2.2. Drugs-induced animal models
There are many drugs and their metabolites that are widely used to develop hepatic injury in animal models (Table 2). These models are used for analyzing the protection, effectiveness, and development of hepatoprotective medicines. These are as follows:
2.2.1. Acetaminophen: Also known as paracetamol, it is widely used as an antipyretic and analgesic and over-the-counter drug. Over usage of acetaminophen causes liver damage and can be used in experimental models to induce hepatotoxicity. Acetaminophen-induced liver cirrhosis occurs at sole dose of 2g/kg given by oral route and sole dose of 300mg/kg by intraperitoneal injection in rats and mice (45, 46). By microsomal enzymatic action, a metabolite N-acetyl-p-benzoquinoneimine is produced which is extremely reactive in nature. The binding of this metabolite with the proteins results in lipid peroxidation and reduction in glutathione levels. Further, the mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) activation disorient mitochondrial function and elevate oxidative stress resulting in hepatic fibrosis and necrosis. Chronic administration of acetaminophen induces autophagy in hepatocytes in the liver of mice, which might accelerate hepatic cell death (47, 48).
2.2.2. Isoniazid: It is an anti-tuberculosis drug and is also called isonicotinic acid hydrazide (INH). It induces hepatic fibrosis at 200 mg/kg and 400 mg/kg with oral gavages for the duration of one week in mice and rats respectively (49, 50). Monoacetyl hydrazine is generated after metabolism by cytochrome P450, which causes depletion of ATP in hepatocytes, and an imbalance of free radicals and antioxidant enzymes. This leads to the development of liver cirrhosis in the animal model (51). Rifampicin in combination with isoniazid interferes with bilirubin uptake isoniazid clearance from hepatic sinusoidal cells, resulting in hyperbilirubinemia, and jaundice without affecting hepatocytes. When used together, the toxic metabolites i.e. hydrazine and cytochrome 2E1 production causes oxidative stress and hepatocellular injury. Therefore, INH (50mg/kg) and rifampicin (100mg/kg) daily dose for a week can induce hepatotoxicity in rodents (52). Rabbits receiving this combined dose showed elevation in serum alanine aminotransferase (ALT) and serum alanine aspartate (AST) measures. The cytochrome P450 activation, co-transportation of sodium taurocholate, and bile formation indicate that rifampicin enhanced hepatotoxicity caused by isoniazid (53).
2.2.3. Tetracycline: Tetracycline induces liver injury at a single dose of 200 mg/kg by intraperitoneal route in mice and rats (54, 55). Tetracycline act mainly via enhanced fatty acid uptake, fatty acid β-oxidation, triglycerides and cholesterol synthesis, inhibition of microsomal triglyceride transfer protein activity and hepatic lipoprotein inhibition (56). It causes micro-vesicular steatosis by increasing fatty acid infusion, followed by oxidative stress due to lipid overload and attack on functioning proteins. The excess lipid in the hepatic cytoplasm activates the long-chain fatty-acid-Coenzyme A ligase-1 and stimulates the formation of triacylglycerol or fatty acid transportation into mitochondria. This speeds up β-oxidation and causes mitochondrial respiration chain stress along with high levels of ROS. This leads to microvesicular steatosis, and hepatic cirrhosis in rodents through the breakdown of fatty acids (57).
2.2.4. Doxorubicin: Doxorubicin, an anthracycline derivative used as an antineoplastic agent in the treatment of leukemia, induces hepatic fibrosis at 15 mg/kg injection via intraperitoneal route once a week for 4 weeks in rats (58, 59). It has been reported that doxorubicin induces hepatotoxicity in off-target cells in individuals who have been on a therapeutic regimen for cancer. This drug causes oxidative stress in liver cells leading to reactive oxygen generation, mitochondrial dysfunction, and hepatic fibrosis and necrosis. The oxidative stress is mediated via nuclear factor erythroid2-related factor-2/ heme oxygenase-1(Nrf-2/HO-1) pathway. There is inflammation due to the ROS generation. There are elevated levels of malondialdehyde via Sirtuin 1/forkhead box protein 1/Nuclear factor-ҡB (SIRT1/FOXO1/NF-κB) signaling pathway. Mitochondrial dysfunction via deregulation of electron transport chain, diminished ATP production and slashed hepatic stellate cells and sinusoidal epithelial cells result in hepatic cellular damage (60). Doxorubicin is metabolized by the NADPH-cytochrome P450 reductase by the stimulation of a single-electron initiation reaction. This leads to the generation of free radicals like hydrogen peroxide and superoxide which reduces the antioxidant level and elevates the reactive oxygen species resulting in hepatic vacuolation, bile duct hyperplasia, and hepatic fibrosis (61).
2.2.5. Cisplatin: Cisplatin is an alkylating anti-neoplastic agent used conventionally in the treatment of breast, uterine, ovarian, and neuroblastoma tumors. It acts through membrane peroxidation, interference in protein synthesis, and DNA damage (62). Simultaneously, it was reported to cause hepatotoxicity by neutrophil infiltration, hyperplasia, fibrosis, and hepatocellular necrosis (63). The antioxidant parameters such as superoxide dismutase (SOD) and catalase (CAT) and glutathione peroxidase (GSH) have been used for the evaluation of liver functioning (64, 65).
2.2.6. Tamoxifen: It is an antiestrogen and is used for the treatment of breast cancer. It induces hepatic injury at 45 mg/kg dose daily administered by intraperitoneal route for 6 days in rats. Tamoxifen, after metabolism, forms reactive hydroxyl radicals and lipid peroxyl radicals that contribute to oxidative stress. Mitochondrial and cell membrane dysfunction results in hepatic fibrosis (66, 67).
2.2.7. Carbamazepine: Carbamazepine induces hepatic cirrhosis at dose of 400 mg/kg administered orally for a period of 4 days and a dose of 800 mg/kg administered orally on the fifth day in mice (68, 69). It leads to the generation of reactive metabolites by cytochrome P450. An imbalance between antioxidant enzymes and free radicals causes lipid peroxidation which stimulates macrophages. Release of pro-inflammatory chemokines result an inflammatory response leading to liver cirrhosis (70).
2.2.8. Sodium Valproate: Sodium valproate induces liver cirrhosis at a dose of 500 mg/kg administered daily by oral gavage for 2 weeks in rats and mice (71, 72). It alters the expression pattern of genes related to lipid transport leading to change in synthesis of triacylglycerol, cholesterol and the metabolism of fatty acids. Abnormal lipid metabolism and interference in the β-oxidation result in mitochondrial malfunctioning and the release of free radicals. Free radicals lead to a lipid peroxidation chain reaction and a reduction in the antioxidant levels, thus inducing hepatic cirrhosis (73).
2.2.9. CNS stimulants: Cannabis, cocaine, heroin, methamphetamine and anabolic androgenic steroids are other compounds reported to cause hepatocellular carcinoma in individuals with adenoma or cirrhosis. These agents successfully induce metabolism-associated fatty liver disease. The Kava (Piper methysticum) has been reported to have the highest hepatotoxic potential and can be used in rodents to evaluate hepatocellular carcinoma (74, 75).