Stimulation of Cultured H9 Human Embryonic Stem Cells with Thyroid Stimulating Hormone Does Not Lead to Formation of Thyroid-Like Cells

Abstract

The sodium-iodine symporter (NIS) is expressed on the cell membrane of many thyroid cancer cells, and is responsible for the radioactive iodine accumulation. However, treatment of anaplastic thyroid cancer is ineffective due to the low expression of NIS on cell membranes of these tumor cells. Human embryonic stem cells (ESCs) provide a potential vehicle to study the mechanisms of NIS expression regulation during differentiation. Human ESCs were maintained on feeder-independent culture conditions. RT-qPCR and immunocytochemistry were used to study differentiation marker expression, 125I uptake to study NIS function. We designed a two-step protocol for human ESC differentiation into thyroid-like cells, as was previously done for mouse embryonic stem cells. First, we obtained definitive endoderm from human ESCs. Second, we directed differentiation of definitive endoderm cells into thyroid-like cells using various factors, with thyroid stimulating hormone (TSH) as the main differentiating factor. Expression of pluripotency, endoderm and thyroid markers and 125I uptake were monitored throughout the differentiation steps. These approaches did not result in efficient induction of thyroid-like cells. We conclude that differentiation of human ESCs into thyroid cells cannot be induced by TSH media supplementation alone and most likely involves complicated developmental patterns that are yet to be understood.

Effect of Chromatin Structure on the Extent and Distribution of DNA Double Strand Breaks Produced by Ionizing Radiation; Comparative Study of hESC and Differentiated Cells Lines

“…To verify this in our cell cultures, we stained H9 and H14 hESC, HT1080 fibrosarcoma cells, and IMR90 primary fibroblasts for heterochromatin marker H3K9me3. We also differentiated our H9 and H14 hESC towards the endodermal lineage using activin A and bFGF as previously described [ 15 ], to directly compare these cells to the differentiated lineages.…”

“…Then, the cells were maintained in mTeSR1 medium at 5% CO 2 and 37 °C for two days with medium changed each day. Starting from day 3, cells in culture were maintained in differentiation media: DMEM/F12 (Stemcell Technologies, Vancouver, BC, Canada) supplemented with 5% KSR (knockout serum replacement) (Invitrogen, Grand Island, NY, USA), 100 ng/mL Activin A (Stemcell Technologies), 4 ng/mL bFGF as was described previously in [ 15 ].…”

Normal vs cancer thyroid stem cells: the road to transformation

“…34,39-41 D'Amour et al 42 reported similar results using human cells, observing that in the presence of FGF-2 highdose activin A induced a massive differentiation of ESCs into transcription factor SOX-17 + /Foxa2 + definitive endodermal cells ( Figure 4 and Table 1). Interestingly, Onyshchenko et al 43 used two methods in order to differentiate human ESCs in thyroid follicular cells: (i) an one-step protocol that aims at a direct differentiation through TSH stimulation by avoiding the intermediate endoderm formation and (ii) a two-step protocol with an intermediate passage in endodermal cells, which foresees the TSH stimulation in combination with activin A and FGF-2. In both cases, we were unable to obtain an efficient generation of differentiated cells that express specific thyroid markers.…”