Presently, bone marrow is considered as a prime source of mesenchymal stem cells; however, there are some drawbacks and limitations. Compared with other mesenchymal stem cell (MSC) sources, gingiva‐derived mesenchymal stem cells (GMSCs) are abundant and easy to obtain through minimally invasive cell isolation techniques. In this study, MSCs derived from gingiva and bone marrow were isolated and cultured from mice. GMSCs were characterized by osteogenic, adipogenic and chondrogenic differentiation, and flow cytometry. Compared with bone marrow MSCs (BMSCs), the proliferation capacity was judged by CCK‐8 proliferation assay. Osteogenic differentiation was assessed by ALP staining, ALP assay and Alizarin red staining. RT‐qPCR was performed for ALP, OCN, OSX and Runx2. The results indicated that GMSCs showed higher proliferative capacity than BMSCs. GMSCs turned more positive for ALP and formed a more number of mineralized nodules than BMSCs after osteogenic induction. RT‐qPCR revealed that the expression of ALP, OCN, OSX and Runx2 was significantly increased in the GMSCs compared with that in BMSCs. Moreover, it was found that the number of CD90‐positive cells in GMSCs elevated more than that of BMSCs during osteogenic induction. Taking these results together, it was indicated that GMSCs might be a promising source in the future bone tissue engineering.

Adipose tissue-derived stromal cells, termed ASCs, play an important role in regenerative applications. They resemble mesenchymal stem cells owing to their inexhaustibility, general differentiation potential, and plasticity and display a series of cell-specific and cluster-of-differentiation (CD) marker profiles similar to those of other somatic stem cells. Variations in phenotypes or differentiation are intimately associated with CD markers. The purpose of our study was to exhibit distinct populations of ASCs with differing characteristics for osteogenic differentiation. The primary cell batch of murine-derived ASCs was extracted from subcutaneous adipose tissue and the cells were sorted for the expression of the surface protein molecules CD90 and CD105 using flow cytometry. Each cell population sorted for CD90 and CD105 was analyzed for osteogenic potency after cell culture. The results suggested that ASCs exhibit distinct populations with differing characteristics for osteogenic differentiation: unsorted ASCs stimulated comparable mineralized nodule formation as bone marrow stromal cells (BMSCs) in osteogenic medium and viral transfection for BMP2 accelerated the formation of mineralized nodules in CD90 and/or CD105 positive ASCs with observation of decrease in CD105 expression after 14 days. Future studies assessing different immunophenotypes of ASCs should be undertaken to develop cell-based tissue engineering.

The treatment of bone defects still presents complex problems, although various techniques have been developed. The periosteum is considered a good source of osteogenic precursor cells for new bone formation. It can be collected easily in the clinical setting and is less invasive to the donor site. However, the murine skull periosteum has a poor cellular component, and growth is very slow, making it important to identify a culture method for efficient growth. In the present study, we used threedimensional cell migration with atelocollagen and gelatin media and found that both were effective for promoting the proliferation of periosteum-derived cells. Moreover, atelocollagen medium is expected to provide an added benefit as a scaffold structure in the ambient temperature of the human body. The selection of a proper surface marker for osteogenesis is imperative for bone regeneration. CD90 is a mesenchymal stem cell marker. Periosteum-derived cells sorted with CD90 showed higher proliferative capacity and osteogenic potential than that of unsorted periosteum-derived cells in vivo and in vitro. Thus, periosteum-derived cells sorted with CD90 are expected to be a good source for bone regeneration. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:227-234 SIGNIFICANCE Periosteum-derived cells showed higher proliferative capacity and osteogenic potential. Periosteum can be collected easily in the clinical setting and is less invasive to the donor site. Thus, periosteumderived cells can be expected to be a good source for bone regeneration.

Low-intensity pulsed ultrasound (LIPUS) has demonstrated its positive effects on osteogenic differentiation of mesenchymal stem cells and the proliferation and differentiation of osteoblasts, negative effects on osteoclast growth, and promotion of angiogenesis, leading to improvement of the tissue perfusion. Heat-shock proteins (HSPs) are initially identified as molecules encouraged and expressed by heat stress or chemical stress to cells and involved in the balance between differentiation and apoptosis of osteoblasts. However, it remains unclear if the effect of LIPUS on osteoblast differentiation could involve HSP expression and contribution. In this study, mouse calvarial osteoblasts were exposed to LIPUS at a frequency of 3.0 MHz by 30 mW/cm 2 for 15 min or to 42°C heat shock for 20 min at day 3 of cell culture and examined for osteogenesis with pursuing induction of HSP27, HSP70, and HSP90. LIPUS as well as heat shock initially upregulated HSP90 and phosphorylation of Smad1 and Smad5, encouraging cell viability and proliferation at 24 h, enhancing mineralized nodule formation stronger by LIPUS after 10 days. However, HSP27, associated with BMP2-stimulated p38 mitogen-activated protein kinase during osteoblast differentiation, was downregulated by both stimulations at this early time point. Notably, these two stimuli maintained Smad1 phosphorylation with mineralized nodule formation even under BMP2 signal blockage. Therefore, LIPUS might be a novel inducer of osteoblastic differentiation through a noncanonical signal pathway. In conclusion, LIPUS stimulation enhanced cell viability and proliferation as early as 24 h after treatment, and HSP90 was upregulated, leading to dense mineralization in the osteoblast cell culture after 10 days.

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