pAUC) for specificity ≥80% for each parameter of Corvis and final D value of Belin/Ambrosio Enhanced Ectasia Display (BAD) were compared. Results Forty-two eyes of 42 patients (21 patients with keratoconus and 21 normal subjects) were included. Both groups were age matched (p=0.760). The ROC analysis showed that the final D value of BAD had the highest AUC (0.994) and pAUC (0.194). Maximum inverse radius had the highest AUC (0.954) but a relatively lower pAUC (0.158), while DA ratio 2 had the second highest AUC (0.946) together with the highest pAUC (0.177) among Corvis parameters. There was no significant difference between AUC and pAUC of BAD compared with those of DA ratio 1 (p≥0.162) and DA ratio 2 (p≥0.208). Conclusions The results of our study suggest that Corvis measurements have the potential to differentiate keratoconus and normal eyes. The diagnostic ability of novel parameters on Corvis was comparable to Pentacam.

Genetic associations for keratoconus: a systematic review and meta-analysis

Genetic associations for keratoconus could be useful for understanding disease pathogenesis and discovering biomarkers for early detection of the disease. We conducted a systematic review and meta-analysis to summarize all reported genetic associations for the disease. We searched in the MEDLINE, Embase, Web of Science, and HuGENET databases for genetic studies of keratoconus published from 1950 to June 2016. The summary odds ratio and 95% confidence intervals of all polymorphisms were estimated using the random-effect model. Among 639 reports that were retrieved, 24 fulfilled required criteria as eligible studies for meta-analysis, involving a total of 53 polymorphisms in 28 genes/loci. Results of our meta-analysis lead to the prioritization of 8 single-nucleotide polymorphisms (SNPs) in 6 genes/loci for keratoconus in Whites. Of them 5 genes/loci were originally detected in genome-wide association studies, including FOXO1 (rs2721051, Pu2009=u20095.6u2009×u200910−11), RXRA-COL5A1 (rs1536482, Pu2009=u20092.5u2009×u200910−9), FNDC3B (rs4894535, Pu2009=u20091.4u2009×u200910−8), IMMP2L (rs757219, Pu2009=u20096.1u2009×u200910−7; rs214884, Pu2009=u20092.3u2009×u200910−5), and BANP-ZNF469 (rs9938149, Pu2009=u20091.3u2009×u200910−5). The gene COL4A4 (rs2229813, Pu2009=u20091.3u2009×u200910−12; rs2228557, Pu2009=u20094.5u2009×u200910−7) was identified in previous candidate gene studies. We also found SNPs in 10 genes/loci that had a summary P valueu2009<u20090.05. Sensitivity analysis indicated that the results were robust. Replication studies and understanding the roles of these genes in keratoconus are warranted.

Comparison of Corneal Dynamic and Tomographic Analysis in Normal, Forme Fruste Keratoconic, and Keratoconic Eyes

PURPOSEnTo investigate and compare the diagnostic ability of corneal tomography and dynamic corneal response to differentiate between normal eyes and those with forme fruste keratoconus and keratoconus.nnnMETHODSnCorneal tomography was performed using Pentacam (Pentacam HR; Oculus Optikgeräte, Wetzlar, Germany). Corneal deformation response was captured via Corvis ST (Optikgeräte) using a beta version of Corvis software. Classification analysis between normal eyes and eyes with forme fruste keratoconus and between normal and keratoconic eyes was evaluated using receiver operating characteristic curves. The area under the ROC curve (AUC) and partial AUC (pAUC) for each classifying parameter were compared.nnnRESULTSnTwenty-one patients with forme fruste keratoconus in one eye and clinically evident keratoconus in the fellow eye and 38 normal individuals were recruited. Overall, 21 eyes with forme fruste keratoconus and 18 eyes with keratoconus were compared with 73 normal eyes. The mean age of the participants was comparable between groups. Comparative analysis between Pentacam and Corvis ST parameters showed significantly lower AUC and pAUC for Corvis ST parameters in differentiating keratoconic from normal eyes (P ≥ .049). However, comparable AUC and pAUC was observed between the Corvis Biomechanical Index (AUC = 0.785; pAUC = 0.079) and D value of the Belin/Ambrósio Enhanced Ectasia Display (AUC = 0.757; pAUC = 0.068) (P ≥ .477) for detection of forme fruste keratoconus with sensitivities of 63.2% and 52.6%, given a common specificity of 80.3%.nnnCONCLUSIONSnThe current study showed the feasibility of use of non-tomographical parameters obtained from the Corvis ST for differentiating normal eyes and those with forme fruste keratoconus and keratoconus. The diagnostic ability of the Corvis ST was comparable to that of the Pentacam for differentiating normal eyes and eyes with forme fruste keratoconus. [J Refract Surg. 2017;33(9):632-638.].

Shift in progression rate of keratoconus before and after epithelium-off accelerated corneal collagen crosslinking

PURPOSEnTo evaluate the shift in keratoconus progression rate after corneal collagen crosslinking (CXL).nnnSETTINGnChinese University of Hong Kong Eye Centre, Hong Kong, China.nnnDESIGNnProspective case series.nnnMETHODnAccelerated epithelium-off CXL was performed in patients with progressive keratoconus. Optical coherence tomography-based corneal measurements were recorded. The data collected after the first postoperative month were used for analysis. The main outcome measure was the rate of progression of steep keratometry (K), flat K, average K, and best-fit sphere (BFS) using linear mixed-effects models. Progression was defined based on a significant slope against time in these models.nnnRESULTSnForty-seven eyes (38 patients; mean age 27.6xa0yearsxa0±xa06.5 [SD]) were included. The median number ofxa0preoperative visits and postoperative visits was 3 and 5, respectively.xa0A significant reduction in the progression rate of the anterior average K (0.063xa0±xa00.138 diopter [D]/mo to -0.022xa0±xa00.029 D/mo) and posterior average K (-0.011xa0±xa00.025 D/mo to 0.000xa0±xa00.009xa0D/mo) occurred (both Pxa0<xa0.001). On univariate analysis, the baseline anterior average Kxa0had the strongest association with the progression rate of the anteriorxa0average K after CXL. On multivariate analysis, the preoperative progression rate of the posterior BFS was significantly associated with postoperative progression rate of the anterior (Pxa0<xa0.001) and posteriorxa0average K (Pxa0≤xa0.05).nnnCONCLUSIONSnA significant reduction in the progression ratexa0of average K occurred after accelerated epithelium-off CXL. The reduction was associated with the baseline anteriorxa0average K and preoperative progression of posterior BFS.

Comparative evaluation of progression rate in keratoconus before and after collagen crosslinking

Purpose To compare the rate of disease progression in keratoconus before and after corneal collagen crosslinking (CXL). Methods 145 eyes were followed without CXL (no-CXL group) for a median duration of 31 months whereas 45 eyes were followed up for 41 months before (pre-CXL) and after (post-CXL) accelerated, epithelium-off crosslinking. Progression was defined based on significant slope found in linear mixed effect models against time. Swept-source optical coherence tomography was used for measurement of anterior steep keratometry, anterior flat keratometry (Ant Kf), anterior average keratometry (Ant Avg K); posterior steep keratometry, posteriorflat keratometry (Post Kf), posterior average keratometry (Post Avg K) and corneal thickness. Results The patients in pre-CXL group were significantly younger (26.3±5.48 years) compared with the patients in no-CXL group (32.7±10.24 years) (P=0.004). Significant differences were observed during baseline examination for all parameters (P≤0.035) between pre-CXL and no-CXL groups except Ant Cyl and Post Cyl. During observation period, statistically significant differences were noted between pre-CXL and no-CXL groups in the progression rate of Ant Kf, Ant Avg K, Post Kf and Post Avg K (P≤0.045). After CXL, the progression rate in post-CXL group was comparable to that in no-CXL group. All corneal parameters remained stable in no-CXL group throughout the follow-up period. Conclusions Serial tomographic examination is useful to document disease progression before and after CXL. In our study, a decrease in progression rate of corneal parameters was noted after CXL. In cases with stable corneal parameters over time, careful monitoring can be considered instead of collagen crosslinking.

Comparison of Corneal Tomography and a New Combined Tomographic Biomechanical Index in Subclinical Keratoconus

PURPOSEnTo investigate and compare the diagnostic ability of corneal tomography and biomechanical and combined parameters for detection of corneal ectasia.nnnMETHODSnConsecutive patients with subclinical keratoconus (SCKC) and age-matched controls were included. Only one eye from each patient was selected for analysis. The final D value from the Belin/Ambrósio Enhanced Ectasia Display (BAD) was obtained from the Pentacam (Oculus Optikgeräte, Wetzlar, Germany). The tomographic biomechanical index (TBI) was derived from the Pentacam and Corvis ST (Oculus Optikgeräte). Classification analysis between normal and subclinical keratoconus (SCKC) was evaluated using receiver operating characteristic (ROC) curves. The area under the ROC curve (AUC) and partial AUC (pAUC) with specificity of 80% or greater were compared.nnnRESULTSnTwenty-three eyes with SCKC and 37 normal eyes were included. All Pentacam-derived parameters (P < .001) and all but two Corvis ST-derived parameters (P < .020) were significantly different between normal and SCKC eyes. A significant difference was found in the final D value (P ≤ .020) and TBI (P ≤ .040) between normal and SCKC eyes. For differentiating normal and SCKC eyes, TBI and BAD final D value demonstrated the highest AUC (0.925 and 0.786, respectively) and pAUC (0.150 and 0.088, respectively). TBI demonstrated 84.4% sensitivity and 82.4% specificity using a cut-off of 0.16. Comparative analysis between these parameters showed that AUC and pAUC of TBI were significantly higher than all parameters from Pentacam (P ≤ .032).nnnCONCLUSIONSnIn the current study, combined use of tomographic and biomechanical parameters demonstrated a higher capability in differentiating normal and SCKC eyes when compared to tomographic analysis alone. [J Refract Surg. 2018;34(9):616-621.].

Histological and microRNA Signatures of Corneal Epithelium in Keratoconus

PURPOSEnTo illustrate the histopathology of keratoconic corneal epithelia and its micro-ribonucleic acid (miRNA) regulation.nnnMETHODSnCorneal epithelia were collected from 27 patients with keratoconus and 26 normal patients after surgery or by impression cytology. The miRNA profile was determined using miRNA microarray. The biological roles of miRNA target genes were delineated by gene ontology and pathway analyses. The expressions of significant miRNAs were validated using TaqMan polymerase chain reaction (PCR), whereas protein localization and expression of the miRNA target genes were examined by immunofluorescence and immunoblotting analyses.nnnRESULTSnHistological assessment showed that corneal epithelia in patients with keratoconus were thinner with loosely packed cells compared to normal patients. Microarray analysis revealed that 12 miRNAs were significantly downregulated in keratoconic corneal epithelia. Gene ontology analysis demonstrated that the predicted miRNA target genes participated in cell junction, cell division, and motor activity, whereas pathway analysis highlighted the involvement of syndecan-mediated signaling pathway. TaqMan PCR validated the altered expression of six miRNAs in corneal epithelia from surgery (hsa-miR-151a-3p, hsa-miR-138-5p, hsa-miR-146b-5p, hsa-miR-194-5p, hsa-miR-28-5p, and hsa-miR-181a-2-3p) and four miRNAs in squamous corneal epithelial samples collected from impression cytology (hsa-miR-151a-3p, hsa-miR-195-5p, hsa-miR-185-5p, and hsa-miR-194-5p). In addition, higher S100A2 expression was found in the epithelial basal cell layer of keratoconic corneal epithelia.nnnCONCLUSIONSnThe miRNA and histological analyses in this study demonstrated structural and biological changes in keratoconic corneal epithelia, broadening the understanding of keratoconus pathology. In addition, impression cytology is useful to collect corneal epithelial tissues for gene expression analysis. [J Refract Surg. 2018;34(3):201-211.].

Analysis of multiple genetic loci reveals MPDZ-NF1B rs1324183 as a putative genetic marker for keratoconus

Objective To investigate the associations between 16 single-nucleotide polymorphisms (SNPs) in 14 genetic loci and keratoconus in an independent Chinese cohort. Methods This cross-sectional, case-control association study included a Chinese cohort of 133 patients with keratoconus and 371 control subjects. In a recent meta-analysis study, we identified association of 16 SNPs in 14 gene loci with keratoconus. In this study, we genotyped these 16 SNPs in all the patients and controls and analysed their association with keratoconus, its clinical severities and progression profiles. We also analysed the genotype-phenotype correlation between individual SNPs and steep keratometry, flat keratometry (Kf), average keratometry (Avg K) and best-fit sphere diameter (BFS) of the anterior and posterior corneal surface. Results Among the 16 selected SNPs, rs1324183 in the MPDZ-NF1B locus showed a significant association with keratoconus (OR=2.22; 95%u2009CI 1.42 to 3.45, p=4.30×10–4), especially severe keratoconus (OR=5.10, 95%u2009CI 1.63 to 15.93, p=0.005). The rs1324183 A allele was positively associated with anterior Kf (p=0.008), anterior Avg K (p=0.017), posterior Kf (p=0.01) and negatively associated with apex pachymetry (p=0.007) and anterior BFS (p=0.023) in keratoconus. The other 15 SNPs had no significant association with keratoconus or genotype-phenotype correlations. Conclusions This study confirmed the association of SNP rs1324183 in MPDZ-NF1B with keratoconus and revealed the association of this SNP with keratoconus severity and corneal parameters. It is thus a putative genetic marker for monitoring the progression of keratoconus to a severe form and facilitating early intervention.

Association of the PAX6 gene with extreme myopia rather than lower grade myopias

Aims To investigate the association of the paired box gene 6 (PAX6) with different severities of myopia. Methods A total of four haplotype-tagging single-nucleotide polymorphisms (SNPs; rs2071754, rs3026354, rs3026390 and rs628224) and two previously reported SNPs (rs644242 and rs662702) in the PAX6 gene were analysed in a Hong Kong Chinese cohort of 1288 myopia subjects (including 252 extreme myopia, 277 high myopia, 393 moderate myopia and 366 mild myopia) and 791 no myopia controls. Allelic association analyses were performed for individual SNPs in different subgroups of myopia and in combined myopia, followed by a meta-analysis of our current data with reported data on PAX6 in myopia. Results The association of tagging SNPs rs2071754 and rs644242 with extreme myopia could not withstand multiple correction (rs2071754: OR=1.25, P value=0.031; rs644242: OR=1.33, P value=0.032). In the meta-analysis, rs644242 showed an enhanced, significant association with extreme myopia (OR=1.27, 95%u2009CI 1.10 to 1.46, P value=0.001; I2=0%). In contrast, there was no significant association between the PAX6SNPs and high, moderate or mild myopia. No linear correlation was found between the PAX6SNPs and axial length. Conclusion This study provides additional evidence suggesting that the PAX6 SNP rs644242 is associated with extreme myopia but not lower grade myopia. Thus, PAX6 may be implicated in the development or progression into severe myopia. Further longitudinal studies are warranted.