Cytosine-Phosphate-Guanine (CpG)-Oligodeoxynucleotides Confer Protection Against Septic AKI by Enhancing LSK Cell Mobilization in the Spleen

Background: We have shown that Toll-like receptor 9 (TLR9) -IL-17 pathway contributed to the development of septic acute kidney injury (AKI). CpG-oligodeoxynucleotides (ODNs) are specific ligands for TLR9 and have been reported to improve survival and reduce bacterial titers when administered prior to sepsis. We found that CpG-ODN induced marked splenomegaly in mice, peaking around day 7. This study aimed to elucidate the mechanism of CpG–ODN–mediated protection, focusing on the role of splenic responses. Methods: Male C57BL/6J, TLR9 knockout (KO), and IL-17 KO mice were treated intraperitoneally with CpG-ODN or vehicle 7 days before sepsis induction by cecal ligation and puncture (CLP). Blood urea nitrogen (BUN) was measured 18 hours after CLP and survival was subsequently monitored. Lineage- Sca-1+ c-Kit+ (LSK) cells, representing hematopoietic stem cell-enriched populations, were quantified by flow cytometry in spleens harvested 7 days after CpG-ODN administration. To investigate the role of the spleen and LSK cells in survival, mice underwent splenectomy or total body irradiation for LSK cell depletion prior to CpG-ODN treatment and CLP. Results: CpG-ODN induced significant splenomegaly (mean 191mg in CpG-ODN group vs. 71mg in vehicle group, P < 0.0001) and increased splenic LSK cell counts (mean 1,052,854 vs. 56,683 cells, P < 0.0001). These changes were not observed in TLR9 KO or IL-17 KO mice, indicating involvement of TLR9- and IL–17–dependent pathways. CpG-ODN pretreatment significantly improved survival after CLP and reduced BUN levels (39.6 vs. 62.7 mg/dL, P = 0.00065). In contrast, no survival benefit of CpG-ODN pretreatment was observed in TLR9 KO or IL-17 KO mice. Furthermore, the survival advantage conferred by CpG-ODN was lost in mice that underwent splenectomy or total body irradiation for LSK cell depletion, suggesting that the mobilization of LSK cells in the spleen is essential for the protective effect. Conclusion: Pretreatment with CpG-ODN improves sepsis survival and preserves kidney function, potentially through TLR9- IL-17-dependent immune pathways and mobilization of splenic LSK cells. These findings highlight a novel immunomodulatory strategy that may have implications for improving sepsis outcomes.