Corneal metabolic biomarkers for moderate and high myopia in human.

This study aimed to identify the corneal metabolic biomarkers for moderate and high myopia in human. We enrolled 221 eyes from 221 subjects with myopia to perform the femtosecond laser small incision lenticule extraction (SMILE) surgery. Among these, 71 eyes of 71 subjects were enrolled in the low myopic group, 75 eyes of 75 subjects in the moderate myopic group and 75 eyes of 75 subjects in the high myopic group. The untargeted metabolomics analysis was performed to analyze the corneal tissues extracted during the SMILE surgery using an ultra-high-performance liquid chromatography (UHPLC) coupled to a quadrupole time-of-flight (Q-TOF) mass spectrometry (MS). The one-way analysis of variance (ANOVA) was used to identify the different metabolites among the three myopic groups, the orthogonal partial least-squares discriminant analysis (OPLS-DA) model was used to reveal the different metabolites between moderate myopia and low myopia, and between high myopia and low myopia. The Venn gram was used to find the overlapped metabolites of the three datasets of the different metabolites. The stepwise multiple linear regression analysis was used to determine the metabolic molecules associated with manifest refractive spherical equivalents (MRSE). The Receiver Operating Characteristics (ROC) analysis was performed to reveal the corneal biomarkers for moderate and high myopia. The hub biomarker was further selected by the networks among different metabolites created by the Cytoscape software. A total of 1594 metabolites were identified in myopic corneas. 321 metabolites were different among the three myopic groups, 106 metabolites were different between high myopic corneas and low myopic corneas, 104 metabolites were different between moderate myopic corneas and low myopic corneas, and 30 metabolic molecules overlapped among the three datasets. The multivariate linear regression analysis revealed the myopic degree was significantly influenced by the corneal levels of azelaic acid, arginine-proline (Arg-Pro), 1-stearoyl-2-myristoyl-sn-glycero-3-phosphocholine, and hypoxanthine. The ROC curve analysis showed that azelaic acid, Arg-Pro and hypoxanthine were effective in discriminating low myopia from moderate to high myopia with the area under the curve (AUC) values as 0.982, 0.991 and 0.982 for azelaic acid, Arg-Pro and hypoxanthine respectively. The network analysis suggested that Arg-Pro had the maximum connections among these three biomarkers. Thus, this study identified azelaic acid, Arg-Pro and hypoxanthine as corneal biomarkers to discriminate low myopia from moderate to high myopia, with Arg-Pro serving as the hub biomarker for moderate and high myopia.

Clinical Feasibility and Safety of Scleral Collagen Cross-Linking by Riboflavin and Ultraviolet A in Pathological Myopia Blindness: A Pilot Study.

INTRODUCTION: To investigate the feasibility and safety of scleral ultraviolet A (UVA) cross-linking (scleral CXL) on pathologically blindness. METHODS: This was a prospective, observational clinical study. Five patients with monocular blindness due to pathological myopic maculopathy were enrolled. Eyes with best corrected visual acuity (BCVA) under 0.05 were defined as experimental eyes. The fellow eyes were defined as control eyes. Patients first underwent posterior scleral reinforcement (PSR) surgery in the control eye. Thereafter, scleral CXL surgery was performed in the experimental eye on the same day. Visual acuity, BCVA, slit lamp biomicroscopic examination, intraocular pressure measurement, corneal specula microscopies, axis length measurement, funduscopy with pupil dilation, color fundus photography, full-field flash electroretinography, optical coherence tomography, and color Doppler flow imaging were performed at baseline, 1 week, 1 month, 3 months, 6 months, and 12 months after surgery. RESULTS: No signs of inflammation were observed after operation and throughout the follow-up period. Retinoschisis was improved, while choroidal neovascularization fibrosis and retinal and choroidal atrophy were unchanged after scleral CXL. There were no statistically significant differences in the ophthalmic artery, central retinal artery, and posterior ciliary artery parameters of color Doppler flow imaging or in retinal thickness, within experimental and control eyes, at baseline, 1 week, 1 month, 3 months, or 12 months (P > 0.05). CONCLUSIONS: This pilot study verified the feasibility and safety of scleral CXL on human blindness. The UVA-CXL on the sclera of human eyes seems to have the same effect as PSR in preventing progressive pathological myopia in the future. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR2100042422).

Safety and Long-term Scleral Biomechanical Stability of Rhesus Eyes after Scleral Cross-linking by Blue Light.

Purpose: To assess the safety and long-term scleral biomechanical stability of rhesus eyes after blue light scleral CXL by investigating the biomechanical and microstructural changes.Methods: Seven rhesus monkeys (14 eyes) were observed in this study. All right eyes received blue light scleral CXL at the superior temporal equatorial sclera, and the left eyes served as controls. Biological ocular parameters were followed up to 1 year after scleral CXL. Stress-strain measurements of three rhesus sclera were measured, three rhesus retinas were examined histologically by H&E and TUNEL staining. And the microstructure of both the sclera and retina were observed by transmission electron microscopy at 1 year.Results: As for the retinal thickness, choroidal thickness, flow density of retinal superficial vascular networks and flash electroretinography (f-ERG) results, no significant differences were observed between the paired eyes at 1 year (P >.05). At the same time, the scleral collagen fibril distribution was much tighter, and the scleral biomechanical properties were significantly increased in the experimental eyes. However, apoptotic cells and retinal ultrastructural changes could still be found in the retina of the experimental eyes.Conclusion: This study demonstrates that blue light scleral CXL could effectively increase the scleral stiffness of the rhesus eye for at least 1 year, but ultrastructural change was still observed in the retina of scleral CXL eye. Therefore, the long-term intraocular safety of the blue light scleral CXL technique for preventing myopia progression should be investigated further.

Evaluation of the Safety and Long-term Scleral Biomechanical Stability of UVA Cross-linking on Scleral Collagen in Rhesus Monkeys.

PURPOSE: To investigate the changes of retinal and choroidal parameters, scleral biomechanical strength, and ocular histopathology after scleral ultraviolet-A (UVA) cross-linking (CXL) in rhesus monkeys eyes, and to evaluate the safety and long-term biomechanical stability of scleral CXL for preventing myopia from progressing further in clinic. METHODS: Six 3-year-old male rhesus monkeys (12 eyes) were randomized to receive UVA-CXL procedures applied on the superotemporal equatorial sclera. Optical coherence tomography (OCT) and optical coherence tomography angiography (OCTA) were used for examination before and 1 week and 1, 3, 6, and 12 months after CXL. The stress-strain behaviors of equatorial scleral strips were analyzed 12 months postoperatively by a biomaterial tester. Hematoxylin-eosin and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining were performed 12 months postoperatively. RESULTS: For central retinal thickness, choroidal thickness, and flow density of central retinal superficial vascular networks, no statistical difference was noted between CXL eyes and control eyes at 12 months postoperatively (P > .05). The biomechanical stability of sclera was increased. The scleral stress and Young modulus at 8% strain corresponded to 184% and 183%, respectively, of the control values at 12 months (each P < .001). No retinal damage was detected on histology in scleral CXL eyes. There was no obvious difference between scleral CXL eyes and control eyes by hematoxylin-eosin and TUNEL staining (P > .05). CONCLUSIONS: Scleral CXL with riboflavin/UVA in rhesus monkey eyes could strengthen the biomechanical properties of scleral tissues and maintain the stability for 12 months postoperatively. The UVA-CXL on the sclera of rhesus monkey eyes seems to be effective and safe. [J Refract Surg. 2020;36(10):696-702.].

Ocular safety evaluation of blue light scleral cross-linking in vivo in rhesus macaques.

PURPOSE: To investigate the safety of blue light scleral cross-linking (SXL) by evaluating changes in biological parameters in the retina and choroid in the eyes of rhesus macaques (Macaca mulatta). METHODS: Fifteen 3-year-old macaques (30 eyes) were randomly divided into three groups (n = 5). SXL was performed via riboflavin (0.5%) and blue light (460 nm) at the location of the equatorial sclera. Right eyes served as experimental eyes, and left eyes as control eyes. One quadrant of each right eye was irradiated in group A, two quadrants of each right eye and one quadrant of each left eye were irradiated in group B, and two quadrants of each right eye were irradiated in group C. Optical coherence tomography, optical coherence tomography angiography, and flash electroretinography (f-ERG) examinations were performed at baseline and 1 week, 1 month, 3 months, and 6 months after SXL. Additionally, retinal tissue alterations were detected via transmission electron microscopy at 1 week postoperatively. RESULTS: There were no significant differences between experimental eyes and control eyes in retinal thickness, vessel density of retinal superficial capillary plexus, and choroid thickness in any of the groups at any of the time points investigated (p > 0.05). Significant reductions in f-ERG parameters were detected 1 week postoperatively in the experimental eyes of groups A and C (p < 0.05), but they gradually recovered, and there was no significant difference 1 month postoperatively (p > 0.05). Ultrastructural changes were evident in the retinal layers of SXL eyes. In group B, there were no significant differences between the right and left eyes at any of the follow-up time points investigated. CONCLUSIONS: Blue light SXL can cause transient retina damage. The f-ERG parameters reductions and retinal ultrastructural changes were found at early stage, even though there were not significant changes in retinal thickness, vessel density of retinal superficial capillary plexus, and choroid thickness after blue light SXL. The long-term intraocular safety of the blue light SXL technique should be investigated further.

Early Changes of Ocular Biological Parameters in Rhesus Monkeys After Scleral Cross-linking With Riboflavin/Ultraviolet-A.

PURPOSE: To evaluate the ocular biological parameter difference between scleral corneal cross-linking (CXL) and control eyes in rhesus monkeys by using a rebound tonometer, A-scan ultrasonography, retinoscopy, optical coherence tomography, and electroretinography (ERG). METHODS: Six rhesus monkeys were used in this study, with ages ranging from 3 to 3.5 years. One eye of each rhesus monkey was randomly selected to receive riboflavin/ultraviolet-A CXL in the temporal quadrant of the equatorial sclera and the contralateral eye served as an intra-individual control. The ocular biological parameters were repeatedly measured in both eyes of the monkeys before scleral CXL and 1 week, 1 month, and 3 months postoperatively. RESULTS: The intraocular pressure, refractive state, total axial length, and axial dimensions of the anterior chamber, crystalline lens, vitreous chamber, and central corneal thickness were not statistically significantly different between the control and cross-linked specimens at the different time periods (each P > .05). No obvious changes in the waveform of the standard full-field ERGs were observed in the control and cross-linked specimens. There were no statistically significant differences between the control and cross-linked specimens in the dark-adapted 0.01 ERG, the dark-adapted 3.0 ERG, the light-adapted 3.0 ERG, and the amplitudes of the a-wave and b-wave for the different time periods (each P >.05). CONCLUSIONS: The scleral CXL laboratory technique might not significantly affect the ocular biological parameters of the rhesus monkey in the early postoperative period, but long-term effects and histological changes still need to be investigated further. [J Refract Surg. 2019;35(5):333-339.].

Lenticule Thickness Accuracy and Influence in Predictability and Stability for Different Refractive Errors after SMILE in Chinese Myopic Eyes.

Purpose: To evaluate the differences between the estimated and measured lenticule thickness and relationships in myopic eyes with different refractive errors after small-incision lenticule extraction (SMILE). Methods: This prospective study included a total of 190 eyes (96 patients) with a manifest refraction spherical equivalent (MRSE) of -6.16 ± 1.71 D. The measured lenticule thickness was obtained from digital spectral-domain optical coherence tomography image analysis. The estimated lenticule thickness was obtained from the software program. Comparative statistics and linear regression analyses were performed. Results: At 1 month, the estimated lenticule thickness was 113.68 ± 19.35 µm and the measured lenticule thickness was 96.9 ± 17.5 µm. The difference between the estimated and measured lenticule thickness was 13.63 ± 9.1 µm (p < 0.001) thicker. The difference was lower than the mean data in the moderate myopia group (9.7 ± 6.4 µm) and larger in the high myopia group (12.3 ± 8.8 µm) and the super-high myopia group (17.9 ± 6.9 µm). The discrepancy between the estimated and measured lenticule thickness was significantly dependent on the MRSE (p < 0.001). MRSE only showed significant differences between the three and six-month follow-up visit in the super-high myopic group (p = 0.03). Conclusion: The measured lenticule thickness was thinner than the estimated lenticule thickness for SMILE in most eyes. The higher the refraction, the larger the difference after SMILE surgery. The mismatch between the estimated and measured thickness did not influence the refractive precision and stability of different refractive errors.

Consistent comparison of angle Kappa adjustment between Oculyzer and Topolyzer Vario topography guided LASIK for myopia by EX500 excimer laser.

AIM: To evaluate and compare the uniformity of angle Kappa adjustment between Oculyzer and Topolyzer Vario topography guided ablation of laser in situ keratomileusis (LASIK) by EX500 excimer laser for myopia. METHODS: Totally 145 cases (290 consecutive eyes )with myopia received LASIK with a target of emmetropia. The ablation for 86 cases (172 eyes) was guided manually based on Oculyzer topography (study group), while the ablation for 59 cases (118 eyes) was guided automatically by Topolyzer Vario topography (control group). Measurement of adjustment values included data respectively in horizontal and vertical direction of cornea. RESULTS: Horizontally, synclastic adjustment between manually actual values (dxmanu) and Oculyzer topography guided data (dxocu) accounts 35.5% in study group, with mean dxmanu/dxocu of 0.78±0.48; while in control group, synclastic adjustment between automatically actual values (dxauto) and Oculyzer topography data (dxocu) accounts 54.2%, with mean dxauto/dxocu of 0.79±0.66. Vertically, synclastic adjustment between dymanu and dyocu accounts 55.2% in study group, with mean dymanu/dyocu of 0.61±0.42; while in control group, synclastic adjustment between dyauto and dyocu accounts 66.1%, with mean dyauto/dyocu of 0.66±0.65. There was no statistically significant difference in ratio of actual values/Oculyzer topography guided data in horizontal and vertical direction between two groups (P=0.951, 0.621). CONCLUSION: There is high consistency in angle Kappa adjustment guided manually by Oculyzer and guided automatically by Topolyzer Vario topography during corneal refractive surgery by WaveLight EX500 excimer laser.

Prevalence of transforming growth factor ß-induced gene corneal dystrophies in Chinese refractive surgery candidates.

PURPOSE: To determine the prevalence of the transforming growth factor (TGF) ß-induced gene corneal dystrophies in refractive surgery candidates in China. SETTING: Five hospitals in China. DESIGN: Prospective case series. METHOD: Refractive surgical candidates from 5 preselected eye hospitals/centers in China were recruited after providing informed consent. All patients had slitlamp biomicroscopy and collection of a buccal swab as a source of DNA for screening of the TGF ß-induced gene for the 5 most common mutations associated with Reis-Bückler corneal dystrophy, Thiel-Behnke corneal dystrophy, granular corneal dystrophy type 1, granular corneal dystrophy type 2, and lattice corneal dystrophy type 1. RESULTS: Of the 2068 refractive surgery candidates analyzed, 4 had corneal opacities in both eyes on slitlamp examination. Screening for the TGF ß-induced gene found the heterozygous p.R124H mutation associated with granular corneal dystrophy type 2 in each of the 4 individuals with corneal opacities as well as in a fifth individual who did not have any corneal opacities, for a prevalence of 0.24%. Exacerbation of dystrophic corneal deposition developed after laser refractive surgery in 2 individuals who did not have preoperative TGF ß-induced gene screening. CONCLUSIONS: The prevalence of the TGF ß-induced gene corneal dystrophies in Chinese refractive surgery candidates was estimated to be approximately 0.24%. Genetic testing is recommended to identify and exclude from candidacy all individuals with a TGF ß-induced gene dystrophy before elective keratorefractive surgery to avoid causing accelerated postoperative dystrophic deposition.

Effect of a Single Nucleotide Polymorphism in the LAMA1 Promoter Region on Transcriptional Activity: Implication for Pathological Myopia.

PURPOSE: To elucidate the role of the protein coding laminin α1 (LAMA1) gene in pathological myopia (PM) at the transcriptional level. To achieve this, the binding affinity of single nucleotide polymorphism (SNP) rs2089760-located on the LAMA1 promoter gene-to human fetal scleral fibroblast (HFSF) nucleoprotein was investigated and its effect on LAMA1 transcriptional initiation activity was analyzed. METHODS: Binding interactions of the HFSF nucleoprotein and biotin-labeled SNP rs2089760 probe were investigated by amplifying the LAMA1 promoter gene and performing overlap extension polymerase chain reaction (PCR) to obtain the G/A mutation of LAMA1 SNP rs2089760. Recombinant adenovirus vectors, Ad5f11p-pLAMA1SNPa-Luc2, Ad5f11p-pLAMA1SNPg-Luc2, and Ad5f11p-CMV-RLuc, were constructed. Fluorescence intensity ratios of firefly luciferase (FLuc) and renilla luciferase (RLuc) vectors were measured 48 h after HFSF infection. RESULTS: Both specific and mutant probes banded precisely with HFSF nucleoprotein. The intensity value of the mutant probe was significantly lower than that of the specific probe (p < 0.05). HFSFs were successfully infected by the recombinant adenoviruses. The FLuc/RLuc fluorescence intensity ratio of Ad5f11p-pLAMA1SNPa-Luc2 (0.0238 ± 0.0009) was significantly lower than that of Ad5f11p-pLAMA1SNPg-Luc2 (0.0281 ± 0.0015) (p < 0.05). CONCLUSIONS: It is highly likely that SNP rs2089760 in the LAMA1 promoter region is located at the transcription factor binding site. The SNP rs2089760 G > A mutation reduces transcription factor binding ability and transcriptional initiation activity, and negatively regulates gene transcription of LAMA1. We suggest that LAMA1 SNP rs2089760 plays an important role in the development of PM.