Relieving the symptoms of dry eye disease: update on lubricating eye drops containing hydroxypropyl-guar.

Hydroxypropyl-guar (HPG) is a thickening agent first added to lubricating eye drops in 2003. This agent, which enhances viscosity, has been used in the SYSTANE® family of lubricant eye drops (Alcon Laboratories, Inc., Fort Worth, TX, USA). HPG forms a partially linked gel with borate to prolong the retention of demulcents, such as polyethylene glycol and propylene glycol, on the eye. This helps to protect the ocular surface, thereby reducing the symptoms of dry eye disease (DED). The definition of DED has evolved with advances in research, leading to changes in HPG-containing eye care solutions. This article reviews current knowledge on the use of HPG-containing lubricating eye drops in the management of DED.

Effect of Artificial Tear Formulations on the Metabolic Activity of Human Corneal Epithelial Cells after Exposure to Desiccation.

Artificial lipid-containing tear formulations are developed to reduce tear evaporation by the restoration of a deficient tear lipid layer. Artificial tear formulations that prevent cell desiccation will result in ocular surface protection and the maintenance of cell metabolic activity. During dehydration, cells undergo the process of loss of metabolic activity and subsequently cell death. This work describes a method for assessing the efficacy of artificial tear formulations. The metabolic dye (i.e., alamarBlue) changes from a low fluorescent molecule resazurin to a fluorescent molecule resorufin in viable cells. The biological performance of an artificial tear formulation is measured as the ability of the formulation to (a) maintain cell viability and (b) provide cell protection from desiccation. Growth media and saline are used as controls for the cell viability/desiccation tests. Cells are incubated with test solutions for 30 min and then desiccated for 0 or 5 min at 37 °C and 45% relative humidity. Cell metabolic activity after initial exposure and after cell desiccation is then determined. The results show the comparative effects of eye drop formulations on cell metabolic activity and desiccation protection. This method can be used to test dry eye formulations that are designed to treat individuals with evaporative dry eye.

Preclinical Evaluation of a New Hydroxypropyl-Guar Phospholipid Nanoemulsion-Based Artificial Tear Formulation in Models of Corneal Epithelium.

PURPOSE: To evaluate the effect of hydroxypropyl-guar anionic phospholipid nanoemulsion (HP-guar nanoemulsion), a new artificial tear formulation for treatment of dry eye disease (DED), in corneal epithelium models. METHODS: Cultured human corneal epithelial cells were used to assess (1) hydration protection and hydration retention protection against desiccation, and (2) cell recovery after benzalkonium chloride (BAC) damage. Corneal epithelium permeability was measured by 5,6-carboxyfluorescein (CF) uptake in intact rabbit eyes. Lubricity was determined using simulated blinking in bovine pericardium-pericardium tribological experiments; elastic filament strength was measured using an extensional rheometer. Comparator arms included vehicle/control and the microemulsion [Systane® Balance (SYSB)]. RESULTS: Cell hydration protection was 39.5%, 7.1%, and -0.1%, and surface hydration retention was 32.6%, 11.0%, and -1.2% with HP-guar nanoemulsion, SYSB, and vehicle, respectively, after desiccation. After 48 h, cell recovery from BAC exposure (relative fluorescence units ± SD) was faster with HP-guar nanoemulsion treatment (2.66 ± 0.2) than SYSB (2.76 ± 0.2) and vehicle (3.11 ± 0.4). The CF permeability (ng CF/g) decreased in rabbit cornea treated with HP-guar nanoemulsion (9.6 ± 2.3) than those with SYSB (13.12 ± 2.8) or BAC-exposed cornea (22.6 ± 5.1). HP-guar nanoemulsion demonstrated greater lubricity and polymer filament break-up time than SYSB and vehicle. In all assessments, HP-guar nanoemulsion showed significant improvement versus vehicle/control (P < 0.05); outcomes were better with HP-guar nanoemulsion versus microemulsion. CONCLUSIONS: The HP-guar nanoemulsion promotes greater moisture retention, protection, improved cell barrier function, and increased elastic filament strength in corneal epithelium models. The potential clinical benefits of HP-guar nanoemulsion needs to be evaluated in patients with DED, in future studies.

Effects of a Hyaluronic Acid/Hydroxypropyl Guar Artificial Tear Solution on Protection, Recovery, and Lubricity in Models of Corneal Epithelium.

PURPOSE: Hydroxypropyl guar (HPG) and hyaluronic acid (HA) have been individually shown to improve dry eye symptoms. The purpose of this in vitro study was to assess the potential benefits of a new lubricant eye drop formulation containing the demulcents propylene glycol and polyethylene glycol and an HA/HPG dual polymer in models of the human corneal epithelium. METHODS: Cultured human corneal epithelial or corneal-limbal epithelial cells were treated with the HA/HPG dual-polymer formulation or single-polymer formulations containing either HPG or HA. Desiccation protection by cell hydration and surface retention was assessed using cell viability assays. Sodium fluorescein permeability, transepithelial resistance, and cell viability assays were conducted using pretreated cells exposed to a surfactant/detergent insult to evaluate cell and cell barrier protection. Surface lubricity was assessed in tribological experiments of pericardium-pericardium friction. RESULTS: Hydration protection against desiccation and protection by surface retention were significantly greater with the HA/HPG formulation versus HPG or HA (P<0.001) alone and with HPG versus HA (P ≤ 0.016). Fluorescein permeability and transepithelial resistance assays demonstrated significantly better cell and barrier protection from surfactant insult with HA/HPG versus the single-polymer formulations (P ≤ 0.01). After insult, there were markedly more viable cells evident with HA/HPG compared with HPG or HA alone. HA/HPG and HPG reduced surface friction to a greater extent than HA (P ≤ 0.02) and maintained lubricity after the formulations were rinsed away. CONCLUSIONS: HA/HPG provided effective hydration and lubrication and demonstrated prolonged retention of effect. HA/HPG may potentially promote desiccation protection and retention on the ocular surface.

Photochromic bacteriorhodopsin mutant with high holographic efficiency and enhanced stability via a putative self-repair mechanism.

The Q photoproduct of bacteriorhodopsin (BR) is the basis of several biophotonic technologies that employ BR as the photoactive element. Several blue BR (bBR) mutants, generated by using directed evolution, were investigated with respect to the photochemical formation of the Q state. We report here a new bBR mutant, D85E/D96Q, which is capable of efficiently converting the entire sample to and from the Q photoproduct. At pH 8.5, where Q formation is optimal, the Q photoproduct requires 65 kJ mol(-1) of amber light irradiation (590 nm) for formation and 5 kJ mol(-1) of blue light (450 nm) for reversion, respectively. The melting temperature of the resting state and Q photoproduct, measured via differential scanning calorimetry, is observed at 100 °C and 89 °C at pH 8.5 or 91 °C and 82 °C at pH 9.5, respectively. We hypothesize that the protein stability of D85E/D96Q compared to other blue mutants is associated with a rapid equilibrium between the blue form E85(H) and the purple form E85(-) of the protein, the latter providing enhanced structural stability. Additionally, the protein is shown to be stable and functional when suspended in an acrylamide matrix at alkaline pH. Real-time photoconversion to and from the Q state is also demonstrated with the immobilized protein. Finally, the holographic efficiency of an ideal thin film using the Q state of D85E/D96Q is calculated to be 16.7%, which is significantly better than that provided by native BR (6-8%) and presents the highest efficiency of any BR mutant to date.

Degradation study of carnosic acid, carnosol, rosmarinic acid, and rosemary extract (Rosmarinus officinalis L.) assessed using HPLC.

Rosemary, whose major caffeoyl-derived and diterpenoid ingredients are rosmarinic acid, carnosol, and carnosic acid, is an important source of natural antioxidants and is being recognized increasingly as a useful preservative, protectant, and even as a potential medicinal agent. Understanding the stability of these components and their mode of interaction in mixtures is important if they are to be utilized to greatest effect. A study of the degradation of rosmarinic acid, carnosol, carnosic acid, and a mixture of the three was conducted in ethanolic solutions at different temperatures and light exposure. As expected, degradation increased with temperature. Some unique degradation products were formed with exposure to light. Several degradation products were reported for the first time. The degradation products were identified by HPLC/MS/MS, UV, and NMR. The degradation of rosemary extract in fish oil also was investigated, and much slower rates of degradation were observed for carnosic acid. In the mixture of the three antioxidants, carnosic acid serves to maintain levels of carnosol, though it does so at least in part at the cost of its own degradation.

Light-induced retinal degeneration is prevented by zinc, a component in the age-related eye disease study formulation.

Mineral supplements are often included in multivitamin preparations because of their beneficial effects on metabolism. In this study, we used an animal model of light-induced retinal degeneration to test for photoreceptor cell protection by the essential trace element zinc. Rats were treated with various doses of zinc oxide and then exposed to intense visible light for as long as 8 h. Zinc treatment effectively prevented retinal light damage as determined by rhodopsin and retinal DNA recovery, histology and electrophoretic analysis of DNA damage and oxidized retinal proteins. Zinc oxide was particularly effective when given before light exposure and at doses two- to four-fold higher than recommended by the age-related eye disease study group. Treated rats exhibited higher serum and retinal pigment epithelial zinc levels and an altered retinal gene expression profile. Using an Ingenuity database, 512 genes with known functional annotations were found to be responsive to zinc supplementation, with 45% of these falling into a network related to cellular growth, proliferation, cell cycle and death. Although these data suggest an integrated and extensive regulatory response, zinc induced changes in gene expression also appear to enhance antioxidative capacity in retina and reduce oxidative damage arising from intense light exposure.

Mechanism of spectral tuning in green-absorbing proteorhodopsin.

The absorption spectrum of green proteorhodopsin (GPR) is pH-dependent, exhibiting either red-shifted (low pH) or blue-shifted (high pH) absorption maxima. We examine the molecular basis of the pH-dependent spectral properties of green proteorhodopsin by using homology modeling and molecular orbital theory. Bacteriorhodopsin (BR) and sensory rhodopsin II (SRII) are compared as homology templates. The model of GPR generated by using BR as the homology parent is better than that generated by using SRII on the basis of the potential energy, relative stability to dynamics, and ability to rationalize pH effects. MNDO-PSDCI (molecular neglect of differential overlap with partial single- and double-configuration interaction) calculations provide insight into the spectroscopic properties of GPR and help rule out the viability of the SRII-based model. The proximity of His 75 to the quadrupole residues (LYR, D97, D227, and R94) in the BR-based model provides a good model for both the low- and high-pH spectral states of GPR. The observation that BR is a better structural model for GPR than SRII is in contrast to our previous study of BPR, which observed that SRII was the better homology parent [Hillebrecht, J. R. (2006) Biochemistry 45, 1579-1590]. The implications of this observation are discussed.

Structure, function, and wavelength selection in blue-absorbing proteorhodopsin.

The absorption maximum of blue proteorhodopsin (BPR) is the most blue-shifted of all retinal proteins found in archaea or bacteria, with the exception of sensory rhodopsin II (SRII). The absorption spectrum also exhibits a pH dependence larger than any other retinal protein. We examine the structural origins of these two properties of BPR by using optical spectroscopy, homology modeling, and molecular orbital theory. Bacteriorhodopsin (BR) and SRII are used as homology parents for comparative purposes. We find that the tertiary structure of BPR based on SRII is more realistic with respect to free energy, dynamic stability, and spectroscopic properties. Molecular orbital calculations including full single- and double-configuration interaction within the chromophore pi-electron system provide perspectives on the wavelength regulation in this protein and indicate that Arg-95, Gln-106, Glu-143, and Asp-229 play important, and in some cases pH-dependent roles. A possible model for the 0.22 eV red shift of BPR at low pH is examined, in which Glu-143 becomes protonated and releases Arg-95 to rotate up into the binding site, altering the electrostatic environment of the chromophore. At high pH, BPR has spectroscopic properties similar to SRII, but at low pH, BPR has spectroscopic properties more similar to BR. Nevertheless, SRII is a significantly better homology model for BPR and opens up the question of whether this protein serves as a proton pump, as commonly believed, or is a light sensor with structure-function properties more comparable to those of SRII. The function of BPR in the native organism is discussed with reference to the results of the homology model.