Neuroprotective Effects And Mechansism Of Tamoxifen On Irradiation Induced Brain Injury In Vitro And In Vivo

Objective:Microglial cells, the immune cells in the CNS, are significantly activated after whole brain irradiation and produce a large number of pro-inflammatory cytokines which are detrimental to brain tissues and the main reason of cognitive impairment. Tamoxifen has been found to be a radiosensitizer and a potent neuroprotective agent in both transient and permanent experimental ischemic stroke, also in a spinal cord injury model. In the present study, we sought to determine the influence of tamoxifen on inflammatory response induced by the irradiated microglia in vitro, and provide the experimental basis for the neuroprotection of tamoxifen in the brain injury induced by whole brain irradiation in vivo.Materials and Methods:BV-2 cells (a murine microglial cell line) were irradiated by various radiation doses (2-10 Gy), and the expression of inflammatory factor IL-1βand TNF-αexcreted by irradiated microglia was determined by ELISA. BV-2 cells were divided into the control group, irradiation group, tamoxifen group. BV2 murine microglia cells were exposed to a single dose of 10 Gy and 24 h later the conditioned culture medium of irradiated microglia was added into the astroglial and primary neuronal culture, respectively. For tamoxifen group, incubation BV-2 cells with tamoxifen (1μM) for 45 min was carried out to determine the possible neuroprotective mechansism of tamoxifen on irradiation induced brain injury.Results:Our results showed that irradiating BV-2 cells (a murine microglial cell line) with various radiation doses (2-10 Gy) led to the increase in IL-1βand TNF-αexpression determined by ELISA, and the conditioned culture medium of irradiated microglia with lOGy radiation dose initiated astroglial activation and decreased the number of neuronal cells in vitro. Incubation BV-2 cells with tamoxifen (1μM) for 45 min significantly inhibited the radiation-induced microglial inflammatory response.Conclusion:Tamoxifen can decrease the irradiation-induced brain damage including reactive astrogliosis and neuronal death via attenuating the microglial inflammatory response in vitro. Objective:It has been shown that tamoxifen is a potent neuroprotective agent in both transient and permanent experimental ischemic stroke, also in a spinal cord injury model. In the section one of this study, we also found that tamoxifen exerts a powerful anti-inflammatory role in BV2 microglial cells induced by X irradiation. In the section two, we investigated to determine whether tamoxifen exerts neuron-protective effect on brain injury induced by whole brain irradiation in adult rats.Materials and Methods:All animals were divided into three groups:control group, whole brain irradiation (WBI) group and tamoxifen treatment group. Irradiation induced brain injury model was carried out by a single dose of 15 Gy was given at a dose rate of 3 Gy/min using a 6 MV Elekta Precise X-ray linear accelerator. All rats were killed at 1d and 3d post irradiation respectively and then the samples were harvested for further use. Microglial activation, astroglial activation and neuronal apoptosis were observed by immunofluorescence. Expression level of OX-42 and GFAP protein in each group was semi-quantified by western blot analysis. The brain tissue water content and Evans blue content after whole brain irradiation were measured.Results:After irradiation, several astrocytes became activated at day 3 after WBI in the cerebral cortex regions with reactive, hypertrophied morphology and increase in OX-42/GFAP immunoreactivity. In addition, many TUNEL-positive neurons were observed in the area CA1 hippocampus after WBI. Furthermore, measurement of brain water content and Evans blue extravasations were significantly increased. When treated with tamoxifen, activation in microglia and astrocyte, neuronal apoptosis were attenuated significantly after irradiation. And the increase in brain water content and Evans blue extravasations was attenuated markedly in the tamoxifen treatment group. Conclusion:We consider that tamoxifen significantly alleviated disruption of BBB permeability and tissue edema formation, attenuated microglial activation, reactive astrogliosis and neuronal cell death after WBI in rats. In turn, the future therapeutic strategies can be designed to achieve efficient compensation of chronic cognitive impairment after whole brain irradiation.