This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.  The optimal dosage of baicalein was 10 μM. DKK-1 significantly reversed the effects of baicalein on hPDLCs. The effects are exerted by activating the Wnt/β-catenin signaling pathway. Page 3 of 29A c c e p t e d M a n u s c r i p t ABSTRACT Objective: Periodontium regeneration is one of the most important processes for periodontitis therapy. Human periodontal ligament cells (hPDLCs) play a vital role in the repair and regeneration of periodontal tissues. Our study aimed to investigated the mechanisms underlying the promotion of hPLDCsosteogenic differentiation by baicalein. Design: hPDLCs were obtained from periodontal ligament (PDL) tissues by primary culture. The MTT assay was used to determine the growth curves of hPDLCs treated with different concentrations of baicalein (1.25, 2.5, 5, or 10 μM). Alkaline phosphatase (ALP) staining and Alizarin red S staining were performed to assess osteogenic differentiation of hPDLCs administered baicalein. Osteogenic differentiation-related gene and protein expression levels and Wnt/β-catenin pathway signal changes were assessed by qRT-PCR and Western blotting analysis. Results:The results showed that baicalein decreased the growth of hPDLCs slightly and increased ALP activity and calcium deposition in a dose-dependent manner. The expression of runt-related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), Osterix (OSX) and osteocalcin (OCN) were elevated after baicalein administration. Moreover, baicalein strongly activated the Wnt/β-catenin pathway and up-regulated the expression of β-catenin, lymphoid enhancer factor 1 (LEF1) and Cyclin D1. Dickkopf-related protein 1 (DKK-1) significantly reversed the effects of baicalein on hPDLCs. Conclusions: Our findings indicated that baicalein enhanced the osteogenic differentiation of hPDLCs via the activation of the Wnt/β-catenin signaling pathway, which may represent a potential candidate for periodontitis therapy.

These results indicate that nicotine prevents the increased osteogenic potential of hPDLCs induced by cyclic tensile stress by binding to an α7 nicotinic acetylcholine receptor and activating the canonical Wnt pathway.

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Mechanical strain plays an important role in bone formation and resorption during orthodontic tooth movement. The mechanism has not been fully studied, and the process becomes complex with increased amounts of periodontal patients seeking orthodontic care. Our aims were to elucidate the combined effects of proinflammatory cytokines and intermittent cyclic strain (ICS) on the osteogenic capacity of human periodontal ligament cells. Cultured human periodontal ligament cells were exposed to proinflammatory cytokines (interleukin-1β 5 ng/mL and tumor necrosis factor-α 10 ng/mL) for 1 and 5 days, and ICS (0.5 Hz, 12% elongation) was applied for 4 h per day. The autocrine of inflammatory cytokines was measured by enzyme-linked immunosorbent assay. The expression of osteoblast markers runt-related transcription factor 2 and rabbit collagen type I was determined using real-time polymerase chain reaction and Western blot. The osteogenic capacity was also detected by alkaline phosphatase (ALP) staining, ALP activity, and alizarin red staining. We demonstrated that ICS impaired the osteogenic capacity of human periodontal ligament cells when incubated with proinflammatory cytokines, as evidenced by the low expression of ALP staining, low ALP activity, reduced alizarin red staining, and reduced osteoblast markers. These data, for the first time, suggest that ICS has a negative effect on the inductive inhibition of osteogenicity in human PDL cells mediated by proinflammatory cytokines.