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International Journal of Bioprinting Droplet-based bioprinting of tumor spheroids

3. Tumor spheroids fabricated by body, fabricated by this technology in this section (see
droplet-based bioprinting Figure 3 and Table 2).

The in vitro tumor models have their origins in 2D cell 3.1. Neuroblastoma spheroids
culture and have been developed since the 1900s. Initially, Neuroblastoma is an extracranial tumor with significant
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it was established with a sufficiently simple subunit system heterogeneity. One notable feature of neuroblastoma
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via culturing monolayer tumor cells on a flat petri dish or is migration. Owing to these properties, it is very
culture flask. However, 2D cell culture cannot recapitulate difficult to treat this disease. Moreover, limited by cost
the interactions of cells in three dimensions, and flat and reproducibility, a high-throughput method capable
monolayer cells cause changes in their morphology, of fabricating the microenvironment to simulate the
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affecting the gene expression and drug testing results. migration process of tumor cells is still lacking. Utama
9,55
Tissues have a certain 3D structure, within which signaling et al. fabricated neuroblastoma spheroids with bioprinting
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molecules (VEGF-A, TNF-α, IL-6, etc.), biomaterials (Figure 3A). In their method, SK-N-BE(2) cells with a
(collagen, laminin, fibronectin, hyaluronan, proteoglycan, concentration of up to 250 million cells/ml were embedded
etc.), and cells interact. 56,57 To match the environment in into alginate hydrogel matrix cup structure to form
vivo, a 3D culture system is needed. spheroids, which showed an average diameter of ~200 μm
after ~72-h culture. The viability of cells exceeded 95% after
Spontaneous aggregation of cells to form a cell bioprinting, and more importantly, high-concentration
cluster with a certain and stable 3D structure, defined bioink rich in nutrients could be printed around the
as a spheroid, is a typical 3D culture model. Spheroids spheroids to build a tumor microenvironment for disease
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recapitulate the structure organization and biological modeling. Compared with manual preparation, bioprinted
features of tumor tissues in vivo. During the formation spheroids were more compact and exhibited a uniform cell
of spheroids after bioprinting, the extracellular matrix distribution. In addition, they observed that the secretion
(ECM) can provide an essential microenvironment for of Ki67 protein was on the periphery of the spheroids after
spheroid growth. Scaffolds are widely utilized to support 6-day culture, and ~5% of the HIF1α-positive cells were
the ECM to form 3D spatial structures. The exchange of at the inner part of the spheroids. Approximately 0.4% of
nutrients and metabolites within the ECM can facilitate the apoptotic cells (caspase-3-positive) were also observed.
the proliferation and differentiation of tumor cells, and 3.2. Renal spheroids
accelerate the formation of tumor spheroids. Besides, Epithelial cells are the barrier that protects internal tissues
it is found that drug resistance and sensitivity of tumor and organs and secretes cytokines to regulate the function
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spheroids within the scaffold are significantly higher than of other cells. In recent years, the intrinsic connection
those without the scaffold. 59,60 However, the scaffold may between epithelial cells and the early formation of tumors
hinder the free movement of tumor cells and confine the has been demonstrated. For instance, the decrease of
expansion of tumor spheroids. 61,62 Meanwhile, low diffusion adhesion proteins secreted by epithelial cells leads to a
efficiency of drugs can introduce notable measurement contribution of the invasion and migration of tumor cells.
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errors in the final results of anti-cancer drug screening. Generally, the loss of epithelial cell polarity contributes
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Thus, while fabricating spheroids, the usage of ECM is to the formation of tumors. Trondle et al. employed a
mainly dependent on the type of tumor spheroid and its piezoelectric inkjet bioprinter to fabricate renal tubular
application. epithelial cell spheroids with iREC cells (Figure 3B). They
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In recent years, many methods have been proposed demonstrated that there was no spheroid formation when
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to fabricate spheroids, which have been widely utilized in the bioink concentration was below 5 × 10 cells/ml. By
tumor disease modeling and anti-cancer drug screening. changing bioink concentration, they fabricated spheroids
Successful fabrication of spheroids with controlled size with a controlled diameter range of 100–150 μm and an
and morphology can be achieved using many strategies, average viability of 92%. Within 7-day culture, the diameter
such as hanging drops, agitation-based method, non- of spheroid continuously decreased. Meanwhile, spheroids
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formed a closed epithelial cell layer and a hollow lumen, and
adhesive microwells, and external-force-driven (acoustic, the spheroids could be utilized to assemble the nephron-like
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magnetic, dielectrophoretic) technologies. 66-70 These tubules. Compared to 2D model, the expression of kidney-
methods can fabricate tumor spheroids but usually suffer specific gene signatures was significantly enhanced, and
from a lack of precision, throughput, and automation. the expression of kidney-specific transmembrane proteins,
Droplet-based bioprinting exhibits lots of advantages transporters, and regulators was increased.
in tumor spheroid fabrication, and we review the tumor
spheroids, associated with distinct parts in the human 3.3. Liver tumor spheroids

Volume 10 Issue 1 (2024) 112 https://doi.org/10.36922/ijb.1214