Structural Passivation Engineering of Montmorillonite Nanoclay Attenuates the ROS-HIF-1α-Ferroptosis Axis for...

Montmorillonite nanoclay holds promise for nanomedicine, yet its structure-dependent biosafety remains poorly defined. Raw montmorillonite (MMT_Raw) was shown to induce excessive reactive oxygen species (ROS) generation, leading to hypoxia-inducible factor-1α (HIF-1α) stabilization and activation of ferroptosis-associated transcriptional programs, thereby identifying oxidative stress as the central cytotoxic mechanism. To elucidate structure-biosafety relationships, structurally distinct montmorillonite variants were prepared via ultrasonication, acid etching, and ball milling. Integrated physicochemical characterization and cell viability data analysis reveal a quantitative structure-toxicity correlation, in which reduced crystallinity and surface hydroxyl density are associated with enhanced cellular compatibility. Among these, ball-milled montmorillonite (MMT_Ball) exhibits the highest biosafety, improving cell viability by 3.2-fold through attenuation of ROS accumulation and suppression of downstream HIF-1α-ferroptosis signaling. Collectively, this study establishes a structure-regulated oxidative stress-ferroptosis framework and highlights structural passivation as an effective strategy for biosafety optimization of clay-based nanomaterials.