Nocodazole Inhibits HFF Cell Cycle And Rotenone Promotes Senescence Of HFF Cells

Objective:The HFF cell line carrying the mAG-hGeminin-GFP reporter plasmid was constructed and the effect of Nocodazole on the cell cycle of HFF was investigated by Nocodazole intervention.To study the effects of Rotenone on the senescence of HFF cells,explore the molecular mechanisms underlying Rotenone's promotion of senescence.Methods:1.The mAG-hGeminin-GFP plasmid was constructed,packaged with lentivirus,infected with 293T cells,and 293T culture medium was collected to infect HFF cells.2.Thymidine-Nocodazole was sequentially administered to treat mAG-hGeminin-GFP-HFF cells,and the dynamic changes of cell cycle were observed by flow cytometry.3.Different concentrations of Rotenone-treated HFF cells were observed under microscope for morphological changes of cells at various time points and counted.4.Western blot analysis showed changes in P21 and P16 protein expression in HFF cells.Results:1.Successfully constructing HFF cells carrying the mAG-hGeminin-GFP reporter plasmid;2.As the drug withdrawal time prolongs,more cells are arrested in the G1-S phase,GFP+cells are less and less,and senescent cells are enriched in GFP+:3.Compared with the Rotenone dosing group,the cells in the dosing group increased in volume,the nucleus was pyknosis,the boundary was unclear,and the cells died in large numbers;4.Compared with the control group,the number of cells in the Rotenone 10?M administration group was sigruficantly decreased(P<0.0001),and the number of cells in the Rotenone 20?M administration group was significantly decreased(P<0.0001).The drug-administered group was compared with each other and there were significant statistics.Learning difference(P<0.01);5.Compared with the control group,the expression of P16 and P21 protein in the drug-administered group was increased(P<0.05).Conclusion:HFF cells carrying the mAG-hGeminin-GFP reporter plasmid were successfully established,and the dynamic changes of the cell cycle were visualized for easier understanding and observation.Sequential administration of Thymidine-Nocodazole blocks most of the HFF cells in the Gl-S phase,leading to cell senescence.Rotenone may promote HFF cell senescence through the P21/P16 signaling pathway.Figure[6]table[2]reference[59]...