In order to identify and characterize thyroid stem/progenitor cells, we have used Side Population (SP) cells that are characterized by their ability to efflux the vital dye Hoechst 33342 when analyzed by flow cytometry, due to expression of the ATP binding cassette (ABC)-dependent transporter ABCG2. SP cells are thought to be highly enriched for stem/progenitor cell activity. In fact, we demonstrated that thyroid SP cells have stem/progenitor-like properties, and a half of the SP cells express Stem cell antigen 1 (Sca1). Partial thyroidectomy was used to model acute thyroid injury, which may require thyroid stem/progenitor cells to actively participate in the regeneration of injured thyroid. Partial thyroidectomy increased the population of thyroid SP cells. Thyroid SP cells were used to establish thyroid SP cell line that can be continuously cultured over many passages. This suggests that SP cells may have self-renewal capacity, one of the critical characteristics of stem/progenitor cells. When SP-derived cells were subjected to FACS analysis, all SP cells expressed Sca1 in a very homogenous fashion. When SP-derived cells were cultured in low serum, they started expressing thyroid differentiation marker genes such as thyroglobulin and a transcription factor NKX2-1 that is critical for the expression of thyroid differentiation marker genes, while Sca1 expression decreased. Thus, Sca1 may be a thyroid stem/progenitor cell marker, and SP-derived cells can be used to understand the nature of stem/progenitor cells of the thyroid. Since we use partial thyroidectomy as a tool to study thyroid regeneration, it is important to understand what type of changes occur in the thyroid after partial thyroidectomy. We have analyzed the thyroid 2 weeks after partial thyroidectomy. We hypothesize that at 2 weeks post-partial thyroidectomy, the thyroid may be in the beginning of repair process after the initial immune response subsides, before the onset of fibrosis. Histological examination of the thyroid indicated the presence of a proliferative center in the central area of the left or right thyroid lobe regardless of partial thyroidectomy, which occasionally had micro-follicles with BrdU positive cells. After partial thyroidectomy, the number of BrdU-positive cells and cells with clear or faintly eosinophilic cytoplasm markedly increased in the central area of the thyroid lobe and the overlapping area close to the cutting edge of partial thyroidectomy. Electron microscopic analysis demonstrated that clear-looking cells have scant cytoplasmic components, some of which retain characteristic features of C cells such as the presence of neuroendocrine granules, while others retain follicular cell-specific features such as directly facing a lumen with microvilli at the apical side of the cell. Some of these cells were BrdU-positive and expressed FOXA2, the definitive endoderm lineage marker. Microarray followed by Pathway analysis revealed that the expression of genes involved in embryonic development and cancer, two processes where stem cells play roles, was affected in the thyroid after partial thyroidectomy. These results suggested that cells previously C cells or follicular cells, may be altered after partial thyroidectomy to become immature cells, or immature cells that might be derived from stem/progenitor cells of thyroid, are on their way to differentiate into C cells or follicular cells. These immature cells participate in the repair and/or regeneration of thyroid, a process similar to embryonic thyroid development and/or cancer.
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