Three-dimensional bioprinting in ophthalmic care

Three-dimensional (3D) bioprinting is widely used in ophthalmic clinic, including in diagnosis, surgery, prosthetics, medications, drug development and delivery, and medical education. Articles published in 2011–2022 into bioinks, printing technologies, and bioprinting applications in ophthalmology were reviewed and the strengths and limitations of bioprinting in ophthalmology highlighted. The review highlighted the trade-offs of printing technologies and bioinks in respect to, among others, material type cost, throughput, gelation technique, cell density, cell viability, resolution, and printing speed. There is already widespread ophthalmological application of bioprinting outside clinical settings, including in educational modelling, retinal imaging/visualization techniques and drug design/testing. In clinical settings, bioprinting has already found application in pre-operatory planning. Even so, the findings showed that even with its immense promise, actual translation to clinical applications remains distant, but relatively closer for the corneal (except stromal) tissues, epithelium, endothelium, and conjunctiva, than it was for the retina. This review similarly reflected on the critical on the technical, practical, ethical, and cost barrier to rapid progress of bioprinting in ophthalmology, including accessibility to the most sophisticated bioprinting technologies, choice, and suitability of bioinks, tissue viability and storage conditions. The extant research is encouraging, but more work is clearly required for the push towards clinical translation of research

Tissue Engineering the Cornea: The Evolution of RAFT.

Corneal blindness affects over 10 million people worldwide and current treatment strategies often involve replacement of the defective layer with healthy tissue. Due to a worldwide donor cornea shortage and the absence of suitable biological scaffolds, recent research has focused on the development of tissue engineering techniques to create alternative therapies. This review will detail how we have refined the simple engineering technique of plastic compression of collagen to a process we now call Real Architecture for 3D Tissues (RAFT). The RAFT production process has been standardised, and steps have been taken to consider Good Manufacturing Practice compliance. The evolution of this process has allowed us to create biomimetic epithelial and endothelial tissue equivalents suitable for transplantation and ideal for studying cell-cell interactions in vitro

Mimicking the Hierarchical Organization of Natural Collagen: Toward the Development of Ideal Scaffolding Material for Tissue Regeneration

Biological materials found in living organisms, many of which are proteins, feature a complex hierarchical organization. Type I collagen, a fibrous structural protein ubiquitous in the mammalian body, provides a striking example of such a hierarchical material, with peculiar architectural features ranging from the amino acid sequence at the nanoscale (primary structure) up to the assembly of fibrils (quaternary structure) and fibers, with lengths of the order of microns. Collagen plays a dominant role in maintaining the biological and structural integrity of various tissues and organs, such as bone, skin, tendons, blood vessels, and cartilage. Thus, "artificial" collagen-based fibrous assemblies, endowed with appropriate structural properties, represent ideal substrates for the development of devices for tissue engineering applications. In recent years, with the ultimate goal of developing three-dimensional scaffolds with optimal bioactivity able to promote both regeneration and functional recovery of a damaged tissue, numerous studies focused on the capability to finely modulate the scaffold architecture at the microscale and the nanoscale in order to closely mimic the hierarchical features of the extracellular matrix and, in particular, the natural patterning of collagen. All of these studies clearly show that the accurate characterization of the collagen structure at the submolecular and supramolecular levels is pivotal to the understanding of the relationships between the nanostructural/microstructural properties of the fabricated scaffold and its macroscopic performance. Several studies also demonstrate that the selected processing, including any crosslinking and/or sterilization treatments, can strongly affect the architecture of collagen at various length scales. The aim of this review is to highlight the most recent findings on the development of collagen-based scaffolds with optimized properties for tissue engineering. The optimization of the scaffolds is particularly related to the modulation of the collagen architecture, which, in turn, impacts on the achieved bioactivity

Keeping an eye on decellularized corneas: a review of methods, characterization and applications

The worldwide limited availability of suitable corneal donor tissue has led to the development of alternatives, including keratoprostheses (Kpros) and tissue engineered (TE) constructs. Despite advances in bioscaffold design, there is yet to be a corneal equivalent that effectively mimics both the native tissue ultrastructure and biomechanical properties. Human decellularized corneas (DCs) could offer a safe, sustainable source of corneal tissue, increasing the donor pool and potentially reducing the risk of immune rejection after corneal graft surgery. Appropriate, human-specific, decellularization techniques and high-resolution, non-destructive analysis systems are required to ensure reproducible outputs can be achieved. If robust treatment and characterization processes can be developed, DCs could offer a supplement to the donor corneal pool, alongside superior cell culture systems for pharmacology, toxicology and drug discovery studies

Optimisation and characterisation of human corneal stromal models

The native corneal structure is highly organised and unified in architecture with structural and functional integration which mediates its transparency and mechanical strength. Two of the most demanding challenges in corneal tissue engineering are the replication of the native corneal stromal architecture and the preservation of stromal cell phenotype which prevents scar-like tissue formation. A concerted effort in this thesis has been devoted to the generation of a functional human corneal stromal model by the manipulation of chemical, topographical and cellular cues. To achieve this, previously built non-destructive, online, real-time monitoring techniques, micro-indentation and optical coherence tomography (OCT), which allow for the mechanical and contraction properties of corneal equivalents to be monitored, have been improved. These macroscopic parameters have been cross-validated by histological, imunohistochemical, morphological and genetic expression analysis

Optimisation and characterisation of human corneal stromal models

The native corneal structure is highly organised and unified in architecture with structural and functional integration which mediates its transparency and mechanical strength. Two of the most demanding challenges in corneal tissue engineering are the replication of the native corneal stromal architecture and the preservation of stromal cell phenotype which prevents scar-like tissue formation. A concerted effort in this thesis has been devoted to the generation of a functional human corneal stromal model by the manipulation of chemical, topographical and cellular cues. To achieve this, previously built non-destructive, online, real-time monitoring techniques, micro-indentation and optical coherence tomography (OCT), which allow for the mechanical and contraction properties of corneal equivalents to be monitored, have been improved. These macroscopic parameters have been cross-validated by histological, imunohistochemical, morphological and genetic expression analysis

Tuneable 3D biocompatible scaffolds for biological and biophysical solid-tumour microenvironment studies; applications in Ovarian Cancer

Recently, three-dimensional (3D) tumour models mimicking the tumour microenvironment and reducing the use of experimental animals have been developed generating great interest to appraise tumour response to treatment strategies in cancer therapy. As tumours have distinct mechanics compared to normal tissues, biomaterials have also been utilized in 3D culture to model the mechanical properties of the tumour microenvironment, and to study the effects of extracellular matrix (ECM) mechanics on tumour development and progression. Mechanical cues regulate various cell behaviours through mechanotransduction, including proliferation, migration, and differentiation. In the context of cancer, both stromal cells (cancer associated fibroblasts) and tumour cells remodel the ECM and change its mechanical properties, and the altered mechanical niche in turn is likely to influence tumour progression. In this study, bovine derived collagen type I and Jellyfish derived marine collagen sources, were tested as biomaterial candidates for cancer studies, moulded to porous scaffolds with tuneable mechanical properties. The resulting interconnected network of collagen fibre constructs, fabricated using lyophilisation provide good control of scaffolding architecture, pore sizes range, high porosity levels, high level of cell viability and low production cost. Importantly these sponge scaffolds were, in the form of 3D models, compatible with a host of cellular and molecular biology assays used to investigate mechanical and biological effects of collagen crosslinking and (hyaluronic acid) HA inclusion on both fibroblasts and ovarian cancer cells. Stromal cells and cancer cells respond differently to the altered stiffness of their local microenvironment. Fibroblasts, once activated with TGF1, converge toward a ‘senescent-like phenotype’, blocking migration and matrix remodelling and promote tumour progression, probably through the secretion of tumour-promoting signals, in stiffer mechanical environments. Cancer cells, of both epithelial and mesenchymal phenotype, respond to increased local matrix stiffness by increasing proliferation while, at the same time, becoming more susceptible to treatment. Mechanically informative scaffolds resemble the physical characteristics of both normal and pathological ovarian tissue mechanics, where ovarian cancer originates. Physical changes observed in the later stage of ovarian cancer disease progression may therefore be fundamental for the increased cancer proliferation that drives metastatic progression, however opening an interesting window for cancer treatment. Bio-physical inclusive models not only lead the path to unveil complex interactions of biophysical and biological signals in the tumour microenvironment, but they represent a highly informative and effective platform to test novel target therapies with effective costs and high throughput. They can accommodate coculture systems and potentially patients-derived cell cultures, providing a platform to test current and new drugs and to evaluate drug efficacy following a precision medicine approach