Birch pollen allergen Bet v 1 binds to and is transported through conjunctival epithelium in allergic patients.

BACKGROUND: Previous work in type-I pollen allergies has mainly focused on lymphocytes and immune responses. Here, we begin to analyse with a systems biology view the differences in conjunctival epithelium obtained from healthy and allergic subjects. METHODS: Transcriptomics analysis combined with light and electron microscopic analysis of birch pollen allergen Bet v 1 located within conjunctival epithelial cells and tissues from birch allergic subjects and healthy controls was carried out. RESULTS: Bet v 1 pollen allergen bound to conjunctival epithelial cells within minutes after the exposure even during the nonsymptomatic winter season only in allergic, but not in healthy individuals. Light- and electron microscopy showed that Bet v 1 was transported through the epithelium within lipid rafts/caveolae and reached mast cells only in allergic patients, but not in healthy individuals. Transcriptomics yielded 22 putative receptors expressed at higher levels in allergic epithelium compared with healthy specimens. A literature search indicated that out of these receptors, eight (i.e. 37%) were associated with lipid rafts/caveolae, which suggested again that Bet v 1 transport is lipid raft/caveola-dependent. CONCLUSIONS: We show a clear difference in the binding and uptake of Bet v 1 allergen by conjunctival epithelial cells in allergic vs healthy subjects and several putative lipid raft/caveolar receptors were identified, which could mediate or be co-transported with this entry. The application of discovery driven methodologies on human conjunctival epithelial cells and tissues can provide new hypotheses worth a further analysis to the molecular mechanisms of a complex multifactorial disease such as type-I birch pollen allergy.

Corneal recovery after lasik for high myopia: a 2-year prospective confocal microscopic study.

AIM: To quantify human corneal recovery after moderate to high myopic laser in situ keratomileusis (LASIK) in a 2-year prospective follow-up study. METHODS: Fifteen eyes of 15 patients (mean refraction -10.1 (SD 2.4) D) were examined preoperatively and postoperatively at day 1, 5 days, 2 weeks, 1, 3 and 6 months and 2 years. Biomicroscopy, visual acuity and refraction were examined prior to imaging studies. An in vivo tandem scanning confocal microscope was used to obtain images from the central cornea. Subbasal nerve density was measured as the total length of nerve trunks in confocal image per mm2. Keratocyte density was calculated manually from stromal sublayers. The thickness of the altered keratocyte zone was measured on both sides of the LASIK interface. RESULTS: At the end of the follow-up, all patients had a 20/20 BCVA, and nine of 15 patients were within +/-0.5 D of the intended correction. The total corneal thickness remained unaltered, but epithelial hyperplasia was seen at 2 years. Keratocyte density in the anterior stroma and posterior to the flap interface showed a slight decrease during the follow-up. Subbasal nerve density decreased 82% in 5 days after LASIK. A gradual increase was observed from 2 weeks postoperatively, but even 2 years after the operation the nerve density was only 64% from the preoperative values. CONCLUSIONS: Subbasal nerve fibre density shows a gradual recovery throughout the follow-up. However, only three subjects showed totally regenerated subbasal nerve fibres at 2 years. This may correlate with the observed decrease in the density of the most anterior keratocytes. Corneal remodelling seemed to continue for at least 2 years.

Human tear fluid PDGF-BB, TNF-alpha and TGF-beta1 vs corneal haze and regeneration of corneal epithelium and subbasal nerve plexus after PRK.

The aim was to determine the association of tear fluid cytokine levels and post-PRK corneal haze evaluated by in vivo confocal microscopy. In addition, the possible association between subbasal neural regeneration and haze formation, or epithelial regeneration were investigated. Twenty eyes of 20 patients (16 women and four men, age 30.7 +/- 7.5 years, range 21-48 years) underwent a myopic PRK. The spherical equivalent (SE) of the intended correction was -4.7 +/- 1.5 D (range -2.75 to -9.00 D). ELISA-methods were used to assess tear fluid concentrations of TGF-beta1, PDGF-BB and TNF-alpha pre-operatively, and post-operatively on day 2 and at 3 months. Tear fluid flow in the collection capillary was recorded, and rates of cytokine release (= tear fluid flow-corrected concentrations) were calculated. In vivo confocal microscopy was performed at 3 months to evaluate the corneal morphology and to determine numerical haze estimate. There was wide interindividual variation between pre-operative and post-operative concentrations and rates of release of TGF-beta1, PDGF-BB and TNF-alpha. Subepithelial haze was observed in all corneas and the mean haze estimate was 506 +/- 401 U (100-1410 U). However, no association was found between tear fluid cytokine levels and post-PRK haze. Regenerating subbasal nerve plexus was found in 18 out of 20 corneas; in two corneas it was absent or could not be visualized due to subepithelial haze. The density of the subbasal nerve fiber bundles had a positive correlation with the epithelial thickness (Pearson correlation, r = 0.56, P = 0.011), but not with the haze estimate or the thickness of the haze area. At 3 months post-PRK, haze could be observed in all patients. The results suggest that tear fluid cytokine analysis, as measured, may not be suitable for screening the potential candidates for haze formation. We did not find any correlation between haze and regeneration of subbasal nerve plexus, but we demonstrated that the regeneration of subbasal nerve plexus might have significant influence on regulation of epithelial healing.

Corneal morphology and sensitivity in lattice dystrophy type II (familial amyloidosis, Finnish type).

PURPOSE: To describe the corneal abnormalities and to measure different modalities of corneal sensitivity in corneal lattice dystrophy type II (familial amyloidosis, Finnish type, also known as gelsolin-related amyloidosis and originally as Meretoja syndrome). METHODS: Twenty eyes of 20 patients were examined by in vivo confocal microscopy and noncontact gas esthesiometry. RESULTS: Pleomorphism of, and dense deposits between or posterior to, the basal epithelial cells were frequently observed, as well as a reduction of long nerve fiber bundles in the subbasal nerve plexus. The anterior stroma was altered in most cases, with fibrosis and abnormal extracellular matrix. In 15 corneas, thick anterior and midstromal filaments, corresponding to lattice lines, and in 11 corneas, thin undulated structures were observed. The average mechanical sensitivity threshold of 12 subjects was increased, and in the remaining 8 subjects there was no response, even to the highest intensity of stimuli used. Three patients did not respond to CO(2), 11 to heat, and 2 to cold, but those patients who responded had normal thresholds. Patients with more long nerve fiber bundles per confocal microscopic image had better mechanical and cold sensitivity than patients with fewer nerve fiber bundles. CONCLUSIONS: Lattice lines seem to be related to amyloid material and not to corneal nerves. However, the subbasal nerve density appears reduced, which results mainly in a decrease in mechanical and, to a lesser extent, thermal sensitivity. The location of stromal filaments and undulated structures changes with increasing age.

Laminin synthesis and the adhesion characteristics of immortalized human corneal epithelial cells to laminin isoforms.

We have studied the synthesis of laminins (Ln) and determined the specific integrins mediating the adhesion of immortalized human corneal epithelial cells to mouse Ln-1, and human Lns-5 and -10. Immunofluorescence microscopy of the cells demonstrated integrin alpha(2), alpha(3), alpha(6), beta(1)and beta(4)subunits, integrins alpha(6)and beta(4)being found in a typical 'leopard-skin' like manner. Immunoprecipitation studies showed that the cells produced alpha 3, beta 3 and gamma 2 chains of Ln-5, but not Lns-1 or -10. In culture Ln-5 was found as small plaques beneath the adhering cells within 1 hr, while in 4 hr widely spread Ln-5 plaques were observed in colocalization with beta(4)integrin subunit. By using a quantitative cell adhesion assay and function-blocking monoclonal antibodies we showed that integrin beta(1)subunit plays a role in mediating corneal epithelial cell adhesion to mouse Ln-1. However, none of the available function-blocking antibodies to integrin alpha-subunits inhibited the adhesion. Integrin alpha(3)beta(1)complex mediated the adhesion of corneal epithelial cells to human Lns-5 and -10. Integrin complex alpha(3)beta(1), as well as laminin alpha(3)chain, was also shown to mediate cell adhesion to newly produced endogenous Ln-5. The present results show that integrin alpha(3)beta(1)complex mediates the adhesion of corneal epithelial cells to Lns-5 and -10, while a yet unknown integrin alpha subunit appears to play a role in the adhesion to Ln-1. The results also show that among corneal basement membrane laminins, Ln-5 is synthetized by epithelial cells while Ln-10 may be a product of keratocytes.

Photorefractive keratectomy in ophthalmic residents.

PURPOSE: To find out how ophthalmologists themselves experience the correction of myopia after photorefractive keratectomy. Visuomotor functions were of special interest. METHODS: Four ophthalmology residents and one medical engineer underwent photorefractive keratectomy for myopia. Objective measurements including refraction, corneal topography, perimetry, contrast sensitivity, pattern visual evoked potentials, in vivo confocal microscopy, and a car driving simulator test were performed preoperatively, postoperatively, and at 6 months. Subjective evaluation was reported. RESULTS: Performing ophthalmological examinations and microsurgery without spectacles was easier postoperatively and was appreciated by the four ophthalmology residents. Minimal haze formation, good accuracy, and normal performance in the car driving simulator were also observed. Visual fields, contrast sensitivity, and pattern visual evoked potentials did not show changes. Negative observations included postoperative pain for 2 to 4 days, dry eye symptoms, a period of anisometropia between operations, and hypersensitivity of the lids. CONCLUSIONS: The four ophthalmic residents were satisfied with the outcome of their refractive surgery. Low to moderate myopic correction did not affect the objective measurements of high and low contrast sensitivity, pattern visual evoked potentials, or simulated car driving in dark illumination.

Laser in situ keratomileusis flap margin: wound healing and complications imaged by in vivo confocal microscopy.

PURPOSE: To examine the healing response of laser in situ keratomileusis flap margin in vivo. METHODS: Forty-three eyes of 43 patients who had undergone myopic (n = 39) or hyperopic (n = 4) laser in situ keratomileusis were examined once after surgery. The flap margin was imaged by in vivo confocal microscopy at various depths, and the wound healing response, flap alignment, and complications were evaluated. Ten eyes were examined on day 3 postoperatively, 13 eyes at 1 to 2 weeks, 10 eyes at 1 to 2 months, five eyes at 3 months, and five eyes at 6 months or later. RESULTS: At 3 days after laser in situ keratomileusis, the surface epithelium and basal epithelium appeared normal. Keratocyte activation was strongest at 1 to 2 weeks and 1 to 2 months, and an increased amount of haze was observed correspondingly. Intrastromal epithelial cells forming a plug could occasionally be perceived in the wound gape. Wound constriction was completed in most cases by 3 to 6 months or later. Good alignment was observed in 12 of 43 flaps (27.9%) and moderate and poor alignment in 17 of 43 flaps (39.5%) and 13 of 43 flaps (30.2%), respectively. Poor alignment was not associated with lamellar epithelial ingrowth. Epithelial ingrowth was associated with dense haze at the interface. Diffuse lamellar keratitis was imaged in two corneas after hyperopic laser in situ keratomileusis. CONCLUSIONS: The laser in situ keratomileusis incision wound at the flap margin appears to heal after the sequence observed in incisional wounds in nonhuman primates. Complications, such as lamellar epithelial in growth and diffuse lamellar keratitis, were often observed, particularly after hyperopic laser in situ keratomileusis.

Postoperative inflammation, microbial complications, and wound healing following laser in situ keratomileusis.

Although the biology of corneal wound healing is only partly understood, healing after photorefractive keratectomy (PRK) and laser in situ keratomileusis (LASIK) differs in many respects, and the mechanisms appear to be differently controlled. There is less of an inflammatory and healing response after LASIK, but a longer period of sensory denervation. The cellular, molecular, and neural regulatory phenomena associated with postoperative inflammation and wound healing are likely to be involved in the adverse effects after LASIK, such as flap melt, epithelial ingrowth, and regression. Interface opacities in the early postoperative period include diffuse lamellar keratitis (DLK), microbial keratitis, epithelial cells, and interface opacities. Diffuse lamellar keratitis (sands of the Sahara syndrome) describes an apparently noninfectious diffuse interface inflammation after lamellar corneal surgery probably caused by an allergic or a toxic inflammatory reaction. Noninfectious keratitis must be distinguished from microbial keratitis to avoid aggressive management and treatment with antimicrobial drugs. Microbial keratitis is a serious complication after LASIK, but a good visual outcome can be achieved following prompt and appropriate treatment.

Corneal structure and sensitivity in type 1 diabetes mellitus.

PURPOSE: Corneal wound healing is impaired in diabetic cornea. The purpose of this study was to examine patients with type 1 diabetes mellitus for changes in corneal morphology and to correlate corneal sensitivity, subbasal nerve morphology, and degree of polyneuropathy with each other. METHODS: Forty-four eyes of 23 patients with diabetes and nine control eyes were included. Corneal sensitivity was tested with a Cochet-Bonnet esthesiometer (Luneau, Paris, France), and corneal morphology and epithelial and corneal thickness were determined by in vivo confocal microscopy. The density of subbasal nerves was evaluated by calculating the number of long subbasal nerve fiber bundles per confocal microscopic field. The degree of polyneuropathy was evaluated using the clinical part of the Michigan Neuropathy Screening Instrument (MNSI) classification, and retinopathy was evaluated using fundus photographs. RESULTS: A reduction of long nerve fiber bundles per image was noted to have occurred already in patients with mild to moderate neuropathy, but corneal mechanical sensitivity was reduced only in patients with severe neuropathy. Compared with control subjects the corneal thickness was increased in patients with diabetes without neuropathy. The epithelium of patients with diabetes with severe neuropathy was significantly thinner than that of patients with diabetes without neuropathy. CONCLUSIONS: Confocal microscopy appears to allow early detection of beginning neuropathy, because decreases in nerve fiber bundle counts precede impairment of corneal sensitivity. Apparently, the cornea becomes thicker in a relatively early stage of diabetes but does not further change with the degree of neuropathy. A reduction in neurotrophic stimuli in severe neuropathy may induce a thin epithelium that may lead to recurrent erosions.

The effects of experimental tear film removal on corneal surface regularity and barrier function.

PURPOSE: To evaluate corneal surface regularity and asymmetry, corneal thickness, barrier function, and contrast sensitivity after experimental removal of the precorneal tear layer. DESIGN: Prospective, clinic-based, nonrandomized (self-controlled) comparative trial. PARTICIPANTS: Six eyes of six healthy volunteers (three males, three females; age range, 29-40 years). METHODS: A precorneal tear lesion was created by pressing a sterile Biopore (Millipore, Bedford, MA) Teflon membrane against the central cornea. Corneal topography with both the Topographic Modeling System (TMS-1; Computed Anatomy, Tomey Technology, Cambridge, MA) and the Orbscan (Orbscan Inc., Salt Lake City, UT) were performed before the lesion was created and 30 seconds, 1 hour, and 4 hours after the lesion was created. Surface regularity and surface asymmetry indices were evaluated by the TMS-1 topography system. Maximum and minimum keratometric readings, corneal fluorescein staining, contrast sensitivity, and corneal thickness were evaluated before and after the tear lesion. Cytologic membranes were stained for MUC4 mucin using an indirect immunofluorescent staining technique. Confocal microscopy was performed to evaluate the integrity of the corneal epithelium in two eyes. Analysis of variance with polynomial contrasts was used to examine time trends of the outcome variables. MAIN OUTCOME MEASURES: The change in corneal surface regularity and asymmetry indices, corneal thickness, permeability to fluorescein dye, and contrast sensitivity before and after the lesion was made were compared. RESULTS: The corneal epithelium in the area of the lesion showed intense fluorescein staining 30 seconds postlesion but appeared normal by 4 hours. Confluent, homogeneous staining for MUC4 mucin was observed on the membranes used to create the lesion in all cases. The surface regularity index measured with the TMS-1 increased after the lesion was created and decreased toward normal by 4 hours (P = 0.017). Corneal thickness measured by the Orbscan instrument significantly increased in the central (P = 0.001), superior (P = 0.006), inferotemporal (P < 0.001) and superotemporal (P = 0.001) cornea immediately following the lesion and returned to normal by 4 hours. The lesion caused a decrease in visual acuity at 6.30%, 4% and 2.5% contrast sensitivities 1 hour postlesion and these measurements returned to prelesion values by 4 hours (P = 0.085, P = 0.005, P = 0.043). CONCLUSIONS: The precorneal tear layer serves as a permeability barrier and is essential for maintaining a smooth quality optical surface.

Immunolocalization of transforming growth factor-beta1 and tenascin in human secondary cataract.

PURPOSE: This study was carried out to investigate the distribution of transforming growth factor-beta1 (TGF-beta1) and the extracellular matrix (ECM) glycoprotein tenascin in secondary cataract and anterior subcapsular cataract. METHODS: Twenty-four pseudophakic human eyes with secondary cataract, obtained at autopsy 1 d to 10 yr (mean 2.4 yr) after cataract surgery, were studied. Additionally, a specimen from an anterior subcapsular cataract was included. Immunohistochemistry was used to localize TGF-beta1 and tenascin in secondary cataract and in anterior subcapsular cataract. RESULTS: Polyclonal antibody to TGF-beta1 immunolabelled spindle-shaped cells in the plaques of secondary cataract in all eyes. Instead, the cells present in Soemmering's ring cataract were not labelled. The ECM in the plaques of secondary cataract was immunoreactive for tenascin in all eyes. In anterior subcapsular cataract spindle-shaped cells and ECM showed similar immunoreactivity. CONCLUSION: Spindle-shaped cells that are immunolabelled with TGF-beta1 and ECM showing tenascin-like-immunoreactivity are present in secondary cataract and in anterior subcapsular cataract, thus implicating a possible role in secondary cataract.

Autosomal recessive cornea plana: in vivo corneal morphology and corneal sensitivity.

PURPOSE: Autosomal recessive corneal plana (RCP) is a rare corneal anomaly with unknown pathogenesis and a high incidence in Finland. The aim was to examine corneal sensitivity and the morphology of different corneal layers and subbasal nerves in RCP patients. METHODS: Three patients with a diagnosed autosomal recessive cornea plana were examined. Corneal sensitivity to different modalities of stimulation was tested in four corneas using noncontact esthesiometry. Tissue morphology of three corneas was evaluated, and in two corneas thickness of corneal layers was measured using in vivo confocal microscopy. RESULTS: Corneas of RCP patients appear to have mechanosensory, polymodal, and cold-sensitive nerve terminals. RCP patients had normal sensation thresholds for chemical, heat, and cold stimulation but a high threshold for mechanical stimulation. Their capacity to discriminate increasing intensities of stimulus was reduced, except for cold stimuli. Thickness of the epithelial layer was reduced, whereas total corneal and stromal thicknesses were slightly reduced or close to normal values. In all cases Bowman's layer was absent. Subbasal nerves had abnormal branching patterns. The arrangement of anterior keratocytes was altered, showing clustered and irregularly shaped nuclei. Increased backscattering of light in confocal microscopy through focusing (CMTF) profiles was observed throughout the stroma. Epithelial and endothelial cells appeared to be regular in shape. CONCLUSIONS: The present study revealed qualitative and quantitative alterations in corneal sensitivity, cellular morphology, and the thickness of corneal layers in RCP patients.

Effects of oleoresin capsicum pepper spray on human corneal morphology and sensitivity.

PURPOSE: To examine the potential harmful effects on corneal structure, innervation, and sensitivity of a spray containing the neurotoxin capsaicin (oleoresin capsicum, OC). METHODS: Ten police officers who volunteered for the study were exposed to OC. Clinical signs were assessed. Corneal sensitivity was measured using a Cochet-Bonnet or a noncontact esthesiometer that provides separate measurements of mechanical, chemical, and thermal sensitivity. Tear fluid nerve growth factor (NGF) was measured. Corneal cell layers and subbasal nerves were examined by in vivo confocal microscopy. The subjects were examined before application and 30 minutes, 1 day, 1 week, and 1 month after OC exposure. RESULTS: OC spray produced occasional areas of focal epithelial cell damage that healed within 1 day. Each eye showed conjunctival hyperemia and in two subjects, mild chemosis. All except one eye had unchanged best corrected visual acuity (BCVA). A transient decrease (day 1) of mechanical sensitivity was observed with the Cochet-Bonnet esthesiometer. With the gas esthesiometer, mechanical sensitivity remained below normal values for 7 days. Chemical sensitivity to CO2 was high for as much as 1 day and decreased below normal 1 week later, whereas sensitivity to cold was unaffected. Two subjects had measurable tear NGF that increased after exposure. Basal epithelial cell morphology suggested temporary corneal epithelial swelling, whereas keratocytes, endothelial cells, and subbasal nerves remained unchanged. CONCLUSIONS: Although OC causes immediate changes in mechanical and chemical sensitivity that may persist for a week, a single exposure to OC appears harmless to corneal tissues. The changes are possibly associated with damage of corneal nerve terminals of mainly unmyelinated polymodal nociceptor fibers.

Confocal microscopy of a patient with irregular astigmatism after LASIK reoperations and relaxation incisions.

PURPOSE: Laser-assisted in situ keratomileusis (LASIK) is widely used for correcting refractive errors. If the predicted refractive result is not achieved after the first operation, a re-operation can be performed by ablating more stromal tissue after reopening the flap. The goal of this study was to analyze, by using in vivo confocal microscopy, the morphologic changes associated with repeated LASIKs. METHODS: Clinical examination, computed corneal topography, and real-time in vivo confocal microscopy were performed on both eyes of a 50-year-old patient with induced irregular astigmatism leading to decreased best-corrected vision in the left eye after LASIK. The left cornea had been operated on 5 times (LASIK with two reoperations followed by two relaxing incisions), and the right cornea twice (LASIK with one reoperation). RESULTS: Microfolds at the level of the Bowman's layer and highly reflective particles at the flap interface were observed in both corneas. The subbasal nerve plexus was severed in the left eye. In addition, we identified epithelial material in the flap margin and inside one of the two relaxing incisions placed inferotemporally. CONCLUSION: Repeated LASIKs may stretch the flap and result in microfolding at the Bowman's layer. This and deposition of particles in the flap interface may increase with the number of reoperations, challenging the healing response. Microfolding and occurrence of foreign material in the interface may add to the irregular astigmatism and poor visual outcome after LASIK. Clinical in vivo confocal microscopy offers new possibilities for the assessment of ultrastructural changes after corneal refractive surgery.

In vivo confocal microscopy of patients with corneal recurrent erosion syndrome or epithelial basement membrane dystrophy.

OBJECTIVE: To characterize morphologic changes in corneas of patients with recurrent erosion syndrome or epithelial basement membrane dystrophy using in vivo confocal microscopy. DESIGN: Observational case series PARTICIPANTS: Fourteen eyes of eight patients with diagnosed epithelial basement membrane dystrophy and 13 eyes of seven patients with recurrent erosion syndrome were examined. METHODS: Slit-lamp examination and in vivo confocal microscopy. The pathologic findings are presented as digitized images obtained from video tape recorded during the confocal microscopy. MAIN OUTCOME MEASURES: The morphology of corneal surface epithelial cells, basal epithelial cells, subbasal nerve plexus, Bowman's layer, stromal keratocytes, and endothelium was analyzed. RESULTS: The surface epithelium was intact in all but two eyes. One cornea (a basement membrane disorder with clinically visible dots) had multinucleate surface epithelial cells, and one eye with recurrent corneal erosions showed a freely floating surface epithelium sheet in the tear fluid. Patients in both groups showed islets of highly reflective cells with presumed intracellular deposits surrounded by normal cells in the basal epithelial cell layer. The basal epithelial cell area also showed other pathologic changes, including drop-shaped configurations, streaks, or ridges. Folding of the Bowman's layer was also observed in both groups. Anterior keratocytes showed signs of activation (highly reflective nuclei with visible processes) in some of the patients regardless of the clinical diagnosis, and in recurrent erosions even increased deposition of abnormal extracellular matrix in the anterior stroma was suspected. Posterior corneal keratocytes and endothelium appeared normal when examined. The subbasal nerve plexus showed various pathologic changes, such as short or strangely shaped nerve fiber bundles, decreased numbers of long nerve fiber bundles, only faintly visible long nerve fiber bundles (instead of the normally observed long parallel running interconnected bundles), or increased amounts of Langerhans cells, but only one patient (with recurrent erosion syndrome) lacked the subbasal nerve plexus. CONCLUSIONS: In vivo confocal microscopy of corneas with recurrent erosions or epithelial basement membrane dystrophy showed deposits in basal epithelial cells, subbasal microfolds and streaks, damaged subbasal nerves, or altered morphology of the anterior stroma. Confocal microscopy cannot replace biomicroscopy in making a specific diagnosis, but it sometimes helps the diagnosis in corneas that appear normal under a biomicroscope.

Corneal stromal changes induced by myopic LASIK.

UNLABELLED: PURPOSE. Despite the rapidly growing popularity of laser in situ keratomileusis (LASIK) in correction of myopia, the tissue responses have not been thoroughly investigated. The aim was to characterize morphologic changes induced by myopic LASIK in human corneal stroma. METHODS: Sixty-two myopic eyes were examined once at 3 days to 2 years after LASIK using in vivo confocal microscopy for measurement of flap thickness, keratocyte response zones, and objective grading of haze. RESULTS: Confocal microscopy revealed corneal flap interface particles in 100% of eyes and microfolds at the Bowman's layer in 96.8%. The flaps were thinner (112 +/- 25 microm) than intended (160 microm). The keratocyte activation in the stromal bed was greatest on the third postoperative day. Patients with increased interface reflectivity due to abnormal extracellular matrix or activated keratocytes at > or = 1 month (n = 9) had significantly thinner flaps than patients with normal interface reflectivity (n = 18; 114 +/- 12 versus 132 +/- 22 microm, P = 0.027). After 6 months the mean density of the most anterior layer of flap keratocytes was decreased. CONCLUSIONS: Keratocyte activation induced by LASIK was of short duration compared with that reported after photorefractive keratectomy. The flaps were thinner than expected, and microfolds and interface particles were common complications. The new findings such as increased interface reflectivity associated with thin flaps and the apparent loss of keratocytes in the most anterior flap 6 months to 2 years after surgery may have important clinical relevance.

Effect of myopic LASIK on corneal sensitivity and morphology of subbasal nerves.

PURPOSE: To investigate whether the morphology of the subbasal nerves corresponds to corneal sensitivity after laser in situ keratomileusis (LASIK). METHODS: In a case series study, 59 patients were examined at 2 to 4 hours, 3 days, 1 to 2 weeks, 1 to 2 months, 3 months, or 6 or more months after undergoing LASIK for myopia, by using a Cochet-Bonnet esthesiometer and an in vivo confocal microscope, and were compared with control subjects. Corneal sensitivity and confocal images of subbasal nerves were obtained centrally and 2 mm nasally and temporally. Subbasal nerve fiber bundles (NFBs) were grouped as follows: corneas with no nerve images; corneas with short (<200 microm), unconnected NFBs; corneas with long (> or =200 microm) NFBs without interconnections; and corneas with long NFBs with interconnections. RESULTS: Corneal sensitivity was at its lowest at 1 to 2 weeks after LASIK. Sensitivity of the hinge area was higher than temporal or central areas at every time point. At 6 or more months the sensitivity values were comparable with the values observed in control subjects. The central area showed mainly short, unconnected subbasal NFBs, even at 6 months. In general, the temporal area presented with long NFBs from 3 months onward, whereas the nasal area showed long NFBs at every time point. CONCLUSIONS: The results suggest that the corneal areas with no nerve images or short, unconnected NFBs are associated with lower sensitivities than corneal areas with long NFBs with or without interconnections. In vivo confocal microscopy reveals LASIK-induced alterations of subbasal nerve morphology and thus enables a direct comparison of corneal sensory innervation and sensitivity.