Shikonin, an active ingredient of Lithospermum erythrorhizon, exerts anti-inflammatory and antibacterial effects, and promotes wound healing. We investigated whether shikonin stimulated gingival tissue wound healing in human gingival fibroblasts (hGF). In addition, we evaluated the effects of shikonin on the mitogen-activated protein kinase (MAPK) signaling pathway, which has an important role in wound healing. hGF were subjected to primary culture using gingiva collected from patients. The cells were exposed to/treated with Shikonin at concentrations ranging from 0.01 to 100 μM. The optimal concentration was determined by cell proliferation and migration assays. Type I collagen and fibronectin synthesis, the gene expression of vascular endothelial growth factor (VEGF) and FN, and the phosphorylation of Extracellular signal-regulated kinase (ERK) 1/2 were investigated. Identical experiments were performed in the presence of PD98059 our data suggest, a specific ERK 1/2 inhibitor. Shikonin significantly promoted hGF proliferation and migration. Shikonin (1 µM) was chosen as the optimal concentration. Shikonin promoted type I collagen and FN synthesis, increased VEGF and FN expression, and induced ERK 1/2 phosphorylation. These changes were partially suppressed by PD98059. In conclusion, Shikonin promoted the proliferation, migration, type I collagen and FN synthesis, and expression of VEGF and FN via ERK 1/2 signaling pathway in hGFs. Therefore, shikonin may promote periodontal tissue wound healing.

Periodontitis is an inflammatory lesion in the periodontal tissue. The behavior of human periodontal ligament stem cells (hPDLSCs), which play an important role in periodontal tissue regeneration, is restricted by the influence of inflammatory mediators. Photobiomodulation therapy exerts anti-inflammatory effects. The purpose of this study was to investigate the effects of light-emitting diode (LED) irradiation on the inflammatory responses of hPDLSCs. The light source was a red LED (peak wavelength: 650 nm), and the total absolute irradiance was 400 mW/cm². The inflammatory response in hPDLSCs is induced by tumor necrosis factor (TNF)-α. Adenosine triphosphate (ATP) levels and pro-inflammatory cytokine (interleukin [IL]-6 and IL-8) production were measured 24 h after LED irradiation, and the effects of potassium cyanide (KCN) were investigated. LED irradiation at 6 J/cm² significantly increased the ATP levels and reduced TNF-α-induced IL-6 and IL-8 production. Furthermore, the inhibitory effect of LED irradiation on the production of pro-inflammatory cytokines was inhibited by KCN treatment. The results of this study showed that high-intensity red LED irradiation suppressed the TNF-α-stimulated pro-inflammatory cytokine production in hPDLSCs by promoting ATP synthesis. These results suggest that high-intensity red LED is a useful tool for periodontal tissue regeneration in chronically inflamed tissues.

Enamel matrix derivative (EMD) is applied for periodontal therapy. We created a synthetic amelogenin peptide (SP) derived from EMD, and have previously investigated the biological function of SP. However, it is unknown whether SP affects odontoblastic differentiation. In this study, we tested the effects of SP in the odontoblast-like cells, KN-3 cells. KN-3 cells were cultured with SP (0 to 1000 ng/mL) and then cultured for 3, 8, 24, or 48 h in order to determine the effects of SP on cell proliferation and detect its optimum concentration. KN-3 cells were treated with SP in odontogenic differentiation medium cultured for 3 or 7 days. Odontogenic markers were measured by the detection of alkaline phosphatase (ALP) activity and dentin sialo phosphoprotein (DSPP) expression, the calcified nodule formation, and calcium deposition. The addition of SP significantly promoted cell proliferation at 100 ng/mL, generating the greatest change in cell proliferation. SP also showed increased odontogenic expression markers and mineralization. These results suggest that SP, derived from EMD, could have potential for application in dental pulp capping.

Shikonin is a naturally occurring compound known for its anti-inflammatory properties and wound-healing effects. The increased production of reactive oxygen species (ROS) causes oxidative stress and influences the progression of periodontal disease. MiR-146a, a type of microRNA, is reported to be involved in the control of inflammation. This research seeks to examine the anti-inflammatory and antioxidant properties of shikonin on human gingival fibroblasts (hGFs) under inflammation induced by Interleukin (IL)-1β, which is associated with the progression of periodontitis and to further investigate the effect of shikonin on miR-146a expression. Cell proliferation, scratching, and toxicity tests were performed, and the shikonin concentration was set at 0.1 µM. The levels of ROS, miR-146a, and inflammatory cytokines were measured to examine the effects of shikonin on inflammation. The addition of shikonin reduced IL-6 by approximately 20% and reduced IL-8 by approximately 30% compared to IL-6 and IL-8 production and expression induced by IL-1β. MiR-146a expression increased 3.38-fold under IL-1β-induced inflammation but showed a decreasing trend with the addition of shikonin. These results suggest that IL-1β-induced inflammation and oxidative stress are regulated by shikonin in hGFs and are associated with the expression of miR-146a.

Enamel matrix derivative (EMD) is used for periodontal tissue regeneration therapy. We designed a synthetic amelogenin peptide (SP) derived from EMD, and have previously investigated the biological function of SP. However, it is unknown whether SP affects odontoblastic differentiation. In the present study, we investigated the effects of SP in odontoblast-like cells, KN-3 cells. KN-3 cells were treated with SP (0, 1, 10, 100, or 1000 ng/mL) and then cultured for 3, 8, 24, or 48 hours, in order to determine the effects of SP on cell proliferation and detect its optimum concentration. To investigate the effect of SP on odontogenic differentiation, KN-3 cells were treated with SP in odontogenic differentiation medium cultured for 3 or 7 days. Odontogenic differentiation was performed by measuring alkaline phosphatase (ALP) activity, the mRNA expression of dentin sialophosphoprotein (DSPP), the formation of calcified nodules, and calcium deposition into the extracellular matrix. The addition of SP significantly promoted KN-3 cell proliferation; a concentration of 100 ng/ml generated the greatest change in cell proliferation. SP also showed increased expression of markers of odontogenic differentiation and mineralization. These results suggest that SP, derived from EMD, could be a potential for applicate to the dental pulp capping.