H.V. Unadkat

An algorithm-based topographical biomaterials library to instruct cell fate

It is increasingly recognized that material surface topography is able to evoke specific cellular responses, endowing materials with instructive properties that were formerly reserved for growth factors. This opens the window to improve upon, in a cost-effective manner, biological performance of any surface used in the human body. Unfortunately, the interplay between surface topographies and cell behavior is complex and still incompletely understood. Rational approaches to search for bioactive surfaces will therefore omit previously unperceived interactions. Hence, in the present study, we use mathematical algorithms to design nonbiased, random surface features and produce chips of poly(lactic acid) with 2,176 different topographies. With human mesenchymal stromal cells (hMSCs) grown on the chips and using high-content imaging, we reveal unique, formerly unknown, surface topographies that are able to induce MSC proliferation or osteogenic differentiation. Moreover, we correlate parameters of the mathematical algorithms to cellular responses, which yield novel design criteria for these particular parameters. In conclusion, we demonstrate that randomized libraries of surface topographies can be broadly applied to unravel the interplay between cells and surface topography and to find improved material surfaces.

Analysis of high-throughput screening reveals the effect of surface topographies on cellular morphology

Surface topographies of materials considerably impact cellular behavior as they have been shown to affect cell growth, provide cell guidance, and even induce cell differentiation. Consequently, for successful application in tissue engineering, the contact interface of biomaterials needs to be optimized to induce the required cell behavior. However, a rational design of biomaterial surfaces is severely hampered because knowledge is lacking on the underlying biological mechanisms. Therefore, we previously developed a high-throughput screening device (TopoChip) that measures cell responses to large libraries of parameterized topographical material surfaces. Here, we introduce a computational analysis of high-throughput materiome data to capture the relationship between the surface topographies of materials and cellular morphology. We apply robust statistical techniques to find surface topographies that best promote a certain specified cellular response. By augmenting surface screening with data-driven modeling, we determine which properties of the surface topographies influence the morphological properties of the cells. With this information, we build models that predict the cellular response to surface topographies that have not yet been measured. We analyze cellular morphology on 2176 surfaces, and find that the surface topography significantly affects various cellular properties, including the roundness and size of the nucleus, as well as the perimeter and orientation of the cells. Our learned models capture and accurately predict these relationships and reveal a spectrum of topographies that induce various levels of cellular morphologies. Taken together, this novel approach of high-throughput screening of materials and subsequent analysis opens up possibilities for a rational design of biomaterial surfaces.

Fabrication of cell container arrays with overlaid surface topographies.

This paper presents cell culture substrates in the form of microcontainer arrays with overlaid surface topographies, and a technology for their fabrication. The new fabrication technology is based on microscale thermoforming of thin polymer films whose surfaces are topographically prepatterned on a micro- or nanoscale. For microthermoforming, we apply a new process on the basis of temporary back moulding of polymer films and use the novel concept of a perforated-sheet-like mould. Thermal micro- or nanoimprinting is applied for prepatterning. The novel cell container arrays are fabricated from polylactic acid (PLA) films. The thin-walled microcontainer structures have the shape of a spherical calotte merging into a hexagonal shape at their upper circumferential edges. In the arrays, the cell containers are arranged densely packed in honeycomb fashion. The inner surfaces of the highly curved container walls are provided with various topographical micro- and nanopatterns. For a first validation of the microcontainer arrays as in vitro cell culture substrates, C2C12 mouse premyoblasts are cultured in containers with microgrooved surfaces and shown to align along the grooves in the three-dimensional film substrates. In future stem-cell-biological and tissue engineering applications, microcontainers fabricated using the proposed technology may act as geometrically defined artificial microenvironments or niches.

Development of multilayer constructs for tissue engineering

The rapidly developing field of tissue engineering produces living substitutes that restore, maintain or improve the function of tissues or organs. In contrast to standard therapies, the engineered products become integrated within the patient, affording a potentially permanent and specific cure of the disease, injury or impairment. Despite the great progress in the field, development of clinically relevantly sized tissues with complex architecture remains a great challenge. This is mostly due to limitations of nutrient and oxygen delivery to the cells and limited availability of scaffolds that can mimic the complex tissue architecture. This study presents the development of a multilayer tissue construct by rolling pre‐seeded electrospun sheets [(prepared from poly (l‐lactic acid) (PLLA) seeded with C2C12 pre‐myoblast cells)] around a porous multibore hollow fibre (HF) membrane and its testing using a bioreactor. Important elements of this study are: 1) the medium permeating through the porous walls of multibore HF acts as an additional source of nutrients and oxygen to the cells, which exerts low shear stress (controllable by trans membrane pressure); 2) application of dynamic perfusion through the HF lumen and around the 3D construct to achieve high cell proliferation and homogenous cell distribution across the layers, and 3) cell migration occurs within the multilayer construct (shown using pre‐labeled C2C12 cells), illustrating the potential of using this concept for developing thick and more complex tissues. Copyright

A modular versatile chip carrier for high-throughput screening of cell–biomaterial interactions

The field of biomaterials research is witnessing a steady rise in high-throughput screening approaches, comprising arrays of materials of different physico-chemical composition in a chip format. Even though the cell arrays provide many benefits in terms of throughput, they also bring new challenges. One of them is the establishment of robust homogeneous cell seeding techniques and strong control over cell culture, especially for long time periods. To meet these demands, seeding cells with low variation per tester area is required, in addition to robust cell culture parameters. In this study, we describe the development of a modular chip carrier which represents an important step in standardizing cell seeding and cell culture conditions in array formats. Our carrier allows flexible and controlled cell seeding and subsequent cell culture using dynamic perfusion. To demonstrate the application of our device, we successfully cultured and evaluated C2C12 premyoblast cell viability under dynamic conditions for a period of 5 days using an automated pipeline for image acquisition and analysis. In addition, using computational fluid dynamics, lactate and BMP-2 as model molecules, we estimated that there is good exchange of nutrients and metabolites with the flowing medium, whereas no cross-talk between adjacent TestUnits should be expected. Moreover, the shear stresses to the cells can be tailored uniformly over the entire chip area. Based on these findings, we believe our chip carrier may be a versatile tool for high-throughput cell experiments in biomaterials sciences.

Microfabrication techniques in materiomics

Scope: This chapter deals with an overview of basic microfabrication techniques. The goal is to explain to the reader how such techniques can be utilized in the field of materiomics. The basic processes used in microfabrication including photolithography, etching, electron beam lithography and micromoulding are explained. Some classic examples of these techniques as applied to materiomics are also shown. Furthermore, possible uses of such techniques, and the development and application of hybrid techniques to be able to answer fundamental questions about biological behaviour of materials, are also suggested. Basic principles of microfabrication: Introduction: Techniques used to fabricate structures or devices smaller than 100 µm are commonly referred to as microfabrication techniques. Initially meant for the electronics industry, they have found a wide range of applications in diverse fields such as chemical engineering and the life sciences. Since the early 1990s, the application of microfabrication technologies in the area of chemical and biological analysis has been termed ‘micro total analysis systems’ (µTAS) (1). Microfabricated devices meant for µTAS initially offered the advantage of sample analysis on the microscale, but over the years, the evolution of these technologies has led to the facilitation of sample preparation, fluid handling, separation systems, cell handling and cell culturing in an integrated manner (1). The application of microtechnologies for the fabrication of devices or systems to study material properties benefits from cost efficiency, high performance, precision-based design flexibility, miniaturization and automated analysis. Miniaturization involves the convergence of multiple disciplines, such as fluid dynamics, material sciences, engineering and the life sciences, that need to join expertise in order to design functional systems. Moreover, these devices can be used to evaluate biological behaviour in vitro and can help us to test thousands of different biomaterials and surface properties without the complexity related to in vivo assays.

High-throughput screening of cell-surface topographic interactions

In the present study we describe a high throughput screening approach to screen the biological interactions of cells with surface topographies created on biomaterial substrates.