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International Journal of Bioprinting Droplet-based bioprinting of tumor spheroids

with changes to matrix microenvironments. Controlled frequently used cell lines in the field of OSCC study. The
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fabrication of hESC spheroids is challenging. Faulkner- extracted CAL27 cell line possesses different mutations
Jones et al. successfully employed microvalve-based due to the influence of donor’s dietary habits. Recently,
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bioprinting technology to achieve the fabrication of hESC researchers have attempted to construct in vitro oral tumor
spheroids, leveraging the low shear stress to cells during diffusion and metastasis model utilizing 3D bioprinting
printing (Figure 3E). After 3-day culture, these hESC technology. Chen et al. utilized an acoustic bioprinter,
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spheroids showed a viability of over 89% and remained which adopts the nozzle-free printing technology, to
positive for the Oct-4 pluripotency marker, indicating good print high-concentration CAL27 cells (1 × 10 cells/ml)
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pluripotency and differentiation ability of hESCs. They (Figure 3H). These cell clusters grew into spheroids with
found that each printed droplet contained ~28 cells, and a diameter of ~100 μm. They also fabricated tumor
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the bioprinting ensured that spheroids had a controllable spheroid-stroma models by printing (CAL27)–CAFs co-
size and sphericity. This fabrication by bioprinting was culture microenvironments to mimic the tumor invasion
automated and demonstrated high throughput with ~40 in vivo. After 3-day co-culture, CAFs were recruited and
spheroids per second, guaranteeing downstream analysis of aggregated, and then a dense 3D network was formed
tumor formation and drug testing. surrounding the CAL27 spheroids. Tumor cell migration
and invasion significantly occurred on day 5.
3.5. Breast tumor spheroids
The pathogenesis of breast cancer still remains obscure. 4. Applications of tumor models with
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Many genes (BRCA1/2, HER2, EGFR, c-Myc, etc.) have
been reported to be related to the development of breast spheroids
cancer. Besides, tumor microenvironment (stochastic While tumor spheroids have been applied for many
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theory) and breast cancer stem cells (CSCs) are suggested applications such as in vitro simulation of tumor cell
as the most likely causes. To understand and treat this proliferation, invasion, hypoxia, and necrosis, this area is
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disease, a reliable and high-throughput platform for still in its infancy. The function of spheroids is impacted
fabrication of breast tumor spheroids is needed. Ling by their size, sphericity, compactness as well as the
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et al. prepared bioink by mixing the MCF-7 cells with surrounding environment. That said, spheroids need to
gelatin matrix at a concentration of 1 × 10 cells/ml and be controlled for direct usage in drug testing and also can
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fabricated breast tumor spheroids by printing MCF-7 cells be co-cultured in a more complicated microenvironment
with microvalve-based bioprinting technology into the to model disease. In vivo, the formation of tumors is a
hydrogel microwells (Figure 3F). Owing to low damage complex and long-term process, and its functions are
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to cells, obtained spheroids exhibited good cell viability induced by the interactions between tumor cells and
(above 95% after 7 days) and morphology. 86 surrounding normal histiocytes, blood vessels, immune
3.6. Colorectal tumor spheroids cells, fibroblasts, bone marrow-derived inflammatory
Colorectal cancer (CRC) is one of the most widespread cells, various signal molecules, and ECM, which are
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malignant tumors worldwide. SW620 and SW480, named as tumor microenvironment (TME). TME
isolated from CRC tumors, are the widely utilized CRC provides an environment for tumor growth, and in
tumor cell lines. Influenced by intrinsic factors (genetic response, tumor further changes this microenvironment
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and epigenetic alterations) and lifestyle factors, CRC by releasing cell signaling molecules as they grow. In
is heterogeneous and exhibits different levels of drug this section, we aim to give a review of representative
resistance. Johnson et al. fabricated the CRC spheroids applications of tumor spheroids.
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by printing the gelatin–alginate bioink containing SW620 4.1. Modeling tumor heterogeneity
cells (Figure 3G). The cell clusters grew into spheroids Heterogeneity is one typical characteristic of tumor cells.
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with a diameter of ~200 μm after 12-day culture. These Different from cancer progenitor cell, tumor tissue exhibits
spheroids were round and tightly packed, indicating strong
cell–cell interactions. Hypoxia cores were also observed distinct levels of gene expression, metabolism, motility,
in large spheroids. The cell density in the core area was proliferation, and metastasis. To fabricate a spheroid
significantly lower than that in the external area, owing to mimicking the tumor heterogeneity, considerations include
hypoxia. These spheroids can be utilized for anti-cancer control of the morphology and structure, and sensitive
drug screening. and accurate measurement of parameters (oxygen content,
protein synthesis, and cytokine expression) in TME.
3.7. Oral tumor spheroids Multicellular spheroids are needed to explore the influences
Oral squamous cell carcinoma (OSCC) is the most of cell composition on spheroid function. Zhang et al.
common oral cancer. CAL27 is one of the most fabricated the liver spheroids with HepG2 and HSC cells,
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Volume 10 Issue 1 (2024) 115 https://doi.org/10.36922/ijb.1214