Ana Carolina Vieira

ELECTRICAL SIGNALING IN CONTROL OF OCULAR CELL BEHAVIORS

Epithelia of the cornea, lens and retina contain a vast array of ion channels and pumps. Together they produce a polarized flow of ions in and out of cells, as well as across the epithelia. These naturally occurring ion fluxes are essential to the hydration and metabolism of the ocular tissues, especially for the avascular cornea and lens. The directional transport of ions generates electric fields and currents in those tissues. Applied electric fields affect migration, division and proliferation of ocular cells which are important in homeostasis and healing of the ocular tissues. Abnormalities in any of those aspects may underlie many ocular diseases, for example chronic corneal ulcers, posterior capsule opacity after cataract surgery, and retinopathies. Electric field-inducing cellular responses, termed electrical signaling here, therefore may be an unexpected yet powerful mechanism in regulating ocular cell behavior. Both endogenous electric fields and applied electric fields could be exploited to regulate ocular cells. We aim to briefly describe the physiology of the naturally occurring electrical activities in the corneal, lens, and retinal epithelia, to provide experimental evidence of the effects of electric fields on ocular cell behaviors, and to suggest possible clinical implications.

Ionic Components of Electric Current at Rat Corneal Wounds

Background Endogenous electric fields and currents occur naturally at wounds and are a strong signal guiding cell migration into the wound to promote healing. Many cells involved in wound healing respond to small physiological electric fields in vitro. It has long been assumed that wound electric fields are produced by passive ion leakage from damaged tissue. Could these fields be actively maintained and regulated as an active wound response? What are the molecular, ionic and cellular mechanisms underlying the wound electric currents? Methodology/Principal Findings Using rat cornea wounds as a model, we measured the dynamic timecourses of individual ion fluxes with ion-selective probes. We also examined chloride channel expression before and after wounding. After wounding, Ca2+ efflux increased steadily whereas K+ showed an initial large efflux which rapidly decreased. Surprisingly, Na+ flux at wounds was inward. A most significant observation was a persistent large influx of Cl−, which had a time course similar to the net wound electric currents we have measured previously. Fixation of the tissues abolished ion fluxes. Pharmacological agents which stimulate ion transport significantly increased flux of Cl−, Na+ and K+. Injury to the cornea caused significant changes in distribution and expression of Cl− channel CLC2. Conclusions/Significance These data suggest that the outward electric currents occurring naturally at corneal wounds are carried mainly by a large influx of chloride ions, and in part by effluxes of calcium and potassium ions. Ca2+ and Cl− fluxes appear to be mainly actively regulated, while K+ flux appears to be largely due to leakage. The dynamic changes of electric currents and specific ion fluxes after wounding suggest that electrical signaling is an active response to injury and offers potential novel approaches to modulate wound healing, for example eye-drops targeting ion transport to aid in the challenging management of non-healing corneal ulcers.

Characterization of Novel O-Glycans Isolated from Tear and Saliva of Ocular Rosacea Patients

O-Glycans in saliva and tear isolated from patients suffering from ocular rosacea, a form of inflammatory ocular surface disease, were profiled, and their structures were elucidated using high resolution mass spectrometry. We have previously shown that certain structures, particularly sulfated oligosaccharides, increased in the tear and saliva of rosacea patients. In this study, the structures of these glycans were elucidated using primarily tandem mass spectrometry. There were important similarities in the glycan profiles of tears and saliva with the majority of the structures in common. The structures of the most abundant species common to both tear and saliva, which were also the most abundant species in both, were elucidated. For sulfated species, the positions of the sulfate groups were localized. The majority of the structures were new, with the sulfated glycans comprising mucin core 1- and core 2-type structures. As both saliva and tear are rich in mucins, it is suggested that the O-glycans are mainly components of mucins. The study further illustrates the strong correspondence between the glycans in the tear and saliva of ocular rosacea patients.

Specific ion fluxes generate cornea wound electric currents

The corneal epithelium generates a significant trans-epithelial potential (TEP) which aids in maintaining cornea water balance and transparency. Injury to the cornea causes a short circuit of the TEP at the wound. The TEP in the intact epithelium around the wound acts like a battery, powering significant ion flux and electric current at the wound. These circulating endogenous currents generate an electric field orientated towards the wound, with the wound the cathode. Many cell types, including human corneal epithelial cells and keratinocytes, migrate to the cathode at physiological electric field strengths. Indeed, the electric signal is a powerful stimulator of cell migration which appears to over-ride other cues such as chemotaxis and wound void. These wound fields also have a dynamic timecourse of change after wounding. It has been assumed that wound electric fields are produced by passive leakage of ions from damaged cells and tissue. Could these fields be actively maintained and regulated as an active wound response? What are the molecular, ionic and cellular mechanisms underlying the wound electric currents?

Rosai-Dorfman disease manifesting as an epibulbar ocular tumour.

The authors report a rare case of extranodal Rosai–Dorfman disease without systemic involvement. A 14‐year‐old girl presented with a slowly progressive, non‐tender, well‐circumscribed, perilimbal nodule in the right eye. Excisional biopsy was performed and histopathologic and immunohistochemical analysis established the diagnosis. Systemic evaluation revealed no other masses. After complete excision of the epibulbar lesion, the patient is asymptomatic and remains on regular follow up with no signs of recurrence. Rosai–Dorfman disease is an infrequent, benign histiocytic disorder that can rarely manifest as an exclusive extranodal mass. We report a case of solitary epibulbar Rosai–Dorfman disease, successfully treated with complete surgical excision of the lesion.

The safety and efficacy of linezolid and daptomycin as an additive in Optisol-GS against methicillin-resistant Staphylococcus aureus.

Purpose: To evaluate the efficacy of adding either linezolid or daptomycin to Optisol-GS donor storage medium in reducing methicillin-resistant Staphylococcus aureus (MRSA) contamination of donor corneas. Methods: Optisol-GS was supplemented with either linezolid at 2×, 4×, or 10× minimum inhibitory concentration (MIC) or daptomycin and calcium at 5× or 50× MIC. Unsupplemented control groups were also used. Gentamicin-sensitive and gentamicin-resistant isolates of MRSA were added, and vials were refrigerated for 48 hours followed by sampling for viable colony counts immediately upon removal from refrigeration and after warming to room temperature for 3 hours. Safety studies of Optisol-GS supplemented with 50× MIC daptomycin and calcium were performed by evaluating the central corneal thickness and endothelial cell density of the donor cornea. Stability of daptomycin in Optisol-GS at storage was also tested. Results: No added benefit was observed with linezolid supplementation to Optisol-GS against gentamicin-sensitive MRSA, with reduction in viable colony counts by >90% in all groups. No benefit was observed with linezolid supplementation against gentamicin-resistant MRSA, with the majority of inocula remaining viable in all groups. Viable counts of gentamicin-sensitive MRSA and gentamicin-resistant MRSA were effectively reduced with both 5× MIC and 50× MIC daptomycin supplementation. 50× MIC daptomycin–supplemented Optisol-GS had no appreciable effect on the central corneal thickness or endothelial cell density of the donor cornea and was stable at storage for 14 days. Conclusions: The addition of daptomycin to Optisol-GS significantly increases the anti-MRSA activity of the medium without any apparent negative effects on donor corneal tissue.