A wide range of clinical applications are reported for doxorubicin (DOX), yet both people and research animals experience substantial tissue damage when exposed to DOX, according to Sheibani et al. [28]. Hanumegowda and Davis, [29] and Eguchi et al. [30], have reported that the cytotoxicity of DOX is associated with the generation of acrolein during drug metabolism which resulted to lipid peroxidation, excess reactive oxygen species (ROS) production, reduced antioxidant defense system and a severe inflammatory response via neutrophil attraction. Dietary antioxidants have recently drawn more attention for their function in protecting the liver from DOX-induced toxicity due to the minimal side effects of utilization [31]. Strong antioxidant qualities and cytoprotective efficiency against a variety of diseases induced by free radicals are attributes of lutein, according to Maiuolo et al. [32]. Thus, in order to evaluate lutein's potential protective mechanism against doxorubicin-induced liver damage in male Wistar rats, this experiment was conducted.
The structure and functionality of biological membranes can be harmed by the chemical process known as lipid peroxidation. It is caused by lipids being attacked by free radicals [33, 34]. The present investigation demonstrated increased levels of hepatic malondialdehyde, a lipid peroxidation marker. This implies that doxorubicin can generate reactive oxygen species (ROS) in the liver that attack lipids affixed to the hepatic cell membrane, leading to peroxidative damage. In keeping with earlier research, animals given doxorubicin also showed noticeably higher hepatocellular nitrite levels. Belenchev et al. [35] state that the production of extremely cytotoxic peroxynitrite, which is the result of the interaction between superoxide anions and nitric oxide, mediates nitrergic damage. Asiwe et al., [36], hypothesized that increased nitrite levels in the liver brought on by doxorubicin therapy may have caused the release of glutamate and pro-inflammatory cytokines. Thus, an environment that encouraged hepatocellular injury might have been produced. Interestingly, lutein pretreatment significantly decreased liver nitrite and MDA levels, indicating cytoprotection against lipid peroxidative damage. One of the key mechanisms underlying the cytotoxic effects of DOX is the overproduction of reactive oxygen species (ROS), which in turn causes enhanced lipid peroxidation and the consequent inhibition of the antioxidant defense system. This report's findings, which are consistent with earlier studies [19, 37], indicated that DOX administration increased oxidative stress. This was shown in the liver tissues by a significant decrease in the levels of GST, SOD, catalase activities as well as GSH level. The overproduction of ROS mediated by doxorubicin may inactivate catalase, GST, and SOD, resulting in a decrease in their activity. Conversely, GSH depletion was previously thought to be caused by the direct conjugation of DOX and its metabolites (acrolein) with free or protein-bound SH groups, which interfered with the antioxidant functions [37, 38]. As mentioned earlier, GSH deficiency weakens the body's defenses against free radical-induced cellular damage, which can result in necrotic cell death [39]. The pretreatment with lutein showed remarkable anti-oxidative action, correcting the poor antioxidant status in liver tissues by significantly raising the GSH level, SOD, catalase and GST activities. Notwithstanding, due to its chemical composition, lutein has a considerable scavenging activity, which is noteworthy [40]. Additionally, it has been shown that lutein can repair the cell's deficient thiol status by promoting the de novo synthesis of GSH [41]. Still, the increase in GSH after lutein injection might be part of the defense against DOX-induced liver injury.
The activities of alkaline phosphatase (ALP), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), and alanine transaminase (ALT) were significantly elevated after doxorubicin treatment. Due to its propensity to increase cell membrane permeability and release these enzymes into the bloodstream, hepatocellular necrosis may be the cause of these consequences. Similar results were found in the serum of rats exposed to lead acetate by Asiwe et al. [42] and Asiwe et al. [43]. According to supplementary research, the principal factor responsible for liver damage in rats receiving doxorubicin is an increase in serum levels of LDH, AST, ALT, and ALP [20, 42]. The capacity of lutein to preserve membranes may be the reason why pretreatment with lutein greatly decreased the high levels of AST, ALT, ALP, and LDH. As a biological defensive mechanism against substances that cause disease, Maheshwari et al. [44], have connected immune cell activation generated by oxidative damage to inflammatory responses. In this study, it was discovered that exposure to doxorubicin raised levels of pro-inflammatory cytokines (TNF-α and IL-6). Nevertheless, lutein pretreatment significantly reduced the levels of TNF-α and IL-6 in the liver. This is consistent with a previous study that found that lutein can prevent NF-kB activation, a protein that causes the production of inflammatory cytokines. Numerous studies have connected oxidative stress and inflammatory reactions to mitochondria-mediated cell death [45]. The primary theory is that excessive ROS production leading to DNA uncoupling releases cytochrome-C into the mitochondria, which starts the signaling cascade that ends in cell death. According to this study, doxorubicin treatment significantly reduced Bcl-2 and increased caspase-3 activity, which in turn activated the apoptotic process. But as previous studies have shown, lutein administration significantly decreased caspase-3 activity while increasing liver Bcl-2.
Recent research has demonstrated that dysregulated cellular processes, specifically autophagy and the mTOR pathway, are responsible for the hepatocellular damage caused by doxorubicin. Autophagy, a cellular breakdown mechanism that exposes intracellular entities to lysosomal degradation, is essentially necessary for maintaining cellular homeostasis [46]. Many liver disorders have been linked to dysregulation of Beclin-1-mediated autophagy, an important regulator of autophagy that acts as an initiator [10]. Also, Ballesteros-Álvarez and Andersen [47], have observed intricate relationships between autophagy and the mTOR pathway, which is a crucial regulator of cell growth and metabolism. In this investigation, doxorubicin exposure significantly increased beclin-1 levels and lowered mTOR levels in the liver, indicating a dysregulated autophagic mechanism [10]. The pretreatment of lutein inhibited Beclin-1 and activated mTOR, protecting hepatocellular components and restoring normal hepatocellular activity.
Upon histological inspection, the effects of doxorubicin exposure were observed, including substantial eosinophilic infiltration, inflammation, and liver cell shrinkage and necrosis. Additionally, doxorubicin therapy caused interstitial collagen fibrils to deposit and a significant infiltration of mono nuclear cells into the liver tissues which was consistent with the reports of El-Horany et al., [48]. Furthermore, doxorubicin therapy dramatically reduced liver weight, indicating liver tissue mass necrosis and atrophy. However, following lutein pretreatment, the fibrotic area significantly reduced and the liver's histoarchitecture appeared normal. Furthermore, there was a discernible improvement in the liver's weight, indicating that liver cells were regenerating and tissue mass was being restored. Other studies have demonstrated improvements in oxidative stress and restoration of the antioxidant defense in tissues, which may account for the improved liver tissue structure brought about by lutein therapy [49, 50].