Comparing the Effects of Propofol and Thiopental on Human Renal HEK-293 Cells With a Focus on Reactive Oxygen Species (ROS) Production, Cytotoxicity, and Apoptosis: Insights Into Dose-Dependent Toxicity

Abstract

Objectives: Propofol and thiopental are widely used as hypnotic, sedative, antiepileptic, and analgesic agents in general anesthesia and intensive care; however, their side effects remain unknown. They are used for long periods and at high doses for sedation in total intravenous anesthesia (TIVA) and intensive care units. Long-term and high-dose use of these drugs can lead to accumulation in plasma and tissues, resulting in high drug concentrations and increasing the risk of potential toxicity (e.g., nephrotoxicity). In our study, the cytotoxic and apoptotic effects of propofol and thiopental on kidney cells (HEK-293) and their effects on the formation of reactive oxygen species (ROS) when used in high doses were investigated and compared in vitro.

Materials and methods: The half-maximal inhibitory concentration (IC₅₀) of each drug in HEK-293 cells was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. The apoptotic effects were assessed at two different doses of each drug using the annexin V method. Morphological examinations were conducted using the acridine orange/ethidium bromide method, and intracellular ROS levels were determined by flow cytometry.

Results: The IC₅₀ values of propofol and thiopental for HEK-293 cells were 206.59 μg/ml and 109.68 μg/ml, respectively. Compared to the control group, thiopental at ≥25 μg/ml and propofol at ≥50 μg/ml exhibited cytotoxicity. Additionally, propofol exhibited significantly lower cytotoxic effects than thiopental did.

Conclusion: Our study showed that both propofol and thiopental exerted significant cytotoxic effects on HEK-293 cells at concentrations exceeding clinical levels, primarily by increasing intracellular ROS levels and inducing apoptosis. Future research in this area will deepen our understanding of these mechanisms and improve patient safety in clinical anesthesia practice.

Keywords: anesthesia; apoptosis; cytotoxicity; human kidney cells (hek-293); propofol-thiopental; reactive oxygen species (ros).

Figure 1. % changes in the viability of HEK-293 cells treated with different concentrations of propofol and thiopental for 24 hours

The data obtained are shown as mean ± standard deviation. ^*P < 0.001 vs control group for propofol. ^#P < 0.001 vs control group for thiopental. ^*#P < 0.001 vs thiopental group for propofol. IC_⁵⁰: half-maximal inhibitory concentration

Figure 2. Morphological (a) and AO/EB fluorescence (b) staining on HEK-293 cells at 20× magnification

Live cells in green, apoptotic cells in orange, and necrotic cells in red. AO/EB: acridine orange/ethidium bromide

Figure 3. Flow cytometric annexin V analysis of the apoptotic effects of drugs on HEK-293 cells

FITC: fluorescein isothiocyanate

Figure 4. Flow cytometric investigation of the effects of propofol and thiopental on intracellular free radical (ROS) formation of HEK-293 cells

ROS: reactive oxygen species

Figure 5. Graphical abstracts

IC₅₀: half-maximal inhibitory concentration; AO/EB: acridine orange/ethidium bromide; ROS: reactive oxygen species; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; ATCC: American Type Culture Collection