Corneal dendritic cells in diabetes mellitus: A narrative review.

Diabetes mellitus is a global public health problem with both macrovascular and microvascular complications, such as diabetic corneal neuropathy (DCN). Using in-vivo confocal microscopy, corneal nerve changes in DCN patients can be examined. Additionally, changes in the morphology and quantity of corneal dendritic cells (DCs) in diabetic corneas have also been observed. DCs are bone marrow-derived antigen-presenting cells that serve both immunological and non-immunological roles in human corneas. However, the role and pathogenesis of corneal DC in diabetic corneas have not been well understood. In this article, we provide a comprehensive review of both animal and clinical studies that report changes in DCs, including the DC density, maturation stages, as well as relationships between the corneal DCs, corneal nerves, and corneal epithelium, in diabetic corneas. We have also discussed the associations between the changes in corneal DCs and various clinical or imaging parameters, including age, corneal nerve status, and blood metabolic parameters. Such information would provide valuable insight into the development of diagnostic, preventive, and therapeutic strategies for DM-associated ocular surface complications.

The effect of high-fat diet-induced metabolic disturbance on corneal neuroimmune features.

PURPOSE: The highly innervated cornea is susceptible to nerve loss secondary to systemic diseases such as diabetes and metabolic disturbances caused by high-fat diet. In this study, we characterize the effect of high-fat diet on the mouse corneal neuroimmune phenotype, including changes to corneal nerve density and resident immune cells, alongside the clinical assessment of corneal thickness and endothelial cell density. METHODS: Male C57Bl6/J mice, aged 10 weeks, were fed a high-fat diet (60 kcal% fat, 5.2 kcal/g) or control diet (10 kcal%, 3.8 kcal/g) for 16 weeks. At the study endpoint, metabolic parameters (HbA1c, weight, fasting glucose, body fat) were measured to confirm metabolic disturbance. Clinical imaging of the anterior segment was performed using optical coherence tomography to measure the corneal epithelial and stromal thickness. Corneal sensory nerves were visualized using flatmount immunostaining and confocal microscopy. The topographical distribution and density of sensory nerves (BIII-tubulin+), intraepithelial CD45+ and MHC- II+ cells, stromal macrophages (IBA1+CD206+) and endothelial cells (ZO-1+) were analysed using FIJI. RESULTS: High-fat diet mice had significantly higher blood HbA1c, higher body weight, a higher percentage of body fat and elevated fasting glucose compared to the control diet mice. Corneal epithelial and stromal thickness was similar in both groups. The sum length of the basal nerve plexus was lower in the central and peripheral cornea of mice fed a high-fat diet. In contrast, the sum length of superficial nerve terminals was similar between groups. Epithelial immune cell density was two-fold higher in the central corneas of high-fat diet mice compared to control diet mice. IBA1+CD206+ macrophage density was similar in the anterior stroma of both groups but was significantly higher in the posterior stroma of the peripheral cornea in the high-fat diet mice compared to controls. The percentage of nerve-associated MHC-II+ cells in the epithelium and stroma was higher in HFD mice compared to controls. Endothelial cell density was similar in the corneas of high-fat diet mice compared to controls. CONCLUSION: Together with corneal neuropathy, corneal immune cells in mice fed a high-fat diet were differentially affected depending on their topographical distribution and location within cornea, and appeared in closer proximity to epithelial and stromal nerves, suggesting a local neuroimmune disruption induced by systemic metabolic disturbance.

Diurnal Control of Sensory Axon Growth and Shedding in the Mouse Cornea.

Purpose: The circadian clock plays an important role in the expression and regulation of various genes and cellular processes in the body. Here, we study diurnal regulation of the growth and shedding of the sensory axons in the mouse cornea. Methods: Male and female BALB/cN mice were euthanized 90 minutes before and after the lights are turned on and off; at 5:30 AM, 8:30 AM, 5:30 PM, and 8:30 PM. Nerve terminal growth, shedding and overall axon density were assessed at these four time points using confocal imaging after staining axons in en face whole mount corneas with antibodies against ßIII tubulin, GAP43, and L1CAM. In addition, corneal epithelial cell proliferation, thickness, and desquamation were assessed using ki67, LAMP1, Involucrin, and ZO1. Results: Nerve terminal shedding took place between 5:30 AM and 8:30 AM and correlated positively with the timing of apical cell desquamation. After shedding the tips of the nerve terminals, axonal growth increased as indicated by increased axonal GAP43 expression. At 5:30 PM and 8:30 PM before and after the lights are turned off, cell proliferation was reduced, and epithelial thickness was maximal. Conclusions: Intraepithelial corneal nerve growth and shedding are under diurnal control regulated by the time of day and whether lights are on or off. Axons extend during the day and are shed within 90 minutes after lights are turned on. The data presented in this article shed light on the potential role that circadian clock plays in corneal pain and discomfort.

Axonal debris accumulates in corneal epithelial cells after intraepithelial corneal nerves are damaged: A focused Ion Beam Scanning Electron Microscopy (FIB-SEM) study.

The intraepithelial corneal nerves (ICNs) that innervate the corneal epithelium are maintained through interactions with corneal epithelial cells and the extracellular matrix they produce. One to several axons bundle together within the basal cell layer and extend parallel to the ocular surface or branch and extend apically. Here we use 3-dimentional (3D) ultrastructural reconstructions of control and trephine injured mouse corneal epithelium and stroma produced using Focused Ion Beam Scanning Electron Microscope (FIB-SEM) to determine whether corneal epithelial or immune cells resident in the epithelium remove axonal debris and degrade it in their lysosomes after trephine injury to the cornea. We demonstrate that axonal fragments are internalized in the corneal epithelium and accumulate within electron dense structures consistent with lysosomes 3 h after trephine injury in both epithelial and immune cells located among the basal cells of the trephine injured cornea. Confocal imaging showed fewer CD45+ immune cells within the corneal epithelium after trephine injury compared to controls. The resolution obtained using FIB-SEM also allowed us to show that the presence of sensory axons at the basal aspect of the epithelial basal cells close to the anterior aspect of the epithelial basement membrane (EBM) is associated with a focal reduction in EBM thickness. In addition, we show using FIB-SEM and confocal imaging that superficial trephine injuries that do not penetrate the stroma, damage the integrity of anterior stromal nerves. These studies are the first to look at the mouse cornea following nerve injury using FIB-SEM.

Efficacy of topical microemulsion of fatty acids of the ω-3 series on the sub-epithelial corneal nerves regeneration after epithelium-off corneal collagen cross-linking for keratoconus.

PURPOSE: To evaluate efficacy of a microemulsion of fatty acids of the ω-3 series on the regeneration of the sub-epithelial corneal nerve plexus in patients with keratoconus after epi-off cross-linking. METHODS: In this prospective study, we recruited 40 patients, 18 females, mean age 28 years (range 22-37), who were randomly divided in two groups. Group A, 20 patients, after cross-linking were treated with a microemulsion of fatty acids of the ω-3 series. Group B were treated with hyaluronic acid (0.15%)-based tear substitute. Nerve tortuosity, reflectivity and density were examined with in vivo confocal microscopy. Ocular surface disease index (OSDI) questionnaire at the preoperative and at each follow-up visit (1, 3 and 6 months) after treatment was completed. RESULTS: No significant difference between the two groups was noted at 1 month in terms of nerve density and OSDI. A statistically significant difference between the two groups was detected at 3 months in terms of nerve fibers density (6 ± 0.82 in Group A and 1 ± 0.51 in Group B, P = 0.0001). Reflectivity and tortuosity of the fibers did not show significant differences between the two groups at any time point. At 1 month, OSDI average value in group A and in group B was 31.5 ± 1.94 and 30 ± 1.96, at 3 months 13 ± 1.71 and 28 ± 1.83, and at 6 months 10.5 ± 1.87 and 9.0 ± 1.81, respectively. CONCLUSION: The use of a microemulsion of fatty acids appears to ensure a faster regeneration of nerve fibers in patients undergoing epi-off cross-linking.

Remodeling of Substance P Sensory Nerves and Transient Receptor Potential Melastatin 8 (TRPM8) Cold Receptors After Corneal Experimental Surgery.

Purpose: To investigate changes in corneal nerves positive to substance P (SP) and transient receptor potential melastatin 8 (TRPM8) and gene expression in the trigeminal ganglia (TG) following corneal surgery to unveil peripheral nerve mechanism of induced dry eye-like pain (DELP). Methods: Surgery was performed on mice by removing the central epithelial and anterior stromal nerves. Mice were euthanized at different times up to 15 weeks. Immunostaining was performed with TRPM8, SP, or protein gene product 9.5 (PGP9.5) antibodies, and epithelial nerve densities were calculated. The origin of TRPM8- and SP-TG neurons were analyzed by retrograde tracing. Gene expression in TG was studied by real-time PCR analysis. Results: SP-positive epithelial corneal nerves were more abundant than TRPM8 and were expressed in different TG neurons. After injury, epithelial nerve regeneration occurs in two distinct stages. An early regeneration of the remaining epithelial bundles reached the highest density on day 3 and then rapidly degraded. From day 5, the epithelial nerves originated from the underlying stromal nerves were still lower than normal levels by week 15. The SP- and TRPM8-positive nerve fibers followed the same pattern as the total nerves. TRPM8-positive terminals increased slowly and reached only half of normal values by 3 months. Corneal sensitivity gradually increased and reached normal values on day 12. Corneal injury also induced significant changes in TG gene expression, decreasing trpm8 and tac1 genes. Conclusions: Abnormal SP expression, low amounts of TRPM8 terminals, and hypersensitive nerve response occur long after the injury and changes in gene expression in the TG suggest a contribution to the pathogenesis of corneal surgery-induced DELP.

The Effects of Diabetes and High-Fat Diet on Polymodal Nociceptor and Cold Thermoreceptor Nerve Terminal Endings in the Corneal Epithelium.

Purpose: There is a substantial body of evidence indicating that corneal sensory innervation is affected by pathology in a range of diseases. However, there are no published studies that have directly assessed whether the nerve fiber density of the different subpopulations of corneal sensory neurons are differentially affected. The present study explored the possibility that the intraepithelial nerve fiber density of corneal polymodal nociceptors and cold thermoreceptors are differentially affected in mice fed with a high-fat high cholesterol (HFHC; 21% fat, 2% cholesterol) diet and in those that also have diabetes. Methods: The mice were fed the HFHC diet for the duration of the experiment (up to 40 weeks). Mice in the diabetes group had hyperglycaemia induced with streptozotocin after 15 weeks on the HFHC diet. Age-matched control animals were fed a standard diet. All corneal nerve fibers were labeled with a pan neuronal antibody (antiprotein gene product 9.5), and polymodal nociceptors and cold thermoreceptors were labeled with antibodies directed against transient receptor potential cation channel, subfamily V, member 1 and transient receptor potential cation channel subfamily M member 8, respectively. Results: The mice fed a HFHC diet and those that in addition have hyperglycemia have similar reductions in corneal nerve fiber density consistent with small fiber neuropathy. Importantly, both treatments more markedly affected the intraepithelial axons of cold thermoreceptors than those of polymodal nociceptors. Conclusions: The results provide evidence that distinct subpopulations of corneal sensory neurons can be differentially affected by pathology.

Contact lens wear and the diabetic corneal epithelium: A happy or disastrous marriage?

Diabetes mellitus is an epidemic in the US and abroad. With the advent of new contact lens technology, the use of contact lenses as glucose sensors in lieu of the traditional finger stick is quickly becoming realized. This has the potential to rapidly expand the contact lens market into this growing patient population. The independent cellular and physiological effects of contact lens wear and diabetes on the corneal epithelium have been described. However, little evidence exists to date to support whether there is increased risk associated with contact lens wear in diabetes. The focus of this review is to discuss what is known about the cellular effects of contact lenses on the corneal epithelium, the pathophysiological changes in the corneal epithelium that occur in diabetes, and whether an increased risk for corneal epithelial damage and/or infection may negatively impact safety in diabetic contact lens wearers. Available data indicates that there are inherent risks associated with contact lens wear in diabetics. Importantly, eye care practitioners fitting contact lenses in the diabetic patient need to carefully consider the duration of disease, the level of glycemic control, the presence of retinopathy, and the patient's overall health.

Developmental analysis of SV2 in the embryonic chicken corneal epithelium.

Intraepithelial corneal nerves (ICNs) help protect the cornea as part of the blink reflex and by modulating tear production. ICNs are also thought to regulate the health and homeostasis of the cornea through the release of trophic factors. Disruption to these nerves can lead to vision loss. Despite their importance little is known about how corneal nerves function and even less is known about how the cornea is initially innervated during its embryonic development. Here, we investigated the innervation of the embryonic chicken cornea. Western blot and immunohistochemistry were used to characterize the localization of the synaptic vesicle marker SV2, a molecule thought to be involved in the release of trophic factors from sensory nerves. The data show that both SV2 and synaptotagmin co-localize to ICNs. Nerves in the conjunctiva also contained SV2 and synaptotagmin, but these were localized to below the basal layers of the conjunctiva epithelium. SV2 isolated from corneal epithelium migrates in western blot at a heavier weight than SV2 isolated from brain, which suggests a role in vesicle targeting, as the deglycosylating enzyme PnGase does not affect corneal SV2.

Reduced intraepithelial corneal nerve density and sensitivity accompany desiccating stress and aging in C57BL/6 mice.

Dry Eye disease causes discomfort and pain in millions of patients. Using a mouse acute desiccating stress (DS) model we show that DS induces a reduction in intraepithelial corneal nerve (ICN) density, corneal sensitivity, and apical extension of the intraepithelial nerve terminals (INTs) that branch from the subbasal nerves (SBNs). Topical application of 0.02% Mitomycin C (MMC) or vehicle alone has no impact on the overall loss of axon density due to acute DS. Chronic dry eye, which develops progressively as C57BL/6 mice age, is accompanied by significant loss of the ICNs and corneal sensitivity between 2 and 24 months of age. QPCR studies show that mRNAs for several proteins that regulate axon growth and extension are reduced in corneal epithelial cells by 24 months of age but those that regulate phagocytosis and autophagy are not altered. Taken together, these data demonstrate that dry eye disease is accompanied by alterations in intraepithelial sensory nerve morphology and function and by reduced expression in corneal epithelial cells of mRNAs encoding genes mediating axon extension. Précis: Acute and chronic mouse models of dry eye disease are used to evaluate the pathologic effects of dry eye on the intraepithelial corneal nerves (ICNs) and corneal epithelial cells. Data show reduced numbers of sensory nerves and alterations in nerve morphology, sensitivity, corneal epithelial cell proliferation, and expression of mRNAs for proteins mediating axon extension accompany the pathology induced by dry eye.

Acute corneal epithelial debridement unmasks the corneal stromal nerve responses to ocular stimulation in rats: implications for abnormal sensations of the eye.

It is widely accepted that the mechanisms for transducing sensory information reside in the nerve terminals. Occasionally, however, studies have appeared demonstrating that similar mechanisms may exist in the axon to which these terminals are connected. We examined this issue in the cornea, where nerve terminals in the epithelial cell layers are easily accessible for debridement, leaving the underlying stromal (axonal) nerves undisturbed. In isoflurane-anesthetized rats, we recorded extracellularly from single trigeminal ganglion neurons innervating the cornea that are excited by ocular dryness and cooling: low-threshold (<2°C cooling) and high-threshold (>2°C) cold-sensitive plus dry-sensitive neurons playing possible roles in tearing and ocular pain. We found that the responses in both types of neurons to dryness, wetness, and menthol stimuli were effectively abolished by the debridement, indicating that their transduction mechanisms lie in the nerve terminals. However, some responses to the cold, heat, and hyperosmolar stimuli in low-threshold cold-sensitive plus dry-sensitive neurons still remained. Surprisingly, the responses to heat in approximately half of the neurons were augmented after the debridement. We were also able to evoke these residual responses and follow the trajectory of the stromal nerves, which we subsequently confirmed histologically. The residual responses always disappeared when the stromal nerves were cut at the limbus, suggesting that the additional transduction mechanisms for these sensory modalities originated most likely in stromal nerves. The functional significance of these residual and enhanced responses from stromal nerves may be related to the abnormal sensations observed in ocular disease.NEW & NOTEWORTHY In addition to the traditional view that the sensory transduction mechanisms exist in the nerve terminals, we report here that the proximal axons (stromal nerves in the cornea from which these nerve terminals originate) may also be capable of transducing sensory information. We arrived at this conclusion by removing the epithelial cell layers of the cornea in which the nerve terminals reside but leaving the underlying stromal nerves undisturbed.

[Botulinum toxin A induced protective ptosis for the treatment of recurrent epithelial defects in neurotrophic keratopathy]. / Medikamentöse Oberlidptosis in der Therapie der rezidivierenden Epitheldefekte infolge neurotropher Keratopathie.

In this article, a case of recurrent epithelial defects in neurotrophic keratopathy is described. Multiple abrasions of the corneal epithelium with a therapeutic contact lens, corneal stitches, and amniotic membrane transplantation in combination with artificial tears brought only short-term success. However, a botulinum toxin A induced protective ptosis could finally achieve permanent epithelial closure. As this case shows, protective ptosis can be a promising approach in spite of multiple previous ineffective therapeutic efforts.

Intraepithelial dendritic cells and sensory nerves are structurally associated and functional interdependent in the cornea.

The corneal epithelium consists of stratified epithelial cells, sparsely interspersed with dendritic cells (DCs) and a dense layer of sensory axons. We sought to assess the structural and functional correlation of DCs and sensory nerves. Two morphologically different DCs, dendriform and round-shaped, were detected in the corneal epithelium. The dendriform DCs were located at the sub-basal space where the nerve plexus resides, with DC dendrites crossing several nerve endings. The round-shaped DCs were closely associated with nerve fiber branching points, penetrating the basement membrane and reaching into the stroma. Phenotypically, the round-shaped DCs were CD86 positive. Trigeminal denervation resulted in epithelial defects with or without total tarsorrhaphy, decreased tear secretion, and the loss of dendriform DCs at the ocular surface. Local DC depletion resulted in a significant decrease in corneal sensitivity, an increase in epithelial defects, and a reduced density of nerve endings at the center of the cornea. Post-wound nerve regeneration was also delayed in the DC-depleted corneas. Taken together, our data show that DCs and sensory nerves are located in close proximity. DCs may play a role in epithelium innervation by accompanying the sensory nerve fibers in crossing the basement membrane and branching into nerve endings.

How Blink Anomalies Can Contribute to Post-LASIK Neurotrophic Epitheliopathy.

: Post-LASIK (laser in situ keratomileusis) symptoms and signs of dry eye have multiple causes. For example, tear osmolarity and the concentration of inflammatory mediators can increase because of reduced aqueous production and increased evaporative loss as a result of lower blink rates and increased interblink intervals. The tear concentration of inflammatory mediators can also increase because of surgical trauma, wound healing, comorbid systemic and ocular diseases, and the use of punctal plugs. Studies that examine only mechanical sensitivity of the cornea cannot detect changes in chemical sensitivity, which can persist longer. Symptoms may be partly attributed to sensitization of the traumatized corneal or lid wiper sensory nerves by inflammatory mediators. Increased lid wiper sensitivity could increase awareness of blinks, especially if ocular surface lubricity is reduced. Incomplete blinks have been found to represent 10 to 22% of the total number of blinks. Loss of neural stimuli and lower blink rates increase the significance of incomplete blinks that approximately double related interblink intervals and tear evaporation. The most common location of post-LASIK epitheliopathy is the inferior area of the cornea, which is overexposed by incomplete blinks. The relevance of incomplete blinking to post-LASIK epitheliopathy is supported by the relative absence of this complication in the similarly neurotrophically disadvantaged upper corneal areas for which blink rates and other tear functions usually appear to be adequate to prevent epitheliopathy, which stains. Occupational or leisure-time activities such as computer use and reading, which have been found to reduce blink rates and blink completeness, appear to be significant risk factors for symptoms and signs of dry eye. Apart from reducing symptoms and signs of dry eye, prophylactic and post-LASIK blink exercises to reduce incomplete blink rates and associated overexposure of the ocular surface may also contribute to more accurate refractive outcomes through faster wound healing.

Poly(ADP-Ribose) Polymerase Inhibition Improves Corneal Epithelial Innervation and Wound Healing in Diabetic Rats.

PURPOSE: We evaluated the effect of poly(ADP-ribose) polymerase (PARP) inhibition by using 1,5-isoquinolinediol (ISO) on corneal epithelial innervation in diabetic rats. METHODS: ISO (3 mg/kg, intraperitoneal) or vehicle was administered to rats with diabetes induced by streptozotocin for 4 weeks. Epithelial innervation, epithelial wound healing, and corneal sensation were evaluated in diabetic rats (DM rats), diabetic rats treated with ISO (DM-ISO rats), and nondiabetic (non-DM) rats. The density of epithelial innervation was calculated separately as nerve terminals and sub-basal nerve plexus by analyzing the images of whole-mount corneas. Healed areas of epithelial defect were measured at 0, 18, and 36 hours after creating a 4-mm wound on the cornea. Corneal sensitivity test was conducted using a Cochet-Bonnet handheld esthesiometer. Additionally, PARP1 and poly(ADP-ribosyl)ated polymers (pADPr) as its products, were identified in trigeminal ganglions (TGs) by Western blot analysis and immunofluorescence staining. RESULTS: In DM rats, the density of nerve terminals (5.57% ± 0.94%) and sub-basal nerve plexus (22.08 ± 1.78 mm/mm(2)) was significantly reduced in comparison with that in DM-ISO rats (8.64% ± 1.42%, 30.82 ± 2.01 mm/mm(2), respectively) and non-DM rats (9.02 ± 1.14%, 34.77 ± 4.45 mm/mm(2), respectively). The percentages of healed area of the epithelial defects at 18 and 36 hours were significantly smaller in DM rats (23.8 ± 5.2%, 53.2 ± 4.6%, respectively) than in DM-ISO rats (43.2 ± 1.4%, 75.8 ± 2.2%, respectively) and non-DM rats (48.1 ± 8.6%, 86.1 ± 3.3%, respectively). Corneal sensitivity decreased in DM rats (51.1 ± 0.3 mm) but not in DM-ISO rats (57.8 ± 0.2 mm). There were no differences between parameters in DM-ISO rats and those in non-DM rats. CONCLUSIONS: Diabetic corneas showed loss of epithelial innervation, resulting in delayed epithelial healing and decreased corneal sensitivity. Inhibition of poly(ADP-ribose) polymerase (PARP) with 1,5-isoquinolinediol alleviated these diabetes-induced alterations in the corneal epithelium in the diabetic rats.

Neurotrophic keratopathy due to dorsolateral medullary infarction (Wallenberg syndrome): case report and literature review.

BACKGROUND: Dorsolateral medullary infarction (Wallenberg syndrome) is rare in clinical practice; however, the subsequent corneal lesions are more uncommon. To our knowledge, only one such case was previously reported. We report a similar case with successful treatment and recovery, and analyse both cases to address the clinical features and outcomes of such syndrome. CASE PRESENTATION: A 43-year-old male presented with neurotrophic keratopathy one month after sustaining dorsolateral medullary infarction. The patient underwent amniotic membrane transplantation twice. Two-year follow-up observation revealed changes in nerve fibers and epithelial cells of corneal by laser confocal microscopy. CONCLUSION: By studying both cases, we confirm that neurotrophic keratopathy could be as a delayed-onset complication of Wallenberg syndrome. The recognition that neurotrophic keratopathy can follow dorsolateral medullary infarction could prevent the clinical misdiagnosis.

Effect of glycemic control on corneal nerves and peripheral neuropathy in streptozotocin-induced diabetic C57Bl/6J mice.

We sought to determine the impact that duration of hyperglycemia and control has on corneal nerve fiber density in relation to standard diabetic neuropathy endpoints. Control and streptozotocin-diabetic C57Bl/6J mice were analyzed after 4, 8, 12, and 20 weeks. For the 20-week time point, five groups of mice were compared: control, untreated diabetic, and diabetic treated with insulin designated as having either poor glycemic control, good glycemic control, or poor glycemic control switched to good glycemic control. Hyperglycemia was regulated by use of insulin-releasing pellets. Loss of corneal nerves in the sub-epithelial nerve plexus or corneal epithelium progressed slowly in diabetic mice requiring 20 weeks to reach statistical significance. In comparison, slowing of motor and sensory nerve conduction velocity developed rapidly with significant difference compared with control mice observed after 4 and 8 weeks of hyperglycemia, respectively. In diabetic mice with good glycemic control, average blood glucose levels over the 20-week experimental period were lowered from 589 ± 2 to 251 ± 9 mg/dl. All diabetic neuropathy endpoints examined were improved in diabetic mice with good glycemic control compared with untreated diabetic mice. However, good control of blood glucose was not totally sufficient in preventing diabetic neuropathy.

The impact of type 1 diabetes mellitus on corneal epithelial nerve morphology and the corneal epithelium.

Diabetic corneal neuropathy can result in chronic, sight-threatening corneal pathology. Although the exact etiology is unknown, it is believed that a reduction in corneal sensitivity and loss of neurotrophic support contributes to corneal disease. Information regarding the relationship between nerve loss and effects on the corneal epithelium is limited. We investigated changes in the corneal epithelium and nerve morphology using three-dimensional imaging in vivo and in situ in a streptozotocin-induced diabetic mouse model. Streptozotocin-treated mice showed increased levels of serum glucose and growth retardation consistent with a severe diabetic state. A reduction in the length of the subbasal nerve plexus was evident after 6 weeks of disease. Loss of the subbasal nerve plexus was associated with corneal epithelial thinning and a reduction in basal epithelial cell density. In contrast, loss of the terminal epithelial nerves was associated with animal age. Importantly, this is the first rodent model of type 1 diabetes that shows characteristics of corneal epithelial thinning and a reduction in basal epithelial cell density, both previously have been documented in humans with diabetic corneal neuropathy. These findings indicate that in type 1 diabetes, nerve fiber damage is evident in the subbasal nerve plexus before terminal epithelial nerve loss and that neurotrophic support from both the subbasal nerve plexus and terminal epithelial nerves is essential for the maintenance of corneal epithelial homeostasis.

Effect of macrophage migration inhibitory factor on corneal sensitivity after laser in situ keratomileusis in rabbit.

PURPOSE: To investigate the effect of macrophage migration inhibitory factor (MIF) on corneal sensitivity after laser in situ keratomileusis (LASIK) surgery. METHODS: New Zealand white rabbits were used in this study. A hinged corneal flap (160-µm thick) was created with a microkeratome, and -3.0 diopter excimer laser ablation was performed. Expressions of MIF mRNA in the corneal epithelial cells and surrounding inflammatory cells were analyzed using reverse transcription polymerase chain reaction at 48 hours after LASIK. After LASIK surgery, the rabbits were topically given either 1) a balanced salt solution (BSS), 2) MIF (100 ng/mL) alone, or 3) a combination of nerve growth factor (NGF, 100 ug/mL), neurotrophine-3 (NT-3, 100 ng/mL), interleukin-6 (IL-6, 5 ng/mL), and leukemia inhibitory factor (LIF, 5 ng/mL) four times a day for three days. Preoperative and postoperative corneal sensitivity at two weeks and at 10 weeks were assessed using the Cochet-Bonnet esthesiometer. RESULTS: Expression of MIF mRNA was 2.5-fold upregulated in the corneal epithelium and 1.5-fold upregulated in the surrounding inflammatory cells as compared with the control eyes. Preoperative baseline corneal sensitivity was 40.56 ± 2.36 mm. At two weeks after LASIK, corneal sensitivity was 9.17 ± 5.57 mm in the BSS treated group, 21.92 ± 2.44 mm in the MIF treated group, and 22.42 ± 1.59 mm in the neuronal growth factors-treated group (MIF vs. BSS, p < 0.0001; neuronal growth factors vs. BSS, p < 0.0001; MIF vs. neuronal growth factors, p = 0.815). At 10 weeks after LASIK, corneal sensitivity was 15.00 ± 9.65, 35.00 ± 5.48, and 29.58 ± 4.31 mm respectively (MIF vs. BSS, p = 0.0001; neuronal growth factors vs. BSS, p = 0.002; MIF vs. neuronal growth factors, p = 0.192). Treatment with MIF alone could achieve as much of an effect on recovery of corneal sensation as treatment with combination of NGF, NT-3, IL-6, and LIF. CONCLUSIONS: Topically administered MIF plays a significant role in the early recovery of corneal sensitivity after LASIK in the experimental animal model.

Effect of insulin-like growth factor-1 on corneal surface ultrastructure and nerve regeneration of rabbit eyes after laser in situ keratomileusis.

To explore the effect of insulin-like growth factor-1 (IGF-1) on corneal surface ultrastructure and nerve regeneration in rabbit models after laser in situ keratomileusis (LASIK). Forty-two healthy New Zealand white rabbits were divided into two groups, the IGF-1 group and the control group, and LASIK surgery was performed. The corneal surface ultrastructure was observed by transmission electron microscopy, and the nerve regeneration was evaluated by counting the newly regenerated nerves at 1 d, 1 w, 2 w, 1 m, 3 m and 6 m after surgery. Dry eye parameters, including the Schirmer I test and tear break-up time, were examined at all time points. The examination of corneal ultrastructure showed that the number of corneal epithelial microvilli in the IGF-1 group was significantly higher than that in the normal saline (NS) group except in the second postoperative week (p<0.05). The observation of corneal nerve regeneration showed that the number of regenerated nerve fibers in the IGF-1 group was higher than the control group at all time points (p<0.05). The parameters of dry eye were significantly higher in the IGF-1 group compared to the control group at all time points except at 1d and 6m after LASIK. IGF-1 can effectively accelerate the early repair of corneal surface ultrastructure and nerve regeneration after LASIK and relieve dry eye symptoms in rabbit eyes.