AccScience Publishing / IJB / Online First / DOI: 10.36922/IJB026270284
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RESEARCH ARTICLE
Early Access

Fabrication of prevascularized co-culture spheroids and their 3D bioprinting application

Daniel S. Alt1 V. Ivy Petsinger1 Oju Jeon2 Aixiang Ding2 Eben Alsberg1,2,3,4,5,6,*
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1 Department of Biomedical Engineering, Case Western Reserve University, Cleveland, OH, 44106 , United States of America
2 Department of Biomedical Engineering, University of Illinois Chicago, Chicago, IL, 60612 , United States of America
3 Department of Orthopaedic Surgery, Case Western Reserve University, Cleveland, OH, 44106 , United States of America
4 National Center for Regenerative Medicine, Case Western Reserve University, Cleveland, OH, 44106 , United States of America
5 Departments of Orthopaedic Surgery, Mechanical and Industrial Engineering, and Pharmacology and Regenerative Medicine, University of Illinois Chicago, Chicago, IL, 60612 , United States of America
6 Jesse Brown Veterans Affairs Medical Center (JBVAMC), Chicago, IL 60612 , United States of America
Received: 1 July 2026 | Revised: 27 August 2026 | Accepted: 2 September 2026 | Published online: 7 September 2026
© 2026 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
Abstract

While the field of tissue engineering holds great promise to address the critical shortage of tissues/organs for transplantation, a key challenge that remains is vascularizing large constructs. Presented herein is a strategy to develop a prevascular network within engineered tissues to facilitate early anastomosis with host vasculature and perfusion. A co-culture system was established that uses endothelial cells and supporting mesenchymal stem cells to generate aggregates of cells containing a network of prevascular cord and plexus-like structures reminiscent of embryonic vasculogenesis. Aggregates comprised of 50,000 cells were 537 µm in diameter, just beyond the limit of oxygen diffusion and thus appropriate for a vascularization study. Fibroblast growth factor 2 (FGF-2) was then shown to be a sufficient media supplement to enhance early vasculogenesis within the first week. A bioink of these prevascular aggregates in fibrinogen was then 3D printed into a thrombin-containing hydrogel slurry temporary support, resulting in fibrin encapsulated prevascular aggregates. These prevascular structures were shown to persist for 7 days of tissue culture. A minimum pre-printing culture time of 7 days was demonstrated to maintain prevascular structures post-printing. This system allows for the precise spatial control of prevascularized constructs that may ultimately enhance microvascularization upon transplantation in vivo.

Keywords
Tissue engineering
Prevascularization
Vasculogenesis
Aggregates
Bioink
Slurry
3D printing
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International Journal of Bioprinting, Electronic ISSN: 2424-8002 Print ISSN: 2424-7723, Published by AccScience Publishing