Objectives/Hypothesis Stem cells are known to proliferate at a slow rate in adult organs, and thus slow-cycling cells exhibiting pluripotency are considered tissue-specific stem cells in some organs. Slow-cycling cells in the vocal fold (VF) have not been well documented. Here we sought to clarify the distributions and characteristics of slow-cycling cells in rat VFs. Methods We applied double-labeling technique to detect the distribution of slow-cycling cells. We injected the exogenous proliferation marker 5-bromo-2'-deoxyuridine (BrdU) into Sprague-Dawley rats. After a chasing period, VFs were immunostained with antibodies to BrdU and the second endogenous proliferation marker, Ki-67. BrdU (+) Ki-67(+) cells were regarded as slow-cycling cells and counted by VF regions. To reveal slow-cycling cells' characteristic, their immunophenotypes were histologically investigated and their kinetics in injured VFs were evaluated. Results Most slow-cycling cells were detected in the basal layer of the epithelium. Slow-cycling cells in the epithelium displayed a low positive ratio of E-cadherin and CK5 and a high positive ratio of vimentin and CD31 as compared with the other epithelial cells. The expression of S100A4 was low in slow-cycling cells of the lamina propria and the macula flava. FGFR1, HAS1, HAS2, and HAS3 were not detected in the slow-cycling cells. A time-dependent reduction of slow-cycling cells was observed in injured VFs. Conclusion Most slow-cycling cells resided in the epithelium, exhibiting various phenotypes in a relatively undifferentiated condition, and they are suspected to contribute to the tissue repair of the injured VFs.
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