Human mesenchymal stem cells are capable of differentiating into various cell types and are useful for applications in regenerative medicine. Previous studies have indicated that stem cells can be isolated from dental pulp, and dental pulp exfoliated from deciduous teeth has become a useful alternative for dental tissue engineering because of its higher proliferation rate. For clinical application, it is necessary to culture cells in vitro and to obtain sufficient numbers of cells as quickly as possible. Furthermore, it is necessary to confirm the change in cells cultured in vitro in order to ensure that they have suitable characteristics. In this study, to investigate the changes in human deciduous dental pulp cells cultured with serum-free media, we analyzed cell proliferation, cell morphology and gene expression changes by microarray analysis. We found a high cell proliferation rate in cells that were cultured in STK2 that is a serum-free medium, and cells tended to remain in close contact with one another in cell morphology. In addition, 3248 genes were expressed at >2-fold higher levels in dental pulp cells from deciduous teeth cultured with STK2 compared with medium containing fetal bovine serum for 3 days.

Human mesenchymal stem cells can differentiate into various cell types and are useful for applications in regenerative medicine. Previous studies indicated that dental pulp exfoliated from deciduous teeth is a valuable alternative for dental tissue engineering because it contains stem cells with a relatively high proliferation rate. For clinical application, it is necessary to rapidly obtain a sufficient number of cells in vitro and maintain their undifferentiated state; however, the abundance of stem cells in the dental pulp tissue is limited. Non-thermal atmospheric pressure plasma (NTAPP) has been applied in regenerative medicine because it activates cell proliferation. Here, we examined the effects of NTAPP to activate the proliferation of human deciduous dental pulp fibroblast-like cells (hDDPFs) in vitro. Compared with untreated cells, NTAPP increased cell proliferation by 1.3-fold, significantly upregulated well-known pluripotent genes for stemness (e.g., Oct4, Sox2, and Nanog), and activated the expression of stem cell-specific surface markers (e.g., CD105). Overall, NTAPP activated the proliferation of various mesodermal-derived human adult stem cells while maintaining their pluripotency and stemness. In conclusion, NTAPP is a potential tool to expand the population of various adult stem cells in vitro for medical applications.

IntroductionThe increase in lifestyle-related diseases following the diversification of eating habits, including dyslipidemia, cardiovascular diseases, and obesity has become a social problem in developed countries. In particular, cardiovascular disease, which causes arteriosclerosis, is one of the leading causes of death in Japan 1 . Therefore, the prevention or amelioration of dyslipidemias, such as hypercholesterolemia, would help to reduce the risk of cardiovascular diseases.There is growing interest in hypocholesterolemic effect of various food components, especially polyphenols, such as tea catechin 2 . Polyphenols are present in a wide range of food products derived from plants such as vegetables, fruits, and beans. Procyanidins are composed of catechin and epicatechin oligomers and exert many health benefits, such as cancer prevention, cardiovascular protection, and diabetes prevention 3 . In a recent study, absorption of low molecular weight procyanidins, including dimers, was observed in rats 4 , while the bioavailability of high molecular

Dental pulp plays an important role in tooth vitality. Previous studies have indicated that stem cells can be isolated from dental pulp, and dental pulp exfoliated from deciduous teeth has become a useful alternative in dental tissue engineering because of its higher proliferation rate. However, heat stress can cause irritation in the dentin/pulp complex, and little is known about the thermotolerance of pulp cells and their response to heat stress. Heat shock protein (HSP) is induced by heat and various other stresses. It also reportedly takes part in causing apoptosis when wounded dental pulp cells heal. In this study, we investigated the responses of dental pulp cells to heat stress using reverse transcriptase-polymerase chain reaction and immunoblotting analysis. Cells were subjected to a temperature of 427C for 1, 2, 7, and 14 h, and HSP70 was determined at each time point. HSP70 expression was weak before heat treatment, but heat shock markedly induced HSP70 expression. This suggests that heat stress raises the wound healing ability of dental pulp cells. The mechanisms by which heat stress induces HSP expression may be an important link between apoptosis and wound healing. Therefore, an understanding of the mechanisms underlying HSP expression in dental pulp is important for the development of new therapeutic strategies. role in anti-apoptotic processes 2,3) and promotion 4) , and are thought to function in the protection and recovery of cells from environmental and pathological stress 5). They are named according to their molecular weight. For example, HSP60, HSP70, and HSP90 refer to families of heat shock proteins on the order of 60, 70, and 90 kilodaltons in size, respectively. HSP70, which serves as a molecular chaperone to maintain homeostasis, is physiologically expressed in response to stimulation. In addition, HSP70 is expressed in dental pulp and plays an important role during dentin formation 6). Moreover, it is said that HSPs take part in the recovery of human pulp fibroblasts (HPFs). However, there are no reports concerning human deciduous dental pulp fibroblasts (HDPFs) and HSP. Little information is available on the expression of HSP70 in the pulp of deciduous teeth. The pulp

Appropriate heat shock results in the production of heat shock proteins (HSPs) whose expression and phosphorylation contribute to repair of damaged proteins, cell proliferation, and cell recovery from shock stimuli. However, there is no information regarding the expression of HSPs in human deciduous dental pulp fibroblast-like cells (hDDPF) in response to mild short-term heat shock. The aim of this study was to investigate the cellular effects of mild short-term heat shock on hDDPF. Cells were subjected to heat shock at 43°C-49°C for 15 min, and cell proliferation was assessed by 3-(4,5-dimethylthiazol-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. mRNA and protein expressions of HSP27, 70, and 90 were detected by reverse transcription PCR and western blot analysis, respectively. Phosphorylation of the AKT and ERK signaling pathways of HSP production was evaluated by western blotting. Heat shock at 43°C for 15 min increased the cell proliferation rate and the mRNA expressions of HSP27, 70, and 90 in hDDPF. Moreover, protein expression of HSP70 was significantly enhanced 24 h after heat shock, and the phosphorylation of AKT was also confirmed. Because HSP70 is critical in tissue repair and regeneration, mild short-term heat shock may enhance tissue repair in hDDPF.