Responsiveness to Thyroid Hormone is Enhanced in Rat Hepatocytes Cultured as Spheroids Compared with that in Monolayers: Altered Responsiveness to Thyroid Hormone Possibly Involves Complex Formed on Thyroid Hormone Response Elements

We previously reported that the expression of type I iodothyronine 5′-deiodinase (5′DI) gene was increased by 3,3,′,5-triiodothyronine (T₃) in isolated rat hepatocytes when cultured as spherical aggregates (spheroids), whereas this effect was greatly attenuated in conventional monolayer cultures. In the current study, we examined whether the enhanced T₃ responsiveness in spheroid cultures extends to other T₃-responsive genes. As observed for 5′DI, T₃ increased spot 14, malic enzyme and fibronectin messenger RNAs (mRNAs) by fourfold to fivefold in spheroid cultures, while the effect in monolayer cultures was blunted. This difference in T₃ responsiveness was also observed when T₃-responsive reporters consisting of the luciferase gene under the control of triiodothyronine response element (TRE) were introduced into hepatocytes using a replication-defective adenovirus vector. These results suggest that the factors required for T₃-dependent transcriptional activation are preserved in spheroid cultures and that they must exert their effect by interacting with TRE. Maximal binding capacity of nuclear T₃ receptor was not different between monolayer and spheroid cultures while the expression of retinoid X receptora (RXRa) mRNA was higher in spheroid cultures compared with that in monolayers. The difference in RXRa mRNA expression, together with enhanced proteolytic cleavage in monolayers that we demonstrated recently, may account for the difference in T₃ responsiveness between the two hepatocyte culture systems.