[The crosstalk between canonical and noncanonical Wnt signaling pathway in osteoblast differentiation of periodontal ligament stem cells in inflammatory microenvironments].
Objective:To investigate the crosstalk between canonical Wnt/β-catenin and noncanonical Wnt/Ca2+pathway in osteoblast differentiation process of periodontal ligament stem cell (PDLSC) in inflammatory microenvironments.Methods:PDLSCs were obtained from human healthy individuals(H-PDLSC) and patients with periodontitis(P-PDLSC). The H/P-PDLSCs were transfected with β-catenin siRNA. Cell morphology was observed under fluorescent microscope and transfection efficiency was easured by Western blotting after transfection of PDLSC. The mRNA expressions of Runt-related transcription factor 2(Runx2), β-catenin and nemo like kinase(NLK) were detected by real time PCR, the protein expressions of calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) and NLK were examined by Western blotting and the CaMK Ⅱ was observed by immunofluorescence staining, respectively.Results:The β-catenin expressions in H/P-PDLSCs were inhibited specifically and efficiently by treatment of β-catenin-siRNA for 24 h. After a 3-day-osteogenic process, results of real-time quantitative PCR showed that the Runx2 mRNA expression in P-PDLSC siRNA β-catenin transfected group(4.553 ± 0.659) was significantly higher than that in P-PDLSC empty plasmid control group(1.918 ± 0.315) (P=0.000). A similar trend was observed in the NLK mRNA expression tests(7.341 ± 1.331 vs. 5.664 ± 0.792) (P=0.030). Accordingly, the protein expression levels of CaMK Ⅱ, NLK were higher in P-PDLSC siRNA β-catenin transfected group than that in P-PDLSC empty plasmid control group in osteogenic differentiation condition for 3 days. CaMKⅡ was more strongly induced in P-PDLSC siRNA β-catenin group than that in P-PDLSC empty plasmid control group after PDLSC cultured in osteogenic medium for 3 days.Conclusions:Both canonical Wnt/β-catenin and noncanonical Wnt/Ca2+pathway could regulate the osteogenic differentiation potential of P-PDLSC. Suppression of β-catenin by siRNA promoted osteogenic differentiation via increasing noncanonical Wnt signaling pathway of PDLSC in inflammatory microenvironments.
Institute of Stomatology, General Hospital of Chinese PLA, Beijing 100853, China.
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N
Liu
Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing 100190, China.
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HG
Shi
Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing 100190, China.
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W
Zhang
Institute of Stomatology, General Hospital of Chinese PLA, Beijing 100853, China.
B
B
Gu
D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D002454:Cell Differentiation; D050659:Core Binding Factor Alpha 1 Subunit; D010006:Osteoblasts; D010012:Osteogenesis; D010513:Periodontal Ligament; D013234:Stem Cells; D060449:Wnt Signaling Pathway; D051176:beta Catenin
D050659:Core Binding Factor Alpha 1 Subunit; D051176:beta Catenin; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2