Calcium oxalate (CaOx) stones are a prevalent urological condition with an unclear etiology. We previously found that matrix metalloproteinases (MMPs) might play a pathogenic role in stone formation. Fibroblasts, the primary source of MMPs, reside in the renal interstitium; however, the urine lithogenic environment act directly upon renal epithelium. We hypothesize that exosomes mediate the interaction between epithelial cells and fibroblasts during stone formation.

NRK-52E cells (a rat renal epithelial tubular cell line) were treated with CaOx monohydrate (COM) crystals for 24 hours. Exosomes were isolated from treated (COM-Exo) and untreated (Ctrl-Exo) cells and subsequently cocultured with NRK-49F cells (a rat renal fibroblast line) for 24 hours. RNA-Seq was performed on NRK-49F cells treated with either Ctrl-Exo or COM-Exo. MMP expression and related signaling pathways were assessed using qPCR and Western blotting. Additionally, Ctrl-Exo or COM-Exo were subcapsularly injected into rat kidneys during modeling, with crystal deposits examined via von Kossa staining. A comparative proteomic analysis was conducted to identify which exosomal protein exerts biological effects.

RNA-Seq revealed that differentially expressed genes between NRK-49F treated with Ctrl-Exo and COM-Exo were primarily associated with extracellular matrix organization and the NF-κB signaling pathway. We verified that COM-Exo treatment activated the NF-κB pathway, leading to enhanced expression of MMPs, specifically MMP-3 and MMP-9, compared to Ctrl-Exo treatment. Additionally, rats preinjected with COM-Exo exhibited significantly increased kidney crystal deposition. Proteomic analysis identified a higher concentration of CD147 in COM-Exo compared to Ctrl-Exo. We further silenced and overexpressed CD147 in NRK-52E cells, isolating exosomes with varying CD147 levels for incubation with NRK-49F cells. We found that increased CD147 content in exosomes corresponded with higher activation levels of NF-κB pathway and elevated MMPs expression.

Our work revealed an exosome-mediated intercellular communication between renal epithelial cells and fibroblasts that promotes stone formation. NRK-52E cells exposed to COM crystals released more exosomal CD147 comparing with normal cells. These exosomes were taken up by NRK-49F cells, resulting in upregulation of MMP expression via the NF-κB pathway, ultimately exacerbating kidney crystal deposition.