(460d) Algorithmic Design of Lymph Node-Inspired 3D Microprinted Cell Scaffolds

Background: Immune cells, such as T cells, interact with a range of physical networks within the human body. For instance, networks formed by interconnected fibroblastic reticular cells (FRCs) in lymph nodes are a key determinant of immune responses and play a critical role in guiding immune cells through 3D space to exchange information. Deciphering how such microscale networks physically interact with immune cells carries the potential of improving cancer immunotherapies and lymph node tissue engineering. However, the influence of network topology and geometry on immune cell migration and decision-making remains elusive, as conventional study methods typically involve cell culture on 2D surfaces. While 3D scaffold fabrication methods exist, many of them lack the high precision required to control network architecture at the microscale. Thus, there is a need for alternative design and fabrication approaches to create 3D culture platforms enabling control of microarchitectural complexity.

Methods: We employed a nature-inspired engineering approach [1–3], where we took inspiration from the small-world topology of FRC networks and created an algorithmic design pipeline to generate 3D printable cell scaffolds. We used a random geometric graph model to create the scaffold skeleton network, before skinning it with a mesh and smoothing in computer-aided design. The networks were 3D microprinted in an acrylic resin using two-photon polymerisation direct laser writing (2PP DLW). To demonstrate biocompatibility, we seeded the scaffolds with FRCs and used confocal imaging to visualise cell-scaffold interactions.

Results: Network analysis showed that our scaffolds were able to recapitulate the lattice-like, small-world organisation of FRC networks. We found that the original network model would generate unprintable structures in the form of floating islands and topologically inaccurate networks with intersecting edges. These issues were remedied with a modified algorithm that placed additional spatial constraints on the generation process. From our optimisation study, we found that the laser power used in 2PP DLW was crucial to the structural integrity of the 3D microprinted scaffolds. Confocal imaging results showed that FRCs successfully adhered to the scaffolds and infiltrated their interior.

Conclusions: In this work, we have developed a novel approach combining algorithmic design, graph theory and 3D microprinting in the creation of 3D cell scaffolds. Our preliminary study showed that the printed scaffolds were suitable for FRC culture. We envision that this platform would allow immunological studies that not only consider geometric characteristics of microenvironmental networks, but also their topological properties. Specifically, the platform could potentially be used as an in vitro artificial lymph node or adapted to investigate how T cells migrate through complex microenvironments in cancer immunotherapies.

References:

[1] M.-O. Coppens, Nature-Inspired Chemical Engineering for Process Intensification, Annu Rev Chem Biomol Eng. 12 (2021) 187–215. https://doi.org/10.1146/annurev-chembioeng-060718-030249.

[2] E.C. Goldfield, M.-O. Coppens, Developmental bioengineering: recapitulating development for repair, Mol Syst Des Eng. 5 (2020) 1168–1180. https://doi.org/10.1039/D0ME00062K.

[3] M.H.W. Chin, E. Gentleman, M.-O. Coppens, R.M. Day, Rethinking Cancer Immunotherapy by Embracing and Engineering Complexity, Trends Biotechnol. 38 (2020) 1054–1065. https://doi.org/10.1016/j.tibtech.2020.05.003.