Evaluation of dose-dependent hepatic injury induced by copper oxide nanoparticles in male Wistar rats
Abstract
This study was conducted to evaluate the hepatotoxic effects of copper oxide (CuO) nanoparticles following repeated oral exposure in male Wistar rats. Seventy-two healthy male rats were randomly divided into four groups (n = 18). Group I served as control, whereas Groups II, III, and IV received 1/10th, 1/20th, and 1/40th of the LD50 dose of CuO nanoparticles, respectively, for 45 days. Six animals from each group were sacrificed at 15, 30, and 45 days of exposure for liver-related evaluation. Body weight, serum biochemical markers of liver function, oxidative stress indices, inflammatory mediators, and liver histopathology were assessed. CuO nanoparticle exposure produced dose-and duration-dependent toxic effects. Rats of Groups II and III showed reduced body weight gain during the later stages of exposure. Serum biochemical analysis showed significantly increased AST and ALT levels, particularly in Groups II and III at 30 and 45 days. Oxidative stress analysis demonstrated increased malondialdehyde levels and decreased superoxide dismutase activity in treated animals, indicating enhanced lipid peroxidation and reduced antioxidant defense. The levels of TNF-alpha and IL-1 were also elevated in treated groups, suggesting activation of inflammatory pathways. Histopathological examination of the liver revealed congestion, hepatocyte degeneration, bile duct dilatation, periportal fibrosis, focal necrosis, and inflammatory cell infiltration, with more severe lesions in higher dose groups. The findings of the present study indicate that repeated oral exposure to CuO nanoparticles induces significant hepatic injury in male Wistar rats through oxidative stress and inflammatory mechanisms. The severity of liver damage increased with higher dose and longer duration of exposure.
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Introduction
Nanotechnology has become an important component of modern medicine, agriculture, food processing, electronics, and pharmaceutical industries [1,2] . Among different metallic nanoparticles, copper oxide nanoparticles are widely utilized because of their antimicrobial, catalytic, semiconducting, and antifungal properties [3,4] . Among various nanoparticles, copper oxide (CuO) nanoparticles are extensively used due to their antimicrobial, catalytic, and industrial applications. However, their increasing use has raised concerns regarding their potential toxic effects on living systems.
Copper is an essential trace element involved in various physiological functions, but excess copper can lead to toxicity through the generation of reactive oxygen species (ROS) [5] . Due to their small size and increased surface area, CuO nanoparticles can penetrate biological membranes, accumulate in tissues, and induce cellular damage [6] . The liver is considered one of the major target organs for nanoparticle accumulation because it plays a central role in metabolism, detoxification, and excretion [7] . The liver is one of the principal organs involved in metabolism, detoxification, and excretion; therefore, it is highly susceptible to xenobiotic-induced injury. Nanoparticles entering the circulation are commonly accumulated in hepatic tissue, where they may trigger oxidative damage, inflammatory reactions, and cellular degeneration.
Copper oxide nanoparticles can generate excessive reactive oxygen species, alter membrane permeability, disturb mitochondrial function, and induce lipid peroxidation [8,9] . Therefore, the present study was designed to investigate the clinicopathological, biochemical, oxidative, inflammatory, and histopathological changes induced by repeated oral administration of CuO nanoparticles in male Wistar rats. However, information regarding the dose-dependent and time-dependent hepatic alterations following prolonged exposure remains limited. Therefore, the present investigation was undertaken to study the toxicopathological effects of CuO nanoparticles on the liver of male Wistar rats.
Materials and Methods
Experimental location
The study was conducted under controlled laboratory conditions using healthy male Wistar rats maintained according to standard animal husbandry practices.
Preparation and dose selection of CuO nanoparticles
A pilot study was conducted before initiation of the experiment to determine the LD50 of CuO nanoparticles.
Based on the LD50 value, three sublethal dose levels were selected for repeated oral administration.
Experimental animals
A total of 72 healthy adult male Wistar rats were used in the study. The animals were maintained under standard laboratory conditions with free access to feed and water throughout the experiment.
Experimental design
The rats were randomly divided into four groups: The CuO nanoparticles were administered orally for 45 days.
Parameters studied
The following parameters were evaluated at 15, 30, and 45 days of exposure: Clinical signs and body weight changes Organ weight of liver Serum biochemical parameters including AST, ALT, creatinine, and BUN Histopathological examination of liver tissue
Sample collection
At each interval of 15, 30, and 45 days, six rats from each group were sacrificed. Blood samples were collected for hematological and biochemical analysis. Liver tissue samples were collected for oxidative stress studies and histopathological examination.
Statistical analysis
The data obtained were analyzed statistically to determine significant differences among the control and treated groups at different intervals of exposure.
Results
Clinical signs and body weight changes
No obvious clinical signs of toxicity were observed up to the third week of experimentation in any treatment group. However, from the fourth week onward, rats in Groups II and III exhibited decreased feed intake, emaciation, drowsiness, and rough body coat. Body weight gain was reduced significantly in Groups II and III during the later stages of exposure. The mean values of body weight (in grams) of rats in different Groups (I, II, III and IV) at various intervals are depicted fig no 1.
Histopathological findings
The liver of control rats showed normal hepatic architecture throughout the study period. On day 15, Group II showed moderate congestion and inflammatory cell infiltration, while Groups III and IV exhibited normal histology. Histopathological finding of seminal vesicle in the rat of experimental groups (II, III and IV) are illustrated in
Discussion
The present investigation demonstrated that oral administration of copper oxide nanoparticles caused progressive and dose-dependent liver injury in male Wistar rats. The toxic changes became more prominent with increasing dose and duration of exposure. The reduction in feed intake and body weight gain in treated groups may be attributed to reduced appetite, altered metabolism, and general systemic stress associated with nanoparticle exposure. Such findings indicate that prolonged exposure to copper oxide nanoparticles adversely affects normal physiological functions. Serum biochemical analysis revealed marked increases in AST and ALT activities in treated animals [10,11] . These enzymes are normally present within hepatocytes, and their elevated concentration in blood indicates membrane damage and leakage from injured liver cells. Therefore, the rise in these enzymes suggests that CuO nanoparticles produce direct hepatocellular injury. Overall, the study indicates that CuO nanoparticles can produce severe hepatic injury after prolonged administration and that oxidative stress and inflammation are the principal mechanisms involved in the pathogenesis of toxicity.
Conclusion
The present study concluded that CuO nanoparticles induce significant hepatotoxicity in male Wistar rats following repeated oral exposure. The toxic effects were more severe at higher doses and longer exposure durations. CuO nanoparticles caused alterations in body weight, hematological parameters, liver function biomarkers, oxidative stress markers, inflammatory mediators, and histopathological architecture of the liver. The major mechanism involved in hepatic toxicity appears to be excessive production of reactive oxygen species leading to oxidative stress, inflammation, and cellular damage. Therefore, prolonged exposure to CuO nanoparticles should be carefully monitored due to their potential to induce serious liver injury.
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