Highly Biomimetic Proregenerative Scaffold for Personalized Nerve Repair

Project Information

ProRegScaffold

Grant agreement ID: 101028152

DOI

10.3030/101028152

Project terminated on 1 September 2022

EC signature date 20 April 2021

Start date 1 August 2021

End date 31 July 2023

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 191 852,16

EU contribution

€ 191 852,16

Coordinated by

KAROLINSKA INSTITUTET
Sweden

Periodic Reporting for period 1 - ProRegScaffold (Highly Biomimetic Proregenerative Scaffold for Personalized Nerve Repair)

Reporting period: 2021-08-01 to 2023-07-31

Summary of the context and overall objectives of the project

Peripheral nerve injuries, which are common and can cause debilitating sensory sensing and motor dysfunction, pose challenges for tissue engineering and regenerative medicine as neural functions depend on their complex 3D structures and anisotropic mechanical properties. Peripheral nerve tissues include complicated nerve fibers with intricate microscale topography and highly hierarchical geometric structures ranging from micrometer to centimeter levels. Artificial scaffolds are promising alternatives to current gold-standard autograft methodologies without availability limitations and donor site morbidity. A desirable characteristic of these artificial scaffolds is the ability to allow the ingress of cells and control their subsequent differentiation and proliferation within the scaffolds. However, the current engineering technologies are limited in creating hierarchically ordered structures ranging from centimeter to micrometer spatial resolution. Additionally, increased extracellular calcium ion (Ca2+) levels can increase mitochondrial motility, which is necessary for the regulation of Wallerian degeneration and thus accelerate nerve repair. Ca2+ has been implicated as a key messenger in the guidance of developing axons. The concentration of extracellular Ca2+ can mediate the axonal growth cone and its turning behavior. A local, controlled accumulation of released Ca2+ has been shown beneficial for long-term peripheral nerve regeneration.

The incorporation of ice-templating and 3D printing technology was implemented to fabricate personalized scaffolds for nerve repair, which can effectively guide the extension of neurites at the single-cell level. The channel sizes of the scaffolds could be easily tuned via annealing in the presence of removable molecules, which addresses the demands for anisotropic scaffolds with precisely tunable porosity. Compared with reported scaffolds and methods, the scaffolds fabricated during this project show high spatial resolution and designed geometry that mimic the complex constructures of native nerve tissues. Furthermore, I developed a NIR-responsive drug loading system based on gelatin/polypyrrole (GB@PPy) nanoparticles for controllable Ca2+ release. Under NIR irradiation, integrated GB@PPy was able to efficiently release Ca2+ to accelerate the growth of neurons.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The following summarizes the main research tasks and results:
1. Channel size of the 3D chitosan matrix can be mediated by ice growth regulators.
1.1 Large-scale manufacturing of scaffolds could thus be implemented by moving the cooling center forward to retain the distance between the advancing ice/water interface and the cooling center.
1.2 The channel size could be precisely regulated by annealing the frozen chitosan matrices containing 0.1 M NaCl and 0.1 M Na2SO4 at −3 °C for 1 h after the unidirectional freezing.
2. The as prepared scaffolds showed anisotropic mechanical properties by filling the channels of the chitosan matrix with different hydrogel fillers. The oriented hydrogels attenuate friction in one direction, thus enhancing the toughness and strength of the overall scaffolds.
3. Hybrid scaffolds can guide the outgrowth of axons at a single-cell level. The single cell alignment can be facilitated by implementing a low adhesive skeleton structure embedded with a filler mediating cell attachment.
4. The prepared scaffolds can guide the growth of different types of neural cells. I demonstrated the versatility of the scaffolds to guide the alignment of these various types of neural cells, including dorsal root ganglia cells (ND7/23), SH-SY5Y cells and Schwann cells.
5. The integration of the ice-templating stereolithography technique showed great potential to fabricate personalized multi-lumen scaffolds.
6. I developed GB@PPy nanoparticles to serve as a responsive drug loading system. I then developed polypyrrole-modified gelatin type B nanoparticles (GB@PPy) preloaded Ca2+ to regulate local concentration of this ion.

Two first-author publications related to this project are in preparation for submission. I am the first author of a review paper on noble metal nanoparticles (Geng et al. Accounts of Chemical Research) and the first author of a paper that has been submitted to Angewandte Chemie related to NIR responsive nanomaterials (manuscript ID, 202217118). These publications will acknowledge European Commission funding and comply with EU open access policies. To disseminate the outcomes of the project, I presented a talk entitled “Highly biomimetic scaffolds for personalized nerve repair” in the quarter seminar on 06/05/2022. I participated in the KI 5th Biomedicum-BioClinicum Young Researchers Symposium (09/11/2021) and attended the lecture “Overcoming barriers: local delivery in the central nervous system” (21/06/2022). I presented the progress of this project several times in internal Stevens group meetings (25/08/2021, 24/11/2021, 22/09/2021, 16/03/2022, and 08/06/2022). I have begun my new position as an Assistant Professor at Tsinghua University, China, whereby I will continue to disseminate the results of my research and exploit the applications of the techniques developed in this project.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The state-of-art techniques of nerve tissue engineering focus on mimicking the anisotropic structures of natural nerve tissues via engineering biomaterials that resemble native tissues. Ice-templating holds promise to fabricate 3D matrices with ordered architectures. However, its potential to elicit highly personalized scaffolds without compromising precision at the micro- and macro-level has yet to be fully developed. This project developed versatile fabrication techniques by combining ice-templating methodology and 3D printing technology to manufacture scaffolds that guide the growth of neurons at the single-cell level. Conjugation of the ice-templating method and 3D printing technique serves as a potentially ideal method to precisely prepare scaffolds to meet personalized requirements. This technique will potentially enhance the medical industry by fabricating scaffolds that look realistic and replicate natural human nerve tissues. I tested a library of compounds that could be used to further control the size distribution of the channels. Tuning the porosity of artificial scaffolds the traditional way is tedious because they need complicated processes or the fabrication of templates. The library of compounds and the ice-templating method provide the freedom to design shape- and pore-controlled scaffolds for personalized treatment. Such a versatile fabrication technique is distinct from the conventional methodologies used in scaffold manufacture; wherein large-scale manufacturing could be easily conducted by printing customized molds.
Photothermal induced drug release has been demonstrated in successful trials. The utilization of NIR irradiation, in particular, the second near-infrared (NIR-Ⅱ) to control drug release in vivo makes it possible to control the pharmacokinetics in a non-invasive format. These semiconductive polymer coated gelatin nanoparticles intrinsically possessed excellent photothermal stability, which was superior to some commercially available small molecules that are fragile to light irradiation. This MSCA project has thus provided a technique that will complement traditional methods to achieve precise nerve repair.

Schematic illustration of ProRegScaffold fabrication with the ice-templating technique