Periodic Reporting for period 1 - NextSkins (Living Therapeutic and Regenerative Materials with Specialised Advanced Layers)
Periodo di rendicontazione: 2022-11-01 al 2023-10-31
The overall aim of NextSkins is to generate platform technologies for ELMs with layers of specialised properties and functions. This goal is to be achieved by new bio-encapsulation strategies and spatiotemporal control of material deposition over scales by engineered multicellular consortia. Two proof-of-concept ELMs will be delivered: a Living Therapeutical Skin (LTS), which is a hydrogel-based ELM of cellulose hosting sense-and-respond cells and covered with biomaterials layer preventing dehydration, and a Living Regenerative Skin (LRS), which is a mineral-polymer composite hosting bacterial spores and encapsulated in a mechanically sensitive biolayer for self-repair and local tuning of mechanical properties.
During the first year of the project, several synthesis biology tools have been developed. The yeast (S. cerevisiae) and Komagateibacter toolkits (YTK and KTK respectively) were distributed among project partners. An automated workflow, aligned with these genetic toolkits, was established. In addition, key proteins vital for Living Therapeutic Skin (LTS) structure (hydrophobins, ELPs, CBDs, SpyTag, and SpyCatcher domains) have been effectively secreted or surface-displayed by yeast. Significant progress has been made in the synthesis of hyaluronic acid and acetylated cellulose by K. rhaeticus with strain development now underway. For the Living Regenerative Skin (LRS) material, a genetically engineered B. subtilis strain with an IPTG inducible GFP gene has been used to enable visualization through GFP fluorescence.
We also worked on bimolecular engineering to push the bio-encapsulations strategies further. This has resulted in successful expression of three bacterial enzymes in S. cerevisiae. The expression system was based on the standard YTK kit. For optimising expression we have implemented a high throughput screening method with a yeast display procedure and executed it in an inter-lab setup with the FACS (fluorescent assisted cell sorting) device. Silk proteins have been expressed and we have analysed thin film formation at the air-water interface and tested different methods for film stabilisation for films on a solid support.
In terms of materials fabrication, although only some of the genetically engineered strains are ready, we still grew initial prototypes for the LTS and LRS materials. These materials are not functional yet, but they allow to demonstrate some of the expected material and biological properties.
To assess how potential users would experience the LTS material, we have created the initial mimics for LTS that simulate their potential tactile and responsive characteristics. These prototypes were utilized in user studies involving AD patients to gather preliminary feedback. Subsequently, the prototypes have undergone further development for presentation at the workshop with Advisory Board members, where we discussed the feasibility, desirability, and potential social impact of the application ideas.
To assess the potential applications of the LRS material, we have conducted a literature and practice review on the use of biomineralization in design. This resulted in a taxonomy, which we will use for selecting application ideas in the next phase.
We also worked on bimolecular engineering to push the bio-encapsulations strategies further. This has resulted in successful expression of three bacterial enzymes in S. cerevisiae. The expression system was based on the standard YTK kit. For optimising expression we have implemented a high throughput screening method with a yeast display procedure and executed it in an inter-lab setup with the FACS (fluorescent assisted cell sorting) device. Silk proteins have been expressed and we have analysed thin film formation at the air-water interface and tested different methods for film stabilisation for films on a solid support.
In terms of materials fabrication, although only some of the genetically engineered strains are ready, we still grew initial prototypes for the LTS and LRS materials. These materials are not functional yet, but they allow to demonstrate some of the expected material and biological properties.
To assess how potential users would experience the LTS material, we have created the initial mimics for LTS that simulate their potential tactile and responsive characteristics. These prototypes were utilized in user studies involving AD patients to gather preliminary feedback. Subsequently, the prototypes have undergone further development for presentation at the workshop with Advisory Board members, where we discussed the feasibility, desirability, and potential social impact of the application ideas.
To assess the potential applications of the LRS material, we have conducted a literature and practice review on the use of biomineralization in design. This resulted in a taxonomy, which we will use for selecting application ideas in the next phase.