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 7 days post PTK. The greatest number of inflammatory cells were observed 3 days after wounding. The highest number of myofibroblasts and the greatest degree of fibrosis occurred 21 days after wounding. While all clinical parameters returned to normal values 400 days 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.

Incorporation of biophysical and biochemical cues into the design of biomaterials is an important strategy for tissue engineering, the design of biomedical implants and cell culture. Hydrogels synthesized from poly (ethylene glycol) diacrylate (PEGDA) were investigated as a platform to simultaneously present human corneal epithelial cells (HCECs) in vitro with topography and adhesion peptides to mimic the native physical and chemical attributes of the basement membrane underlying the epithelium in vivo. Hydrogels synthesized from aqueous solutions of 20% PEGDA (MW of 3400 g/mol) prevented non-specific cell adhesion and were functionalized with the integrin-binding peptide Arg-Gly-Asp (RGD) in concentrations from 5 to 20 mM. The hydrogels swelled minimally after curing and were molded with ridge and groove features with lateral dimensions from 200 nm to 2000 nm and 300 nm depth. HCECs were cultured on topographic surfaces functionalized with RGD and compared to control unfunctionalized topographic substrates. HCEC alignment, either parallel or perpendicular to ridges, was influenced by the culture media on substrates promoting non-specific attachment. In contrast, the alignment of HCECs cultured on RGD hydrogels showed substantially less dependence on the culture media. In the latter case, the moldable RGD-functionalized hydrogels allowed for decoupling the cues from surface chemistry, soluble factors, and topography that simultaneously impact HCEC behavior.

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.

Citation: Yáñez-Soto B, Leonard BC, Raghunathan VK, et al. Effect of stratification on the surface properties of corneal epithelial cells. Invest Ophthalmol Vis Sci. 2015;56:8340-8348. DOI:10.1167/iovs.15-17468 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.