Ionising radiation exposure can lead to acute haematopoietic radiation syndrome. Despite significant advancements in the field of radioprotection, no drugs with high efficacy and low toxicity have yet been approved by the Food and Drug Administration. FG-4592, as a proline hydroxylase inhibitor, may play an important role in radioprotection of the haematopoietic system. Mice were peritoneal injected with FG-4592 or normal saline. After irradiation, the survival time, body weight, peripheral blood cell and bone marrow cell (BMC) count, cell apoptosis, pathology were analysed and RNA-sequence technique (RNA-Seq) was conducted to explore the mechanism of FG-4592 in the haematopoietic system. Our results indicated that FG-4592 improved the survival rate and weight of irradiated mice and protected the spleen, thymus and bone marrow from IR-induced injury. The number of BMCs was increased and protected against IR-induced apoptosis. FG-4592 also promoted the recovery of the blood system and erythroid differentiation. The results of RNA-Seq and Western blot showed that the NF-κB signalling pathway and hypoxia-inducible factor-1 (HIF-1) signalling pathway were upregulated by FG-4592. Meanwhile, RT-PCR results showed that FG-4592 could promote inflammatory response significantly. FG-4592 exhibited radioprotective effects in the haematopoietic system by promoting inflammatory response and targeting the NF-κB, HIF signalling pathway.

Intestinal stem cells (ISCs) are responsible for intestinal homeostasis and are important for the regeneration of damaged intestine. We established an ionizing radiation (IR)-induced intestinal injury model and observed that Gelsolin KO mice had increased radiosensitivity. The deletion of Gelsolin aggravated intestinal damage and reduced the number of ISCs after lethal IR. The intestinal organoid experiments showed that Gelsolin deletion inhibited ISCs function after IR. Notably, RNA sequencing and RT-PCR results showed IL-17 signaling pathway was down-regulated and Th17 cells differentiation was inhibited in Gelsolin KO mice. Moreover, recombinant IL-17 A ameliorated IR-induced intestinal injury and promoted ISCs regeneration. To figure out the role of Gelsolin in Th17 cells differentiation, flow cytometry was used and we found that Gelsolin targets Th17 cells functionality via the p-STAT3/RORγt axis. By establishing the co-culture system, we proved that Th17 cells promoted self-renewal and budding abilities in Gelsolin-deficient organoids. Finally, we found that Gelsolin was protective against DSS-induced colitis and that this protective effect was not specific or limited to the IR induced intestinal injury model. Based on these results, we proved Gelsolin maintained the regeneration of ISCs by sustaining Th17 cells functions via the p-STAT3/RORγt axis.

Background: Intestinal tissue is extremely sensitive to ionizing radiation (IR), which is easy to cause intestinal radiation sickness, and the mortality rate is very high after exposure. Recent studies have found that intestinal immune cells and intestinal stem cells (ISCs) may play a key role in IR-induced intestinal injury. Methods: C57BL6 mice matched for age, sex and weight were randomly grouped and intraperitoneal injected with PBS, Scleroglucan (125.0 mg/kg) or Anti-mouse IL-17A -InVivo (10 mg/kg), the number of mice in each group was n >= 3.Survival time, body weight, pathology, organoids and immune cell markers of the mice after IR (10.0 Gy) were compared, and the mechanism of action in intestinal tissues was verified by transcriptome sequencing. Results: Scleroglucan has significant radiation protective effects on the intestine, including improving the survival rate of irradiated mice, inhibiting the radiation damage of intestinal tissue, and promoting the proliferation and differentiation of intestinal stem cells (ISCs). The results of RNA sequencing suggested that Scleroglucan could significantly activate the immune system and up-regulate the IL-17 and NF-kappa B signaling pathways. Flow cytometry showed that Scleroglucan could significantly up-regulate the number of Th17 cells and the level of IL17A in the gut. IL-17A provides radiation protection. After intraperitoneal injection of Scleroglucan and Antimouse IL-17A -InVivo, mice can significantly reverse the radiation protection effect of Scleroglucan, downregulate the molecular markers of intestinal stem cells and the associated markers of DC, Th1 and Th17 cells, and up-regulate the associated markers of Treg and Macrophage cells. Conclusion: Scleroglucan may promote the proliferation and regeneration of ISCs by regulating the activation of intestinal immune function mediated by IL-17 signaling pathway and play a protective role in IR-induced injury.

Intestinal stem cells (ISCs) play a crucial role in maintaining the equilibrium and regenerative potential of intestinal tissue, thereby ensuring tissue homeostasis and promoting effective tissue regeneration following injury. It has been proven that targeting Toll-like receptors (TLRs) can help prevent radiation-induced damage to the intestine. In this study, we established an intestinal injury model using IR and evaluated the effects of CL429 on ISC regeneration both in vivo and in vitro. Following radiation exposure, mice treated with CL429 showed a significant increase in survival rates (100% survival in the treated group compared to 54.54% in the control group). CL429 also showed remarkable efficacy in inhibiting radiation-induced intestinal damage and promoting ISC proliferation and regeneration. In addition, CL429 protected intestinal organoids against IR-induced injury. Mechanistically, RNA sequencing and Western blot analysis revealed the activation of the Wnt and Hippo signaling pathways by CL429. Specifically, we observed a significant upregulation of YAP1, a key transcription factor in the Hippo pathway, upon CL429 stimulation. Furthermore, knockdown of YAP1 significantly attenuated the radioprotective effect of CL429 on intestinal organoids, indicating that CL429-mediated intestinal radioprotection is dependent on YAP1. In addition, we investigated the relationship between TLR2 and YAP1 using TLR2 knockout mice, and our results showed that TLR2 knockout abolished the activation of CL429 on YAP1. Taken together, our study provides evidence supporting the role of CL429 in promoting ISC regeneration through activation of TLR2-YAP1. And further investigation of the interaction between TLRs and other signaling pathways may enhance our understanding of ISC regeneration after injury.

Accidental radiation exposure causes the acute lethal damage of hematopoietic system and gastrointestinal tract1,2. By establishing an ionizing radiation (IR) induced injury model, we found macrophage-activating lipopeptide-2 (MALP-2) exhibited significant radioprotective effects in mice. MALP-2 improved the survival of irradiated mice, inhibited the radiation-induced gastrointestinal tract damage. Through intestinal organoid experiments, we found that MALP-2 protected the intestinal organoid against IR-induced injury. Next, we identified the differentially expressed genes (DEGs) between PBS and MALP-2 groups based on the RNA sequencing result3. And the RNA-seq results showed that MALP-2 increased the levels of interleukin 6 (IL-6), IL-12, G-CSF, GM-CSF, TNF-α, CCL-3, PGE-2 and SOD2. KEGG enrichment analysis showed that DEGs were significantly enriched in Toll-like receptor signaling pathway and NF-κB signaling pathway. In line with these observations, the expression level of IL-6 and GM-CSF were increased by using flow cytometry. Moreover, MALP-2 protected WT mice from IR induced death but had no radioprotective effects on the TLR2 KO and IL-6 KO mice, suggesting that the radioprotection of MALP-2 was mediated by activating TLR2/IL-6 axis. In conclusion, our data suggested that the MALP-2 could induce significant radioprotective effects and MALP-2 might be a potential radioprotective agent.

Intestinal stem cells (ISCs) are responsible for intestinal tissue homeostasis and are important for the regeneration of damaged intestinal epithelia. Through the establishment of ionizing radiation (IR) induced intestinal injury model, we found that the radiosensitivity of the intestine of GSN KO mice increases significantly. It means that GSN played a key role in the process of radiation-induced intestinal injury. The deletion of GSN aggravated the radiation damage of the intestine and decreased the survival rate of intestinal crypts and the number of ISCs after lethal IR in vivo. Through intestinal organoid experiments, we found that the deletion of GSN inhibited the proliferation and differentiation of ISCs after IR. Remarkably, the results of RNA sequencing and RT-PCR showed that IL-17 signaling pathway was down-regulated and Th17 cells differentiation was inhibited in GSN KO mice. Moreover, recombinant IL-17A ameliorated IR-induced intestinal injury and promoted the ISCs regeneration in GSN KO mice and GSN-deficient organoids. To figure out the role of GSN in Th17 cells differentiation, flow cytometry was used and we found that Gelsolin targets Th17 cells functionality via p-STAT3/RORγt signaling pathway. Finally, by establishing the co-culture system of intestinal organoids and Th17 cells, we found that decreased self-renewal and budding efficiency in GSN-deficient organoids were rescued by Th17 cells. In conclusion, we demonstrated that GSN maintained the regeneration of ISCs by sustaining Th17 cells functions via p-STAT3/RORγt axis.Funding: This study was supported in part by the grants from Youth Training Program (No. 21QNPY034), Shanghai Sailing Program (No. 19YF1459100), and National Natural Science Foundation of China (No. 81903260, No. 81872559, No. 82173459, No.81372932). These support the experimental research and data collection of the subject.Declaration of Interest: The authors confirmed that there are no conflicts of interest.Ethical Approval: All animal experiments conformed to the National Institute of Health Guide for the Care and Use of Laboratory Animals' (NIH Publication No. 85-23, National Academy Press, Washington, DC, revised 1996), with the approval of the Laboratory Animal Center of the Naval Medical University, Shanghai.