In-vitro evaluation of the evaporation retardation by Meibomian lipids in homogeneous and non-homogeneous evaporation.

HYPOTHESIS: An important function of the Tear Film Lipid Layer (TFLL) is the retardation of evaporation. We propose two micro-scaled systems to quantify the influence of the TFLL in evaporation for single patients, which may contribute as an improvement on the diagnosis of Meibomian Gland Dysfunctions (MGD). EXPERIMENTS: Meibum was extracted from 10 patients with hypersecretory MGD and 9 healthy controls. The lipids were placed over water, and the evaporation was determined in the case of homogeneous evaporation over a surface (pendant drop), and the case where the evaporation depends on a pinned triple contact line (meniscus). FINDINGS: For the homogeneous case, the presence of Meibum reduced evaporation in 30%, although there was no significant difference between controls and MGD patients. However, evaporation induced by menisci was 25 % higher in MGD patients. Our results contribute to the evidence of the inhibition of evaporation by Meibum. Our study also suggests that the evaporation induced by contact points may be a more relevant model to measure differences in evaporation due to the composition of Meibum. This model may also have connotations in the occurrence of internal stresses in the tear film, inducing its instability.

Particle/wall electroviscous effects at the micron scale: comparison between experiments, analytical and numerical models.

We report a experimental study of the motion of 1 µm single particles interacting with functionalized walls at low and moderate ionic strengths conditions. The 3D particle's trajectories were obtained by analyzing the diffracted particle images (point spread function). The studied particle/wall systems include negatively charged particles interacting with bare glass, glass covered with polyelectrolytes and glass covered with a lipid monolayer. In the low salt regime (pure water) we observed a retardation effect of the short-time diffusion coefficients when the particle interacts with a negatively charged wall; this effect is more severe in the perpendicular than in the lateral component. The decrease of the diffusion as a function of the particle-wall distancehwas similar regardless the origin of the negative charge at the wall. When surface charge was screened or salt was added to the medium (10 mM), the diffusivity curves recover the classical hydrodynamic behavior. Electroviscous theory based on the thin electrical double layer (EDL) approximation reproduces the experimental data except for smallh. On the other hand, 2D numerical solutions of the electrokinetic equations showed good qualitative agreement with experiments. The numerical model also showed that the hydrodynamic and Maxwellian part of the electroviscous total drag tend to zero ash→ 0 and how this is linked with the merging of both EDL's at close proximity.

Tissue and cellular biomechanics during corneal wound injury and repair.

Corneal wound healing is an enormously complex process that requires the simultaneous cellular integration of multiple soluble biochemical cues, as well as cellular responses to the intrinsic chemistry and biophysical attributes associated with the matrix of the wound space. Here, we document how the biomechanics of the corneal stroma are altered through the course of wound repair following keratoablative procedures in rabbits. Further we documented the influence that substrate stiffness has on stromal cell mechanics. Following corneal epithelial debridement, New Zealand white rabbits underwent phototherapeutic keratectomy (PTK) on the right eye (OD). Wound healing was monitored using advanced imaging modalities. Rabbits were euthanized and corneas were harvested at various time points following PTK. Tissues were characterized for biomechanics with atomic force microscopy and with histology to assess inflammation and fibrosis. Factor analysis was performed to determine any discernable patterns in wound healing parameters. The matrix associated with the wound space was stiffest at 7days post PTK. The greatest number of inflammatory cells were observed 3days after wounding. The highest number of myofibroblasts and the greatest degree of fibrosis occurred 21days after wounding. While all clinical parameters returned to normal values 400days after wounding, the elastic modulus remained greater than pre-surgical values. Factor analysis demonstrated dynamic remodeling of stroma occurs between days 10 and 42 during corneal stromal wound repair. Elastic modulus of the anterior corneal stroma is dramatically altered following PTK and its changes coincide initially with the development of edema and inflammation, and later with formation of stromal haze and population of the wound space with myofibroblasts. Factor analysis demonstrates strongest correlation between elastic modulus, myofibroblasts, fibrosis and stromal haze thickness, and between edema and central corneal thickness. STATEMENT OF SIGNIFICANCE: Tissue biomechanics during the course of corneal wound healing is documented for the first time through atomic force microscopy, and is correlated with advanced clinical imaging and immunohistochemistry. Parameters obtained from the study are applied in a multivariate statistical model to cluster the data for better classification and monitor the wound repair process. Elastic modulus of the anterior corneal stroma is dramatically altered following wounding and correlates initially with the development of edema and inflammation, and later with formation of stromal haze and population of the wound space with myofibroblasts. Importantly, the occurrence of myofibroblasts is preceded by changes in tissue mechanics, which is important to consider in light of crosslinking procedures applied to treat corneal diseases.

Fogging Control on LDPE/EVA Coextruded Films: Wettability Behavior and Its Correlation with Electric Performance.

The transformation of fog at a non-visible water layer on a membrane of low-density polyethylene (LDPE) and ethylene-vinyl acetate (EVA) was evaluated. Nonionic surfactants of major demand in the polyolefin industry were studied. A kinetic study using a hot fog chamber showed that condensation is controlled by both the diffusion and permanency of the surfactant more than by the change of the surface energy developed by the wetting agents. The greatest permanency of the anti-fog effect of the LDPE/EVA surface was close to 3000 h. The contact angle results demonstrated the ability of the wetting agent to spread out to the surface. Complementarily, the migration of nonionic surfactants from the inside of the polymeric matrix to the surface was analyzed by Fourier transform infrared (FTIR) microscopy. Additionally, electrical measurement on the anti-fogging membrane at alternating currents and at a sweep frequency was proposed to test the conductivity and wetting ability of nonionic surfactants. We proved that the amphiphilic molecules had the ability to increase the conductivity in the polyolefin membrane. A correlation between the bulk electrical conductivity and the permanency of the fogging control on the LDPE/EVA coextruded film was found.

Species variation and spatial differences in mucin expression from corneal epithelial cells.

Mucins are large glycoproteins expressed by epithelial cells of both the conjunctiva and cornea, and principle components of the glycocalyx. They are thought to play an important role in determining the interactions between the cornea/conjunctiva and the overlying tear film. The purpose of this study was to characterize the membrane-associated corneal mucin expression pattern from multiple species commonly used in ophthalmic research and drug development to better define the biochemical attributes of the ocular surface. Humans, rhesus macaques and dogs were found to have a very similar pattern of mucin expression, with mucin 16 (MUC16) being the most prevalent mucin transcript. In contrast, the rabbit had a unique mucin expression pattern with all mucin transcripts expressed at relatively similar levels. To determine if there were spatial differences in expression, peripheral and central corneal epithelium were individually isolated and evaluated for mucin expression. In all species examined, MUC1, MUC4 and MUC16 had higher peripheral corneal expression when compared with central, which reached statistical significance in MUC1 (rhesus and dog). The data demonstrated variation in corneal epithelial membrane-associated mucin expression between species, with the rabbit having a distinct expression pattern. These differences may be reflective of the environment, pathogen exposure or tear film dynamics of the respective species. The species differences, as well as regional mucin expression patterns, characterized in this study further define the biochemical composition of the ocular surface and may play an important role in tear film stability.

Transport of Colloids along Corners: Visualization of Evaporation-Induced Flows beyond the Axisymmetric Condition.

Nonhomogeneous evaporation fluxes have been shown to promote the formation of internal currents in sessile droplets, explaining the patterns that suspended particles leave after the droplet has dried out. Although most evaporation experiments have been conducted using spherical-cap-shaped drops, which are essentially in an axisymmetric geometry, here we show an example of nonhomogeneous evaporation in asymmetric geometries, which is visualized by following the motion of colloidal particles along liquid fingers forming a meniscus at square corners. It is found that the particle's velocity increases with the diffusive evaporation factor [Formula: see text] for the three tested fluids: water, isopropyl alcohol (IPA), and ethanol (EtOH). Here, [Formula: see text] is the vapor diffusivity in air, RH is the relative amount of vapor in the atmosphere, and cs is the saturated vapor concentration. We observed that in IPA and EtOH the internal currents promote a 3D spiral motion, whereas in water the particle's trajectory is basically unidirectional. By adding 0.25 critical micelle concentration (CMC) of sodium dodecyl sulfate (SDS) surfactant in water, a velocity blast was observed in the whole circulation flow pattern, going from [Formula: see text] to nearly [Formula: see text] in the longitudinal velocity component. To assess the effect of breaking the axisymmetric condition on the evaporation flux profile, we numerically solved the diffusive equation in model geometries that preserve the value of the contact angle θ but introduce an additional angle ϕ that characterizes the solid substrate. By testing different combinations of θ and ϕ, we corroborated that the evaporation flux increases when the substrate and the gas-liquid curves meet at corners with increasing sharpness.

Effect of Stratification on Surface Properties of Corneal Epithelial Cells.

PURPOSE: The purpose of this study was to determine the influence of mucin expression in an immortalized human corneal epithelial cell line (hTCEpi) on the surface properties of cells, such as wettability, contact angle, and surface heterogeneity. METHODS: hTCEpi cells were cultured to confluence in serum-free medium. The medium was then replaced by stratification medium to induce mucin biosynthesis. The mucin expression profile was analyzed using quantitative PCR and Western blotting. Contact angles were measured using a two-immiscible liquid method, and contact angle hysteresis was evaluated by tilting the apparatus and recording advancing and receding contact angles. The spatial distribution of mucins was evaluated with fluorescently labeled lectin. RESULTS: hTCEpi cells expressed the three main ocular mucins (MUC1, MUC4, and MUC16) with a maximum between days 1 and 3 of the stratification process. Upon stratification, cells caused a very significant increase in contact angle hysteresis, suggesting the development of spatially discrete and heterogeneously distributed surface features, defined by topography and/or chemical functionality. Although atomic force microscopy measurements showed no formation of appreciable topographic features on the surface of the cells, we observed a significant increase in surface chemical heterogeneity. CONCLUSIONS: The surface chemical heterogeneity of the corneal epithelium may influence the dynamic behavior of tear film by "pinning" the contact line between the cellular surface and aqueous tear film. Engineering the surface properties of corneal epithelium could potentially lead to novel treatments in dry eye disease.

Interfacial phenomena and the ocular surface.

Ocular surface disorders, such as dry eye disease, ocular rosacea, and allergic conjunctivitis, are a heterogeneous group of diseases that require an interdisciplinary approach to establish underlying causes and develop effective therapeutic strategies. These diverse disorders share a common thread in that they involve direct changes in ocular surface chemistry as well as the rheological properties of the tear film and topographical attributes of the cellular elements of the ocular surface. Knowledge of these properties is crucial to understand the formation and stability of the preocular tear film. The study of interfacial phenomena of the ocular surface flourished during the 1970s and 1980s, but after a series of lively debates in the literature concerning distinctions between the epithelial and the glandular origin of ocular surface disorders during the 1990s, research into this important topic has declined. In the meantime, new tools and techniques for the characterization and functionalization of biological surfaces have been developed. This review summarizes the available literature regarding the physicochemical attributes of the ocular surface, analyzes the role of interfacial phenomena in the pathobiology of ocular surface disease, identifies critical knowledge gaps concerning interfacial phenomena of the ocular surface, and discusses the opportunities for the exploitation of these phenomena to develop improved therapeutics for the treatment of ocular surface disorders.

The influence of substrate topography on the migration of corneal epithelial wound borders.

Currently available artificial corneas can develop post-implant complications including epithelial downgrowth, infection, and stromal melting. The likelihood of developing these disastrous complications could be minimized through improved formation and maintenance of a healthy epithelium covering the implant. We hypothesize that this epithelial formation may be enhanced through the incorporation of native corneal basement membrane biomimetic chemical and physical cues onto the surface of the keratoprosthesis. We fabricated hydrogel substrates molded with topographic features containing specific bio-ligands and developed an in vitro wound healing assay. In our experiments, the rate of corneal epithelial wound healing was significantly increased by 50% in hydrogel surfaces containing topographic features, compared to flat surfaces with the same chemical attributes. We determined that this increased healing is not due to enhanced proliferation or increased spreading of the epithelial cells, but to an increased active migration of the epithelial cells. These results show the potential benefit of restructuring and improving the surface of artificial corneas to enhance epithelial coverage and more rapidly restore the formation of a functional epithelium.

Arrays of topographically and peptide-functionalized hydrogels for analysis of biomimetic extracellular matrix properties.

Epithelial cells reside on specialized extracellular matrices that provide instructive cues to regulate and support cell function. The authors have previously demonstrated that substrate topography with dimensions similar to the native extracellular matrix (submicrometer and nanoscale features) significantly impacts corneal epithelial proliferation and migration. In this work, synthetic hydrogels were modified with both topographic and biochemical cues, where specified peptide ligands were immobilized within nanopatterned hydrogels. The efficient, systematic study of multiple instructive cues (peptide, peptide concentration, topographic dimensions), however, is contingent on the development of higher throughput platforms. Toward this goal, the authors developed a hydrogel array platform to systematically and rapidly evaluate combinations of two different peptide motifs and a range of nanoscale topographic dimensions. Specifically, distinct functional pegylated peptide ligands, RGD (GGGRGDSP) and AG73 (GRKRLQVQLSIRT), were synthesized for incorporation into an inert hydrogel network. Elastomeric stencils with arrays of millimeter-scale regions were used to spatially confine hydrogel precursor solutions on elastomeric stamps with nanoscale patterns generated by soft lithography. The resulting topographically and peptide-functionalized hydrogel arrays were used to characterize single cell migration. Epithelial cell migration speed and persistence were governed by both the biochemical and topographical cues of the underlying substrate.