calpain and Chemical-and-Drug-Induced-Liver-Injury

Calpain is a Ca(2+)-regulated cytosolic cysteine protease that exists mainly in two isoforms and mediates crucial cellular functions, including rearrangement of cytoskeletal proteins, transport of the glucose transporter GLUT4, and protein cleavage to activate various receptors and pro-enzymes. Unintentional activation or functional loss of intracellular calpain has been implicated in several pathologies, including neurodegenerative diseases, traumatic brain and spinal cord injuries, cataracts and ischemia-associated injuries. Furthermore, polymorphism in the gene encoding calpain-10 has been associated with increased risk of type 2 diabetes. Recent studies have revealed a novel role for calpain in the progression of toxicant-induced liver damage. Evidence suggests that calpain leaking out of necrotic hepatocytes is highly activated in the extracellular milieu and hydrolyzes proteins in the plasma membrane of neighboring cells leading to progression of injury. Experimental intervention with calpain inhibitors substantially mitigates progression of liver injury initiated by toxicants, thereby preventing acute liver failure, and toxicant-induced animal death, pointing to a new potential therapeutic strategy against acute toxicities.

Topics: Animals; Calpain; Chemical and Drug Induced Liver Injury; Disease Progression; Humans

Microcystins produced by cyanobacteria are potent and specific hepatotoxins; however, the mechanisms of microcystin-induced hepatotoxicity have not been fully elucidated. The induction of free radical formation and mitochondrial alterations are two major events found in microcystin-treated cultured rat hepatocytes. The mitochondrial alterations, i.e. loss of mitochondrial membrane potential and mitochondria permeability transition are now recognized as key steps in apoptosis. The activation of calpain and Ca(2+)/calmodulin-dependent protein kinase II is believed to be critical in the microcystin-induced apoptotic process.

Topics: Animals; Apoptosis; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Calpain; Caspases; Chemical and Drug Induced Liver Injury; Cyanobacteria; Cysteine Proteinase Inhibitors; Enzyme Activation; Hepatocytes; Humans; Intracellular Membranes; Marine Toxins; Microcystins; Mitochondria, Liver; Models, Biological; Molecular Structure; Oxidative Stress; Peptides, Cyclic; Permeability; Rats; Reactive Oxygen Species

Atorvastatin (ATO), a 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor, is used widely for the treatment of hypercholesterolemia and hypertriglyceridemia. Application of this drug has now been made somehow limited because of ATO associated several acute and chronic side effects. The present study has been carried out to investigate the dose-dependent hepatic tissue toxicity in ATO induced oxidative impairment and cell death in mice. Administration of ATO enhanced ALT, ALP level, increased reactive oxygen species (ROS) production and altered the pro oxidant-antioxidant status of liver by reducing intracellular GSH level, anti-oxidant enzymes activities and increasing intracellular lipid peroxidation. Our experimental evidence suggests that ATO markedly decreased mitochondrial membrane potential, disturbed the Bcl-2 family protein balance, enhanced cytochrome c release in the cytosol, increased the levels of Apaf1, caspase-9, -3, cleaved PARP protein and ultimately led to apoptotic cell death. Besides, ATO distinctly increased the phosphorylation of p38, JNK, and ERK MAPKs, enhanced Caspase12 and calpain level. Histological studies also support the dose-dependent toxic effect of ATO in these organs pathophysiology. These results reveal that ATO induces hepatic tissue toxicity via MAPKs, mitochondria and ER dependent signaling pathway, in which calcium ions and ROS act as the pivotal mediators of the apoptotic signaling.

Topics: Animals; Apoptosis; Atorvastatin; Biomarkers; Calcium Signaling; Calpain; Caspase 12; Chemical and Drug Induced Liver Injury; Dose-Response Relationship, Drug; Endoplasmic Reticulum Stress; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Liver; Male; MAP Kinase Signaling System; Mice; Mitochondria, Liver; Oxidative Stress; Phosphorylation; Protein Processing, Post-Translational; Random Allocation; Reactive Oxygen Species; Survival Analysis

Systemic exposure to bacterial lipopolysaccharide (LPS, endotoxin) induces hypotension, disseminated intravascular coagulation and neutrophil infiltration in various organs including the lung, kidney and liver. A rat endotoxemic neutrophilic hepatitis model (repeat dose LPS, 10 mg/kg, i.v. 24 hours apart) was developed exhibiting hepatic neutrophil infiltration and mid-zonal hepatic necrosis. The goal of the study was to investigate the role of the intracellular enzyme calpain in the development of neutrophilic hepatitis with midzonal necrosis in this model. A second goal was to compare the observed protective effects of calpain inhibition with a relatively selective inducible nitric oxide synthase (iNOS) inhibitor aminoguanidine (AG) and an inhibitor of coagulation, heparin. When compared to rats administered LPS alone, administration of calpain 1 inhibitor prior to LPS significantly reduced hepatic iNOS expression, hepatic neutrophil infiltration and attenuated midzonal hepatic necrosis. Administration of AG or heparin prior to LPS also decreased liver iNOS expression, hepatic neutrophil infiltration and liver pathology comparable to calpain inhibition. Blood neutrophil activation, as measured by the neutrophil adhesion molecule CD11b integrin, was upregulated in all the LPS treated groups regardless of inhibitor administration. We conclude that amelioration of liver pathology via calpain inhibition is likely dependent on the down-regulation of iNOS expression in the rat model of LPS-mediated hepatitis.

Topics: Animals; Anticoagulants; Blood Coagulation; Blotting, Western; Calpain; Carrier Proteins; Chemical and Drug Induced Liver Injury; Cysteine Proteinase Inhibitors; Cytochrome P-450 CYP2E1; Disease Models, Animal; Endotoxemia; Endotoxins; Enzyme Inhibitors; Flow Cytometry; Glycoproteins; Guanidines; Heparin; Liver; Male; Microfilament Proteins; Necrosis; Neutrophil Activation; Neutrophils; Nitric Oxide Synthase Type II; Rats; Rats, Sprague-Dawley

Liver injury is known to progress even after the hepatotoxicant is long gone and the mechanisms of progressive injury are not understood. We tested the hypothesis that hydrolytic enzymes such as calpain, released from dying hepatocytes, destroy the surrounding cells causing progression of injury. Calpain inhibitor, N-CBZ-VAL-PHE-methyl ester (CBZ), administered 1 h after a toxic but nonlethal dose of CCl(4) (2 ml/kg, ip) to male Sprague Dawley rats substantially mitigated the progression of liver injury (6 to 48 h) and also led to 75% protection against CCl(4)-induced lethality following a lethal dose (LD75) of CCl(4) (3 ml/kg). Calpain leakage in plasma and in the perinecrotic areas increased until 48 h and decreased from 72 h onward paralleling progression and regression of liver injury, respectively, after CCl(4) treatment. Mitigation of progressive injury was accompanied by substantially low calpain in perinecrotic areas and in plasma after CBZ treatment. Normal hepatocytes incubated with the plasma collected from CCl(4)-treated rats (collected at 12 h when most of the CCl(4) is eliminated) resulted in extensive cell death prevented by CBZ. Cell-impermeable calpain inhibitor E64 also protected against progression of CCl(4)-induced liver injury, thereby confirming the role of released calpain in progression of liver injury. Following CCl(4) treatment, calpain-specific breakdown of alpha-fodrin increased, while it was negligible in rats receiving CBZ after CCl(4). Hepatocyte cell death in incubations containing calpain was completely prevented by CBZ. Eighty percent of Swiss Webster mice receiving a lethal dose (LD80) of acetaminophen (600 mg/kg, ip) survived if CBZ was administered 1 h after acetaminophen, suggesting that calpain-mediated progression of liver injury is neither species nor chemical specific. These findings suggest the role of calpain in progression of liver injury.

Topics: Acetaminophen; Animals; Blotting, Western; Calpain; Carbon Tetrachloride; Carrier Proteins; Chemical and Drug Induced Liver Injury; Cysteine Proteinase Inhibitors; Cytochrome P-450 CYP2E1; Cytochrome P-450 CYP2E1 Inhibitors; Dipeptides; Disease Progression; Hepatocytes; Immunohistochemistry; Liver Diseases; Male; Mice; Microfilament Proteins; Necrosis; Random Allocation; Rats; Rats, Sprague-Dawley