A Pre-clinical investigation on hepatotoxicity and osteoporosis induced by amphetamine derivative (methamphetamine and 3, 4-methylenedioximethamphetamine) with associated metabolic pathways using rat

Abstract

Abuse of amphetamine derivative as methamphetamine (Meth, Yaba) or 3, 4-methylenedioximethamphetamine (MDMA, Ecstasy) is on a raising trend globally and Bangladesh is not an exception of this. The long term effects of abusing these drugs include risk of developing permanent brain damage with various psychological, cognitive and behavioral effects that may manifest as depression, anxiety, memory loss and various neuropsychotic disorders. Many studies, survey and case reports have described the clinical manifestations associated with the abuse of these drugs included mental disorder, cardiac disorder, jaundice, tooth decay, etc. But yet surprisingly little is known about the toxic effect of these drugs on vital organs like liver and bone. The purpose of this study was to look insight and reveal the effects of Yaba and Ecstasy on liver and bone using rat model. The present study was conducted to determine the toxic effects of amphetamine derivatives using two representative tablets, Yaba and Ecstasy, where Yaba was subjected to measure liver and bone toxicity and Ecstasy to monitor the cytochrome P450 mediated metabolic pathways in vivo. Along with methamphetamine Yaba also contain a significant amount of caffeine. So the toxic action of Yaba might be due to the contribution of its both components. To clarify this, a separate study was also carried out with caffeine to determine its effect on liver and bone in vivo.The in vivo study performed with Yaba tablet used Long Evans rats for eight weeks to investigate liver and bone toxicities. ALT, AST, ALP level of plasma; AOPP, MDA, NO, Catalase, Glutathione and SOD level of plasma and tissue were measured. Histopathological study of rat liver was done with H&E and Picro Sirius Red staining. Rat tibias were tested by digital X-ray. Study with caffeine was performed following the same schedule as Yaba. The third experiment carried out with Ecstasy (3, 4-methylenedioximethamphetamine, MDMA) used Sprague Dawley rats, pretreated with phenobarbital (PB) or β-naphthoflavone (BNF), to monitor the metabolic pathways of MDMA in liver by measuring urine drug metabolites concentration and liver CYP isozymes content. Liver inflammation was observed in Yaba treated rat liver with significantly elevated plasma level of ALT, AST, and ALP. Plasma and liver tissue level of MDA and NO were significantly increased. AOPP, Catalase, GSH, SOD activity in plasma and liver found unchanged as compared to control. In caffeine treated rat, liver transaminases, ALT, AST, ALP; oxidation end product, AOPP, MDA and oxidative stress indicator, NO increased significantly, Catalase & GSH remained unchanged but SOD decreased significantly as compared to control. Histology of liver tissue showed invasion of inflammatory cells and progressive deposition of collagen fibre in both Yaba and caffeine treated group. Digital X-ray of rat tibia treated with Yaba showed radiolucency not significant as compared to control. In urine analysis of MDMA treated rats, end product of N-demethylation, 3, 4-methylenedioxiamphetamine (MDA) concentration was significantly decreased in PB and BNF treated rats and end product of O-methylation, 4-hydroxy 3-methoxy methamphetamine (HMMA) concentration was significantly increased in PB and BNF treated rats as compared to control. Hepatic enzyme assays showed increased activities of CYP1A1(600-fold), CYP1A2(4-fold), CYP2B(4-fold) and CYP3A(2-fold) related enzymes after 24 hours of MDMA administration in the inducer pretreated rats as compared to control. In PB treated rat urine MDMA concentration decreased significantly. The total percent dose recoveries of MDMA and three metabolites in urine samples was less than 35% of the administered dose. The overall results suggested that methamphetamine tablet Yaba produced inflammation and fibrogenesis in liver. Caffeine, the second ingredient of Yaba showed similar liver toxicity in histochemical study. However, the extent of biochemical changes, notably higher in caffeine treated group than those of Yaba, suggested that methamphetamine in combination with caffeine might follow dissimilar pathways for liver toxicity to that followed by caffeine separately. The effect of Yaba on rat bone turnover was inconclusive. Metabolic pathway study results of 3,4-methylenedioximethamphetamine (MDMA) suggested that PB and BNF induced CYP isozymes might have inhibitory effects on N-demethylation of MDMA to MDA in rats. HHMA, the precursor of HMMA, increased through the process of O-demethylenation of MDMA by induced CYP isozymes. Decreased urinary concentration of MDMA in PB pretreated rats indicated its increased metabolism. The truly low percent recoveries of MDA, HMMA and HMPA in inducer pretreated rats suggested that other major pathways of MDMA metabolism might exist which were activated through inducer pretreatment.