Hydroxyapatite nanoparticles promote mitochondrial-based pyroptosis via activating calcium homeostasis and redox imbalance in vascular smooth muscle cells

Abstract

Hydroxyapatite nanoparticles (HAP) have been widely used in various fields because of their natural biological origin and functional properties. The emerging evidence on their toxicities has attracted research interest. HAP-induced vascular smooth muscle cell (VSMC) damage is a key step in vascular calcification (VC), particularly in patients with chronic kidney disease. However, the injury effects and mechanism of action of HAP on VSMCs have not been extensively investigated. This study comprehensively characterized commercially available HAP and investigated its adverse biological effects in cultured A7R5 cells. In vitro experiments revealed that internalized HAP was localized in lysosomes, followed by the release of Ca2+ owing to the low pH microenvironment. Upon Ca2+ homeostasis, Ca2+ enters the mitochondria, leading to the simultaneous generation of reactive oxygen species (ROS). ROS subsequently attack mitochondrial transmembrane potentials, promote mitochondrial ROS production, and oxidize mitochondrial DNA (Ox-mtDNA). Mitochondrial permeability-transition pores open, followed by the release of more Ox-mtDNA from the mitochondria into the cytosol due to the redox imbalance. This activates NLRP3/caspase-1/gasdermin D-dependent pyroptosis and finally excretes inflammatory factors to induce VC; an antioxidant could rescue this process. It has been suggested that HAP could induce an imbalance in intracellular Ca2+ homeostasis in A7R5 cells, followed by the promotion of mitochondrial dysfunction and cell pyroptosis, finally enhancing VC. To detect the in vivo toxicity of HAP, mice were treated with Cy7-labelled HAP NPs for 24 h. In vivo results also demonstrated that HAP accumulated in the kidneys, accompined with increased Ca concentration, upregulated oxidative stress-related factor and kidney damage. Overall, our research elucidates the mechanism of calcium homeostasis and redox imbalance, providing insights into the prevention of HAP-induced cell death.

Curcumin/L‐OHP co‐loaded HAP for cGAS‐STING pathway activation to enhance the natural immune response in colorectal cancer

“…These results confirmed the successful construction of the Cur/L-OHP@HAP nanocomposite. A previous report showed that HAP exhibited almost complete dissolution of the deposited layer when immersed in an acidic environment, 28 indicating that HAP can undergo biodegradation at a low pH (such as in the TME) and thereby release calcium and loaded drugs. Therefore, we studied the pH-sensitive, drug-release curves of curcumin, L-OHP, and Ca 2+ in vitro.…”

“…Xia et al discovered that intracellular HAP can undergo biodegradation in an acidic environment and thereby release calcium 28 . To explore this, we used a Fluo‐4 AM probe to detect the production of free Ca 2+ in the cytoplasm of CT26 cells.…”

“…Xia et al discovered that intracellular HAP can undergo biodegradation in an acidic environment and thereby release calcium. 28 To explore this, we used a Fluo‐4 AM probe to detect the production of free Ca 2+ in the cytoplasm of CT26 cells. The results revealed almost no green fluorescence in the control group, whereas weak green fluorescence was observed in the HAP‐treated group, which may have reflected calcium‐ion release from HAP into CT26 cells (Figures 3c and S6 ).…”

Nanomaterial-induced pyroptosis: a cell type-specific perspective

“…For instance, Hussain et al reported that multi-walled carbon nanotubes trigger NLRP3 inflammasome activation and consequent cell pyroptosis in primary human bronchial epithelial cells in a concentration- and time-dependent manner ( Hussain et al, 2014 ). Similarly, Xia et al found that hydroxyapatite nanoparticles disrupt Ca 2+ homeostasis in vascular smooth muscle cells; subsequently, they cause mitochondrial dysfunction and cell pyroptosis ( Xia et al, 2022 ). Consistent with previous studies on these NM-induced cellular changes, Du et al’s study revealed how silica nanoparticles initiate mitochondrial ROS production, GSDMD cleavage, and pyroptosis in microglial cells ( Du et al, 2019 ).…”

Phenotypic plasticity of vascular smooth muscle cells in vascular calcification: Role of mitochondria

“…It was reported that histamine-stimulated IP3R mediates Ca 2+ release from the endoplasmic reticulum to mitochondria, and the increased mitochondrial calcium concentration can enhance oxidative phosphorylation by activating four mitochondrial dehydrogenases, improving the metabolic efficiency of VSMCs, and modulate the phenotypic transition of VSMCs ( 116 ), and PKA/Mfn2 can regulate the connection between mitochondria and ER, the overexpression of Mfn2 can increase the functional coupling between the two organelles, which is conducive to the rapid flow of mitochondrial Ca 2+ ( 116 ). Xia et al found that nanosized hydroxyapatite (nano-HAp) can increase mitochondrial Ca 2+ uptake in VSMCs, which in turn leads to excessive ROS production and oxidation of mtDNA (ox-mtDNA), and the release of more ox-mtDNA from mitochondria to the cytoplasm activates NLRP3/caspase-1/gasdermin D and induce VC by secreting inflammatory factors, antioxidants can effectively reverse this process and alleviate the progress of VC ( 33 ). In addition, the anticoagulants sodium citrate and diethyl citrate can also effectively reduce cell necrosis and lysosomal damage from [Ca2+]i overload induced by nano-HAp, avoiding further VC from cell death ( 29 ).…”