For the majority of patients with severe corneal injury or disease, corneal transplantation is the only suitable treatment option. Unfortunately, the demand for donor corneas greatly exceeds the availability. To overcome shortage issues, a myriad of bioengineered constructs have been developed as mimetics of the corneal stroma over the last few decades. Despite the sheer number of bioengineered stromas developed , these implants fail clinical trials exhibiting poor tissue integration and adverse effects in vivo. Such shortcomings can partially be ascribed to poor biomechanical performance. In this review, existing approaches for bioengineering corneal stromal constructs and their mechanical properties are described. The information collected in this review can be used to critically analyze the biomechanical properties of future stromal constructs, which are often overlooked, but can determine the failure or success of corresponding implants.

The corneal endothelium is the innermost layer of the cornea that selectively pumps ions and metabolites and regulates the hydration level of the cornea, ensuring its transparency. Trauma or disease affecting human corneal endothelial cells (hCECs) can result in major imbalances of such transport activity with consequent deterioration or loss of vision. Since tissue transplantation from deceased donors is only available to a fraction of patients worldwide, alternative solutions are urgently needed. Cell therapy approaches, in particular by attempting to expand primary culture of hCECs in vitro, aim to tackle this issue. However, existing cell culture protocols result in limited expansion of this cell type. Recent studies in this field have shown that topographical features with specific dimensions and shapes could improve the efficacy of hCEC expansion. Therefore, potential solutions to overcome the limitation of the conventional culture of hCECs may include recreating nanometer scale topographies (nanotopographies) that mimic essential biophysical cues present in their native environment. In this review, we summarize the current knowledge and understanding of the effect of substrate topographies on the response of hCECs. Moreover, we also review the latest developments for the nanofabrication of such bio-instructive cell substrates.

In the event of disease or injury, restoration of the native organization of cells and extracellular matrix is crucial for regaining tissue functionality. In the cornea, a highly organized collagenous tissue, keratocytes can align along the anisotropy of the physical microenvironment, providing a blueprint for guiding the organization of the collagenous matrix. Inspired by this physiological process, anisotropic contact guidance cues have been employed to steer the alignment of keratocytes as a first step to engineer in vitro cornea-like tissues. Despite promising results, two major hurdles must still be overcome to advance the field. First, there is an enormous design space to be explored in optimizing cellular contact guidance in three dimensions. Second, the role of contact guidance cues in directing the long-term deposition and organization of extracellular matrix proteins remains unknown. To address these challenges, here we combined two microengineering strategies—UV-based protein patterning (2D) and two-photon polymerization of topographies (2.5D)—to create a library of anisotropic contact guidance cues with systematically varying height (H, 0 µm ≤ H ≤ 20 µm) and width (W, 5 µm ≤ W ≤ 100 µm). With this unique approach, we found that, in the short term (24 h), the orientation and morphology of primary human fibroblastic keratocytes were critically determined not only by the pattern width, but also by the height of the contact guidance cues. Upon extended 7-day cultures, keratocytes were shown to produce a dense, fibrous collagen network along the direction of the contact guidance cues. Moreover, increasing the heights also increased the aligned fraction of deposited collagen and the contact guidance response of cells, all whilst the cells maintained the fibroblastic keratocyte phenotype. Our study thus reveals the importance of dimensionality of the physical microenvironment in steering both cellular organization and the formation of aligned, collagenous tissues.

Electrospinning is a powerful method to fabricate structures resembling the fibrous texture of the native extracellular matrix. However, the random fiber deposition of the process hinders a faithful reproduction of the fiber mesh morphology on multiple samples, which raises difficulties in experimental designs to systematically test and assess cell response in vitro. A multi‐replication process to precisely reproduce the fiber morphology on different cell culture substrates is developed. The process involves a decoupling of the fiber structure, material, and porosity by combining the key advantages of electrospinning and imprinting. With this, fiber patterns having a diameter between 0.4 and 2.8 µm are replicated on polycarbonate, polystyrene, poly(methyl methacrylate), and cyclic olefin copolymer films. Identical fiber morphology is, then, obtained on porous films having a pore diameter between 2 and 12 µm. Having full control over these parameters allows the multireplication process to engineer well‐characterized cell microenvironments, which can potentially be used to further investigate complex cell–material interactions.

Corneas In article number 2100972, Stefan Giselbrecht and colleagues review existing approaches for bioengineering corneal stromal constructs. There is a high demand for bioengineered corneal constru...