Size-Dependent Metabolic Reprogramming in A549 Cells Induced by Mesoporous Silica Nanoparticles: Insights from Subcellular Targeting

Background/Objectives: Mesoporous silica nanoparticles (MSNs) are widely investigated as nanocarriers for drug delivery, gene transfer, and bioimaging. However, the mechanisms underlying their size-dependent cytotoxicity at the metabolic level remain incompletely understood. This study aimed to determine whether different-sized MSNs induce distinct patterns of subcellular injury and metabolic reprogramming in lung epithelial cells. Methods: A549 cells were exposed to 80 nm and 600 nm MSNs at 50 and 200 μg/mL for 24 h. Ultrastructural changes were examined by transmission electron microscopy (TEM). Intracellular reactive oxygen species (ROS) and Ca2+ were measured by 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) and Fluo-4 AM fluorescence, respectively. Inflammatory gene expression (IL1B, IL6, TNFA, HIF1A) was quantified by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Untargeted metabolomics were performed using combined gas chromatography–mass spectrometry (GC-MS) and liquid chromatography–mass spectrometry (LC-MS) platforms, followed by principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and MetaboAnalyst-based pathway enrichment. Results: TEM revealed distinct size-dependent subcellular distributions: 80 nm MSNs were predominantly associated with mitochondrial abnormalities, including cristae disruption, swelling, and mitophagy-like features, whereas 600 nm MSNs accumulated in endocytic vesicles with membrane disruption. Metabolomic profiling showed that 80 nm MSNs were associated with TCA cycle blockade—characterized by the accumulation of early intermediates (citrate, oxaloacetate) and the depletion of distal intermediates (fumarate, malate)—with compensatory glycolytic activation (increased glyceraldehyde-3-phosphate and pyruvate) and reduced deoxynucleotide pools (dCDP, dUMP). By contrast, 600 nm MSNs triggered broad nucleotide triphosphate accumulation (ATP, CTP, dGTP, dCTP), amino acid depletion, and robust inflammatory activation, including a ~136-fold increase in IL1B expression and HIF1A transcriptional upregulation. PCA and PLS-DA confirmed distinct size-dependent metabolic phenotypes. Conclusions: MSN size strongly influences subcellular targeting—80 nm particles were predominantly associated with mitochondrial injury while 600 nm particles disrupted endocytic vesicles—driving qualitatively distinct patterns of metabolic reprogramming and inflammatory signaling. These findings establish a correlative mechanistic framework linking particle size to organelle-specific injury and provide candidate metabolic markers for nanotoxicological evaluation.