P.g induces CKD by activating the NF‑κB/NLRP3 pathway in a ferroptosis-dependent manner in the kidney in vivo and in vitro.
As shown in Fig. 6A and B, the CCK-8 and live/dead staining results showed that Fer-1 (5 μmol) protected the viability of SV40-MES-13 cells in the presence of MΦCM. The IHC results indicated that Fer-1 (5 μmol) effectively inhibited ferroptosis in SV40-MES-13 cells induced by MΦCM (Figure 7C-E). The changes in ferroptosis-related markers were also reversed in GMCs, which verified that Fer-1 effectively inhibited ferroptosis in SV40-MES-13 cells induced by MΦCM (Fig. 7F, Supplementary Figure 4A). Similarly, the changes in the mRNA levels of GPX4, NCOA4 and PTGS2 were reversed by Fer-1 treatment (Fig. 7G). Interestingly, the Western blot and IHC results showed that the protein expression of NF-κB, NLRP3, ASC, Caspase 1 and IL-1β was significantly decreased, and the expression levels of IKBα and p-IKBα were increased by 5 μmol Fer-1 in SV40-MES-13 cells that were cocultured with MΦCM (Fig. 7H, I, Supplementary Figure 4B, C). The mRNA levels of NF-κB, NLRP3, ASC, Caspase 1 and IL-1β were also inhibited by Fer-1 (Fig. 7 J and K). These results showed that the ferroptosis inhibitor Fer-1 inhibited the NF-κB/NLRP3 pathway in SV40-MES-13 cells induced by proinflammatory cytokines produced by M1 macrophages.