Apitoxin protects rat pups brain from propionic acid-induced oxidative stress: The expression pattern of Bcl-2 and Caspase-3 apoptotic genes Samah R. Khalil a , Yasmina M. Abd-Elhakim a , Manar E. Selim b,c, *, Laila Y. Al-Ayadhi d a Forensic Medicine and Toxicology Department, Faculty of Veterinary Medicine, Zagazig University, Egypt b Zoology Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia c Zoology Department, Faculty of Science, Ain Shams University, Cairo, Egypt d Physiology Department, Faculty of Medicine, King Saud University, Autism Research and Treatment Centre, AL-Amodi Autism Research Chair, Riyadh, Saudi Arabia 1. Introduction Food additives are commonly used as food preservatives by inhibiting the growth of microorganisms. The safety of such compounds is routinely tested prior to use, and therefore food additives are often considered safe (Parke and Lewis, 1992). Nevertheless, the effects on health are also highly controversial at times. Propionic acid (PPA) is characterized by effective preservative activity, mould inhibition and bactericidal properties, causing PPA to be commonly used and considered an efficient additive to animal feed (stored grains and hay), poultry litter, poultry, and livestock drinking water as well as wheat and dairy products consumed by humans (Zarate et al., 2004). PPA is often employed as a growth promoter for animals raised for food as a way to improve output levels, especially in poultry and pigs (Marcos et al., 2004). The PPA esters are sometimes used as solvents or artificial flavourings as well (Haque et al., 2009). PPA does occur naturally in a few food products, however, primarily in dairy products such as milk, yoghourt and cheese (Fernandez-Garcia and McGregor, 1994). Excessive propionate has potential adverse effects such as propionic acidemia, a neurodevelopmental metabolic disorder (Xu et al., 2012) characterized by elevation of PPA levels in the blood, cerebro-spinal fluid, and most notably in neuronal cells (Feliz et al., NeuroToxicology 49 (2015) 121–131 A R T I C L E I N F O Article history: Received 9 March 2015 Received in revised form 27 May 2015 Accepted 31 May 2015 Available online 3 June 2015 Keywords: Amygdala Bee venom Comet assay Neural cell death Oxidative stress Propionic acid A B S T R A C T The primary aim of this study was to determine the potential modulatory role of the apitoxin (bee venom; BV) against propionic acid (PPA)-induced neurotoxicity. The biochemical responses to PPA exposure in rat pups were assayed, including changes in the antioxidant barrier systems and lipid peroxidation and protein oxidation biomarkers in the brain tissue. DNA damage was measured by single- cell gel electrophoresis and differences in Bcl-2 and Caspase-3 mRNA expression were assessed using real-time PCR. Changes in amygdala complex ultrastructure were visually assessed using electron microscopy. Sixty rat pups were assigned into six groups: a control group, a PPA-treated group, a BV-treated group, a protective co-treated group, a therapeutic co-treated group, and a protective/ therapeutic co-treated group. The results indicate that PPA induced a pronounced increase (64.6%) in malondialdehyde (MDA), and in DNA damage (73.3%) with three-fold increase in protein carbonyl concentration. A significant reduction was observed in the enzyme activities of superoxide dismutase (SOD) (48.7%) and catalase (CAT) (74.8%) and reduced glutathione (GSH) level (52.6%). BV significantly neutralized the PPA-induced oxidative stress effects, especially in the BV protective/therapeutic co- treated group. In this group, GSH levels were restored to 64.5%, and MDA, protein carbonyl levels and tail moment % were diminished by 69.5, 21.1 and 18.8% relative to the control, respectively. Furthermore, while PPA induced significant apoptotic neural cell death, BV markedly inhibited apoptosis by promoting Bcl-2 expression and blocking Caspase-3 expression. BV markedly restored the normal ultrastructural morphology of the amygdala complex neurons. These results conclusively demonstrate that BV administration provides both protective and therapeutic effects in response to the PPA-induced deleterious effects, including oxidative stress, DNA damage, and neuronal death in the brains of rat pups. ß 2015 Published by Elsevier Inc. * Corresponding author at: Faculty of Science, Ain Shams University, Egypt. Tel.: +20 966563588453. E-mail address: manar.selim@hotmail.com (M.E. Selim). Contents lists available at ScienceDirect NeuroToxicology http://dx.doi.org/10.1016/j.neuro.2015.05.011 0161-813X/ß 2015 Published by Elsevier Inc.