easy-manipulation-architectures-protein-based-hydrogels-for-cell

Easy Manipulation of Architectures in Protein-based Hydrogels for Cell Culture Applications

Hydrogels are recognized as promising materials for cell culture applications due to their ability to provide highly hydrated cell environments. The field ...

Ulm University

Here, we present a novel approach to form hydrogels from yeast whole cell protein. Countless hydrogels are available for sophisticated research, but their fabrication is often difficult to reproduce, with the gels being complicated to handle or simply too expensive. The yeast hydrogels presented here are polymerized using a four-armed, amine reactive crosslinker and show a high chemical and thermal resistance. The free water content was determined by measuring swelling ratios for different protein concentrations, and in a freeze-drying approach, pore sizes of up to 100 μm in the gel could be created without destabilizing the 3D network. Elasticity was proofed to be adjustable with the help of atomic force microscopy by merely changing the amount of used protein. Furthermore, the material was tested for possible cell culture applications; diffusion rates in the network are high enough for sufficient supply of human breast cancer cells and adenocarcinomic human alveolar basal epithelial cells with nutrition, and cells showed high viabilities when tested for compatibility with the material. Furthermore, hydrogels could be functionalized with RGD peptide and the optimal concentration for sufficient cell adhesion was determined to be 150 μM. Given that yeast protein is one of the cheapest and easiest available protein sources and that hydrogels are extremely easy to handle, the developed material has highly promising potential for both sophisticated cell culture techniques as well as for larger scale industrial applications.

Beyond bread and beer: whole cell protein extracts from baker's yeast as a bulk source for 3D cell culture matrices.

Different methods to create and manipulate pore sizes in hydrogel fabrication are available, but systematic studies are normally conducted with hydrogels made of synthetic chemical compounds as backbones. In this study, a hydrogel made of natural and abundant protein in combination with different, well-available techniques was used to produce different architectures within the hydrogel matrix. Pore sizes and distribution are compared and resulting hydrogel properties like swelling ratio, resistance towards external stimuli and enzymatic degradation were investigated. Porous hydrogels were functionalized and two cancer cell lines were successfully adhered onto the material. With simple methods, pores with a radius between 10 and 80 μm and channels of 25 μm radius with a length of several hundreds of μm could be created and analyzed with laser scanning confocal microscopy and electron microscopy respectively. Furthermore, the influence of different methods on swelling ratio, enzymatic degradation and pH and temperature resistance was observed.

Evaluation of methods for pore generation and their influence on physio-chemical properties of a protein based hydrogel.

3D cell culture is a helpful approach to study cell-cell interaction in a native-like environment, but is often limited due the challenge of retrieving cells from the material. In this study, we present the use of recombinant lectin B, a sugar-binding protein with four binding cavities, to enable reversible cell integration into a macroporous protein hydrogel matrix. By functionalizing hydrogel precursors with saccharose, lectin B can both bind to sugar moieties on the cellular surface as well as to the modified hydrogel network. Confocal microscopy and flow cytometry analysis revealed cells to be integrated into the network and to adhere and proliferate. Furthermore, the specificity and reversibility was investigated by using a recombinantly produced yellow fluorescent - lectin B fusion protein and a variety of sugars with diverging affinities for lectin B at different concentrations and elution times. Cells could be eluted within minutes by addition of L-fucose to the cell-loaded hydrogels to make cells available for further analysis.

Article	Year
Beyond bread and beer: whole cell protein extracts from baker's yeast as a bulk source for 3D cell culture matrices. Applied microbiology and biotechnology\| PubMed ID: 27864602	2017
Evaluation of methods for pore generation and their influence on physio-chemical properties of a protein based hydrogel. Biotechnology reports (Amsterdam, Netherlands)\| PubMed ID: 28352549	2016
Lectin-mediated reversible immobilization of human cells into a glycosylated macroporous protein hydrogel as a cell culture matrix. Scientific reports\| PubMed ID: 28733655	2017

Dennis Kubiczek

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