ArticleInternational journal of molecular sciences2021
Immortalization and Characterization of Rat Lingual Keratinocytes in a High-Calcium and Feeder-Free Culture System Using ROCK Inhibitor Y-27632.
Article in International journal of molecular sciences, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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2 citing papers in PubMed, 7 citations in OpenAlex.
- Standardized Methodology of Scaffold-Free and Scaffold-Based 3D Epithelial Spheroid Culture for Skin Regenerative Research.Methods and protocols · 2025Article
- Positive In Vitro Effect of ROCK Pathway Inhibitor Y-27632 on Qualitative Characteristics of Goat Sperm Stored at Low Temperatures.Animals : an open access journal from MDPI · 2024Article
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Authors and funding
4 authors at 2 institutions in 2 countries.
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Abstract
Cultured keratinocytes are desirable models for biological and medical studies. However, primary keratinocytes are difficult to maintain, and there has been little research on lingual keratinocyte culture. Here, we investigated the effect of Y-27632, a Rho kinase (ROCK) inhibitor, on the immortalization and characterization of cultured rat lingual keratinocyte (RLKs). Three Y-27632-supplemented media were screened for the cultivation of RLKs isolated from Sprague-Dawley rats. Phalloidin staining and TUNEL assay were applied to visualize cytoskeleton dynamics and cell apoptosis following Y-27632 removal. Label-free proteomics, RT-PCR, calcium imaging, and cytogenetic studies were conducted to characterize the cultured cells. Results showed that RLKs could be conditionally immortalized in a high-calcium medium in the absence of feeder cells, although they did not exhibit normal karyotypes. The removal of Y-27632 from the culture medium led to reversible cytoskeletal reorganization and nuclear enlargement without triggering apoptosis, and a total of 239 differentially expressed proteins were identified by proteomic analysis. Notably, RLKs derived from the non-taste epithelium expressed some molecular markers characteristic of taste bud cells, yet calcium imaging revealed that they rarely responded to tastants. Collectively, we established a high-calcium and feeder-free culture method for the long-term maintenance of RLKs. Our results shed some new light on the immortalization and differentiation of lingual keratinocytes.
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