Johan Germundsson

Standardized Baseline Human Corneal Subbasal Nerve Density for Clinical Investigations With Laser-Scanning in Vivo Confocal Microscopy

PURPOSE We established a baseline value for central corneal subbasal nerve density in a large, healthy cohort. METHODS A total of 106 healthy volunteers (207 eyes) underwent full ophthalmic examination, including laser-scanning in vivo confocal microscopy (IVCM) of the central cornea. Images of the corneal subbasal nerve plexus were acquired and analyzed based on defined criteria. Nerve tracing was performed by two human observers and by a fully automated method. Subbasal nerve density was stratified by eye, observer, tracing method, calculation method, and age group. Association of nerve density with age was examined by linear regression and population distribution was examined by nonlinear regression. RESULTS We analyzed 892 distinct, high quality images of the subbasal nerve plexus (mean, 4.3 images/eye) from 207 eyes. An overall mean central subbasal nerve density of 19 mm/mm(2) was found in 106 subjects aged 15 to 88 years, independent of eye, sex, or nerve tracing method, while the SD was a consistent 4 to 5 mm/mm(2). Subbasal nerve density followed a normal Gaussian distribution, and correlated negatively with age, with a mean decline of 0.25% to 0.30% per year, independent of eye, observer, or nerve tracing method. Moreover, the use of automated tracing techniques and randomized sampling may improve the speed and reproducibility of subbasal nerve density assessment for clinical applications. CONCLUSIONS A baseline human corneal subbasal nerve density has been determined by laser-scanning IVCM using rigorous methods. The methods and results could aid in the future assessment of corneal nerves in various patient populations.

The Role of Bowman's Layer in Corneal Regeneration after Phototherapeutic Keratectomy: A Prospective Study Using In Vivo Confocal Microscopy

PURPOSE To examine the role of Bowmans layer (BL) on the nature of anterior corneal regeneration after excimer laser phototherapeutic keratectomy (PTK). METHODS A cohort of 13 patients underwent PTK to remove either 7 mum of BL for treatment of primary recurrent corneal erosions (RCE; six patients) or complete BL removal (15-mum ablation) to treat RCE or poor vision secondary to map-dot-fingerprint (MDF) dystrophy (seven patients). Clinical examinations and laser-scanning in vivo confocal microscopy (IVCM) were conducted before surgery and at a mean of 4 and 8 months after surgery. RESULTS Total BL removal resulted in a significant decline in subbasal nerve density at 4 months (P = 0.007) that barely recovered to preoperative levels at 8 months (P = 0.055). With BL partially present, subbasal nerve density did not significantly change from preoperative levels. Superficial, wing, and basal epithelial cell density recovered to preoperative levels within 4 months after PTK, regardless of the presence of BL. Subepithelial keratocytes, however, were more densely distributed in corneas without BL relative to those with a partial BL present (P = 0.005), and increased anterior keratocyte reflectivity was noted in all eyes without BL and in no eye with a partial BL present. CONCLUSIONS Subbasal nerve regeneration is delayed and subepithelial keratocyte density and reflectivity remain elevated up to 10 months after total BL removal by PTK. The results provide initial evidence for a possible role of BL in facilitating rapid stromal wound healing and an associated recovery of anterior corneal transparency and the restoration of epithelial innervation after epithelial trauma.

Clinical Outcome and Recurrence of Epithelial Basement Membrane Dystrophy after Phototherapeutic Keratectomy: A Cross-sectional Study

OBJECTIVE To evaluate the outcome of phototherapeutic keratectomy (PTK) treatment of epithelial basement membrane dystrophy (EBMD) patients and to examine clinical and morphologic signs of recurrent dystrophy. DESIGN Cross-sectional, clinic-based study. PARTICIPANTS Fifty-two eyes of 39 patients diagnosed with EBMD who underwent PTK between 2001 and 2008. METHODS Preoperative symptoms, best spectacle-corrected visual acuity (BSCVA), and refraction data were collected. At follow-up, refraction and BSCVA were measured, symptoms were noted, and slit-lamp biomicroscopy and in vivo confocal microscopy (IVCM) were performed. MAIN OUTCOME MEASURES Best spectacle-corrected visual acuity and signs of recurrent EBMD based on symptoms and morphologic features. An assessment of EBMD severity after PTK additionally was considered. RESULTS Mean follow-up time was 43 months (range, 7-100 months). After PTK, BSCVA remained unchanged or improved in 49 (98%) of 51 eyes. Twenty-four (46%) of 52 eyes had recurrence of some form, and recurrence was correlated positively with postoperative time (P<0.001). Symptomatic recurrence occurred in 7 eyes (13%), whereas morphologic recurrence occurred in 21 eyes (40%). Symptoms were coupled with positive IVCM findings in 3 (43%) of 7 cases and with slit-lamp findings in 1 (14%) of 7 cases. Of 17 eyes with morphologic recurrence by IVCM, 9 eyes (53%) were classified as having grade 1 recurrence, 8 eyes (47%) were classified as having grade 2 recurrence, and none were classified as having grade 3 recurrence. Morphologic recurrence was associated with epithelial removal by laser ablation before PTK. CONCLUSIONS Although PTK is an effective method of alleviating the clinical symptoms of EBMD, the dystrophy can recur with time. The relationship between the postoperative development of clinical symptoms and the corneal morphologic features is complex and requires further investigation.

Age-Related Thinning of Bowman's Layer in the Human Cornea In Vivo

PURPOSE To determine the thickness of Bowmans layer (BL) in vivo in a healthy population and to determine its variation with age. METHODS Eighty-two subjects aged 15 to 88 years with clear, healthy corneas were examined bilaterally with laser scanning in vivo confocal microscopy (IVCM). Bowmans layer thickness was determined from IVCM images of anterior and posterior BL boundaries. For a given eye, BL thickness was averaged across four central locations by two independent observers. In addition, central corneal thickness was measured by time-domain optical coherence tomography. RESULTS A significant negative correlation of BL thickness with age was found in right eyes (Pearson r = -0.579, P < 0.0001) and in left eyes (r = -0.558, P < 0.0001). Linear regression analysis yielded a decline in BL thickness of 0.06 μm per year. In 41 older subjects (mean age, 64.4 years), BL thickness was significantly thinner (mean ± SD, 8.6 ± 1.7 μm in right eyes) than that in 41 younger subjects (mean age, 31.6 years) (mean ± SD, 10.7 ± 1.6 μm in right eyes) (P < 0.001). No correlation of corneal thickness with age or of BL thickness with corneal thickness was observed. Strong intereye correlations in BL thickness (r = 0.771, P < 0.0001) and corneal thickness (r = 0.969, P < 0.001) were found. CONCLUSIONS Bowmans layer thins with age in the normal cornea, losing one-third of its thickness between the ages of 20 and 80 years. In vivo measurement of BL thickness by IVCM could aid in clinical assessment and planned treatments of the anterior cornea.

Laser-Scanning in vivo Confocal Microscopy of the Cornea: Imaging and Analysis Methods for Preclinical and Clinical Applications

Laser-Scanning in vivo Confocal Microscopy of the Cornea : Imaging and Analysis Methods for Preclinical and Clinical Applications

Pathologically reduced subbasal nerve density in epithelial basement membrane dystrophy is unaltered by phototherapeutic keratectomy treatment

PURPOSE To investigate the effect of phototherapeutic keratectomy (PTK) treatment on corneal epithelial wing cell and corneal subbasal nerve density in epithelial basement membrane dystrophy (EBMD). METHODS A total of 39 patients with EBMD who underwent PTK treatment, 40 healthy volunteers, and 24 untreated eyes with EBMD were examined with laser-scanning in vivo confocal microscopy (IVCM). Corneal subbasal nerves and epithelial wing cells were manually quantified from IVCM images by two observers, while epithelial wing cells were additionally quantified by a fully automated method. RESULTS Subbasal nerve density was significantly reduced in untreated (10,164 ± 4139 μm/mm(2); n = 24) and PTK-treated (10,624 ± 4479 μm/mm(2); n = 39) EBMD eyes, relative to healthy controls (18,241 ± 4479 μm/mm(2); n = 40) (P < 0.001). Subbasal nerve density in PTK-treated and untreated eyes did not differ (P > 0.05). Epithelial wing cell density did not differ between PTK-treated and untreated EBMD eyes, by either manual or automated analysis; however, epithelial wing cell density in PTK-treated EBMD corneas was significantly reduced (P = 0.008) relative to healthy corneas, by automated cell counting. CONCLUSIONS Subbasal nerve density in EBMD is reduced by 45% and recovers only to the reduced level in the long term after PTK treatment, whereas epithelial wing cell density in EBMD is not affected by PTK in the long term. Fully automated cell analysis from IVCM images could provide an objective, standardized means to quantify and compare corneal cell densities in future studies.

An accurate method to determine Bowman's layer thickness in vivo in the human cornea.

PURPOSE To determine an accurate value for Bowmans layer (BL) thickness in vivo in humans. METHODS Seventeen corneal transplant patients were examined preoperatively by laser-scanning in vivo confocal microscopy (IVCM), and corneal buttons were removed postoperatively and sectioned for light microscopy (LM). Nine corneas with uniformly thick BL by LM were used for thickness measurement. In the uniformly thick samples, probable overestimation of BL thickness in vivo by a first in vivo method (Method 1) led to the development of a revised in vivo method (Method 2). Method 2 was used to measure BL thickness in 20 healthy volunteers. RESULTS In nine patients, mean BL thickness prior to transplantation was 13.7 ± 1.6 μm by IVCM (Method 1) while BL thickness of the removed corneal button was 9.7 ± 1.7 μm by LM (P < 0.001). The correlation of BL thickness between IVCM (Method 1) and LM was poor (P = 0.226). In 20 right eyes of 20 normal corneas, both in vivo methods were used to determine BL thickness. Mean BL thickness by Method 1 was 13.2 ± 1.6 μm and by Method 2 was 9.1 ± 1.4 μm (P < 0.001). BL thickness measurements by both in vivo methods were highly correlated (P < 0.001). CONCLUSION BL thickness by a revised in vivo method was close to LM values in this study and to values reported in fixed tissue in other studies. The authors believe this revised method provides the most accurate estimates of BL thickness in vivo to date.

RGTA in corneal wound healing after transepithelial laser ablation in a rabbit model: a randomized, blinded, placebo-controlled study.

To evaluate the efficacy of the agent RGTA for epithelial, stromal and nerve regeneration after laser‐induced corneal wounding in rabbits.