Post-printing structure formation in bioprinted tissue constructs that mimic the tumor microenvironment

Jun 28, 2022, 2:00 PM
Room: S1

Room: S1


Neagu, Adrian (Victor Babes University of Medicine and Pharmacy Timisoara )


Understanding the key players in cancer progression is essential for the development of effective therapies. Aiming to pinpoint the roles of biochemical and biophysical factors involved in malignancy, tissue engineers developed in vitro cancer models of increasing complexity [1]. Three-dimensional (3D) bioprinting techniques were extensively used in this endeavor [2,3] due to their ability to create biomimetic spatial patterning of several cell types that coexist with cancer cells in the tumor microenvironment (TME), including tumor-associated fibroblasts, immune cells, mesenchymal stem cells, adipocytes, and vascular cells [4]. Nevertheless, bioprinted cells rarely remain where the bioprinter delivers them; they remodel their extracellular matrix and take advantage of their motility to establish firm bonds with other cells and/or biomaterials [5]. Therefore, in the present work, we investigated structure formation in bioprinted models of the TME both experimentally and computationally [6]. We used extrusion-based bioprinters to build models of the TME. SK-BR-3 breast cancer cells dispersed in a hydrogel droplet (106 cells/mL) were embedded in the same hydrogel (Bioink, CELLINK, Sweeden) loaded with tumor associated fibroblasts (TAFs − 5×105 cells/mL) and peripheral blood mononuclear cells (PBMCs − 5×105 cells/mL) harvested from breast cancer patients. The bioprinted tissue constructs were cultured in vitro for two weeks and cryosectioned for histological evaluation. Hoechst staining demonstrated that cells remained viable and remodeled the hydrogel. Hematoxylin and eosin (H&E) staining of histological sections, prepared at various time points, indicated that cells proliferated and formed heterotypic aggregates of malignant and peritumoral cells. To investigate the interactions responsible for the observed phenomena, we built lattice models of the bioprinted constructs and simulated their evolution using Metropolis Monte Carlo methods [6]. The computational model was formulated on a 3D cubic lattice, representing the biological system, at single-cell resolution, in terms of 4 types of particles: tumor cells, peritumoral cells, volume elements of the hydrogel, and volume elements of the cell culture medium. Based on the differential adhesion hypothesis, computer simulations reproduced most features of the experimentally observed structure formation, but did not account for the superficial localization of the heterotypic aggregates. Depending on model parameters, peritumoral cells wrapped or infiltrated cancer cell aggregates, as expected from TAFs and immune cells, respectively. Despite their limited complexity, the tissue constructs developed in this study could be used to establish co-culture conditions for cancer cells, TAFs, and PBMCs. Future investigations should consider model tissues incorporating perfusable channels with endothelial cell lining. Also, the computational model needs to be extended to describe the self-assembly of different cell types present in the native TME.

[1] Bray, L.J., Hutmacher, D.W., Bock, N., Front. Bioeng. Biotechnol. 7, 217 (2019)
[2] Li, J., Parra-Cantu, C., Wang, Z., Zhang, Y.S., Trends Cancer 6, 745–756 (2020)
[3] Datta, P., Dey, M., Ataie, Z., Unutmaz, D., Ozbolat, I.T., Precision Oncology, 4, 18 (2020)
[4] Turley, S.J., Cremasco, V., Astarita, J.L., Nat. Rev. Immunol. 15, 669–682 (2015)
[5] Robu, A., Aldea, R. et al., BioSystems 109, 430–443 (2012)
[6] Bojin, F., et al., Micromachines 12, 535 (2021)


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