| Ionizing radiation refers to all radiation that can cause the ionization of materials.Characteristics of ionizing radiation include short wavelength, high frequency and high energy. It can be categorized into different classes: alpha particles, beta particles and the protons are high-speed charged particles; while neutrons, X-ray and gamma rays are not charged particles. Ionizing radiation is also known as mutagens, excessive amount of radiation can cause radiation sickness, cancer and the death or deformity of fetus.A major paradigm shift in radiation biology in the past decade has resulted from the studies of bystander effect. Radiation-induced bystander effect(RIBE) is defined as the induction of biological effects in cells that are not directly traversed by a charged particle but are in close proximity to cells that are, or have received signals from these irradiated cells. In such a scenario, extranuclear and/or extracellular events may also contribute to the final biological consequences of radiation. Biological effects caused by RIBE include apoptosis or delayed cell death, genetic instability, mutation, changes of gene expressions,inflammation, micronucleus formation and abnormal cell growth. However, RIBE related molecular mechanism(s) is still not very clear so far. It is generally believed that RIBE is caused by gap junction and/or signal transduction through cytokines secreted by directly irradiated cells.Protein kinase C is a family of multifunctional serine and threonine kinases. They are involved in a number of important reactions within cells, including cell differentiation,apoptosis, cell skeleton repair and the regulation of ion channels. Previous reports showed that that PKC may participate in radiation-induced bystander effects, but the detailed mechanism remains unclear. In the current study, we utilized Chinese hamster ovary cell line AL as the study model and explored the role of PKC in the signal transduction pathway that lead to RIBE.The results obtained are as follows:1. Protein kinase C(PKC) translocated from cytosol to cell membrane fraction inbystander cells after 0.5Gy alpha particle irradiation. MAPK(Mitogen-activatedprotein kinase)pathway member ERK(Extracellular regulated protein kinases)was activated and cyclooxygenase-2(COX-2), which has been shown to beclosely related with radiation-induced bystander effect, was up-regulated. Further,CD59-mutation yield was significantly increased in bystander cells as well. Whencells were treated with PKC translocation inhibitor chelerythrine chloride 1 hourbefore irradiation, the translocation of PKC, activation of ERK and up-regulationof COX-2 was attenuated. Mutation yield of CD59 was suppressed as well. Theseresults indicated that translocation and activation of PKC may be the reason ofRIBE;2. The level of TNF alpha in was increased in direct irradiation cells but showed noapparent change in bystander cells. On the other hand, the level of TNF alphareceptors TNFR1 was significantly elevated in bystander cells, implicating thatTNF alpha released from directly irradiation cells may lead to the occurrence ofbystander effect;3. When TNF alpha was directly added into the culture medium, PKC translocatedfrom cytosol to cell membrane fraction. In turn, ERK was activated and COX-2up-regulated. Mutation yield of CD59 was increased as well. Addition of TNFalpha neutralizing antibody attenuated these effects, suggesting that TNF alpha isthe reason of bystander effect.4. We also treated cells with PKC activator PMA(Phorbol 12-myristate 13-acetate)and found out that after 15, 30 and120minutestreatment,thelevelofTNFR1inmembrane fraction was significantly increased. Further analysis demonstrated thatthe activation of PKC accelerated TNFR1 internalization and recycling, However,the effect on recycling was more significant than on TNFR1internalization,whichresulted in a higher amount of the receptor present on cell surface after activationof PKC... |
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