Reprimo (RPRM) as a Potential Preventive and Therapeutic Target for Radiation-Induced Brain Injury via Multiple Mechanisms.

Patients receiving cranial radiotherapy for primary and metastatic brain tumors may experience radiation-induced brain injury (RIBI). Thus far, there has been a lack of effective preventive and therapeutic strategies for RIBI. Due to its complicated underlying pathogenic mechanisms, it is rather difficult to develop a single approach to target them simultaneously. We have recently reported that Reprimo (RPRM), a tumor suppressor gene, is a critical player in DNA damage repair, and RPRM deletion significantly confers radioresistance to mice. Herein, by using an RPRM knockout (KO) mouse model established in our laboratory, we found that RPRM deletion alleviated RIBI in mice via targeting its multiple underlying mechanisms. Specifically, RPRM knockout significantly reduced hippocampal DNA damage and apoptosis shortly after mice were exposed to whole-brain irradiation (WBI). For the late-delayed effect of WBI, RPRM knockout obviously ameliorated a radiation-induced decline in neurocognitive function and dramatically diminished WBI-induced neurogenesis inhibition. Moreover, RPRM KO mice exhibited a significantly lower level of acute and chronic inflammation response and microglial activation than wild-type (WT) mice post-WBI. Finally, we uncovered that RPRM knockout not only protected microglia against radiation-induced damage, thus preventing microglial activation, but also protected neurons and decreased the induction of CCL2 in neurons after irradiation, in turn attenuating the activation of microglial cells nearby through paracrine CCL2. Taken together, our results indicate that RPRM plays a crucial role in the occurrence of RIBI, suggesting that RPRM may serve as a novel potential target for the prevention and treatment of RIBI.

Irradiated microvascular endothelial cells may induce bystander effects in neural stem cells leading to neurogenesis inhibition.

Radiation-induced neurocognitive dysfunction (RIND) has attracted a lot of attention lately due to the significant improvement of the survival of cancer patients after receiving cranial radiotherapy. The detailed mechanisms are not completely understood, but extensive evidence supports an involvement of the inhibition of hippocampal neurogenesis, which may result from radiation-induced depletion of neural stem cells (NSCs) as well as the damage to neurogenic niches. As an important component of neurogenic niches, vascular cells interact with NSCs through different signaling mechanisms, which is similar to the characteristics of radiation-induced bystander effect (RIBE). But whether RIBE is involved in neurogenesis inhibition contributed by the damaged vascular cells is unknown. Thus, the purpose of the present study was to investigate the occurrence of RIBEs in non-irradiated bystander NSCs induced by irradiated bEnd.3 vascular endothelial cells in a co-culture system. The results show that compared with the NSCs cultured alone, the properties of NSCs were significantly affected after co-culture with bEnd.3 cells, and further change was induced without obvious oxidative stress and apoptosis when bEnd.3 cells were irradiated, manifesting as a reduction in the proliferation, neurosphere-forming capability and differentiation potential of NSCs. All these results suggest that the damaged vascular endothelial cells may contribute to neurogenesis inhibition via inducing RIBEs in NSCs, thus leading to RIND.

RPRM negatively regulates ATM levels through its nuclear translocation on irradiation mediated by CDK4/6 and IPO11.

How the ataxia telangiectasia mutated (ATM) protein kinase, a core protein in DNA damage response, is regulated at post-transcription level remains unclear. Here it is identified that protein Reprimo (RPRM) downregulates ATM protein levels, resulting in impaired DNA repair and enhanced cellular radiosensitivity. Mechanistically, although primarily localized in the cytoplasm, RPRM translocates to the nucleus shortly after induced by X-irradiation, interacts with ATM and promotes its nuclear export and proteasomal degradation. The RPRM nuclear translocation involves its phosphorylation at serine 98 mediated by cyclin-dependent kinases 4/6 (CDK4/6), and requires Importin-11 (IPO11). Of importance, IPO11-regulated RPRM nuclear import upon irradiation is essential for its regulation on ATM. Thus, RPRM overexpression and its phosphorylation inhibition sensitize cells to genotoxic agents such as irradiation, whereas RPRM deficiency significantly increases resistance to radiation-induced damage both in vitro and in vivo. These findings establish a crucial regulatory mechanism in which ATM is negatively modulated by RPRM.

Radiation-induced bystander effects may contribute to radiation-induced cognitive impairment.

PURPOSE: Despite being a major treatment modality for brain cancer due to its efficiency in achieving cancer control, radiotherapy has long been known to cause long-term side effects, including radiation-induced cognitive impairment (RICI). Neurogenesis inhibition due to radiation-induced damage in neural stem cells (NSCs) has been demonstrated to be an important mechanism underlying RICI. Radiation-induced bystander effects (RIBEs) denote the biological responses in non-targeted cells after their neighboring cells are irradiated. We have previously demonstrated that RIBEs could play an important role in the skin wound healing process. Therefore, we aimed to investigate whether RIBEs contribute to RICI in this study. MATERIALS AND METHODS: The transwell co-culture method was used to investigate bystander effects in mouse NSCs induced by irradiated GL261 mouse glioma cells in vitro. The proliferation, neurosphere-forming capacity and differentiation potential of NSCs were determined as the bystander endpoints. The exosomes were extracted from the media used to culture GL261 cells and were injected into the hippocampus of C57BL/6 mice. Two months later, the neurogenesis of mice was assessed using BrdU incorporation and immunofluorescence microscopy, and cognitive function was evaluated by the Morris Water Maze. RESULTS: After co-culture with GL261 glioma cells, mouse NSCs displayed inhibited proliferation and reduced neurosphere-forming capacity and differentiation potential. The irradiated GL261 cells caused greater inhibition and reduction in NSCs than unirradiated GL261 cells. Moreover, adding the exosomes secreted by GL261 cells into the culture of NSCs inhibited NSC proliferation, suggesting that the cancer cell-derived exosomes may be critical intercellular signals. Furthermore, injection of the exosomes from GL261 cells into the hippocampus of mice caused significant neurogenesis inhibition and cognitive impairment two month later, and the exosomes from irradiated GL261 cells induced greater inhibitory effects. CONCLUSION: RIBEs mediated by the exosomes from irradiated cancer cells could contribute to RICI and, therefore, could be a novel mechanism underlying RICI.