Toxicity of 4-methylimidazole on isolated brain mitochondria: using both in vivo and in vitro methods

Abstract

4-methylimidazole (4MI) is a compound widely used in various industrial and consumer applications. The most important sources of exposure include chemical caramel coloring, ammoniated molasses, dyes and pigments, rube, cleaning and agricultural chemicals. Toxicity attributed to 4MI in foods has recently become a focus of research. Recent studies showed that 4MI induced adverse changes in various target tissues. Brain is known to be a target organ for 4MI-induced toxicity but its cytotoxic mechanisms have not yet been elucidated. In this study, experiments were divided into two parts: (1) using in vivo methodology, doses of 4MI at 100, 200, or 300 mg/kg were administered orally to mice daily for 14 to obtain brain mitochondria; and (2) utilizing in vitro methodology, brain mitochondria were incubated with 4MI at 400, 800, or 1600 μM concentrations. Subsequently, the neurotoxicity of 4MI was assessed using mitochondrial dysfunction tests, including reactive oxygen species (ROS) formation, mitochondrial membrane potential (MMP) collapse, mitochondrial swelling, and cytochrome c release. Our results from both in vivo and in vitro experiments on isolated brain mitochondria showed a significant decrease in complex II activity and also marked elevation in the ROS formation, MMP collapse, mitochondrial swelling, and enhanced release of cytochrome c. Data indicated that 4MI induced neurotoxicity through the impairment of electron transfer chain especially at complex II and elevated ROS formation leading to subsequent oxidative stress events including mitochondrial membrane depolarization, mitochondrial swelling, and release of cytochrome c, which is the starting point of mitochondrial-mediated apoptosis signaling and neurodegeneration.

Keywords:

• 4-methylimidazole
• isolated mitochondria
• in vivo
• in vitro
• brain

Acknowledgments

The results presented in this article were partly extracted from the thesis of Dr Freshteh Mehri (PhD graduate of Department of Pharmacology and Toxicology, Faculty of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran) who performed her thesis under the supervision of Professor Mehrdad Faizi and Professor Jalal Pourahmad. The investigation was carried out in Professor J. Pourahmad's laboratory at Faculty of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

The research was funded by the Shahid Beheshti University of Medical Sciences, Deputy of Research [grant number 1393-1-94-12883].