Reprint of ""Dynamics of the expression of intermediate filaments vimentin and desmin during myofibroblast differentiation after corneal injury""

Previous studies have suggested that abnormal corneal wound healing in patients after photorefractive keratectomy (PRK) is associated with the appearance of myofibroblasts in the stroma between two and four weeks after surgery. The purpose of this study was to examine potential myofibroblast progenitor cells that might express other filament markers prior to completion of the differentiation pathway that yields alpha-smooth muscle actin (SMA)-expressing myofibroblasts associated with haze localized beneath the epithelial basement membrane after PRK. Twenty-four female rabbits that had -9 diopter PRK were sacrificed at 1 week, 2 weeks, 3 weeks or 4 weeks after surgery. Corneal rims were collected, frozen at -80 degrees C, and analyzed by immunocytochemistry using anti-vimentin, anti-desmin, and anti-SMA antibodies. Double immunostaining was performed for the co-localization of SMA with vimentin or desmin with SMA. An increase in vimentin expression in stromal cells is noted as early as 1 week after PRK in the rabbit cornea. As the healing response continues at two or three weeks after surgery, many stromal cells expressing vimentin also begin to express desmin and SMA. By 4 weeks after the surgery most, if not all, myofibroblasts express vimentin, desmin and SMA. Generalized least squares regression analysis showed that there was strong evidence that each of the marker groups differed in expression over time compared to the other two (p < 0.01). Intermediate filaments - vimentin and desmin co-exist in myofibroblasts along with SMA and may play an important role in corneal remodeling after photorefractive keratectomy. The earliest precursors of myofibroblasts destined to express SMA and desmin are detectible by staining for vimentin at 1 week after surgery. (C) 2009 Elsevier Ltd. All rights reserved.

Effect of Femtosecond Laser Energy Level on Corneal Stromal Cell Death and Inflammation

PURPOSE: To analyze the effects of variations in femtosecond laser energy level on corneal stromal cell death. and inflammatory cell influx following flap creation in a rabbit model. METHODS: Eighteen rabbits were stratified in three different groups according to level of energy applied for flap creation (six animals per group). Three different energy levels were chosen for both the lamellar and side cut; 2.7 mu J (high energy), 1.6 mu J (intermediate energy), and 0.5 mu J (low energy) with a 60 kHz, model II, femtosecond laser (IntraLase). The opposite eye of each rabbit served as a control. At the 24-hour time point after surgery, all rabbits were euthanized and the comeoscleral rims were analyzed for the levels of cell death and inflammatory cell influx with the terminal uridine deoxynucleotidyl transferase dUTP-nick end labeling (TUNEL) assay and immunocytochemistry for monocyte marker CD11b, respectively. RESULTS: The high energy group (31.9 +/- 7.1 [standard error of mean (SEM) 2.9]) had significantly more TUNEL positive cells in the central flap compared to the intermediate (22.2 +/- 1.9 [SEM 0.8], P=.004), low (17.9 +/- 4.0 [SEM 1.6], P <= .001), and control eye (0.06 +/- 0.02 [SEM 0.009], P <= .001) groups. The intermediate and low energy groups also had significantly more TUNEL positive cells than the control groups (P <= .001). The difference between the intermediate and low energy levels was not significant (P=.56). The mean for CD11b-positive cells/400x field at the flap edge was 26.1 +/- 29.3 (SEM 11.9), 5.8 +/- 4.1 (SEM 1.6), 1.6 +/- 4.1 (SEM 1.6), and 0.005 +/- 0.01 (SEM 0.005) for high energy, intermediate energy, low energy, and control groups, respectively. Only the intermediate energy group showed statistically more inflammatory cells than control eyes (P = .015), most likely due to variability between eyes. CONCLUSIONS: Higher energy levels trigger greater cell death when the femtosecond laser is used to create corneal flaps: Greater corneal inflammatory cell infiltration is observed with higher femtosecond laser energy levels. [J Refract Surg. 2009;25:869-874.] doi:10.3928/1081597X-20090917-08

Dynamics of the expression of intermediate filaments vimentin and desmin during myofibroblast differentiation after corneal injury

Previous studies have suggested that abnormal corneal wound healing in patients after photorefractive keratectomy (PRK) is associated with the appearance of myofibroblasts in the stroma between two and four weeks after surgery. The purpose of this study was to examine potential myofibroblast progenitor cells that might express other filament markers prior to completion of the differentiation pathway that yields a-smooth muscle actin (SMA)-expressing myofibroblasts associated with haze localized beneath the epithelial basement membrane after PRK. Twenty-four female rabbits that had -9 diopter PRK were sacrificed at I week, 2 weeks, 3 weeks or 4 weeks after surgery. Corneal rims were collected, frozen at -80 degrees C, and analyzed by immunocytochemistry using anti-vimentin, anti-desmin, and anti-SMA antibodies. Double immunostaining was performed for the co-localization of SMA with vimentin or desmin with SMA. An increase in vimentin expression in stromal cells is noted as early as 1 week after PRK in the rabbit cornea. As the healing response continues at two or three weeks after surgery, many stromal cells expressing vimentin also begin to express desmin and SMA. By 4 weeks after the surgery most, if not all, myofibroblasts express vimentin, desmin and SMA. Generalized least squares regression analysis showed that there was strong evidence that each of the marker groups differed in expression over time compared to the other two (p < 0.01). Intermediate filaments - vimentin and desmin co-exist in myofibroblasts along with SMA and may play an important role in corneal remodeling after photorefractive keratectomy. The earliest precursors of myofibroblasts destined to express SMA and desmin are detectible by staining for vimentin at I week after surgery. (C) 2009 Elsevier Ltd. All rights reserved.

Relationship between Optical Coherence Tomography, Pattern Electroretinogram and Automated Perimetry in Eyes with Temporal Hemianopia from Chiasmal Compression

PURPOSE. To evaluate the relationship between pattern electroretinogram (PERG) amplitude, macular and retinal nerve fiber layer (RNFL) thickness by optical coherence tomography (OCT), and visual field (VF) loss on standard automated perimetry (SAP) in eyes with temporal hemianopia from chiasmal compression. METHODS. Forty-one eyes from 41 patients with permanent temporal VF defects from chiasmal compression and 41 healthy subjects underwent transient full-field and hemifield (temporal or nasal) stimulation PERG, SAP and time domain-OCT macular and RNFL thickness measurements. Comparisons were made using Student`s t-test. Deviation from normal VF sensitivity for the central 18 of VF was expressed in 1/Lambert units. Correlations between measurements were verified by linear regression analysis. RESULTS. PERG and OCT measurements were significantly lower in eyes with temporal hemianopia than in normal eyes. A significant correlation was found between VF sensitivity loss and fullfield or nasal, but not temporal, hemifield PERG amplitude. Likewise a significant correlation was found between VF sensitivity loss and most OCT parameters. No significant correlation was observed between OCT and PERG parameters, except for nasal hemifield amplitude. A significant correlation was observed between several macular and RNFL thickness parameters. CONCLUSIONS. In patients with chiasmal compression, PERG amplitude and OCT thickness measurements were significant related to VF loss, but not to each other. OCT and PERG quantify neuronal loss differently, but both technologies are useful in understanding structure-function relationship in patients with chiasmal compression. (ClinicalTrials.gov number, NCT00553761.) (Invest Ophthalmol Vis Sci. 2009; 50: 3535-3541) DOI:10.1167/iovs.08-3093

Corneal stroma PDGF blockade and myofibroblast development

Myofibroblast development and haze generation in the corneal stroma is mediated by cytokines, including transforming growth factor-beta (TGF-beta), and possibly other cytokines. This study examined the effects of stromal PDGF-beta blockade on the development of myofibroblasts in response to -9.0 diopter photorefractive keratectomy in the rabbit. Rabbits that had haze generating photorefractive keratectomy (PRK, for 9 diopters of myopia) in one eye were divided into three different groups: stromal application of plasmid pCMV.PDGFRB.23KDEL expressing a subunit of PDGF receptor b (domains 2-3, which bind PDGF-B), stromal application of empty plasmid pCMV, or stromal application of balanced salt solution (BSS). The plasmids (at a concentration 1000 ng/mu l) or BSS was applied to the exposed stroma immediately after surgery and every 24 h for 4-5 days until the epithelium healed. The group treated with pCMV.PDGFRB.23KDEL showed lower alpha SMA+ myofibroblast density in the anterior stroma compared to either control group (P <= 0.001). Although there was also lower corneal haze at the slit lamp at one month after surgery, the difference in haze after PDGF-B blockade was not statistically significant compared to either control group. Stromal PDGF-B blockade during the early postoperative period following PRK decreases stromal alpha SMA+ myofibroblast generation. PDGF is an important modulator of myofibroblast development in the cornea. (C) 2008 Elsevier Ltd. All rights reserved.

Agreement between stress intraocular pressure and long-term intraocular pressure measurements in primary open angle glaucoma

The aim of this paper is to study the correlation between the intraocular pressure peaks and fluctuation detected during the water drinking test and the same parameters observed during long-term follow up. This prospective cohort study enrolled 22 eyes of 22 newly diagnosed primary open angle glaucoma patients. After an initial complete ophthalmological examination, patients were started on antiglaucoma medication and returned 4 weeks later to perform the water drinking test. Thereafter, patients were evaluated at least eight times within a period of 6-12 months. The intraocular pressure peaks and fluctuation detected during the water drinking stress test were compared with those observed during regular office visits. Spearman`s correlation coefficient and Bland-Altman Plots were used for statistical analysis. The mean age of participants was 54.3 +/- 8.2 years (+/- SD), 59% were women, and average mean deviation -10.2 +/- 4.5 dB. The mean follow-up period was 8.2 +/- 2.0 months. The average intraocular pressure peaks and fluctuation during the water drinking test were 20.0 +/- 2.9 mmHg and 40 +/- 10%, respectively, and 18.1 +/- 2.8 mmHg and 30 +/- 10% during follow up. Spearman`s correlation coefficients were significant and strong between the intraocular pressure peaks and fluctuation detected during the water drinking test and during the follow-up period (P < 0.001, rho = 0.76 and 0.82, respectively). There was good agreement between the variables. The intraocular pressure peaks and fluctuation detected during the water drinking test showed significant correlation and agreement with the pressures observed during follow-up visits. Stress tests could be used to estimate long-term intraocular pressure variation.

Comparative study of Botox (R) injection treatment for upper eyelid retraction with 6-month follow-up in patients with thyroid eye disease in the congestive or fibrotic stage

Aim To compare morphometric data of the eyelid fissure and the levator muscle function (LF) before and up to 6 months after transcutaneous injection with five units of Botox (R) in patients with upper lid retraction (ULR) from congestive or fibrotic thyroid eye disease (TED). Methods Twenty-four patients with ULR from TED were submitted to transcutaneous injection of 5 units (0.1 ml) of Botox in one eye only. Patients were divided into two groups: 12 with congestive-stage TED (CG), and 12 with fibrotic-stage TED (FG). Bilateral lid fissure measurements using digital imaging and computer-aided analysis were taken at baseline and at regular intervals 2 weeks, 1 month, 3 months and 6 months after unilateral Botox injection. Mean values taken at different follow-up points were compared for the two groups. Results Most patients experienced marked improvement in ULR, with a mean reduction of 3.81 mm in FG and 3.05 mm in CG. The upper eyelid margin reflex distance, fissure height and total area of exposed interpalpebral fissure were significantly smaller during 1 month in CG and during 3 months in FG. Reduction in LF and in the difference between lateral and medial lid fissure measurements was observed in both groups. The treatment lasted significantly longer in FG than in CG. Conclusions A single 5-unit Botox injection improved ULR, reduced LF and produced an adequate lid contour in patients with congestive or fibrotic TED. The effect lasts longer in patients with fibrotic orbitopathy than in patients with congestive orbitopathy.

Browlifting as an Alternative Procedure for Apraxia of Eyelid Opening

Essential blepharospasm is an idiopathic disorder that consists of spontaneous, spasmodic, and involuntary eyelid closure in the absence of ocular disease. Some patients develop an inability to open their eyelids in the absence of orbicularis spasms. These patients have essential blepharospasm combined with apraxia of eyelid opening. Botulinum toxin injections are the treatment of choice for blepharospasm but results may be insufficient, especially in cases associated with apraxia. Apraxia can be treated surgically by levator aponeurosis advancement, frontalis suspension, and upper myectomy. The authors report the first browlift using polypropylene suture to manage eyelid apraxia associated with blepharospasm as an alternative and minimally invasive procedure.

Haze Development After Photorefractive Keratectomy: Mechanical vs Ethanol Epithelial Removal in Rabbits

PURPOSE: To compare mechanical and ethanol epithelial removal with respect to myofibroblast development and haze formation after photorefractive keratectomy (PRK). METHODS: Seventeen rabbits underwent mechanical or ethanol debridement, and the opposite eye of each rabbit served as an unwounded control. In both groups, the epithelium was removed with a spatula and discarded. A -9.00-diopter PRK was performed in each eye. The level of haze in each cornea at 4 weeks was graded at the slit-lamp microscope according to the Fantes scale. Myofibroblast generation was detected with immunocytochemistry for alpha-smooth muscle actin (alpha-SMA) and cells were quantitatively analyzed. RESULTS: No difference was noted between the two groups in alpha-SMA + myofibroblasts 4 weeks after surgery (43.6 +/- 2.0/400X field and 45.7 +/- 4.8/400X field in ethanol and mechanical groups, respectively) (P=.10). A slight difference was noted but did not reach statistical significance with regard to stromal haze between ethanol and mechanical groups (2.0 +/- 0.5 and 2.3 +/- 0.4, respectively, P=.063). The ethanol and mechanical groups were statistically different when compared to controls regarding stromal haze and alpha-SMA+ cells (P <.0001 for all comparisons). CONCLUSIONS:No difference was noted in clinical haze or myofibroblast generation between corneas that had PRK with mechanical,or ethanol epithelial debridement. [J Refract Surg., 2008;24:923-927.]

Clinical Treatment of Ocular Demodex folliculorum by Systemic Ivermectin

PURPOSE: To report clinical outcomes of the treatment of ocular Demodex folliculorum with oral ivermectin. DESIGN: Noncomparative, interventional case series. METHODS: Setting. Institutional. Study Population. Twenty-four eyes of 12 patients (3 male and 9 female; mean age +/- standard deviation, 50.4 +/- 21.0 years) with refractory posterior blepharitis with the presence of D. folliculorum in lash samples were enrolled in this study. Intervention. Patients were instructed to take 1 dose of oral ivermectin (200 mu g/kg). All patients were instructed to repeat the treatment after 7 days. Main outcome measures. Tear meniscus height, Schirmer I test results, noninvasive tear film break-up time (BUT), quantification of the absolute number of D. folliculorum found in the lashes, and corneal fluorescein and rose bengal staining scores were obtained from all patients 1 day before and 28 days after treatment. RESULTS: Statistical improvement was observed in the absolute number of D. folliculorum found in the lashes after the treatment with oral ivermectin. Average values of Schirmer I test results and tear film break-up time improved statistically after the treatment of oral ivermectin. No statistical improvement was observed in average lacrimal meniscus height or value of corneal fluorescein and rose bengal staining after treatment with oral ivermectin. CONCLUSIONS: Ivermectin successfully reduced the number of D. folliculorum found in the lashes of patients with refractory blepharitis. Oral ivermectin may be very useful as a complement in the treatment of D. folliculorum infestation with ocular manifestation, especially in cases of unsuccessful treatment related to patient compliance. (Am J Ophthalmol 2011;151:1030-1034. (C) 2011 by Elsevier Inc. All rights reserved.)

Intraorbital polyacrylamide gel injection for the treatment of anophthalmic enophthalmos

Purpose: To evaluate the use of orbital polyacrylamide gel injection for the correction of anophthalmic enophthalmos. Methods: Noncontrolled clinical trial of 21 patients (14 with ocular implants, 5 with phthisis bulbi, and 2 with dermis-fat graft). Orbital CT was performed to estimate the volume of polyacrylamide gel needed to restore orbital volume. Polyacrylamide gel was injected using a 22-gauge (30 x 0.7 min) needle transcutaneously inserted in the lateral third of the lower eyelid, directed to the orbital muscle cone. A second injection was administered 15 days later. if necessary. CT was repeated 30 days after the last procedure. Exophthalmometry was performed before Bind 90 days after file procedure. Results: The mean total volume injected per orbit was 2.4 +/- 0.7 ml (range 1-3.5 ml). The volume of the enophthalmic orbit increased front 26.9 +/- 5.0 ml to 29.3 +/- 4.9 ml (p < 0.001). The mean difference in exophthalmometry readings was 3.3 +/- 1.6 mm (range, 1.5-8.0 mm) before the procedure and 1.0 +/- 0.9 mm (range, 0.0-3.0 mm) after 3 months (p < 0.001). Adjustment of the ocular prosthesis or fabrication of a new one was necessary in 11 patients (52.4%), and the mean volume of the ocular prosthesis was reduced front 2.0 +/- 0.6 ml to 1.6 +/- 0.6 ml (p = 0.003). All patients were satisfied with the aesthetic results. No serious adverse events were observed. The initial results were maintained 1 year after the procedure. Conclusions: Polyacrylamide gel injection in the orbital space effectively reduces enophthalmos in ocular prosthesis wearers.

Combination micrografting and tattooing in the reconstruction of eyebrows of patients with craniofacial clefts

The absence of an eyebrow, either partial or total, has been observed in patients with craniofacial clefts, such as the Tessier 9 to 13 cleft. Several techniques have been used to improve the appearance of the region, such as island scalp flaps and scalp strip grafting, with limited or marginally satisfactory aesthetic results. The authors report 2 patients with craniofacial clefts in whom a novel technique combining 2 separate surgical approaches, micrografting and tattooing, was used. The use of micrografting with single or double hair units, properly angulated, produces natural-looking and satisfactory results with a minimum of morbidity. Excellent volume and appearance of the eyebrow may be achieved in a single session using this technique. Tattooing performed subsequently over the microimplanted hairs provides the illusion of greater density to the eyebrow, resulting in an appearance closer to normal.

Topical interleukin-1 receptor antagonist inhibits inflammatory cell infiltration into the cornea

Interleukin (IL)-1 alpha and beta are important modulators of many functions of corneal epithelial and stromal cells that occur following injury to the cornea, including the influx of bone marrow-derived inflammatory cells into the stroma attracted by chemokines released from the stroma and epithelium. In this study, we examined the effect of topical soluble IL-1 receptor antagonist on bone marrow-derived cell influx following corneal epithelial scrape injury in a mouse model. C57BL/6 mice underwent corneal epithelial scrape followed by application of IL-1 receptor antagonist (Amgen, Thousand Oaks, CA) at a concentration of 20 mg/ml or vehicle for 24 h prior to immunocytochemical detection of marker CD11b-positive cells into the stroma. In two experiments, topical IL-1 receptor antagonist had a marked effect in blocking cell influx. For example, in experiment 1, topical IL-1 receptor antagonist markedly reduced detectible CD11b-positive cells into the corneal stroma at 24 It after epithelial injury compared with the vehicle control (3.5 +/- 0.5 (standard error of the mean) cells/400x field and 13.9 +/- 1.2 cells/400x field, respectively, p < 0.01). A second experiment with a different observer performing cell counting had the same result. Thus, the data demonstrate conclusively that topical IL-1 receptor antagonist markedly down-regulates CD-11b-positive monocytic cell appearance in the corneal stroma. Topical IL-1 receptor antagonist could be an effective adjuvant for clinical treatment of corneal conditions in which unwanted inflammation has a role in the pathophysiology of the disorder. (c) 2008 Elsevier Ltd. All rights reserved.

Comparison of HRT-3 glaucoma probability score and subjective stereophotograph assessment for prediction of progression in glaucoma

PURPOSE. To assess whether baseline Glaucoma Probability Score (GPS; HRT-3; Heidelberg Engineering, Dossenheim, Germany) results are predictive of progression in patients with suspected glaucoma. The GPS is a new feature of the confocal scanning laser ophthalmoscope that generates an operator-independent, three-dimensional model of the optic nerve head and gives a score for the probability that this model is consistent with glaucomatous damage. METHODS. The study included 223 patients with suspected glaucoma during an average follow-up of 63.3 months. Included subjects had a suspect optic disc appearance and/or elevated intraocular pressure, but normal visual fields. Conversion was defined as development of either repeatable abnormal visual fields or glaucomatous deterioration in the appearance of the optic disc during the study period. The association between baseline GPS and conversion was investigated by Cox regression models. RESULTS. Fifty-four (24.2%) eyes converted. In multivariate models, both higher values of GPS global and subjective stereophotograph assessment ( larger cup-disc ratio and glaucomatous grading) were predictive of conversion: adjusted hazard ratios (95% CI): 1.31 (1.15 - 1.50) per 0.1 higher global GPS, 1.34 (1.12 - 1.62) per 0.1 higher CDR, and 2.34 (1.22 - 4.47) for abnormal grading, respectively. No significant differences ( P > 0.05 for all comparisons) were found between the c-index values ( equivalent to area under ROC curve) for the multivariate models (0.732, 0.705, and 0.699, respectively). CONCLUSIONS. GPS values were predictive of conversion in our population of patients with suspected glaucoma. Further, they performed as well as subjective assessment of the optic disc. These results suggest that GPS could potentially replace stereophotograph as a tool for estimating the likelihood of conversion to glaucoma.

Scanning laser polarimetry with enhanced corneal compensation for detection of axonal loss in band atrophy of the optic nerve

PURPOSE: To compare the abilities of scanning laser polarimetry (SLP) with enhanced corneal compensation (ECC) and variable corneal compensation (VCC) modes for detection of retinal nerve fiber layer (RNFL) loss in eyes with band atrophy (BA) of the optic nerve. DESIGN. Cross-sectional study. METHODS: Thirty-seven eyes from 37 patients with BA and temporal visual field defect from chiasmal compression and 40 eyes from 40 healthy subjects were studied. Subjects underwent standard automated perimetry and RNFL measurements using an SLP device equipped with VCC and ECC. Receiver operating characteristic (ROC) curves were calculated for each parameter. Pearson correlation coefficients were obtained to evaluate the relationship between RNFL thickness parameters and severity of visual field loss, as assessed by the temporal mean defect. RESULTS: All RNFL thickness parameters were significantly lower in eyes with BA compared with normal eyes with both compensation modes. However, no statistically significant differences were observed in the areas under the ROC curves for the different parameters between GDx VCC and ECC (Carl Zeiss Meditec, Inc, Dublin, California, USA). Structure-function relationships also were similar for both compensation modes. CONCLUSIONS: No significant differences were found between the diagnostic accuracy of GDx ECC and that of VCC for detection of BA of the optic nerve. The use of GDx ECC does not seem to provide a better evaluation of RNFL loss on the temporal and nasal sectors of the peripapillary retina in subjects with BA of the optic nerve.