Effects Of Cannabidiol On The Proliferation And Osteogenic Differentiation Of Human Periodontal Ligament Stem Cells

Background:Periodontitis,aging,trauma,and metabolic diseases often cause alveolar bone resorption and bone tissue defects,which can lead to loosening and loss of teeth and is not conducive to various kinds of denture restorations.It has a negative effect on mastication,pronunciation and aesthetics of patients.At present,the commonly used treatment methods still have certain limitations,and how to promote the repair of bone defects has become a research hotspot for scholars.Among them,stem cell therapy that combines seed cells with osteogenic drugs is an attractive option.Periodontal ligament stem cells can differentiate into osteoblasts under certain conditions and have strong anti apoptotic and proliferative capacities,making them excellent seed cells for bone tissue and periodontal tissue regeneration.Studies have found that a variety of osteogenic drugs can significantly promote the osteogenic differentiation of periodontal ligament stem cells and promote bone tissue formation.Cannabidiol,one of the major non-psychotomimetic components from plant Cannabis sativa,has antioxidant,anti-inflammatory,and other pharmacological effects.Recently,it was shown that cannabidiol could effectively promote the osteogenic differentiation of preosteoblasts and mesenchymal stem cells.However,the effect of cannabidiol on the proliferation and osteogenic differentiation of human periodontal ligament stem cells remains unclear.Objective:In this experiment,the effects of different concentrations of cannabidiol on the proliferation and osteogenic differentiation of human periodontal ligament stem cells were studied in order to provide a theoretical basis for the application of cannabidiol in bone and periodontal regeneration.Methods:1.Human periodontal ligament stem cells were isolated and cultured using explant method.Cell identification was performed by morphological observation,flow cytometry analysis,and multidirectional differentiation assay.2.Human periodontal ligament stem cells were divided into control group and cannabidiol group(concentrations were 0.1,0.5,2.5,12.5 ?mol/L,respectively).After 1,3,5,and 7 days of treatment,the cell proliferation ability of each group was detected by CCK-8 assay.3.Human periodontal ligament stem cells were divided into control group and cannabidiol group(concentrations of 0.1,0.5,and 2.5 ?mol/L,respectively).Through performing alkaline phosphatase activity assay,Alizarin Red S staining and semi quantitative analysis,the osteogenic differentiation ability of each group was assessed.4.Human periodontal ligament stem cells were divided into control group and cannabidiol group(concentrations were 0.1,0.5,2.5 ?mol/L,respectively).The effects of different concentrations of cannabidiol on the expression of osteogenic differentiation-related genes COL1,OCN,RUNX2 and ?-catenin in human periodontal ligament stem cells were assessed by q PCR.Results:1.Human periodontal ligament stem cells isolated and cultured using explant method were spindle-shaped and could form spiral arrangement,positively expressed mesenchymal stem cell specific markers CD73,CD90,and CD105,while negatively expressed CD34 and CD45,with the multilineage differentiation ability of osteogenic differentiation and adipogenic differentiation.2.Compared with the control group,0.1-12.5 ?mol/L cannabidiol had no significant effect on the proliferation of human periodontal ligament stem cells in the first 5days.On day 7,0.1 ?mol/L cannabidiol showed no significant effect on the viability of human periodontal ligament stem cells,whereas 0.5 and 2.5 ?mol/L cannabidiol significantly promoted their proliferation,however,12.5 ?mol/L cannabidiol had obvious cytotoxicity.3.Compared with the control group,0.1-2.5 ?mol/L cannabidiol significantly enhanced alkaline phosphatase activity and mineral deposition of human periodontal ligament stem cells,and its promoting effect increased with the increase of the concentration.4.Compared with the control group,0.1-2.5 ?mol/L cannabidiol up-regulated expression level of osteogenic differentiation related genes COL1,OCN,Runx2,and ?-catenin.Conclusion:1.Cannabidiol modulated human periodontal ligament stem cell proliferation in a concentration and time-dependent manner,of which 2.5 ?mol/L has the most significant promoting effect,and 12.5 ?mol/L has cytotoxicity.2.Cannabidiol promotes the osteogenic differentiation of human periodontal ligament stem cells in a concentration dependent manner,with the most significant effect at 2.5 ?mol/L.