The Effects And Mechanism Of Cannabidiol On Photoaging Dermal Fibroblasts

Objective:To investigate the effect of Cannabidiol(CBD)for the therapeutic potential mechanism of against photoaging damage,in UVB-induced dermal fibroblast model.Methods:Human dermal fibroblasts in logarithmic growth phase were treated with UVB dose of 60,120,180,240mJ/cm2.Determined the appropriate UVB radiation dose by detecting the cell proliferation ability,reactive oxygen species and active caspase-3 expression.Detected cell cycle,redox balance,inflammatory factors and extracellular matrix components and other aging phenotypes under this irradiation dose,and performed subsequent CBD administration experiments.Different concentrations of CBD were respectively treated for 6 hours before and after UVB irradiation HDFa,then observed and evaluated the phenotypic changes of aging.The aging-related signal pathways(p53,p21 and p16),MAPK signal pathway,transcription factors AP-1 and NF-κB were detected by RT-PCR to explore the relevant mechanisms.Results:1.180mJ/cm2 UVB irradiation reduced cell proliferation,increased in ROS generation rather than cell death,and displayed a senescence-associated secretory phenotype,including cell cycle arrest,redox balance disorder,Increased expression of MMP-l,and especially the degradation of collagen type I.2.CBD promoted DNA synthesis,and the regulatory effect on p53,p21 and p16 is related to different administrations of CBD.EdU incorporation assay revealed that compared with the UVB irradiation group,the rate of EdU positive cells increased after CBD treatment.Real-time quantitative PCR revealed that,compared with the UVB irradiation group,the mRNA of p53,p21 and p16 in the CBD pretreatment groups remained unchanged.The mRNA of p53,p21 and p16 in the CBD treatment groups were significantly down-regulated.In this experiment,CBD failed to alleviate the cell cycle arrest induced by UVB irradiation.3.CBD regulated redox balance and promoted cell proliferation.After CBD treatment,the level of ROS in the cell decreased.The enzyme activity test revealed that,compared with the UVB irradiation group,CBD pretreatment inhibited the decrease of SOD and CAT enzyme activity,inhibited the increase of GSH-Px enzyme activity,and reduced the MDA content.CBD treatment groups restored SOD and GSH-Px enzyme activity and reduced MDA content.CBD pretreatment groups and treatment groups both promoted cell proliferation.4.CBD inhibited the expression of MMP-1 and promoted the expression of type I collagen.Real-time quantitative PCR revealed that compared with the UVB irradiation group,the mRNA of type I collagen and tropoelastin in the CBD pretreatment groups and the CBD treatment groups were significantly up-regulated,and the mRNA of MMP-1 was significantly down-regulated.TIMP-1 remained unchanged.The results of cellular immunofluorescence and ELISA were consistent with the results of RT-qPCR.5.CBD down-regulated the mRNA of MAPK signaling pathway and transcription factors,and inhibited photoaging-related inflammation.Real-time quantitative PCR revealed that,compared with the UVB irradiation group,CBD down-regulated the mRNA of MAPK signal pathway and transcription factor AP-1 and NF-κB.ELISA showed that CBD inhibited the secretion of downstream inflammatory factors such as TNF-α and IL-6.Conclusions:1.Our study illustrated that exposure of human dermal fibroblasts to UVB at a dose of 180mJ/cm2 induced cell senescence and displayed a senescence-associated secretory phenotype,while the underlying molecular mechanisms,including signaling pathways and transcription factors,remain to be featured further.2.CBD played an anti-oxidant role by restoring the activity of antioxidant enzymes and removing lipid peroxides,and inhibited the secretion of TNF-α and IL-6 by down-regulating the MAPK signaling pathway and transcription factors(AP-1 and NF-κB),while exerted an anti-inflammatory effect in the HDFa photoaging model.In addition,CBD down-regulated the expression of MMP-1,inhibited the degradation of the main components of ECM,and promoted the expression of type I collagen and tropoelastin.3.CBD repaired UVB-induced DNA damage by down-regulating the mRNA of p53,p21 and p16.The effect is related to the way of administration.