Endocrine Abstracts

Thyroid hormones (TH) play a critical role during nervous system development and patients carrying coding variants of THRA or SLC16A2/MCT8 genes present a spectrum of neurological phenotypes presumably resulting from perturbed local TH action during early brain development. Monocarboxylate transporter MCT8 is a well-characterized TH membrane transporter and MCT8 mutations have been associated with the Allan-Herndon-Dudley Syndrome. Recent advances in stem cell biology allow for generation of human cerebral organoids (hCOs) from induced pluripotent stem cells (hiPSC). By recapitulating many aspects of human neocortex development, hCOs provide a tractable model to probe local TH action during cortical neurogenesis. The primary aim of this study was to map the spatiotemporal expression of MCT8 during hCO differentiation. hCOs were generated from healthy hiPSC lines and cultured for up to 10 weeks to a stage corresponding to human cortex development at midgestation. The developmental expression profile of MCT8 protein was analyzed by immunofluorescence staining of cryosections prepared from hCOs of different stages. Expression of SLC16A2 mRNA was analyzed by single molecule fluorescent in situ hybridization (smFISH) and quantitative RT-PCR. Already along with the initial formation of rosette-like structures containing neuronal progenitors, MCT8 protein was detectable in SOX2+/Nestin+ radial glia cells (RGCs). During subsequent development of the ventricular zone, MCT8 immunostaining in RGCs became enhanced in apical endfeet at the ventricular surface. MCT8 expression was also detected in EOMES+ intermediate progenitors and HOPX+ outer RGCs located in the inner and outer subventricular zone, respectively. In addition to the diverse progenitor cell types, we detected robust MCT8 protein expression in TBR1+/CTIP2+ deep layer neurons and at later developmental stages in SATB2+ upper layer neurons. The spatial expression of SLC16A2 mRNA across cortical cell layers (detected by smFISH) was highly concordant with the MCT8 protein expression profile. When comparing the spatial expression profile of SLC16A2 and THRA mRNAs, we observed enhanced THRA probe staining in the cortical plate zone and smFISH confirmed that excitatory neurons express much higher levels of THRA mRNA than neuronal progenitors. Our study shows that MCT8 is expressed at comparable levels in both neuronal progenitors and excitatory neurons during early cortical neurogenesis in hCOs whereas THRA mRNA is clearly expressed at higher levels in neurons compared to progenitors. The similarity of expression profiles in hCOs and human embryonic cortex tissue suggests that hCOs provide a promising system for studies on neuronal effects resulting from targeted disruption of MCT8 function.