Three-dimensional bioprinting of embryonic stem cells directs highly uniform embryoid body formation

Abstract

With the ability to manipulate cells temporarily and spatially into three-dimensional (3D) tissue-like construct, 3D bioprinting technology was used in many studies to facilitate the recreation of complex cell niche and/or to better understand the regulation of stem cell proliferation and differentiation by cellular microenvironment factors. Embryonic stem cells (ESCs) have the capacity to differentiate into any specialized cell type of the animal body, generally via the formation of embryoid body (EB), which mimics the early stages of embryogenesis. In this study, extrusion-based 3D bioprinting technology was utilized for biofabricating ESCs into 3D cell-laden construct. The influence of 3D printing parameters on ESC viability, proliferation, maintenance of pluripotency and the rule of EB formation was systematically studied in this work. Results demonstrated that ESCs were successfully printed with hydrogel into 3D macroporous construct. Upon process optimization, about 90% ESCs remained alive after the process of bioprinting and cell-laden construct formation. ESCs continued proliferating into spheroid EBs in the hydrogel construct, while retaining the protein expression and gene expression of pluripotent markers, like octamer binding transcription factor 4, stage specific embryonic antigen 1 and Nanog. In this novel technology, EBs were formed through cell proliferation instead of aggregation, and the quantity of EBs was tuned by the initial cell density in the 3D bioprinting process. This study introduces the 3D bioprinting of ESCs into a 3D cell-laden hydrogel construct for the first time and showed the production of uniform, pluripotent, high-throughput and size-controllable EBs, which indicated strong potential in ESC large scale expansion, stem cell regulation and fabrication of tissue-like structure and drug screening studies.

3D Bioprinting for Tissue and Organ Fabrication

“…Although challenges still present, with new niches for technological developments on the instrumentation with improved spatial and temporal resolutions as well as optimized bioinks and cell sources for specific organs, it is expected that 3D bioprinting will eventually become one of the most efficient, reliable, and convenient methods to biofabricate tissue constructs in the near future. Combination with the stem cell technologies 124-126 and advanced materials engineering approaches featuring stimuli-responsiveness 127, 128 will further allow temporal evolution of bioprinted tissue constructs that potentially meet the requirements of dynamic tissue remodeling during developmental processes.…”

Bioimpressão e produção de mini-órgãos com células tronco

“…Uma pesquisa realizada por Ouyang et al (2015) mostrou que células-tronco embrionárias foram impressas com sucesso utilizando um hidrogel em macroporo 3D. Segundo os autores, esses resultados indicam um forte potencial de expansão em larga escala para regulação de células-tronco e estrutura da fabricação de tecidos vivos e estudos de triagem de drogas.…”

3D bioprinting technology for regenerative medicine applications

“…This approach may be helpful to generate patient-specific 3D liver constructs using IPSCs for drug screening and organ transplantation [107] . In another study, 3D bioprinting technology was used to create ES cells into 3D hydrogel spheroids to maintain the stem cell pluripotency [109] . These spheroids were made from gelatin and alginate.…”

“…In this method, ES cells laden hydrogel spheroids with controlled size and uniform pluripotency were bioprinted using an extrusion-based 3D bioprinter. The cell spheroids were shown to retain pluripotent stem cell markers such as Oct 4, SSEA-1 and Nanog [109] . In another study, a novel bioink made from ultrashort peptide hydrogels were used to bioprint 3D structures encapsulated with human embryonic stem cells.…”