Beneficial effects of benfotiamine, a NADPH oxidase inhibitor, in isoproterenol-induced myocardial infarction in rats

Abstract

Background: Acute myocardial infarction (AMI) remains the most common cause of morbidity and mortality worldwide. The present study was directed to investigate the beneficial effects of benfotiamine pre- and post-treatments in isoproterenol (ISO)-induced MI in rats.

Methods: Myocardial heart damage was induced by subcutaneous injection of ISO (150 mg/kg) once daily for two consecutive days. Benfotiamine (100 mg/kg/day) was given orally for two weeks before or after ISO treatment.

Results: ISO administration revealed significant changes in electrocardiographic recordings, elevation of levels of cardiac enzymes; creatinine kinase (CK-MB) and troponin-I (cTn-I), and perturbation of markers of oxidative stress; nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, malondialdehyde (MDA), reduced glutathione (GSH), superoxide dismutase (SOD) and glutathione peroxidase (GPx) and markers of inflammation; protein kinase C (PKC), nuclear factor-kappa B (NF-κB) and metalloproteinase-9 (MMP-9). The apoptotic markers (caspase-8 and p53) were also significantly elevated in ISO groups in addition to histological alterations. Groups treated with benfotiamine pre- and post-ISO administration showed significantly decreased cardiac enzymes levels and improved oxidative stress, inflammatory and apoptotic markers compared to the ISO groups.

Conclusion: The current study highlights the potential role of benfotiamine as a promising agent for prophylactic and therapeutic interventions in myocardial damage in several cardiovascular disorders via NADPH oxidase inhibition.

Fig 1

Effect of benfotiamine pre- and post-treatments on serum (A) cTn-I and (B) CK-MB levels in ISO-induced MI in rats. Each bar represents mean ± SEM (n = 8). Statistical analysis was done using One-way ANOVA followed by Tukey’s post-hoc test. *significantly different from the normal group at p< 0.05. @significantly different from the ISO-A group at p< 0.05. #significantly different from the ISO-B group at p< 0.05. ISO-A: ISO pre-treatment control; ISO-B: ISO post-treatment control; ISO-A + Benf.: benfotiamine prophylactic; ISO-A+ Benf.: benfotiamine treatment.

Fig 2

Effect of benfotiamine pre- and post-treatments on myocardial (A) NADPH oxidase activity, (B) MDA, (C) GSH contents, in addition to (D) GPx as well as (E) SOD activity in ISO-induced MI in rats. Each bar represents mean ±SEM (n = 8). Statistical analysis was done using One-way ANOVA followed by Tukey’s post-hoc test. *significantly different from the normal group at p< 0.05. @significantly different from the ISO-A group at p< 0.05. #significantly different from the ISO-B group at p< 0.05. ISO-A: ISO pre-treatment control; ISO-B: ISO post-treatment control; ISO-A+ Benf.: benfotiamine prophylactic; ISO-A+ Benf.: benfotiamine treatment.

Fig 3

Effect of oral benfotiamine pre- and post-treatments on myocardial (A) PKC, (B) MMP-9 and (C) NF-κB contents in ISO-induced MI in rats. Each bar represents mean ± SEM (n = 8). Statistical analysis was done using One-way ANOVA followed by Tukey’s post-hoc test. *significantly different from the normal group at p< 0.05. @significantly different from the ISO-A group at p< 0.05. #significantly different from the ISO-B group at p< 0.05. ISO-A: ISO pre-treatment control; ISO-B: ISO post-treatment control; ISO-A+ Benf.: benfotiamine prophylactic; ISO-A+ Benf.: benfotiamine treatment.

Fig 4

Effect of benfotiamine pre- and post-treatments on myocardial (A) p53 and (B) caspase-8- contents in ISO-induced MI in rats. Each bar represents mean ± SEM (n = 8). Statistical analysis was done using One-way ANOVA followed by Tukey’s post-hoc test. *significantly different from the normal group at p< 0.05. @significantly different from the ISO-A group at p< 0.05. #significantly different from the ISO-B group at p< 0.05. ISO-A: ISO pre-treatment control; ISO-B: ISO post-treatment control; ISO-A+ Benf.: benfotiamine prophylactic; ISO-A+ Benf.: benfotiamine treatment.

Fig 5. Effect of benfotiamine pre- and post-treatments on histological damage in ISO-induced MI in rats.

(A–F) Cardiac specimens were stained with H&E (magnification x200). Normal group (A) showed normal histological structure of cardiac muscle. ISO-A treatment group (B) showed severe cardiac insult with atrophy, muscle shrinkage (s) edema (e) of cardiac muscle, marked inflammatory cells infiltration (n), extensive edema in-between muscle fibers (e), and marked number of apoptotic or degenerated myocytes filament (black arrows). Benfotiamine pre-treatment group (C) showed mostly normal cardiac muscle architecture. ISO-B treatment group (D) revealed shrinkage (s) of cardiac muscle with preserved nuclei all over sections examined, edema in between muscle fibers (e). Benfotiamine post-treatment (E) demonstrated almost normal cardiac muscle with only mild edema in-between muscle fibers. Myocardial score of damage expressed as median (interquartile changes) (F). Each value represents the median value [interquartile range] (n = 6). Statistical analysis was done using non-parametric One-Way ANOVA (Kruskal-Wallis test) followed by Dunn's multiple comparison test. * p < 0.05 vs. normal, @ p < 0.05 vs. ISO-A, # p < 0.05 vs. ISO-B. ISO-A: ISO pre-treatment control; ISO-B: ISO post-treatment control; ISO-A+ Benf.: benfotiamine prophylactic; ISO-A+ Benf.: benfotiamine treatment.