Research On Growth Inhibition And Radiosensitivity Of Biological Clock Gene Bmal1 In Subcutaneous Xenograft Of Human Nasopharyngeal Carcinoma CNE1 In Nude Mice

Objective: To explore the effects of up-and down-regulation of circadian clock gene Bmal1 on the growth and radiation sensitivity of nasopharyngeal carcinoma CNE1xenograft in nude mice.Methods: Produce four group cells with lentiviral transfection which were CNE1overexpression(CNE1OE),CNE1overexpression negative control(OENC),CNE1short hairpin RNAi3(CNE1sh3),CNE1short hairpin RNAi negative control(CNE1shNC).Verify the expression of Bmal1 protein in the four group cells with Western Blot.The xenograft grew after injecting subcutaneously the four group cells respectively.Measure the size of xenograft on each nude mice.Observe volume variation of each xenograft group after radiation with 15 Gy electron ray of 6-Me V.Measure the expression of mRNA and Protein of Bmal1,P53 and P21 in the xenograft by RT-PCR and Western Blot,respectively.Results: CNE1OE group highly expressed Bmal1 protein while CNE1sh3 was opposed compared with the control group according to the Western Blot,Which indicated that the transfection was successful.Xenograft of the nude mice grew well.The volumes of xenograft in the CNE1OE,CNE1OENC,CNE1sh3,CNE1shNC groups were 10.34±7.21,44.71±26.30,211.84±27.36,40.16±17.63 on the 15 th day and 28.16±13.75? 142.91±94.23 ?372.05±110.72?109.91±49.63 one the 18 th day and 54.90±26.47 ? 240.80±81.43 ? 868.49±294.54 ? 269.18±52.88(Unit:mm3)one the 21 th day,respectively.The volume of CNE1OE xenograft was smaller than that of CNE1OENC while CNE1sh3 was bigger than CNE1shNC obviously(P < 0.05).All the volumes of xenograft of CNE1OE,CNE1OENC,CNE1shNC shrank after radiation(t=4.32,5.38,5.16,P<0.05)and the extent of CNE1OE was the highest,while the variation of CNE1sh3 had no statistical significance.The volume of the CNE1OE group shrank 77.2%±7.4% after radiation.The relative amount of mRNA and Protein of Bmal1,P53,P21 was more high in the CNE1OE group than in the CNE1OENC group,while more lower in the CNE1sh3 than in the CNE1shNC group(P<0.05).Conclusion: Overexpression of Bmal1 gene could inhibit the growth of CNE1xenograft in nude mice and enhance the radiation sensitivity of it while interfering Bmal1 gene could promote the growth of it and lead to radiation resistence.We believe that Bmal1 overexpression leads to P53 and P21 overexpression,thereby inhibiting the growth of the xenograft.