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Aims: Accumulating evidence suggests that Porphyromonas gingivalis is closely associated with the development of various chronic inflammatory diseases, particularly periodontitis. This study investigated the antibacterial activity and action mechanism of a novel antimicrobial peptide (AMP), DP7, against P. gingivalis. Methods and Results:The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for DP7 were determined via a broth microdilution method, revealing an MIC of 8 μg ml −1 and MBC of 32 μg ml −1 . Growth inhibition and killing assays confirmed the bactericidal effect of DP7, and treatment with DP7 at MBC eliminated P. gingivalis within 8 h. DP7 had a low cytotoxic effect against human cells. Transmission electron microscopy revealed that DP7 destroyed the bacterial membrane, and confocal laser scanning microscopy revealed its inhibitory effect on P. gingivalis biofilms. Quantitative reverse transcription-polymerase chain reaction revealed DP7-mediated inhibition of several virulence factor genes, partially explaining its antibacterial mechanism.Conclusions: DP7, a novel AMP with low mammalian cytotoxicity, inhibits both planktonic and biofilm forms of P. gingivalis by destroying the bacterial membrane and reducing virulence factor gene expression.Significance and Impact of the Study: DP7 has potential clinical application in the prevention and treatment of P. gingivalis-associated diseases.

Background and objective As an important mediator of intercellular interaction and formation of extracellular bone matrix, porous scaffolds are widely used for bone regeneration. Accumulating evidences demonstrate that microRNA are involved in the regulation of scaffolds-induced bone regeneration. Recently, we revealed that miR-210-3p was highly expressed during osteogenesis induced by HAG. In present study, we further explored the molecular mechanism underlying the effect of miR-210-3p on osteogenic differentiation. Materials and methods In this study, miR-210-3p mimics and inhibitors were synthesized and transfected into MC3T3-E1 cells to explore their effects on osteogenic differentiation. The expression of osteogenic marker (Alp and Runx2) were detected by real-time quantitative PCR (qRT-PCR) and western blotting. After osteogenesis induction for 7 days, Alp staining were used to detected osteoblast differentiation of MC3T3-E1 cells. CCK8 and Transwell assays were performed to detected cell proliferation and migration. Then, top ranking list of target genes of miR-210-3p obtained from TargetScan and the expression of BDNF were detected by qRT-PCR and ELISA. The relationship between miR-210-3p and BDNF was verified by luciferase report assay. Furthermore, the effect of BDNF on osteoblast differentiation was verified by transfecting siRNA or adding BDNF to the culture medium. Results MiR-210-3p mimics markedly suppress osteogenic differentiation, cell migration and cell proliferation of MC3T3-E; nevertheless, silencing of miR-210-3p dramatically enhanced MC3T3-E1 osteogenesis, cell migration and proliferation. Furthermore, luciferase reporter assay verified that brain derived neurotrophic factor (BDNF) is a directly target of miR-210-3p. Moreover, BDNF siRNA significantly decreased the expression levels of ALP and cell migration. The addition of BDNF partially rescued the inhibition of osteogenesis by miR-210-3p. Conclusion miR-210-3p inhibited the osteogenic differentiation via targeting BDNF. Our Results provide a promising target for regulating osteogenic differentiation.

Background and objective Hydroxyapatite scaffolds with different morphologies have been widely used in bone tissue engineering. Moreover, microRNAs (miRNAs) have been proven to be extensively involved in regulating bone regeneration. We developed grooved porous hydroxyapatite (HAG) scaffolds with good osteogenic efficiency. However, little is known about the role of miRNAs in HAG scaffold‐mediated promotion of bone regeneration. The objective of this study was to reveal the mechanism from the perspective of differential miRNA expression. Methods Scanning electron microscopy (SEM) was used to perform the coculture of cells and scaffolds. The miRNA profiles were generated by a microarray assay. A synthetic miR‐129‐5p mimic and inhibitor were used for overexpression or inhibition. The expression of osteogenic marker mRNAs and proteins was detected by quantitative real‐time PCR (qRT–PCR), Western blotting, and immunofluorescence. An ALP activity kit and alizarin red staining (ARS) were used to measure ALP activity and mineral deposition formation. Cell migration ability was examined by wound healing and transwell assays. Protein kinase A (PKA) activity was measured by enzyme‐linked immunosorbent assay (ELISA) after miR‐129‐5p transfection. Target genes were identified by a dual‐luciferase reporter assay. H89 preculture evaluated the cross talk between miR‐129‐5p and PKA activity. Heterotopic implantation models, hematoxylin–eosin (HE), immunohistochemistry staining, and micro‐CT were used to evaluate miR‐129‐5p osteogenesis in vivo. Results miRNAs were differentially expressed during osteogenic differentiation induced by HAG in vitro and in vivo. miR‐129‐5p was the only highly expressed miRNA both in vitro and in vivo. miR‐129‐5p overexpression promoted osteoblast differentiation and cell migration, while its inhibition weakened the effect of HAG. Moreover, miR‐129‐5p activated PKA to regulate the phosphorylation of β‐catenin and cAMP‐response element binding protein (CREB) by inhibiting cAMP‐dependent protein kinase inhibitor alpha (Pkia). H89 prevented the effects of miR‐129‐5p on osteogenic differentiation and cell migration. HE, immunohistochemistry staining and micro‐CT results showed that miR‐129‐5p promoted in vivo osteogenesis of the HAG scaffold. Conclusion The HAG scaffold activates Pka by upregulating miR‐129‐5p and inhibiting Pkia, resulting in CREB‐dependent transcriptional activation and accumulation of β‐catenin and promoting osteogenic marker expression.

ObjectiveA lack of relevant research on Lycium barbarum polysaccharide‐glycoprotein (LBP) application in oral diseases. Here, we focused on the effect of LBP on osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) and periodontitis bone loss.MethodsHuman periodontal ligament stem cells (hPDLSCs) were isolated and identified by flow cytometry. Alkaline phosphatase (ALP) activity, Alizarin Red staining, and combined qPCR and Western blot analyses were performed to elucidate the effects of LBP on the osteogenic potential of hPDLSCs. In vivo experiments were performed with the treatment of LBP in rat periodontal model. MicroCT scanning and histological analysis were conducted to evaluate osteogenesis in situ.ResultsHuman periodontal ligament stem cells (hPDLSCs) were successfully isolated and identified with CD90, CD29, and CD45. LBP enhanced hPDLSCs proliferation and migration and promoted RUNX2, ALP, Collagen I, and Osteocalcin expression through activating the ERK1/2 signaling pathway in vitro. The inflammatory factors, including interleukin 6 (IL‐6) and interleukin 8 (IL‐8) were reduced after LBP treatment. Alveolar bone resorption was significantly decreased in the LBP‐treated groups in vivo, and osteoclast was markedly decreased by LBP application.ConclusionLBP promoted hPDLSC osteogenesis by targeting the ERK1/2 signaling pathway and reverse bone loss by reducing inflammation. These findings provided latent hope for LBP application in periodontal therapy.