The presence of thyroid stem/progenitor cells has not been clearly demonstrated, nor their specific cell surface markers are known. Mice are used as a model animal to study stem/progenitor cells of thyroid, and partial thyroidectomy as a tool to activate their stem/progenitor cells. The latter is based on the hypothesis that a sudden loss of thyroid tissue may activate otherwise dormant stem/progenitor cells to participate in thyroid regeneration. However, how these cells contribute thyroid regeneration remains unclear. In order to address these questions, we used beta-galactosidase reporter mouse in conjunction with partial thyroidectomy and bromodeoxyuridine (BrdU) long label-retaining cell analysis. Beta-galactosidase reporter mouse expresses beta-galactosidase only in differentiated mature thyroid follicular cells, thus allowing tracing the thyroid lineage by examining the expression of beta-galactosidase. In BrdU long label-retaining analysis, BrdU positivity was used as a surrogate marker for stem/progenitor cells due to the fact that BrdU positive stem/progenitor cells do not divide frequently or asymmetrically divide, thus BrdU being retained only in stem/progenitor cells. We demonstrated that cells positive for stem cell antigen 1 (Sca1) and BrdU, but negative for beta-galactosidase and NKX2-1, a marker for thyroid differentiation were found in the non-follicular mesenchymal area 7 days after partial thyroidectomy. Sca1 was previously demonstrated that it is expressed in a half of verapamil-resistant side population (SP) cells of mouse thyroid that exhibit stem-progenitor-like characteristics. Thus, Sca1 can potentially be used as a marker for thyroid stem/progenitor cells. The Sca1- and BrdU-positive, but beta-galactosidase- and NKX2-1-negative cells temporarily co-expressed cytokeratin 14, and were observed in part of follicles by day 35 post-partial thyroidectomy. Cells positive for all four markers, Sca1, BrdU, beta-galactosidase, and NKX2-1 were found 120 days post-partial thyroidectomy. These results suggested that Sca1 and BrdU positive cells may participate in the formation of new thyroid follicles after partial thyroidectomy. The process of thyroid follicular cell regeneration was recapitulated in ex vivo thyroid slice collagen gel culture studies, in which BrdU was added 2 hours before fixation of cells. Sca1 expression was observed in the layer of cells surrounding follicular cells without BrdU expression on day 1 of culture. On day 3, Sca1 and BrdU-positivity was found in non-follicular mesenchymal cells. Follicles formed by day 7;some intrafollicualr cells expressed Sca1 while some were also positive for BrdU. These studies altogether demonstrated that Sca1-positive mesenchymal cells may be involved in the process of thyroid differentiation into mature follicular cells after partial thyroidectomy. The studies will help facilitate research on thyroid stem/progenitor cells and their roles in thyroid diseases, particularly thyroid carcinomas.
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