Speaker
Description
Introduction
Over the past few decades, extensive research has been actively conducted for the fabrication of human tissues to, amongst many applications, understand the effects of a wide range of chemicals on human health and the environment (1). Thus, the need for the development of innovative assessment tools that provide reliable results in identifying and regulating the risks of potentially harmful substances is increasingly recognized. This study presents a precisely arranged fibrous architecture, fabricated via MeltElectroWriting (MEW) as spheroid culture support for 3D in vitro models. This platform facilitates multi-spheroid bioassembly, overcoming single spheroid limitations such as hypoxia while promoting cell-cell communication and adaptive interactions with the surrounding Extracellular-Matrix-like fibers (2). The technique has been employed to recapitulate different human tissues, for this study the targeted tissue was the thyroid.
Methods
The scaffolds are printed with an in-house build MeltElectrowriting device by heating a syringe containing melted Polycaprolactone and extruding the material with air pressure while applying voltage to pull a microscale fiber. Fibers are precisely deposited in the shape of squared boxes, in a variety of sizes between 100 µm and 1 mm. The scaffolds perimeter was reinforced with a 3D printed Poly Lactic Acid ring for better handling and coated in polydopamine for improved cell compatibility (3). Human thyroid epithelial cells (huThyrEC) spheroids are formed by cell aggregation in wells to be a similar size as the boxes and after 3 days of culture- supplied with thyroid-stimulating hormone- are transferred in the scaffold so that each box is occupied by a single spheroid. The assemblies are further cultured for 7 more days followed by evaluating thyroid hormones T3 and T4, thyroid-related protein, metabolomic and transcriptomic changes.
Results
The final scaffolds present a box size of 530 µm and a highly precise architecture, raman analysis confirmed the presence of polydopamine on the surface. Spheroids cultivated in the MEW scaffolds disassembled long the fibers to occupy the box volume and showed significantly reduced hypoxia compared to single spheroid controls that would instead compact. Thyroid hormone secretion was significantly higher in the spheroid groups compared to monolayer with improved results for the MEW ones. From metabolomics it was evident how this tool can bridge the gap between 2D and 3D cultures while transcriptomics resulted in evidence of more upregulated genes in the fibrous environment.
Discussion
Analyzing thyroid hormone levels, protein, metabolomic and transcriptomic changes, provided a comprehensive understanding of cellular responses and metabolic shifts crucial for evaluating thyroid function and response to exposure to harmful or potentially hazardous substances.
Our findings suggest that the microarchitectures platform provides a reliable in vitro model for regulating thyroid function, representing a significant step toward advanced toxicity assessment tools.
References
1) La Merrill MA, Consensus on the key characteristics of endocrine-disrupting chemicals as a basis for hazard identification, DOI:10.1038/s41574-019-0273-8
2)McMaster R, Tailored Melt Electrowritten Scaffolds for the Generation of Sheet-Like Tissue Constructs from Multicellular Spheroids, DOI:10.1002/adhm.201801326
3)Lamberger Z, Streamlining the Highly Reproducible Fabrication of Fibrous Biomedical Specimens toward Standardization and High Throughput, DOI:/10.1002/adhm.202402527
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