Docosahexaenoic acid attenuates oxidative stress and protects human gingival fibroblasts against cytotoxicity induced by hydrogen peroxide and butyric acid

• The various function indicators are examined after treatment with DHA.
• Fibroblasts can be enriched with DHA when its extracellular concentration increases.
• Enrichment of cells with DHA effectively reduces oxidative stress.
• Enrichment of cells with DHA promotes their survival from H2O2-induced cytotoxicity.

ObjectiveThe oxidative burst of the host cells associated with bacterial pathogen infection contributes to the destruction of periodontal tissue. The present study investigates the effect of docosahexaenoic acid (DHA) on human gingival fibroblast (HGF) viability and ROS generation.MethodsThe cell viability by MTT assay, ROS level using H2DCF-DA probe, and protein thiol content were measured in HGFs after 24 h preincubation with different concentrations of DHA followed by treatment with H2O2. The cell death rate was determined by Annexin V/propidium iodide staining, and mitochondrial membrane potential (ΔΨm) was examined by MitoTracker Red probe in H2O2- and butyric acid-treated HGFs. The fatty acid composition of plasma membranes after incubation with DHA was determined by gas chromatography mass spectrometry.ResultsDHA preincubation in a dose-dependent manner increased the viability of HGFs exposed to H2O2 and decreased ROS generation compared to the control cells. In HGFs preincubated with 30 μM DHA, the ΔΨm significantly increased in both H2O2- and butyric acid-treated cells. Moreover, incubation with DHA preserved the protein thiol level as effectively as N-acetylcysteine. Application of 50 μM DHA increased the quantity of viable cells, decreased the number of necrotic cells after H2O2 treatment, and protected HGFs from apoptosis induced by butyric acid. DHA in the plasma membranes of these HGFs represented about 6% of the total amount of fatty acids.ConclusionsThese results demonstrate that enrichment of HGFs with DHA reduces ROS generation and enhances the mitochondrial membrane potential protecting the fibroblasts against cytotoxic factors.