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Rutin is one of the flavonoids found in fruits and vegetables. Recent reports have revealed that rutin is a major player in proliferation and bone development. However, data on how rutin regulates the proliferation of periodontal ligament stem cells (PDLSCs), as well as the differentiation of osteogenic cells are scanty. Here, our findings showed that rutin enhanced PDLSCs proliferation, increased ALP activity, and matrix mineralization. Moreover, rutin significantly promoted the expression of osteogenic genes and elevated phosphorylated AKT and mTOR. Treatment with LY294002 reversed these effects by inhibiting PI3K. We also found that the expression levels of GPR30 were increased by rutin. Interestingly, this upregulation was not altered after the addition of LY294002. In addition, G15, a selective antagonist of GPR30, could reduce the beneficial effects induced by rutin and interfere with the modulation of PI3K/AKT/mTOR signal transduction. Collectively, our findings revealed that rutin increased proliferation and osteogenic differentiation of PDLSCs through GPR30-mediated PI3K/AKT/mTOR signal transduction. Therefore, it could be deduced that rutin as a certain flavonoid possesses therapeutic value for periodontal bone regeneration and tissue engineering. Impact statement In our study, the effects and mechanisms of rutin on the osteogenic differentiation and proliferation of PDLSCs were investigated. Our findings might provide basic knowledge and guidance to understand and use rutin in the bioengineering of the periodontal tissues and regeneration of bones. The following is a short description of the main findings: rutin promotes the osteogenic differentiation and proliferation of PDLSCs; PI3K/AKT/mTOR signal pathway mediates the effects of rutin on PDLSCs; rutin activates PI3K/AKT/mTOR signal pathway via GPR30.

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Quercetin (Quer) is a typical antioxidant flavonoid from plants that is involved in bone metabolism, as well as in the progression of inflammatory diseases. Elevated levels of tumor necrosis factor-α (TNF-α), a typical pro-inflammatory cytokine, can affect osteogenesis. In the present study, TNF-α was used to establish an in vitro model of periodontitis. The effects of Quer on, as well as its potential role in the osteogenic response of human periodontal ligament stem cells (hPDLSCs) under TNF-α-induced inflammatory conditions and the underlying mechanisms were then investigated. Within the appropriate concentration range, Quer did not exhibit any cytotoxicity. More importantly, Quer significantly attenuated the TNF-α induced the suppression of osteogenesis-related genes and proteins, alkaline phosphatase (ALP) activity and mineralized matrix in the hPDLSCs. These findings were associated with the fact that Quer inhibited the activation of the NF-κB signaling pathway, as well as the expression of NLRP3 inflammation-associated proteins in the inflammatory microenvironment. Moreover, the silencing of NLRP3 by small interfering RNA (siRNA) was found to protect the hPDLSCs against TNF-α-induced osteogenic damage, which was in accordance with the effects of Quer. On the whole, the present study demonstrates that Quer reduces the impaired osteogenesis of hPDLSCs under TNF-α-induced inflammatory conditions by inhibiting the NF-κB/NLRP3 inflammasome pathway. Thus, Quer may prove to be a potential remedy against periodontal bone defects.

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Cell sheet technology is a novel tissue engineering technology that has been rapidly developed in recent years. As a novel technology, cell sheet technology is expected to become one of the preferred methods for cell transplantation. The present study investigated the biological effects of rutin on the formation of periodontal ligament stem cell (PDLSC) sheets and their resultant osteogenic properties. The results of Cell Counting Kit-8 (CCK-8) assay demonstrated that a concentration of 1×10−6 mol/l rutin promoted the proliferation of PDLSCs more effectively compared with other designed concentrations. Rutin-modified cell sheets could be induced by complete medium supplemented with 20 µg/ml vitamin C (VC) and 1×10−6 mol/l rutin. Rutin-modified cell sheets appeared thicker and more compact compared with the VC-induced PDLSC sheets, demonstrating more layers of cells (3 or 4 layers), which secreted a richer extracellular matrix (ECM). Furthermore, the improved cell sheets exhibited varying degrees of increases in the mRNA and protein expression of collagen type I (COL1), alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2) and osteopontin (OPN). Combined treatment with VC and rutin promoted the formation of PDLSC sheets and enhanced the osteogenic differentiation potential of the cell sheets. Therefore, rutin-modified cell sheets of PDLSCs are expected to play an important role in the treatment of periodontal tissue regeneration by stem cells.