Abstract
The generation of artificial human thyroid tissues in suspension (low-shear environment, present in simulated microgravity [MG] and generated by a rotary cell culture system [RCCS]), was enhanced by increasing medium kinematic viscosity with a (3% v/v) suspension of extracellular matrix (basement membrane extract [BME]) in serum-free medium to generate artificial human thyroid organoids. Recombinant human keratinocyte growth factor (KGF, 7 ng/mL) facilitated human thyrocyte aggregation and three-dimensional (3-D) differentiation. There was an MG-associated decrease in extractable DNA that was reversed after addition of keratinocyte growth factor (KGF). In simulated MG, the increase in extractable DNA after KGF addition was up to 170% over non-KGF control cultures. In contrast, monolayer cultures in unit gravity showed a maximum DNA increase of 39% after KGF addition. Morphologically, differentiated thyroid neofollicles displayed polarization and were located in close proximity after 2 weeks of culture. Immunogold labeling with antibody to human thyroglobulin (Tg) revealed staining of follicular lumina and secretory vesicles, and a time-dependent increase in human Tg was detected in the culture media. Culture under simulated MG thus allowed direct visualization of KGF-facilitated thyrocyte/extracellular matrix interaction. Such artificial human thyroid organoids—generated in MG and in the presence of KGF—structurally resembled natural thyroid tissue. The above findings may have implications for autoimmune thyroid disease where KGF (if, for example, secreted locally by intraepithelial γδ T cells among other cells) may contribute to thyroid cell growth.
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