Neurotoxic and developmental effects of scented incense stick smoke: Network toxicology and zebrafish model study

Burning incense sticks is a traditional practice in many cultures, especially in Southeast Asia. While it is often regarded as sacred and beneficial, modern incense sticks contain various chemicals that can pose health risks. A GCMS analysis of the ICS revealed potential compounds. Network toxicology revealed that ICS contains compounds violating Lipinski's rule of five, leading to potential neurotoxic effects. Key pathways affected include neuroactive ligand-receptor interaction and calcium signaling, associated with neurodegenerative diseases like Parkinson's and Alzheimer's. Significant genes involved are STAT3, BCL2, and MTOR, emphasizing the chemical hazards of ICS exposure. We investigated the toxicity of ICS using zebrafish (Danio rerio) embryos as a mode. ICS exposure resulted in a dose-dependent increase in toxicity. High concentrations (7 and 14 µg/ml) led to immediate mortality, while lower concentrations (0.1, 0.3, 0.5, and 1 µg/ml) caused developmental defects such as yolk sac edema, skeletal malformations, and pericardial edema. Mortality rates increased with higher concentrations, confirming dose-dependent ICS exposure caused hypoactive locomotion, with reduced distance traveled and velocity toxicity. Higher concentrations of ICS led to increased ROS levels and cellular damage, as evidenced by enhanced staining levels. A dose-dependent increase in lipid peroxidation (DPPP assay) and lipid accumulation (Nile red assay) was observed. Higher ICS concentrations led to significant oxidative damage to lipids and increased lipid deposition. Enzymatic assays showed that ICS exposure significantly decreased the activities of antioxidant enzymes SOD and CAT, indicating impaired antioxidant defense, while increasing LDH activity, signaling tissue damage and cytotoxicity. Gene expression analysis revealed downregulation of SOD1 and CAT genes, upregulation of inflammatory genes TNF-α and IL-1β, and increased expression of the apoptotic gene p53 with decreased expression of Bcl-2 and BDNF. These findings highlight ICS's potential to cause oxidative stress, inflammation, apoptosis, and neurodevelopmental impairments.