# Protective Action of 3,5-Diiodo-L-Thyronine on Cigarette Smoke-Induced Mitochondrial Dysfunction in Human Alveolar Epithelial Cells

## TL;DR

This study shows that 3,5-diiodo-L-thyronine (T2) protects human lung cells from cigarette smoke-induced mitochondrial damage and oxidative stress.

## Contribution

The novel contribution is the demonstration of T2's protective effects on mitochondrial function and oxidative stress in cigarette smoke-exposed alveolar epithelial cells.

## Key findings

- T2 pretreatment prevents cigarette smoke-induced reductions in ATP production and enhances cytochrome c oxidase activity.
- T2 mitigates oxidative stress by increasing SOD activity and reducing superoxide and H2O2 levels.
- Cigarette smoke increases mitochondrial mass, but T2 does not significantly prevent this increase.

## Abstract

Background: Cigarette smoke (CS) is a major risk factor for chronic lung conditions. Oxidative stress and mitochondrial dysfunction play a crucial role in CS-induced pulmonary injury. 3,5-Diiodothyronine (T2) affects energy metabolism, having mitochondria as a major target. However, the underlying mechanisms of T2 related to lung diseases are poorly understood. Aims: To investigate the protective action of T2 on CS-induced mitochondrial dysfunction in an in vitro model of human epithelial alveolar cells. Methods: ATP synthesis and cytochrome c oxidase (COX) activity, as a marker of mitochondrial function, was assessed in A549 cells pretreated with T2 and exposed to CS using a bioluminescence assay and an Oroboros 2k-Oxygraph system, respectively. An evaluation of the oxidative status was conducted by assessing superoxide radical production, superoxide dismutase (SOD) activity, and H2O2 levels. Moreover, we investigated the mitochondrial mass via Mito-Tracker Green (MTG) staining and flow cytometry analysis. Results: CS significantly reduced ATP production. T2 pretreatment was found to prevent CS-induced impairments in ATP synthesis, enhancing COX activity. Additionally, the 2 h T2 pretreatment of CS-exposed cells mitigated CS-induced oxidative stress, thereby enhancing SOD activity and reducing the superoxide anion and H2O2 levels. Finally, MTG labeling was correlated with CS-induced mitochondrial mass gain, which is associated with cell senescence. Unexpectedly, T2 was not able to significantly prevent this mass increment, probably due to its rapid mode of action. Conclusions: Our results provide new insights into the protective effects of T2 against CS-induced mitochondrial damage.