Vitamin D ameliorates particulate matter induced mitochondrial damages and calcium dyshomeostasis in BEAS-2B human bronchial epithelial cells.

Background: Mitochondria is prone to oxidative damage by endogenous and exogenous sources of free radicals, including particulate matter (PM). Given the role of mitochondria in inflammatory disorders, such as asthma and chronic obstructive pulmonary disease, we hypothesized that supplementation of vitamin D may play a protective role in PM-induced mitochondrial oxidative damages of human bronchial epithelial BEAS-2B cells.

Methods: BEAS-2B cells were pretreated with 1,25(OH)₂D₃, an active form of vitamin D, for 1 h prior to 24-hour exposure to PM (SRM-1648a). Oxidative stress was measured by flow cytometry. Mitochondrial functions including mitochondrial membrane potential, ATP levels, and mitochondrial DNA copy number were analyzed. Additionally, mitochondrial ultrastructure was examined using transmission electron microscopy. Intracellular and mitochondrial calcium concentration changes were assessed using flow cytometry based on the expression of Fluo-4 AM and Rhod-2 AM, respectively. Pro-inflammatory cytokines, including IL-6 and MCP-1, were quantified using ELISA. The expression levels of antioxidants, including SOD1, SOD2, CAT, GSH, and NADPH, were determined.

Results: Our findings first showed that 24-hour exposure to PM led to the overproduction of reactive oxygen species (ROS) derived from mitochondria. PM-induced mitochondrial oxidation resulted in intracellular calcium accumulation, particularly within mitochondria, and alterations in mitochondrial morphology and functions. These changes included loss of mitochondrial membrane integrity, disarrayed cristae, mitochondrial membrane depolarization, reduced ATP production, and increased mitochondrial DNA copy number. Consequently, PM-induced mitochondrial damage triggered the release of certain inflammatory cytokines, such as IL-6 and MCP-1. Similar to the actions of mitochondrial ROS inhibitor MitoTEMPO, 1,25(OH)₂D₃ conferred protective effects on mtDNA alterations, mitochondrial damages, calcium dyshomeostasis, thereby decreasing the release of certain inflammatory cytokines. We found that greater cellular level of 1,25(OH)₂D₃ upregulated the expression of enzymatic (SOD1, SOD2, and CAT) and non-enzymatic (GSH and NADPH) antioxidants to modulate cellular redox homeostasis.

Conclusion: Our study provides new evidence that 1,25(OH)₂D₃ acts as an antioxidant, enhancing BEAS-2B antioxidant responses to regulate mitochondrial ROS homeostasis and mitochondrial function, thereby enhancing epithelial defense against air pollution exposure.