Effect of corneal stromal lenticule customization on neurite distribution and excitatory property.

Introduction: Refractive stromal lenticules from Small Incision Lenticule Extraction (SMILE), though usually discarded, hold a potential for various ophthalmic applications, including refractive correction, stromal volume expansion, and biomechanical strengthening of the cornea. Objectives: To investigate the effect of lenticule customization on lenticule neurite length profile and the excitatory response (calcium signaling) and the potential of reinnervation. Methods: Human and porcine stromal lenticules were treated by (1) excimer laser reshaping, (2) ultraviolet A-riboflavin crosslinking (CXL), and (3) decellularization by sodium dodecyl sulfate (SDS), respectively. The overall neurite scaffold immuno-positive to TuJ1 (neuron-specific class III ß-tubulin) expression and population of active neurite fragments with calcium response revealed by L-glutamate-induced Fluo-4-acetoxymethyl ester reaction were captured by wide-field laser-scanning confocal microscopy, followed by z-stack image construction. The NeuronJ plugin was used to measure neurite lengths for TuJ1 (NL-TuJ1) and calcium signal (NL-Ca). Reinnervation of lenticules was examined by the ex vivo grafting of chick dorsal root ganglia (DRG) to the decellularized human lenticules. Differences between groups and controls were analyzed with ANOVA and Mann-Whitney U test. Results: The customization methods significantly eliminated neurites inside the lenticules. NL-TuJ1 was significantly reduced by 84% after excimer laser reshaping, 54% after CXL, and 96% after decellularization. The neurite remnants from reshaping and CXL exhibited calcium signaling, indicative of residual excitatory response. Re-innervation occurred in the decellularized lenticules upon stimulation of the grafted chick embryo DRG with nerve growth factor (NGF 2.5S). Conclusion: All of the lenticule customization procedures reduced lenticule neurites, but the residual neurites still showed excitatory potential. Even though these neurite remnants seemed minimal, they could be advantageous to reinnervation with axon growth and guidance after lenticule reimplantation for refractive and volume restoration of the cornea.

Anterior Segment Optical Coherence Tomography Angiography and Optical Coherence Tomography in the Evaluation of Episcleritis and Scleritis.

Purpose: To evaluate the feasibility of using anterior segment optical coherence tomography (AS-OCT) and AS-OCT angiography (AS-OCTA) in assessing patients with episcleritis and scleritis.Methods: Degree of vascularity [vessel density index (VDI)], measured with AS-OCTA, and sclera thickness [conjunctiva epithelium (CE), conjunctiva/episclera complex (CEC), and episclera/sclera complex (ESC)], measured with AS-OCT were compared.Results: A total of 37 eyes (13 episcleritis, 11 scleritis, 13 controls) were analyzed. VDI was lowest for controls for the various tissue depths (p < .001). Episcleritis versus scleritis revealed a significant difference in VDI at ESC (38.1 ± 11.4% vs 46.4 ± 6.4%; p = .03). Mean sclera thickness was lower in controls for CE (p < .001), CEC (p < .001) but not for ESC (p = .54).Conclusions: The degree of vascularity and tissue thickness were different between episcleritis, scleritis and controls. AS-OCTA and AS-OCT may potentially be useful in evaluating patients with scleral inflammation.

Anterior segment optical coherence tomography angiography for iris vasculature in pigmented eyes.

PURPOSE: To compare anterior segment optical coherence tomography angiography (AS-OCTA) systems in delineating normal iris vessels and iris neovascularisation (NVI) in eyes with pigmented irides. METHODS: Prospective study from January 2019 to June 2019 of 10 consecutive patients with normal pigmented iris, had AS-OCTA scans with a described illumination technique, before using the same protocol in five eyes with NVI (clinical stages 1-3). All scans were sequentially performed using a spectral-domain OCTA (SD-OCTA), and a swept-source OCTA (SS-OCTA, Plex Elite 9000). Images were graded by two masked observers for visibility, artefacts and NVI characteristics. The main outcome measure was iris vessel density measurements comparing SS-OCTA and SD-OCTA systems. RESULTS: The median age of subjects was 28 (20-35) years, and 50% were female. The paired mean difference of iris vessel density measurements was 11.7 (95% CI 14.7 to 8.1; p=0.002), SS-OCTA detecting more vessels than SD-OCTA. The inter-rater reliability for artefact score (κ=0.799, p<0.001) and visibility score (κ=0.722; p<0.001) were substantial. Both AS-OCTA systems were able to detect NVI vessels with a fair agreement (κ=0.588), with clearer NVI characteristics in stage 1/2 compared to stage 3 NVI (mean difference NVI score: 2.7±0.4, p=0.009). CONCLUSION: The SS-OCTA was better able to delineate iris vessels in normal pigmented irides compared to SD-OCTA. Both AS-OCTA systems identified NVI characteristics based on its atypical configuration or location, but further improvements are needed to allow for more accurate objective, serial quantification for clinical use.

Descemet Membrane Endothelial Keratoplasty With a Pull-Through Insertion Device: Surgical Technique, Endothelial Cell Loss, and Early Clinical Results.

PURPOSE: To describe a surgical technique for Descemet membrane endothelial keratoplasty (DMEK) using a pull-through, endothelium-in insertion device, the DMEK EndoGlide. We evaluated the endothelial cell loss (ECL) associated with the EndoGlide-DMEK (E-DMEK) technique in both ex vivo and prospective clinical studies. METHODS: The ex vivo study involved calcein acetoxymethyl staining and preparation of DMEK grafts, which were trifolded endothelium-in, loaded into the EndoGlide, pulled through, and unfolded in imaging dishes. Inverted fluorescent microscopy was performed, and ECL was quantified using trainable segmentation software. The prospective clinical series describes the outcomes of consecutive surgeries using the E-DMEK technique. Grafts were pulled through the EndoGlide with forceps and unfolded in the anterior chamber endothelium-down. Our main outcome measure was ECL in both studies. RESULTS: In the ex vivo study with 9 human donor corneas, mean ECL was 15.2% ± 5.4% (n = 9). In our clinical series of 69 eyes, leading indications for surgery were pseudophakic/aphakic bullous keratopathy (47.8%), previous failed grafts (23.2%), and Fuchs endothelial dystrophy (18.8%). Rebubbling and primary graft failure rates related to E-DMEK were 11.6% and 1.5%, respectively. Among eyes with at least 6 months of follow-up, mean preoperative endothelial cell density was 2772 (range 2457-3448) cells/mm, and postoperative endothelial cell density was 1830 (range 541-2545) cells/mm. Mean ECL was 33.6% (range 7.5-80.4; n = 32) at the 7.1 (range 6-11) months follow-up. CONCLUSIONS: The ex vivo and pilot clinical studies suggest that E-DMEK shows acceptable rates of ECL, with safe and promising early clinical outcomes.

Lamellar Dissection Technique for Descemet Membrane Endothelial Keratoplasty Graft Preparation.

PURPOSE: To describe a novel lamellar dissection technique for Descemet membrane endothelial keratoplasty (DMEK) graft preparation, and to evaluate the rate of endothelial cell loss (ECL) and graft preparation failure associated with this technique. METHODS: We conducted an ex vivo laboratory-based study comparing ECL between the lamellar dissection and peeling techniques. Eight pairs of human donor corneas underwent calcein acetoxymethyl staining-all right eyes underwent the peeling technique and all left eyes underwent the lamellar dissection technique. ECL was quantified by image analysis with trainable segmentation software and compared between groups. We also conducted a retrospective analysis of 161 consecutive DMEK graft preparations by a single surgeon using the lamellar dissection technique from 2010 to 2018. Data on donor characteristics and graft preparation failures were obtained. RESULTS: Baseline donor characteristics were comparable in both arms of the laboratory-based study. Mean (SD) ECL with the lamellar dissection and peeling techniques was 13.8% (4.2%) and 11.2% (6.1%), respectively. There was no significant difference between the two (P = 0.327). In the clinical series, there were 2 graft preparation failures in 161 cases (1.2%). Among cases performed on diabetic donor tissue, the rate of graft preparation failure was 4.7%. CONCLUSIONS: The lamellar dissection technique has a similar rate of ECL compared with the peeling technique for DMEK graft preparation. This technique also has a low rate of graft preparation failure and may be a useful technique for diabetic donor tissue.

Optical coherence tomography angiography for the assessment of choroidal vasculature in high myopia.

AIMS: To assess specific layers of the choroid in highly myopic young adults and to examine their associations with levels of myopia. METHODS: We recruited 51 young myopes (n=91 eyes) from the Singapore Cohort of Risk Factors for Myopia cohort. We performed standardised optical coherence tomography (OCT) and OCT angiography imaging and developed a novel segmentation technique assessing choroidal layers' thickness (overall choroidal thickness (CT), medium-vessel choroidal layer (MVCL) thickness, large-vessel choroidal layer (LVCL)) and vasculature (choroidal vessel density (%), choroidal branch area (CBA, %) and mean choroidal vessel width (MCVW, mm)). RESULTS: We found that eyes with extreme myopia (EM) had thinner vascular layers compared with high myopia (HM), that is, LVCL (36.0±1.5 vs 39.2±1.2 µm, p=0.002) and MVCL (185.5±5.7 vs 198.2±4.6 µm, p=0.014). Overall CT was thinnest in the nasal and inferior quadrants in EM (nasal: 157.1±9.6 vs 187.2±8.3 µm, p<0.001; superior: 236.6±11.1 vs 257.0±9.5 µm, p=0.02; temporal: 228.0±10.6 vs 254.3±8.8 µm, p=0.012; and inferior quadrant: 198.7±10.0 vs 239.8±8.3 µm, p=<0.001) when compared with HM. We also observed significantly more vessel branching in eyes with EM as compared with eyes with HM (CBA, 10.2%±0.7% vs 9.95%±0.8%, p=0.018). CONCLUSIONS: The novel segmentation technique and introduced choroidal parameters may serve as new biomarkers to study disease conditions in myopia.

Optical coherence tomography angiography in diabetic retinopathy: a review of current applications.

BACKGROUND: Diabetic retinopathy (DR) is a leading cause of vision loss in adults. Currently, the standard imaging technique to monitor and prognosticate DR and diabetic maculopathy is dye-based angiography. With the introduction of optical coherence tomography angiography (OCTA), it may serve as a potential rapid, non-invasive imaging modality as an adjunct. MAIN TEXT: Recent studies on the role of OCTA in DR include the use of vascular parameters e.g., vessel density, intercapillary spacing, vessel diameter index, length of vessels based on skeletonised OCTA, the total length of vessels, vascular architecture and area of the foveal avascular zone. These quantitative measures may be able to detect changes with the severity and progress of DR for clinical research. OCTA may also serve as a non-invasive imaging method to detect diabetic macula ischemia, which may help predict visual prognosis. However, there are many limitations of OCTA in DR, such as difficulty in segmentation between superficial and deep capillary plexus; and its use in diabetic macula edema where the presence of cystic spaces may affect image results. Future applications of OCTA in the anterior segment include detection of anterior segment ischemia and iris neovascularisation associated with proliferative DR and risk of neovascular glaucoma. CONCLUSION: OCTA may potentially serve as a useful non-invasive imaging tool in the diagnosis and monitoring of diabetic retinopathy and maculopathy in the future. Future studies may demonstrate how quantitative OCTA measures may have a role in detecting early retinal changes in patients with diabetes.

In vivo corneal endothelium imaging using ultrahigh resolution OCT.

We investigate the influence of optical coherence tomography (OCT) system resolution on high-quality in vivo en face corneal endothelial cell images of the monkey eye, to allow for quantitative analysis of cell density. We vary the lateral resolution of the ultrahigh resolution (UHR) OCT system (centered at 850 nm) by using different objectives, and the axial resolution by windowing the source spectrum. By suppressing the motion of the animal, we are able to obtain a high-quality en face corneal endothelial cell map in vivo using UHR OCT for the first time with a lateral resolution of 3.1 µm. Increasing lateral resolution did not result in a better image quality but a smaller field of view (FOV), and the axial resolution had little impact on the visualization of corneal endothelial cells. Quantitative analysis of cell density was performed on in vivo en face OCT images of corneal endothelial cells, and the results are in agreement with previously reported data. Our study may offer a practical guideline for designing OCT systems that allow for in vivo corneal endothelial cell imaging with high quality.

Optical Coherence Tomography Angiography Imaging to monitor Anti-VEGF treatment of Corneal Vascularization in a Rabbit Model.

Optical coherence tomography angiography (OCTA) is a well-established non-invasive retinal vascular imaging technique. It has been recently adapted to image the anterior segment and has shown good potential to image corneal vascularisation. The purpose of the study is to evaluate the usefulness of OCTA to monitor regression of corneal vessels following anti-VEGF (vascular endothelial growth factor) treatment using a previously established corneal vascularisation rabbit model. The regression of vessels following the treatment with aflibercept and ranibizumab anti-VEGFs using both topical instillation and sub-conjunctival injection was quantified using OCTA and compared with ICGA (indocyanine green angiography). Overall vessel density measurements using OCTA showed good correlation (r = 0.988, p < 0.001) with ICGA, with no significant difference between the two treatment groups (p = 0.795). It was also shown that OCTA provided good repeatability outcomes of the quantitative measurements. Using Bland-Altman plots, vessel growth density values between anti-VEGF treatments were compared to control saline group. It was observed that aflibercept provided longer lasting effect than ranibizumab. We also observed that in both drugs, the topical route of administration topical provided longer regression outcomes compared to one-time sub-conjunctival injection. Thereby, with this pilot study, it was demonstrated that OCTA is a reliable imaging technique to follow-up and monitor corneal vascularisation and its treatment quantitatively.

Three-Dimensional Neurite Characterization of Small Incision Lenticule Extraction Derived Lenticules.

Purpose: We study the density and excitatory response of neurites, and Schwann cells (SCs) in fresh and cryopreserved stromal lenticules derived from small incision lenticule extraction (SMILE). Methods: Human stromal lenticules (n = 23) were immunostained for ß III-tubulin and imaged using spinning disk confocal laser microscopy, followed by three-dimensional reconstruction, to reveal neurite distribution. The lenticule neurite density (LND) was assessed using a validated neurite tracing and length measurement method with NeuronJ. LND was compared among groups of different lenticule thickness (71-165 µm) obtained from -3 to >-6 diopters (D) corrections. SCs were identified by marker expression and the laser effect on SC-neurite interaction was examined under transmission electron microscopy (TEM). Fresh porcine SMILE-lenticules (n = 18) were used for LND comparison among storage conditions and functional excitatory calcium response assay. Results: Using a validated neurite length measurement method, we found an inverse correlation of LND with lenticule thickness. Higher LND was found in thinner lenticules obtained from lower power of correction (r = -0.8925, P < 0.0001), whereas total lenticule neurite lengths did not alter significantly with regards to lenticule thickness. SCs were identified by GAP43 and p75NTR expression and were closely associated with lenticule neurites under TEM. In porcine lenticules, LND and excitatory calcium response were reduced after cold and cryogenic storage, when compared to fresh lenticules. Conclusions: The stromal neurites showed variations in density related to SMILE lenticule thickness and cryopreservation. With the presence of SC support and excitatory response, these neurite residues could retain minimal functionality that might serve as a potential advantage in the event of lenticule implantation.

Optical Coherence Tomography Angiography for Evaluation of Reperfusion After Pterygium Surgery.

PURPOSE: To describe the use of optical coherence tomography angiography (OCTA) to quantitatively monitor the conjunctival graft revascularization after pterygium excision and conjunctival autograft (CAG) transplantation. DESIGN: Prospective, interventional case series. METHODS: Ten patients undergoing pterygium excision and femtosecond laser-assisted CAG transplantation were included. OCTA was performed at 1 week, 1 and 3 months postoperatively at the CAG transplantation site and harvested area. The vessel density at three different depths: conjunctival epithelium or CAG epithelium, conjunctival stroma or CAG stroma, and episclera, was evaluated and quantified. The revascularization rate was assessed and correlated with the postoperative CAG thickness. RESULTS: No intraoperative and postoperative complications occurred. Reperfusion of the CAGs was observed at 1 week, and early reperfusion within the first month accounted for more than half of graft revascularization. The vessel regrowth density was 9.6±2.6 % and 11.1±2.8 % between 1 week and 1 month, and was 9.8±1.8 % and 11.9±1.9 % between 1 and 3 months, at the CAG and underlying episcleral levels, respectively. All the CAGs were well-perfused at 3 months. The vessel regrowth density was significantly and strongly correlated with the changes of CAG thickness in a negative relationship (γ = -0.94, P = 0.019). At the harvested site, the vascular network of episclera was not affected, and the conjunctival vascularization was restored at 1 month. CONCLUSIONS: OCTA is a promising tool to evaluate the vascularization or revascularization of conjunctiva, conjunctival graft and episclera, in a quantitative and serial manner, helping in diseases diagnosis and treatment monitoring. The graft revascularization rate was predictive of postoperative graft deswelling.

Impact of systemic vascular risk factors on the choriocapillaris using optical coherence tomography angiography in patients with systemic hypertension.

We investigated the characteristics of the choriocapillaris flow voids using optical coherence tomography angiography (OCTA) in 85 patients (164 eyes) with hypertension (mean ± SD age, 56 ± 11 years; 45% women; 20% poorly controlled BP; 16% diabetes) who are without ocular diseases and determined possible correlations with systemic vascular risk factors. Data on 24-hour ambulatory blood pressure (BP), serum creatinine, and urine microalbumin/creatinine ratio (MCR) were collected. Estimated glomerular filtration rate (eGFR) was calculated based on CKD-EPI Creatinine Equation. OCTA imaging (6 × 6 mm scans; AngioVue) with quantitative microvascular analysis of the choriocapillaris was performed. Linear regression was used to investigate the association of systemic risk factors with area (percentage), size (pixels) and number of choriocapillaris flow voids. Diabetes (ß = 0.33; 95% CI, 0.02 to 0.63) and daytime systolic BP (ß = -0.13; 95% CI, -0.24 to -0.02) were associated with areas of flow voids. Age (ß = 0.21; 95% CI, 0.06 to 0.36) and daytime diastolic BP (ß = -0.18; 95% CI, -0.34 to -0.02) were associated with size of flow voids. Age (ß = -21.21; 95% CI, -31.79 to -10.63) and nighttime diastolic BP (ß = 13.89; 95% CI, 0.61 to 27.17) were associated with number of flow voids. Kidney parameters were not associated with any features of flow voids. In patients with hypertension, a higher BP appeared to increase blood flow in the choriocapillaris which needs to be considered when using the OCTA to study eye diseases in hypertensives.

Correction to: Vessel density and En-face segmentation of optical coherence tomography angiography to analyse corneal vascularisation in an animal model.

[This corrects the article DOI: 10.1186/s40662-018-0128-8.].

Optical coherence tomography angiography for the anterior segment.

Optical coherence tomography angiography (OCTA) is a rapid and non-invasive technique for imaging vasculature in the eye. As OCTA can produce high-resolution cross-sectional images and allow depth-resolved analysis for accurate localization of pathology of interest, it has become a promising method for anterior segment imaging. Furthermore, OCTA offers a more patient-friendly alternative to the conventional invasive dye-based fluorescent angiography. However, conventional OCTA systems are typically designed and optimized for the posterior segment of the eye, and thus using OCTA for anterior segment imaging can present several difficulties and limitations. In this review, we summarized the recent developments and clinical applications in anterior segment OCTA (AS-OCTA) imaging, such as for the cornea, iris, sclera and conjunctiva. We also compared commercially available OCTA systems, discussed the limitations of adapting current OCTA technology for the anterior segment imaging, and proposed possible future directions for AS-OCTA systems. AS-OCTA provides potential for future clinical applications such as diagnosis of corneal and iris pathologies, pre-operative surgical planning, assessment of new anti-angiogenic therapeutics or evaluation of limbal stem cell deficiency. With further development, OCTA for anterior segment imaging in the clinics may become common in the near future.

Vessel density and En-face segmentation of optical coherence tomography angiography to analyse corneal vascularisation in an animal model.

Background: Optical coherence tomography angiography (OCTA) is a novel non-invasive angiography technology that has recently been extensively studied for its utility in anterior segment imaging. In this study, we compared a split-spectrum amplitude decorrelation angiography (SSADA) OCTA and an Complex OCT signal difference angiography [corrected] (CODAA SD) [corrected] OCTA system to current angiographic technique, indocyanine green angiography (ICGA), to assess corneal vascularisation in an animal model. Methods: We imaged 16 rabbits, (one eye per animal) with corneal vascularisation using SSADA OCTA (AngioVue; Optovue Inc., USA), CODAA OCTA [corrected] (Angioscan; RS-3000 Nidek Co. Ltd., Japan) and ICGA in the same region of interest of the cornea at successive time-points. We then analysed all scanned images for vessel density measurements and used paired t-tests and Bland-Altman plots to examine for significant differences. The en-face segmentation images from each of the OCTA scans were also extracted and were matched at every 50 µm segmentation to be compared for vessel density at the respective depths. Results: Bland-Altman plots revealed a good agreement between all three imaging techniques (P > 0.05) for all vessel density measurements computed, and the ranges of 95% limit of agreement were acceptable from a clinical perspective. No significant difference was reported, with ICGA (µ = 16.52 ± 8.94%) being more comparable to the CODAA [corrected] OCTA (µ = 16.23 ± 9.51%; p = 0.50) than the SSADA OCTA (µ = 17.09 ± 7.34%; p = 0.33) system. Also, a good correlation value (r > 0.9) was obtained when comparing the vessel density measurements of the en-face segmentations between the OCTA systems. Conclusions: Comparable vessel density quantification between the two OCTA systems, and with ICGA was obtained. Segmentation analysis of the vasculature at different depths showed varied performance in the two OCTA systems relative to each other. The implications of the study may help to aid in the development of better OCTA algorithms for the anterior segment and its use in clinical translational research.

Age-related changes of individual macular retinal layers among Asians.

We characterized the age-related changes of the intra-retinal layers measured with spectral-domain optical coherence tomography (SD-OCT; Cirrus high-definition OCT [Carl Zeiss Meditec]. The Singapore Epidemiology of Eye Diseases is a population-based, cross-sectional study of Chinese, Malays and Indians living in Singapore. Iowa Reference Algorithms (Iowa Institute for Biomedical Imaging) were used for intra-retinal layer segmentation and mean thickness of 10 intra-retinal layers rescaled with magnification correction using axial length value. Linear regression models were performed to investigate the association of retinal layers with risk factors. After excluding participants with history of diabetes or ocular diseases, high-quality macular SD-OCT images were available for 2,047 participants (44-89 years old). Most of the retinal layers decreased with age except for foveal retinal nerve fiber layer (RNFL) and the inner/outer segments of photoreceptors where they increased with age. Men generally had thicker retinal layers than women. Chinese have the thickest RNFL and retinal pigment epithelium amongst the ethnic groups. Axial length and refractive error remained correlated with retinal layers in spite of magnification correction. Our data show pronounced age-related changes in retinal morphology. Age, gender, ethnicity and axial length need be considered when establishing OCT imaging biomarkers for ocular or systemic disease.

Novel application of In Vivo Micro-Optical Coherence Tomography to assess Cornea scarring in an Animal Model.

This pilot study uses a micro-optical coherence tomography (micro-OCT) system with ~1 µm axial resolution specifically to image the cornea and corneal scars in vivo. We used an established murine corneal scar model by irregular phototherapeutic keratectomy in ten C57BL/6 mice, with serial imaging using the micro-OCT and compared to anterior segment (AS-OCT) (RTvue, Optovue, Fremont, CA) before and after scar induction. Main outcome was agreement between the AS-OCT and micro-OCT using Bland-Altman plots (95% limits of agreement, LoA).We analysed 10 control eyes and 10 eyes with corneal scars and found that there was good agreement between AS-OCT and micro-OCT (P > 0.05) LOA: lower limit -14 µm (95% CI: -19 to -8.8 µm) upper limit 23 µm (95% CI: 18 to 28.5 µm) in terms of central corneal thickness. There was also good agreement between AS-OCT and micro-OCT in terms of corneal scar measurements (P > 0.5; correlation coefficient >0.99) LOA lower limit -2.1 µm (95% CI: -2.8 to -1.5 µm); upper limit 1.8 µm (95% CI: 1.1 to 2.4 µm). Our pilot study suggests that this novel in vivo micro-OCT imaging technique was able to measure central corneal thickness and scar thickness in agreement with current AS-OCT techniques.

Comparison of Optical Coherence Tomography Angiography to Indocyanine Green Angiography and Slit Lamp Photography for Corneal Vascularization in an Animal Model.

Corneal neovascularization (CoNV) could be treated by novel anti-angiogenic therapies, though reliable and objective imaging tools to evaluate corneal vasculature and treatment efficacy is still lacking. Optical coherence tomography angiography (OCTA) -currently designed as a retinal vascular imaging system- has been recently adapted for anterior-segment and showed good potential for successful imaging of CoNV. However, further development requires an animal model where parameters can be studied more carefully with histological comparison. Our study evaluated the OCTA in suture-induced CoNV in a rabbit model compared to indocyanine green angiography (ICGA) and slit-lamp photography (SLP). Overall vessel density measurements from OCTA showed good correlation with ICGA (0.957) and SLP (0.992). Vessels density by OCTA was higher than ICGA and SLP (mean = 20.77 ± 9.8%, 15.71 ± 6.28% and 17.55 ± 8.36%, respectively, P < 0.05). OCTA was able to depict CoNV similarly to SLP and ICGA, though it could better detect small vessels. Moreover, the depth and growth of vessels could be assessed using en-face and serial-scans. This study validated the OCTA in a rabbit model as a useful imaging tool for translational studies on CoNV. This may contribute to further studies on OCTA for anterior-segment including serial evaluation of emerging anti-angiogenic therapies.

Comparison of anterior segment optical coherence tomography angiography systems for corneal vascularisation.

AIM: To newly describe a spectral-domain (SD) optical coherence tomography angiography (OCTA) for the cornea and directly compare two OCTA system scans of the same eyes with corneal vascularisation. METHODS: Cross-sectional, observational, comparative case series. We performed sequential OCTA scans (10 eyes of 10 subjects with corneal vascularisation,4 scans each eye) repeated using split-spectrum amplitude decorrelation algorithm angiography system (SSADA, AngioVue; Optovue Inc, USA) and SD OCTA (Angioscan; Nidek Co. Ltd, Japan) in the same region of interest. We analysed all scan images for repeatability, image quality and vessel density measurements and compared OCTA systems. RESULTS: We obtained substantial interobserver repeatability in terms of image quality score (κ=0.86) for all 80 OCTA scans (median age 49 years, 50% women). The correlation was moderately good (r=0.721) when comparing vessel density measurements between OCTA systems, but greater in the SSADA compared with SD OCTA system (mean vessel density 20.3±4.9% vs 15.1±4.2%, respectively; p<0.001). CONCLUSION: In this pilot clinical study, we describe successful delineation of corneal vessels with substantial image quality using a new SD OCTA system. The vessel density measurements were greater using the SSADA compared with SD OCTA system in the same area of corneal vascularisation. Further studies are required to confirm the advantages, limitations and differences between these OCTA systems for the anterior segment.

Blue light-mediated transcriptional activation and repression of gene expression in bacteria.

Light-regulated modules offer unprecedented new ways to control cellular behavior in precise spatial and temporal resolution. The availability of such tools may dramatically accelerate the progression of synthetic biology applications. Nonetheless, current optogenetic toolbox of prokaryotes has potential issues such as lack of rapid and switchable control, less portable, low dynamic expression and limited parts. To address these shortcomings, we have engineered a novel bidirectional promoter system for Escherichia coli that can be induced or repressed rapidly and reversibly using the blue light dependent DNA-binding protein EL222. We demonstrated that by modulating the dosage of light pulses or intensity we could control the level of gene expression precisely. We show that both light-inducible and repressible system can function in parallel with high spatial precision in a single cell and can be switched stably between ON- and OFF-states by repetitive pulses of blue light. In addition, the light-inducible and repressible expression kinetics were quantitatively analysed using a mathematical model. We further apply the system, for the first time, to optogenetically synchronize two receiver cells performing different logic behaviors over time using blue light as a molecular clock signal. Overall, our modular approach layers a transformative platform for next-generation light-controllable synthetic biology systems in prokaryotes.