Subbasal Nerve Density Research Articles

In Vivo Evaluation of Corneal Nerves and Epithelial Healing After Treatment With Recombinant Nerve Growth Factor for Neurotrophic Keratopathy

To evaluate the renewal of corneal nerve structure and function in patients with neurotrophic keratopathy (NK) treated with recombinant human nerve growth factor (rhNGF) eye drops. Prospective, interventional, before-and-after case series. This study included 18 patients with NK with a persistent epithelial defect or corneal ulcer, treated with topical rhNGF, and age-matched healthy controls. Patients underwent clinical examination with corneal fluorescein staining, Schirmer 1 tear test, assessment of corneal sensitivity with the Cochet-Bonnet esthesiometer, and morphologic examination of the nerves by invivo confocal microscopy (IVCM) at baseline and at 4 and 8weeks of treatment. IVCM analysis was used to assess corneal sub-basal nerve density, number of nerve branches, and the diameter of nerve fibers. A complete resolution of the epithelial defect was observed in all patients within 8weeks. Schirmer 1 test showed a significant improvement of tear film secretion. Change from baseline in corneal sensation was significant (P < .001) but did not approach that of healthy controls. After 8weeks of treatment, there was a significant increase in the mean nerve density in affected eyes as compared to baseline (P= .007) as well as in the number of nerve branches (P= .008) and nerve fiber diameter (P= .007). Topical treatment with rhNGF was effective in promoting complete corneal healing of persistent epithelial defects and corneal ulcers in patients with NK. This was associated with an improvement of corneal sensitivity and an increase of sub-basal nerve density, diameter, and number of nerve branches, indicating improvement in structure and function of corneal nerves.

Sensory neurons directly promote angiogenesis in response to inflammation via substance P signaling.

Blood vessels and nerves travel together to supply most tissues in the body. However, there is a knowledge gap in the mechanisms underlying the direct regulation of angiogenesis by nerves. In the current study, we examined the regulation of angiogenesis by sensory nerves in response to inflammation using the cornea, a normally avascular and densely innervated ocular tissue, as a model. We used desiccating stress as an inflammatory stimulus in vivo and found that sub-basal and epithelial nerve densities in the cornea were reduced in dry eye disease (DED). We established a co-culture system of trigeminal ganglion sensory neurons and vascular endothelial cells (VEC) and found that neurons isolated from mice with DED directly promoted VEC proliferation and tube formation compared with normal controls. In addition, these neurons expressed and secreted higher levels of substance P (SP), a proinflammatory neuropeptide. SP potently promoted VEC activation in vitro and blockade of SP signaling with spantide I, an antagonist of SP receptor Neurokinin-1, significantly reduced corneal neovascularization in vivo. Spantide I and siRNA knockdown of SP abolished the promotion of VEC activation by DED neurons in vitro. Taken together, our data suggested that sensory neurons directly promote angiogenesis via SP signaling in response to inflammation in the cornea.

Changes in Corneal Dendritic Cell and Sub-basal Nerve in Long-Term Contact Lens Wearers With Dry Eye.

To evaluate the changes of corneal sub-basal nerve (SBN) and dendritic cell (DC) in contact lens (CL) wearers with mild dry eye and their potential relationship. Twenty mild dry eye volunteers who had never worn CLs were recruited for long-term CL wearing. Each subject underwent ocular surface evaluations at baseline and at 1, 4, 12, and 24 weeks, including Ocular Surface Disease Index (OSDI) questionnaire, tear film break-up time (TBUT), and Schirmer I test. In vivo confocal microscopy was used to examine the density, area, number of dendrites, total dendritic length of DC, and SBN densities in central and peripheral corneas. Only right eyes were included. The DCs were activated and peaked at week 4 after wearing CLs. The peripheral DC density increased beginning the first week, whereas the central ones increased by week 4. After 4 weeks, both began to decrease, but still higher than baseline at week 24. The central and peripheral SBN densities decreased. However, the peripheral SBN tended to increase beginning at week 12. In early period, SBN was negatively correlated with DC parameters. After 4 weeks, the correlation changed to be positive. The OSDI increased, whereas the Schirmer I test and TBUT showed no significant change. After wearing CLs, corneal DC were activated and increased, indicating ocular surface inflammation and decreased after week 4. In the early period, increases in DC may lead to decreases in SBN. Upon decrease of DC, the SBN may regenerate.

Corneal Reinnervation and Sensitivity Recovery after Pterygium Excision.

Purpose To evaluate changes in corneal sensitivity and subbasal nerve density after pterygium excision. Methods This prospective trial included 22 eyes with nasal primary pterygium and 18 controls. Corneal sensitivity was evaluated using a Cochet–Bonnet esthesiometer in the nasal, superior, temporal, inferior, and center quadrants of the cornea before surgery and 10 days, 1 month, and 3months after surgery. The central cornea was analyzed using in vivo confocal microscopy (IVCM) before surgery and 1 and 3 months after surgery. Subbasal nerve density and other nerve parameters were analyzed using NeuronJ. Nerve tortuosity was evaluated and graded in individual IVCM scans. The tear film break-up time (TBUT) test and Schirmer's test were performed before surgery, as well as 1 and 3 months after surgery. All the same tests were performed in the controls. Results All affected eyes showed a significant increase in corneal sensitivity in the nasal corneal quadrant after surgery when compared with preoperative data (F = 37.3; P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (F = 37.3; P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (F = 37.3; P < 0.01). Compared with controls, pterygium patients demonstrated decreased corneal subbasal nerve density (Conclusion Pterygium patients demonstrated deteriorated corneal subbasal nerve fibers when compared with healthy controls in terms of nerve length, nerve trunks, and nerve branches. Therefore, pterygium excision improves corneal sensitivity and increases corneal subbasal nerve density.

Changes in corneal innervation and pain responses in fungal keratitis

To characterize changes in the cornea nerve and pain responses in fungal keratitis (FK). A retrospective analysis of in vivo confocal microscopy images of 11 FK corneas was performed, and the results were compared with those for 11 normal corneas. Subbasal corneal nerves were analyzed for total nerve number, main nerve trunk number, branching patterns and tortuosity. C57BL/6 mice were infected with Aspergillus fumigatus. Disease severity was determined through clinical scoring and slit lamp photography. Corneas were harvested at 1, 3, 5, and 7d post infection (p.i.) and assessed for β III tubulin. Corneal mechanical sensitivity thresholds were detected by von Frey test. β-endorphin (β-EP) and µ receptor protein expression was detected through Western blotting. Total nerve number, main nerve trunk number, and nerve branching were significantly lower in FK patients than in controls, but tortuosity was not significantly different. In infected mice, subbasal nerve density decreased from 1d p.i., reaching a minimum at 5d p.i. Clinical scores rose at 1d p.i., peaked at 3d p.i., and decreased at 5d p.i. Mechanical sensitivity thresholds showed the same trends. β-EP and µ receptor protein expression increased after infection. Corneal nerve density is lower in FK patients and Aspergillus fumigatus-infected mice than in controls. Pain sensitivity decreases with postinfection corneal ulcer aggravation. β-EP and µ receptor proteins are both upregulated in infected mouse corneas.

Pain and sensation in dry eye disease

AbstractThe recent definition of dry eye disease (DED) proposed by DEWS II includes, for the first time, neurosensory abnormalities in etiology of the disease. In DED, reduced tear secretion leads to inflammation and corneal nerve damage, which induce sensitization of corneal polymodal and mechano‐nociceptors, and an abnormal activity of cold thermoreceptors thus evoking pain and eye dryness sensations. Moreover, long‐term nerve damage and inflammation, modify the expression of ion channels involved in the stimulus detection (transduction) and excitability of ocular sensory neurons, changing their impulse firing and their connectivity at the CNS, which is the basis of neuropathic pain and dysesthesia referred to the eye surface in DED patients. These altered transducing and encoding ion channels are potential molecular targets to modulate neuropathic nerve activity in DED, currently under study. Although many subjective (surveys and questionnaires, corneal sensitivity measurement) and objective metrics (morphology and density of subbasal corneal nerves and immune cells, blink parameters, biomarkers in tears) are currently used to evaluate pain in DED patients, new instruments and procedures designed to obtain more reliable information on eye sensations and pain in DED need to be developed (SAF‐2017‐83674‐C2‐1‐R and 2‐R, Agencia Estatal de Investigación‐ERDF, Spain‐EU, and PROMETEO/2018/114, Generalitat Valenciana, Spain).

Early phenotypic features of aniridia-associated keratopathy and association with PAX6 coding mutations

PurposeTo investigate corneal phenotype in aniridia-associated keratopathy (AAK) including its earliest manifestations, in relation to PAX6 mutational status. Methods46 subjects (92 eyes) with congenital aniridia from a German registry were examined using slit lamp biomicroscopy, anterior segment optical coherence tomography, contact esthesiometry and in vivo confocal microscopy. Cytogenetic analysis was conducted by Sanger sequencing of PAX6 exons and/or MLPA analysis. Measured parameters included AAK grade, distance-corrected visual acuity (DCVA), central corneal thickness (CCT), corneal sensitivity, subbasal nerve density, mature dendritic cell (DC) density and corneal epithelial phenotype. Results46 subjects (age range: 1–64 years) were examined, including 23 (50%) children under the age of 18. Five subjects (11.1%) with absent PAX6 coding mutation (non-PAX6 cases) had mild AAK (Grade 0–1) into the fourth decade of life and maintained corneal epithelial phenotype, greater subbasal nerve density (16.8 mm/mm2 vs. 3.58 mm/mm2, P = 0.01) and better corneal sensitivity (41 ± 11 mm vs. 28 ± 12 mm, P = 0.03) relative to those with PAX6 coding mutations. In five subjects, corneal endothelial cell density ranged from 3245 to 4399 cells/mm2. Independent of mutational status, an increased CCT, over tenfold increased mature DC density and reduced corneal sensitivity characterized all subjects. ConclusionsPAX6 coding mutations influence AAK phenotype and progression from the earliest stages of life. A minimal keratopathy present in 100% of congenital aniridia cases is independent of the specific mutation and consists of increased corneal thickness, reduced touch sensitivity, and increased ocular surface immune activity.

Clinical and in vivo confocal microscopic features of neuropathic corneal pain

AimsTo describe clinical and in vivo confocal microscopy (IVCM) features of neuropathic corneal pain (NCP) without clinically visible signs.MethodsProspective, observational study of 27 eyes of 14 patients who had continuous...

Degeneration of Corneal Sensation and Innervation in Patients with Facial Paralysis: A Cross-Sectional Study Using in Vivo Confocal Microscopy

ABSTRACTPurpose: Facial paralysis (FP) leads to diverse periocular complications which threats visual acuity and affects corneal nerve functionally and morphologically. This study aims to summarize the clinical ophthalmic outcomes, corneal sensation, and morphological alterations of subbasal nerve and dendritic cells (DCs) in patients with facial paralysis.Methods: We performed a cross-sectional study of 48 consecutive patients with facial paralysis at one tertiary hospital. Forty-eight healthy participants were enrolled as controls. The images of corneal nerves and epithelial DCs were detected by in vivo confocal microscopy (IVCM). Each patient received thorough ophthalmic examination, tear film function tests, corneal fluorescence staining and Cochet-Bonnet esthesiometry test. Clinical and morphologic data were compared with controls.Results: Forty patients (83.3%) showed corneal injuries from punctate epithelial defects to corneal ulcers and scars. Visual impairment and eyelid malposition were observed. Corneal sensitivity remarkably decreased (25.1 ± 23.8 mm) in the affected eyes and was correlated to diminished subbasal nerve density (P = .019, r = 0.387). Numbers of corneal main nerve trunks and branches were significantly reduced (P < .0001) while DCs were increased (P < .0001) in patients with FP when compared with controls. Nerve fiber density showed inverse association with DC density (P = .019, r = −0.389).Conclusions: Ocular complications including corneal erosions, loss of corneal sensation, visual impairment and eyelid malposition have largely affected patients with facial paralysis. Morphological changes of diminished corneal subbasal nerve and increased DCs were detected by IVCM. Corneal epithelial defect, corneal opacity, corneal sensation, dendritic cell density are factors associated with corneal subbasal nerve density. Patients with FP are suggested to have complete ophthalmic evaluation and early instruction on ocular prevention.

Sensory Adaptation to Silicone Hydrogel Contact Lens Wear Is Not Associated With Alterations in the Corneal Subbasal Nerve Plexus.

To evaluate corneal subbasal nerve alterations in contact lens (CL)-naive silicone hydrogel CL wearers and to investigate the relationship between structural subbasal nerve changes with corneal sensitivity. Twenty eyes of 20 neophyte daily silicone hydrogel CL wearers and 20 eyes of age-matched control subjects were recruited for this prospective longitudinal study. Corneal subbasal nerve densities were evaluated using in vivo confocal microscopy. Central corneal tactile sensitivity was measured using Cochet-Bonnet esthesiometers. In vivo confocal microscopy and Cochet-Bonnet esthesiometry were performed before and at the 6-month time point after the initiation of CL wear. Two-way repeated measures analysis of variance, χ, and Wilcoxon signed-rank tests were used for statistical analyses. The mean ages of CL users and control subjects were 21.8 ± 1.8 years (range = 19-24 years) and 21.5 ± 2.3 years (range = 20-24 years), respectively (P = 0.579). Compared with their baseline values, there were no significant changes at the 6-month follow-up period in the mean total subbasal nerve fiber length (2266.6 ± 414.6 vs. 2277.9 ± 405.0 μm/frame; P = 0.432), mean total subbasal nerve branch density (10.4 ± 1.5 vs. 10.5 ± 1.3 nerves/frame; P = 0.655), or the mean long nerve fiber density (4.7 ± 0.7 vs. 4.8 ± 0.7 nerves/frame; P = 0.564) of CL users. Mechanical corneal sensitivity remained unaltered during the 6-month period in CL users (11.2 ± 0.5 vs. 11.2 ± 0.5; P = 1.000). No significant changes were observed in the subbasal nerve plexus or corneal tactile sensitivity of the control subjects during the study interval. Sensory adaptation to CL wear is not mediated through attenuation of the subbasal nerve or reduction of corneal tactile sensitivity in CL-naive users.

Clinical and microstructural changes with different iontophoresis-assisted corneal cross-linking methods for keratoconus.

To compare the clinical and microstructural changes induced by different transepithelial iontophoresis-assisted corneal cross-linking (I-CXL) methods for keratoconus. A total of 42 eyes of 42 patients with progressive keratoconus were divided into two groups. Group A received I-CXL for 5min, while group B received I-CXL for 10min. Visual acuity, optical coherence tomography (OCT), specular microscopy and confocal microscopy were evaluated preoperatively and at 1, 3, 6, and 12mo postoperatively. Twelve months after the operation, uncorrected visual acuity (UCVA) and corrected distance visual acuity (CDVA) were improved in both groups, with a better outcome in the I-CXL 10min group (P=0.025, 0.021, respectively). Kmax values decreased by 0.94±3.00 D in the I-CXL 10min group (P=0.033) but increased by 1.87±3.29 D in the I-CXL 5min group (P=0.012). OCT scans showed that the demarcation line was most visible and substantially deeper in the I-CXL 10min group. Confocal microscopy showed greater anterior stromal keratocyte decreases in the I-CXL 10min group than in the I-CXL 5min group at 3 and 6mo postoperatively (P<0.001); however, anterior stromal keratocytes and subbasal nerve density were not significantly different between the two groups at 12mo postoperatively. I-CXL for 10min more effectively halts the progression of keratoconus than I-CXL for 5min after 12mo of follow-up. However, long-term studies are needed to evaluate the efficacy and safety of I-CXL.

Mapping the entire nerve architecture of the cat cornea.

To provide a complete nerve architecture and neuropeptide distribution in the cat cornea. Two adult domestic cats. The cat corneas were stained with protein gene product (PGP) 9.5 antibody-a pan marker for nerve fibers-and then divided into four quarters and double labeled with calcitonin gene-related peptide (CGRP) or substance P (SP) antibodies. Relative corneal nerve fiber densities and nerve terminals were evaluated in whole mount images by computer-assisted analysis. An average of 21.5±2.1 thick stromal nerves enters the cornea around the limbus where they split into many branches going up to the anterior stroma. Some branches link to each other, but most of them penetrate the basement membrane in the periphery to give origin to subbasal bundles, which run centripetally and merge to form a whirl-like structure (vortex) at the center. These nerve bundles send out many fine terminals that innervate the epithelial cells. Subbasal nerve density and nerve terminals were greater in the center than in the periphery of the cornea. Additionally, CGRP-positive central epithelial nerve fibers and terminals were more abundant than SP-positive nerves and terminals. The architecture of cat corneal nerves shows similarities to human and mouse cornea innervation. This study provides useful data for researchers who use the cat model to assess corneal nerve pathological alterations, as well as in the veterinary field where corneal opacities, ulcerations, and infections damage the nerves and decrease sensitivity.

Long-term evaluation of corneal sub-basal nerve recovery after photorefractive keratectomy and influence of pars plana vitrectomy.

The corneal sub-basal nerve (SBN) plexus is destroyed during photorefractive keratectomy (PRK) and its recovery is still a matter of debate. In vivo confocal microscopy (IVCM) was used to evaluate SBN plexus in 23 patients at a distance of 10-25 years (mean 15.6 years) from myopic PRK. Because 8 out of the 23 PRK patients underwent pars plana vitrectomy (PPV) for rhegmatogenous retinal detachment, IVCM was also performed on those patients 6 months after PPV. Thirteen patients matched for age and myopia served as controls (non-PRK). SBN plexus was markedly reduced after PRK compared with non-PRK eyes and showed a slow, continuous but incomplete recovery up to the end of our follow-up (range 10-25 years). PRK and non-PRK eyes showed a marked reduction in SBN density 6 months after PPV, thus demonstrating a detrimental effect exerted by PPV on SBN plexus.

Corneal Subbasal Nerve Density and Sensitivity After Pars Plana Vitrectomy Using Contact or Noncontact Wide-Angle Viewing Systems.

To assess corneal subbasal nerve density (SBND) and corneal sensitivity (CS) after pars plana vitrectomy (PPV) for rhegmatogenous retinal detachment (RRD) without intraoperative 360-degree laser treatment using contact or noncontact panoramic viewing systems. This is a prospective study of 34 eyes affected by RRD, which underwent PPV using contact (18 eyes) and noncontact (16 eyes) panoramic viewing systems; 12 eyes which underwent scleral buckling, and 17 eyes which underwent cataract surgery. SBND and CS were assessed before surgery and 3 and 6 months postoperatively by in vivo confocal microscopy and Cochet-Bonnet esthesiometry. Compared with baseline values, at 6 months, SBND and CS decreased in both contact PPV (SBND preoperative value: 19.1 ± 3.7 mm/mm; SBND postoperative value: 3.5 ± 1.3 mm/mm; CS preoperative value: 5.1 ± 0.5 cm; CS postoperative value: 1.5 ± 0.4 cm) and in noncontact PPV groups (SBND preoperative value: 19.5 ± 3.8 mm/mm; SBND postoperative value: 8.7 ± 2.3 mm/mm; CS preoperative value: 5.3 ± 0.5 cm; CS postoperative value: 2.5 ± 0.7 cm) (P < 0.001 for all comparisons). SBND reduction was greater in the contact PPV group than in the noncontact PPV group (P < 0.001). By contrast, the scleral buckling and cataract surgery group values were unchanged (P > 0.1 for all comparisons). In multivariate analysis, no significant effect was found for cataract surgery associated with PPV, pseudophakia, surgical time, intraocular pressure, or for laser in horizontal sectors for PPV groups. SBND was highly correlated with CS (r = 0.93). A contact viewing system reduces SBND after PPV more than a noncontact system does.

High fat diet induces pre-type 2 diabetes with regional changes in corneal sensory nerves and altered P2X7 expression and localization

Type 2 diabetes is one of the leading pathologies that increases the risk of improper wound healing. Obesity has become a major risk factor for this disease that is now considered to be the 4th highest cause of preventable blindness according to the World Health Organization. The cornea is the most densely innervated structure in the human body and senses even the slightest injury. In diabetes, decreased corneal sensitivity secondary to diabetic peripheral neuropathy can lead to increased corneal abrasion, ulceration, and even blindness. In this study, a diet induced obesity (DIO) mouse model of pre-Type 2 diabetes was used to characterize changes in sensory nerves and P2X7, a purinoreceptor, a pain receptor, and an ion channel that is expressed in a number of tissues. Since our previous studies demonstrated that P2X7 mRNA was significantly elevated in diabetic human corneas, we examined P2X7 expression and localization in the DIO murine model at various times after being fed a high fat diet. Fifteen weeks after onset of diet, we found that there was a significant decrease in the density of sub-basal nerves in the DIO mice that was associated with an increase in tortuosity and a decrease in diameter. In addition, P2X7 mRNA expression was significantly greater in the corneal epithelium of DIO mice, and the increase in transcript was enhanced in the central migrating and peripheral regions after injury. Interestingly, confocal microscopy and thresholding analysis revealed that there was a significant increase in P2X7 distal to the injury, which contrasted with a decrease in P2X7-expressing stromal sensory nerves. Therefore, we hypothesize that the P2X7 receptor acts to sense changes at the leading edge following an epithelial abrasion, and this fine-tuned regulation is lost during the onset of diabetes. Further understanding of the corneal changes that occur in diabetes can help us better monitor progression of diabetic complications, as well as develop new therapeutics for the treatment of diabetic corneal dysfunction.

Comparison of standard and accelerated corneal cross-linking for the treatment of keratoconus: a meta-analysis.

To compare results between standard and accelerated corneal collagen cross-linking (CXL) for the treatment of progressive keratoconus. We performed literature searches in PubMed, Cochrane Library, Web of Science, ISRCTN registry, ClinicalTrials.gov, and EMBASE for studies comparing conventional Dresden (C-CXL) and accelerated CXL (A-CXL). Outcomes were clinical results and changes in corneal properties. Weighted mean differences were used to evaluate the effects. Here, 22studies with 1158 eyes (C-CXL: 577eyes; A-CXL: 581eyes) were included. At the last follow-up, C-CXL was superior regarding minimum keratometry (p<0.00001) and demarcation line depth (p<0.00001), whereas A-CXL should be favoured when considering minimum corneal thickness (p=0.0005). No differences in uncorrected and corrected distance visual acuity (p=0.09 and 0.98), spherical equivalent (p=0.11), spherical and cylindrical error (p=0.29 and 0.32), maximal and average keratometry (p=0.05 and 0.65), central corneal thickness (p=0.15), corneal biomechanical properties (p≥0.21 respectively), time of reepithelialization (p=0.76), subbasal nerve density (p=0.69), endothelial cell density (p=0.30) and morphology (p≥0.40 respectively) were found among both groups. Consideration of less corneal thinning favours A-CXL, whereas the deeper demarcation line and greater changes in minimum keratometric values in C-CXL may indicate a higher treatment efficacy. Altogether, C-CXL, as well as A-CXL, provides successful results in the strengthening of corneal tissue.

Functional and Morphologic Alterations in Mechanical, Polymodal, and Cold Sensory Nerve Fibers of the Cornea Following Photorefractive Keratectomy.

To define the characteristics and time course of the morphologic and functional changes experienced by corneal sensory nerves after photorefractive keratectomy (PRK). Unilateral corneal excimer laser photoablation was performed in 54 anesthetized 3- to 6-month-old mice; 11 naïve animals served as control. Mice were killed 0, 3, 7, 15, and 30 days after PRK. Excised eyes were placed in a recording chamber superfused at 34°C. Electrical nerve impulse activity of single sensory terminals was recorded with a micropipette applied onto the corneal surface. Spontaneous and stimulus-evoked (cold, heat, mechanical, and chemical stimuli) nerve terminal impulse (NTI) activity was analyzed. Corneas were fixed and stained with anti-β-Tubulin III antibody to measure nerve density and number of epithelial nerve penetration points of regenerating subbasal leashes. Nerve fibers and NTI activity were absent in the injured area between 0 and 7 days after PRK, when sparse regenerating nerve sprouts appear. On day 15, subbasal nerve density reached half the control value and abnormally responding cold-sensitive terminals were recorded inside the lesion. Thirty days after PRK, nerve density was almost restored, active cold thermoreceptors were abundant, and polymodal nociceptor activity first reappeared. Morphologic regeneration of subbasal corneal nerves started shortly after PRK ablation and was substantially completed 30 days later. Functional recovery appears faster in cold terminals than polymodal terminals, possibly reflecting an incomplete damage of the more extensively branched cold-sensitive axon terminals. Evolution of postsurgical discomfort sensations quality may be associated with the variable regeneration pattern of each fiber type.

Proinflammatory Markers, Chemokines, and Enkephalin in Patients Suffering from Dry Eye Disease.

Dry eye symptoms are among the leading complaints in ophthalmology. Dry eye disease (DED) is associated with significant pain affecting quality of life. Cellular and molecular mechanisms underlying ocular pain associated with DED are not fully understood. In this study, we investigated the ocular surface of patients with DED using in vivo confocal microscopy (IVCM) to quantify corneal nerve density and its relation with corneal inflammation. Gene expression of the proinflammatory markers HLA-DR, IL-6, CXCL12, and CCL2 and the receptors CXCR4 and CCR2, as well as PENK (enkephalin precursor), was therefore quantified in conjunctival impression cytology specimens. Thirty-two patients with DED and 15 age-matched controls were included. Subbasal nerve density was significantly lower in DED patients compared to controls. IVCM analysis revealed that DED patients had a significantly higher corneal dendritic cell density compared to controls. Conjunctival impression cytology analysis revealed that HLA-DR, IL-6, CXCR4, and CCL2/CCR2 mRNA levels were significantly increased in DED patients compared to controls, whereas PENK mRNA levels were significantly decreased. Similar results were obtained in vitro on immortalized human conjunctiva-derived epithelial cells challenged with osmotic stress that mimics the DED condition. These results demonstrate that proinflammatory molecules and endogenous enkephalin have opposite gene regulation during DED.

Alterations of Murine Subbasal Corneal Nerves After Environmental Dry Eye Stress.

To investigate the morphologic changes in the corneal subbasal nerve (CSN) plexus in wild-type mice after exposure to environmental dry eye stress (EDES) using in vivo confocal microscopy (IVCM). We examined 22 eyes of 8-week-old wild-type male mice (Balb/c, n = 11). The mice were exposed to an air fan inside a small compartment 5 hours/day for 3 days (EDES). Aqueous tear secretion and corneal epithelial damage were assessed. The CSNs were investigated by laser-scanning IVCM. Density; tortuosity; and reflectivity of CSNs; and dendritic cell (DC) densities were evaluated using semi-automated NeuronJ software. EDES significantly decreased the aqueous tear secretion quantity (P = 0.0019) and significantly increased the corneal fluorescein (P = 0.005) and lissamine green staining scores (P = 0003). The CSN density showed a significant decrease after EDES exposure (before, 2813 ± 762 pixels/frame; after, 1906 ± 896 pixels/frame, P = 0.0071). The tortuosity and the reflectivity grades did not show statistically significant differences after EDES exposure (tortuosity, P = 0.307; reflectivity, P = 0.758). However, the mean DC density showed a significant increase after EDES exposure (before, 12.62 ± 5.94 cells/mm2; after, 15.93 ± 5.30 cells/mm2, P = 0.026). Even short-term exposure to EDES induced alterations in the CSN plexus morphology including decreased subbasal corneal nerve density and increased amount of DCs in mice. The EDES mouse model is a promising model to study the ocular surface and corneal nerve changes associated with dry eye disease.

Corneal nerve regeneration after herpes simplex keratitis: A longitudinal in vivo confocal microscopy study

PurposeTo evaluate the long-term alterations of corneal nerves in patients with herpes simplex virus (HSV) keratitis using in vivo confocal microscopy (IVCM). DesignProspective, longitudinal, cross sectional. MethodsThis study included 16 patients with a history of HSV keratitis and 15 age-matched normal controls. Slit-scanning IVCM was performed in all subjects at baseline and then after a mean follow-up of 37.3 ± 1.7 months in the patient group. Corneal subbasal nerve density and corneal sensation were compared between groups at baseline and follow-up. ResultsAt baseline, the mean subbasal nerve density was significantly lower in both affected eyes (1.4 ± 0.6 mm/mm2) and contralateral unaffected eyes (6.4 ± 0.7 mm/mm2) compared with the controls (14.1 ± 1.6 mm/mm2; all P < .001). At the end of follow-up, the mean nerve density in affected eyes increased to 2.8 ± 0.7 mm/mm2 (P = .006), with no significant change in contralateral unaffected eyes (6.5 ± 1.0 mm/mm2, P = .72). However, both eyes had lower nerve density than controls (all P < .001). Corneal sensation was significantly lower in affected eyes (2.6 ± 0.6 cm) than in the control group (6.0 ± 0.0, P < .001) and showed no significant change at the end of follow-up (2.5 ± 0.6 cm, P = .80). Corneal sensation in contralateral unaffected eyes was not different in comparison with controls at both baseline and follow up (all p > .05). ConclusionsOur results demonstrate that although corneal nerve regeneration occurs in patients with HSV keratitis, this change is not clinically significant and does not results in changes of corneal sensation. Therefore, these patients need to be followed closely for complications of neurotrophic keratopathy and might benefit from neuro-regenerative therapies.