| Theaflavin is an important quality component and functional component of black tea.Recently,theaflavins have been attracted considerable interests because of their potential benefits for human health,including blood lipid-lowering,antioxidation,antiaging and antimutagenicity.UVB-induced cellular damage was established and the theaflavin mixture and four theaflavin monomers were used to intervene the model.The related indicators of epidermal cell senescence were detected and the anti-aging effects of theaflavins were analyzed.It will provide a theoretical basis for the prevention and treatment of skin photoaging with the active ingredient theaflavins in black tea.The main results are as follows:1.The effect of theaflavins mixture and four theaflavin monomers on the proliferation rate of photoaging(Ha CaT:human keratinocytes cells)cells was determined by MTT method.Theaflavins mixture(5-100 ?g/m L)could significantly increase cell viability and promote cell proliferation & the cell activity was gradually enhanced with the theaflavins extract concentration(0-25?g/mL).The four theaflavin monomers(theaflavins,TF,theaflavin-3-gallate,TF3 G,theaflavin-3?-gallate,TF3'G,theaflavin-3,3?-digallate,TFDG)could improve the relative viability of cells and promote cell proliferation in varying degrees and TF3'G had the best effect on improving the activity of HaCaT cells.The EC50 values of TF,TF3 G,TF3 'G and TFDG were obtained as follows respectively: 0.427±0.032,0.196±0.008,0.175±0.023 and 0.278±0.057.The effects of theaflavins monomers on the proliferation rate of photoaging HaCaT cells were in the order: TF3'G>TF3G>TFDG>TF.2.Based on photoaging model,the protective effect of TF3'G on ultraviolet ray B(UVB)induced HaCaT cells was studied.The activity of intracellular antioxidant enzyme(GSH-Px,SOD,CAT),oxidation product(MDA)content and oxidation reaction(LDH),intracellular oxidation level(ROS),changes of mitochondrial membrane potential and senescence related protein mRNA expression were detected.Compared with the model group,the activities of SOD,CAT,GSH-Px in photoaging(HaCaT)cells with TF3'G were significantly increased with the theaflavins extract concentration,while theactivities of LDH,MDA and ROS in photoaging(HaCaT)cells with TF3'G were significantly decreased with the theaflavins extract concentration.The cell membrane potential was gradually increased.A large number of free radicals were produced in UVB induced HaCaT cells,mitochondrial membrane potential was decreased and apoptotic rate was increased.TF3'G could reduce the production of cell free radicals,inhibit the decline of mitochondrial membrane potential and reduce the apoptotic rate.The results showed that TF3'G had significant protective effect on HaCaT induced by UVB radiation.3.Annexin V-FITC kit was used to detect the effect of theafin on apoptosis of photoaging(HaCaT)cells;RT-PCR was used to detect the effect of TF3'G on the mRNA expression levels of SOD,CAT,Bax,Bcl-2,Mfn2,Opal and Caspase-3 in photoaging(HaCaT)cells.Compared with the blank control group,the apoptosis rate in the model group was significantly increased,the mRNA expression levels of SOD,CAT,Bcl-2,Mfn2 and Opa1 in the model group were significantly decreased,and the mRNA expression levels of Bax and Caspase-3 were significantly increased.Compared with the model group,the apoptosis rate of 5,10,20,40 ?M theaflavin monomer TF3'G group was significantly reduced,the mRNA expression levels of SOD,CAT,Bcl-2,Mfn2 were increased in TF3'G group,The increase of Opa1 mRNA expression level was not significant and the mRNA expression levels of Bax and Caspase-3 were decreased.The results showed that TF3'G could promote RNA expression of antioxidant enzymes(SOD,CAT),mitochondrial fusion genes Mfn2 and anti-apoptotic factor Bc1-2,but inhibit the expression of apoptotic factor Bax & reduce the expression of Caspase family.Therefore TF3'G could inhibit the apoptosis of HaCaT cells induced by UVB and protect HaCaT cells induced by UVB. |
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