Dynamic changes of the extracellular matrix during corneal wound healing.

The extracellular matrix (ECM) confers transparency to the cornea because of the precise organization of collagen fibrils and a wide variety of proteoglycans. We monitored the corneal wound healing process after alkali burns in rabbits. We analyzed the location and expression of collagens and proteoglycans, the clinical impact, and the recovery of optical transparency. After the animals received both general and ocular topical anesthesia, the central cornea of the left eye was burned by placing an 8-mm diameter filter paper soaked in 0.5 N NaOH for 60 s. The eyes were evaluated under a surgical microscope at 1, 3, and 6 months after burning. At each time point, the clinical conditions of the burned and control corneas were observed. The arrangement of collagen fibers in the corneal stroma was visualized by Picrosirius-red staining, Gomori's silver impregnation and transmission electronic microscopy. Corneal light transmittance was also measured. Myofibroblasts presence was analyzed by immunohistochemistry. mRNA expression levels of collagen types I and III, lumican, decorin, keratocan and alpha-smooth muscle actin were determined by quantitative real-time polymerase chain reaction. One month after alkali burn, the ECM was disorganized and filled with lacunae containing different types of cells and collagen type III fibers in the wound area. Corneal opacities were present with attendant loss of light transmittance. Collagen and proteoglycan mRNA expression levels were up-regulated. After three months, wound healing progress was indicated by reduced corneal opacity, increased light transmittance, reorganization of collagen fibers and only collagen type I expression levels were at control levels. After six months, the wound area ECM morphology was similar to controls, but transmittance values remained low, denoting incomplete restoration of the stromal architecture. This multidisciplinary study of the stromal wound healing process revealed changes in corneal transmittance, collagen organization, myofibroblasts presence and ECM composition at 1, 3, and 6 months after alkali burning. Documenting wound resolution during the six-month period provided reliable information that can be used to test new therapies.

Influence of age and spectral power distribution on mesopic visual sensitivity.

The ability of the human visual system to detect stimuli at low illumination levels provides awareness of potential risks. The influence of age and spectral power distribution on mesopic spectral sensitivity is analyzed. Two typical light sources are used, a high-pressure sodium lamp (HPS), with a higher content of long wavelengths, and a metal halide lamp (MH), with a higher content of short wavelengths. Two experiments were performed, using a two-channel Maxwellian-view optical system to measure contrast threshold under different experimental conditions. In Experiment 1, three age groups (young, middle-aged and old, n = 2 each), two retinal locations (on-axis and off-axis vision), four background luminances (0.01, 0.07, 0.45, and 3.2 cd/m2), and two photometry systems (photopic and the MES2 systems) were considered. In Experiment 2, contrast threshold measurement was performed with two age groups (young and old, n = 11 each), one retinal location (off-axis vision), one background luminance (0.01 cd/m2), and two photometry systems (photopic and the MES2 systems). In on-axis vision, neither age nor spectral power distribution have an effect on the contrast threshold. In off-axis vision, however, a significant interaction between age and spectral power distribution is obtained, albeit only at 0.01 cd/m2 with an MH lamp. Only at this lowest background luminance was the greater content of short wavelengths of this lamp responsible for higher rod stimulation in off-axis vision, with the subsequent improvement in detection performance in young subjects. However, the effect of diffused light inside the aged eye counteracted the benefits of increased rod sensitivity for the MH lamp.

Effect of eccentricity and light level on the timing of light adaptation mechanisms.

We explored the complexity of the light adaptation process, assessing adaptation recovery (Ar) at different eccentricities and light levels. Luminance thresholds were obtained with transient background fields at mesopic and photopic light levels for temporal retinal eccentricities (0°-15°) with test/background stimulus size of 0.5°/1° using a staircase procedure in a two-channel Maxwellian view optical system. Ar was obtained in comparison with steady data [Vis. Res.125, 12 (2016)VISRAM0042-698910.1016/j.visres.2016.04.008]. Light level proportionally affects Ar only at fovea. Photopic extrafoveal thresholds were one log unit higher for transient conditions. Adaptation was equally fast at low light levels for different retinal locations with variations mainly affected by noise. These results evidence different timing in the mechanisms of adaptation involved.

Influence of background size, luminance and eccentricity on different adaptation mechanisms.

Mechanisms of light adaptation have been traditionally explained with reference to psychophysical experimentation. However, the neural substrata involved in those mechanisms remain to be elucidated. Our study analyzed links between psychophysical measurements and retinal physiological evidence with consideration for the phenomena of rod-cone interactions, photon noise, and spatial summation. Threshold test luminances were obtained with steady background fields at mesopic and photopic light levels (i.e., 0.06-110cd/m(2)) for retinal eccentricities from 0° to 15° using three combinations of background/test field sizes (i.e., 10°/2°, 10°/0.45°, and 1°/0.45°). A two-channel Maxwellian view optical system was employed to eliminate pupil effects on the measured thresholds. A model based on visual mechanisms that were described in the literature was optimized to fit the measured luminance thresholds in all experimental conditions. Our results can be described by a combination of visual mechanisms. We determined how spatial summation changed with eccentricity and how subtractive adaptation changed with eccentricity and background field size. According to our model, photon noise plays a significant role to explain contrast detection thresholds measured with the 1/0.45° background/test size combination at mesopic luminances and at off-axis eccentricities. In these conditions, our data reflect the presence of rod-cone interaction for eccentricities between 6° and 9° and luminances between 0.6 and 5cd/m(2). In spite of the increasing noise effects with eccentricity, results also show that the visual system tends to maintain a constant signal-to-noise ratio in the off-axis detection task over the whole mesopic range.

Tissue reaction after intrastromal corneal ring implantation in an experimental animal model.

PURPOSE: To evaluate corneal wound healing in the hen animal model after additive surgery with an intracorneal ring segment (ICRS). METHODS: We implanted one ICRS in each eye of 76 hens. In control group 1 (n = 22 hens), the stromal channel was prepared but no ICRS was inserted. In control group 2 (n = 2 hens), no surgery was performed. Animals were randomly separated into groups and euthanized after clinical follow-up of 4 and 12 hours, 1, 2, 3, and 7 days, and 1, 2, 3, 4, and 6 months. Corneas were stained with hematoxylin-eosin. Apoptosis was measured by terminal uridine nick end-labeling assays. Cell proliferation and myofibroblast-like differentiation were assayed by BrdU and α-smooth muscle actin immunofluorescence microscopy. Stromal matrix changes were documented by electron microscopy. RESULTS: Epithelial and stromal cell apoptosis around the ICRS-implanted and control group 1 eyes peaked at 12 hours, but continued for 72 hours. In ICRS-implanted eyes, epithelial and stromal proliferation was present at 12 and 24 hours, respectively, and peaked at 7 days and 72 hours, respectively. Some proliferation in the ICRS-implanted group continued through the 6-month follow-up, and myofibroblast-like cells differentiated one to three months after ICRS implantation. The segments rotated within the stroma as the limbal inferior angle approached the epithelium. CONCLUSIONS: Wound healing after ICRS implantation in hen corneas was similar to that of other corneal surgical wounds in stages. However, there were some specific features related to the small size of the epithelial wound and the device permanently implanted inside the cornea.

Integration of an intensified charge-coupled device (ICCD) camera for accurate spectroscopic measurements.

Intensified charge-coupled devices (ICCD) are used in a great variety of spectroscopic applications, some of them requiring high sensitivity and spectral resolution. The setup, configuration, and featuring of these cameras are fundamental issues in order to acquire high quality spectra. In this work a critical assessment of these detectors is performed and the specific configuration, the optical alignment, featuring, and the dark and shot noise are described and analyzed. Spatial response of the detector usually shows a significant lack of spatial homogeneity and a map of interferences may appear in certain ranges of wavelengths, which damages the quality of the recorded spectra. In this work the spectral resolution and the spatial and spectral sensitivity are also studied. The analysis of the dark current reveals the existence of a smooth but clear spatial dependence. As a final conclusion, the spectra registered with the spectrometer equipped with our ICCD camera allow us to explore and measure accurately spectral line shapes emitted by pulsed plasmas in the visible range and particularly in the ultraviolet (UV) range.

Temporal retinal sensitivity in mesopic adaptation.

PURPOSE: Night driving is a complex visual task with important ramifications for driver and pedestrian safety. It is usually performed under mesopic or scotopic conditions and frequently, in the presence of transient glare sources that can adapt parts of the central retina. The objective of this work was to analyze the time response of adaptation for the central 15° of the retina when part of it is exposed to transient or steady mesopic adapting fields. METHODS: Absolute visual thresholds and luminance thresholds when viewing steady and transient adaptation fields were measured for three observers, at temporal retinal eccentricities of 0°-14.5° in steps of 2.9° (subsequently described as 0°, 3°, 6°, 9°, 12° and 15°) using a two-channel Maxwellian view optical system. The adaptation field and stimulus subtended 1.05° and 0.45° respectively. The transient adaptation field was presented with a stimulus onset asymmetry (SOA) of 300 ms. Time course adaptation curves were also measured at 0°, 6° and 9°. RESULTS: The absolute dark adaptation threshold (threshold measured at dark adaptation conditions or L(a)(t) decreases in peripheral retina due to an increasing rod contribution. Luminance thresholds vs eccentricity curves for transient L(SOA300)(t) and steady L(LA)(t) mesopic adaptation fields intersect across the first 15° of the peripheral retina. CONCLUSIONS: While the fovea shows higher sensitivity than the areas of peripheral retina investigated in this study, the speed of adaptation, measured from the visibility loss, is greater for retinal regions between 6° and 9° than for the fovea or retinal eccentricities beyond 9°.

Quantitative and functional influence of surround luminance on the letter contrast sensitivity function.

AIM: To determine the influence of surround luminance on the letter contrast sensitivity function. METHOD: The binocular contrast sensitivities of 31 young and ocularly healthy individuals were measured with letters of sizes calculated to obtain the targeted fundamental frequencies of 3, 10, 20, and 30 c deg(-1), respectively; with surround luminances from 1 to 1000 cd m(-2), always with a test chart background luminance of 200 cd m(-2). RESULTS: The letter contrast sensitivity increased with surround luminance up to 100 cd m(-2) and decreased when surround luminance increased from 100 to 1000 cd m(-2). These increments are larger for higher fundamental spatial frequencies, while decrements are similar for all frequencies. To analyse pupil size influence, results were compared with theoretical predictions obtained by combining different ocular MTFs with a typical neural function, where pupil size decrease leads to letter contrast sensitivity increments and veiling luminance causes the observed decrements. Other possible optical or neural factors that influence these values have also been considered. CONCLUSIONS: Letter contrast sensitivity function depends on surround luminance and this influence should be considered in future standardized directives.