Single clonal glandular stem cells derived from human parotid glands do not attain malignant phenotype during long-term in vitro culture

Mesenchymal stem cells (MSCs) are being intensively investigated as future therapeutics for various human diseases. One of the most important challenges to the clinical application of MSCs is the possibility of malignant transformation during long-term in vitro culturing. However, there have been no reports on the tumorigenicity of salivary gland-derived MSCs following long-term in vitro culturing. Here, we isolated a single clonal glandular stem cells from human parotid gland stem cells (hpGSCs) using a modified sub-fractionation culturing method. The possibility of malignant transformation of these cells following long-term culturing was evaluated under in vitro and in vivo culture conditions. Single clonal glandular stem cells from the human parotid gland have unique multipotent MSCs traits. hpGSCs at passage 18 stained strongly for β-galactosidase expression and the long-term culture of hpGSCs led to a reduction in telomerase activity. hpGSCs could not survive in a soft agar environment and did not cause tumor formation in a xenograft mouse model. In addition, the expression of salivary cancer-related oncogenes was not elevated in hpGSCs following the long-term culture. In conclusion, we demonstrated that there is no possibility of acquiring a malignant transformation during long-term in vitro cell expansion of hpGSCs.