口腔疾病防治 (Jul 2018)
Culture and identification of the osteogenic properties of beagle maxillary sinus membrane stem cells
Abstract
Objective To investigate the osteogenic properties of maxillary sinus membrane stem cells (MSMSCs). Methods Beagle maxillary sinus mucosa was collected, immunomagnetic bead method was applied for isolation of CD146 + cells, and MSMSCs were harvested and cultured from the canine maxillary sinus floor mucosa. The levels of the cell surface antigens CD44, CD146, and CD34 were determined at passage one by flow cytometry. Cells at passage one were cultured in basal medium and osteogenic inductive medium. Real⁃time PCR, immunohistochemical staining, al⁃ kaline phosphatase activity, alizarin red staining and Von Kossa staining were used to investigate the osteogenic proper⁃ ties in vitro. Results The canine MSMSCs were cultured successfully. The results of flow cytometry were positive for CD146 and CD44 expression but negative for CD34 expression. The relative mRNA expression of runt⁃related transcrip⁃ tion factor 2 (RUNX2) (t = 14.44，P < 0.001), osteopontin (OPN) (t = 7.85，P = 0.001) and alkaline phosphatase alka⁃line phosphatase (t = 14.27，P < 0.001) was apparently higher in the osteoinductive medium group than in the basal me⁃ dium group, the differences in relative mRNA expression between the groups were significant. The protein levels of RUNX2 and OPN increased in the osteoinductive medium group. The alkaline phosphatase activity of the MSMSCs in⁃ creased when the cells were cultured in osteoinductive medium; the activity increased to a level that was significantly higher than that in basal medium, particularly at days 3 (t = 8.79, P < 0.001), 7 (t = 9.75，P < 0.001), 14 (t = 12.14， P < 0.001), 21 (t = 19.62，P < 0.001) and 28 (t = 17.53，P < 0.001). Obvious mineralized nodules were observed by aliz⁃ arin red staining or Von Kossa staining. Conclusion Maxillary sinus membrane stem cells exhibit osteogenic ability.
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