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American Journal of Public Health Research
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The Implementation of a Tele-Homecare System with Service Oriented Architecture and HL7 Message Transmission Standard
Ching-Sung Wang

American Journal of Medical and Biological Research
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The Influence of Ethanol on Pyruvate Kinases Activity in Vivo, in Vitro, in Silico
Lelevich Sergey Vladimirovich, Khrustalev Vladislav Victorovich, Barkovsky Eugene Victorovich, Shedogubova Tatyana Aleksandrovna

American Journal of Medical Sciences and Medicine
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Clinical and Histological Features in Wegener Granulomatosis
Li Cavoli Gioacchino, Passantino Rita, Bono Luisa, Tortorici Calogera, Ferrantelli Angelo, Giammarresi Carlo, Rotolo Ugo

American Journal of Infectious Diseases and Microbiology
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The Curse of Lymphatic Filariasis: Would the Continual Use of Diethylcarbamazine Eliminate this Scourge in Papua New Guinea?
Anne Lanham, Lillian Mwanri

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Dear Label_Name,

Greetings! I hope you are well.

I am writing on behalf of an international publishing house, Science and Education Publishing, USA (http://www.sciepub.com).

Science and Education Publishing (SciEP) is a multidisciplinary open access publisher of Journals covering the fields of Science, Technology and Medicine. All SciEP publications are subject to high-quality peer review, editorial and production processes and are freely available online.

Welcome to the April, 2013 edition of SciEP newsletter, providing monthly summaries of articles on science & education related topics of interest to scientist and professionals. The mission of our journals is to provide an open access forum for publishing new findings on issues pertinent to scientific and academic field. We also invite you to submit original research papers, review articles, short reports, or case studies are considered for publication in our Multidisciplinary Journals.

American Journal of Public Health Research
DOI: 10.12691/ajphr-1-1-3

The Implementation of a Tele-Homecare System with Service Oriented Architecture and HL7 Message Transmission Standard
Ching-Sung Wang

American Journal of Medical and Biological Research
DOI: 10.12691/ajmbr-1-1-2

The Influence of Ethanol on Pyruvate Kinases Activity in Vivo, in Vitro, in Silico
Lelevich Sergey Vladimirovich, Khrustalev Vladislav Victorovich, Barkovsky Eugene Victorovich, Shedogubova Tatyana Aleksandrovna

American Journal of Medical Sciences and Medicine
DOI: 10.12691/ajmsm-1-1-1

Clinical and Histological Features in Wegener Granulomatosis
Li Cavoli Gioacchino, Passantino Rita, Bono Luisa, Tortorici Calogera, Ferrantelli Angelo, Giammarresi Carlo, Rotolo Ugo

American Journal of Infectious Diseases and Microbiology
DOI: 10.12691/ajidm-1-1-2

The Curse of Lymphatic Filariasis: Would the Continual Use of Diethylcarbamazine Eliminate this Scourge in Papua New Guinea?
Anne Lanham, Lillian Mwanri

We have released issues of some journals on February 2013.
American Journal of Public Health Research
American Journal of Medical Sciences and Medicine
American Journal of Medical and Biological Research
American Journal of Pharmacological Sciences
American Journal of Clinical Medicine Research
American Journal of Nursing Research
American Journal of Cancer Prevention
American Journal of Medicine Studies
American Journal of Infectious Diseases and Microbiology
American Journal of Cardiovascular Disease Research
American Journal of Biomedical Research
American Journal of Mechanical Engineering
American Journal of Microbiological Research
Journal of Preventive Medicine
American Journal of Food and Nutrition
American Journal of Hypertension Research
American Journal of Sports Science and Medicine
Journal of Cancer Research and Treatment
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The Antioxidant Properties of Lutein Efficiently Mitigate the Cadmium-Induced Oxidative Stress through Restoring the Oxidant: Antioxidant Balance and Enhancing the Total Antioxidative Capacity in Wistar Rats

Mohammed Mubarak*, Mohammed Al-Zharani, Hassan Rudayni, Eman AlmuqriShaikha A. Albatli, Saad Alkahtani, Fahd A. Nasr, Amin A. Al-Doaiss, Mohammed S. Al-eissa

Department of Biology, College of Science, Imam Mohammad Ibn Saud Islamic University (IMSIU), Riyadh 11623, Saudi Arabia

Abstract: Lutein is a xanthophyll carotenoid existing in dark green vegetables. The objective of this study is to test the antioxidant properties of Lutein and evaluate its efficiency in mitigating the cadmium-induced oxidative stress. Eighty experimental rats were randomly allotted into four groups (n=20 each): an untreated control group (Group 1), a lutein-treated group (Group 2), a cadmium-exposed group (Group 3), and a cadmium-lutein group (Group 4). Group 2 rats received daily oral doses of lutein at 200 mg/kg body weight and those in Group 3 were given an aqueous solution of cadmium chloride at a final concentration of 5 mg/kg b.w. per day. Rats in Group 4 were treated with both lutein and cadmium chloride. The haematological and biochemical profiles of the cadmium-exposed rats (Group 3) showed significant alterations compared to the untreated control. The biochemical assessments of the lutein-treated rats (Group 2) exhibited no significant changes compared to the untreated control. Rats in Group 4 (exposed to cadmium and treated with lutein) exhibited increased levels of total proteins and significant increases in the antioxidant markers, including total thiols, glutathione, total antioxidant capacity (TAC), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase. Additionally, this group demonstrated significant decrements in the blood cadmium levels, the oxidation markers (H₂O₂ and malondialdehyde), alkaline phosphatase (ALP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatinine, blood urea nitrogen (BUN), urea, and bilirubin compared to the cadmium group (Group 3). Tissue homogenates prepared from the liver and kidneys of Group 4 rats exhibited parallel results to those demonstrated by the serum biochemical assay. Based on the present results, it could be concluded that the antioxidant properties of lutein significantly alleviate the cadmium-induced oxidative stress.

Keywords: Lutein, cadmium toxicity, oxidative stress, antioxidant, biochemical profile

Received February 23, 2025; Revised March 24, 2025; Accepted April 01, 2025

Citation: Mohammed Mubarak, Mohammed Al-Zharani, Hassan Rudayni, Eman AlmuqriShaikha A. Albatli, Saad Alkahtani, Fahd A. Nasr, Amin A. Al-Doaiss, Mohammed S. Al-eissa, The Antioxidant Properties of Lutein Efficiently Mitigate the Cadmium-Induced Oxidative Stress through Restoring the Oxidant: Antioxidant Balance and Enhancing the Total Antioxidative Capacity in Wistar Rats. Journal of Food and Nutrition Research, 13 (3). 163-171. January 2025. doi:10.12691/jfnr-13-3-6

1. Introduction

Lutein ( C₄₀ H₅₆ O₂ ) is a xanthophyll carotenoid encountered in dark green vegetables, and used on a large scale as a dietary supplement based on its significant health benefits [1].

Carotenoids (tetra-terpenes) are known to be synthesized by some microbes, fungi, and algae as well as higher plants [2]. These phytochemicals include two main chemical groups; hydrocarbon carotenes (β-carotene and lycopene) and the oxygen containing xanthophylls (lutein) [3].

The research carried out on lutein, and carotenoids in general, has approved diverse health benefits and protective effects as demonstrated in the clinical and experimental investigations [4,5]. These investigations revealed a positive impact in multiple conditions of significant medical concern such as cardiovascular diseases, cancer, and neudegenerative diseases. The health promoting effects of lutein are largely ascribed to its antioxidative and anti-inflammatory activities [6].

Lutein has been demonstrated to possess a greater antioxidant efficiency compared to other carotenoids [7].

Oxidative stress is one of the major contributors in the development of many diseases and age-related changes [8,9,10,11]. Oxidative stress is referred to as sequential events that eventually lead to imbalance between Oxidation reactivity (mainly the activities of reactive oxygen species) and efficacy of the endogenous antioxidant system to encounter this hazardous reactivity [12]. Certain metabolites or external toxins could trigger the oxidation reactions and contribute to varied forms of cellular and tissue damage. These oxidative harmful effects can be vigorous enough to cause cell death and significant tissue damage. A controlled level of ROS is essential to accomplish vital cellular metabolic activities conversely the excessive generation of reactive oxygen species results in the status of oxidative stress and the associated tissue damage [13].

The induced excess generation of ROS damages cellular lipids, proteins, and DNA, and cause neurodegenerative disorders, cardiovascular diseases, cancer, diabetes, accelerated aging, and immune dysfunction [14,15]. In other words, oxidative stress could be a significant contributing factor to the development of complications of high medical concern. These actions can be counteracted by the efficient endogenous antioxidant system, which identifies oxidative metabolites, such as free radicals, and alleviate their hazardous effects. [16]. Dysfunction of this defense system can significantly contributes to the development of serious complications [17,18]. The major role of a potent antioxidant is to limit the damage caused by ROS and actively maintain the redox status as a cytoprotective mechanism [19].

Antioxidants that are used as natural dietary supplements such as polyphenols, vitamins, terpenes, and carotenoids are mainly of plant sources [20,21]. Two main categories of these natural bioactive compounds are known; the water-soluble group (hydrophilic group) including vitamin C, glutathione, and Lutein, and the lipid-soluble (hydrophobic group), which encompass vitamins A and E [18]. The hydrophilic antioxidants perform their activities in cytoplasm and blood plasma while the hydrophobic ones function in the cellular membranes. Most of the relevant experimental and clinical studies clearly indicate the role of natural antioxidants in enhancing and maintaining the vital body functions, and slowing the signs of aging-related changes [19,20,21].

Natural antioxidants as dietary supplements have been the focus of many studies and their usage is on the rise as preventive and therapeutic strategies for various major health issues [22,23]. These are exemplified by their beneficial effects in cardiovascular diseases, neurodegenerative disorders, cancer, arthritis, and age-related changes [24,25,26].

The widely used antioxidants including vitamins C and E, selenium, zinc, polyphenols, and carotenoids are often deficient in the regular food sources. This explains the growing demands for antioxidant supplements that support the bioactive processes and help maintain the overall health. The effective antioxidants exert their activities through maintaining the redox status and combating the oxidation reactions [19].

Some highly hazardous toxic heavy metals occur naturally in the environment, among these is cadmium (Cd), which is used in a variety of industries. Exposure to cadmium can be either occupational or non-occupational (environmental). Occupational exposure typically happens through the inhalation of industrial fumes, while non-occupational exposure is often associated with the ingestion of contaminated food and water. The long-term accumulation of this heavy metal results in chronic toxicity with the development of progressive harmful effects in cells and tissues [27]. Cumulative cadmium toxicity is associated with dramatic oxidative stress and tissue damage, especially vital organs such as liver and kidneys.

The objective of the current research work was to evaluate the antioxidant properties of the lutein in male Wistar rats exposed to cadmium toxicity. The employed cadmium-induced oxidative stress represents the target challenge for which the antioxidant properties are tested.

2. Materials and Methods

Ethical Considerations:

The guidelines of care and use of laboratory rats established by the Research Ethics Committee of Imam Mohammad Ibn Saud Islamic University (IMSIU), were strictly followed per institutional and national regulations (LAB-rats-2024-0352).

Type of Sampling and reasons for selection:

The whole Blood, serum harvested from blood samples, and tissue homogenates were selected for investigation in this study. These samples were chosen to reflect changes in the hematological and biochemical profiles of the experimental rats.

Inclusion and Exclusion Criteria:

Inclusion Criteria

All the experimental groups of rats were included in the assays conducted. Blood samples were collected from all experimental rats to perform a hematological assay, which assessed erythrocyte and leukocyte counts, hemoglobin concentration, and packed cell volume. Additionally, the harvested serum samples and tissue homogenates from all experimental rats were subjected to a biochemical assay to estimate the levels of antioxidant and oxidative parameters.

Exclusion Criteria

In the present study, no exclusion criteria were applied. All experimental rats were included in the collection of samples (blood and tissues). Any exclusion criteria could alter the accuracy of the analysis performed.

Experimental rats:

In the present research, eighty adult male Wistar rats, each three months old and weighing between 120 and 215 grams. The rats were sourced from the inbred colonies at the animal house of the College of Pharmacy at King Saud University in Riyadh, Saudi Arabia. The rats were maintained under standard laboratory conditions, with an ambient temperature set at 24 ± 1°C, a 12-hour light-dark cycle, and a relative humidity level ranging from 35% to 70%. Throughout the experimental period, all animal groups were monitored for food intake, abnormal behavior, and mortality.

Lutein:

C₄₀H₅₆O₂ (Xanthophyll, α-Carotene-3,3′-diol, β, ε-carotene, β, ε-carotene-3,3′-diol, Lutein) (Molecular Weight: 568.87, ≥96.0% (HPLC) , CAS Number: 127-40-2) was purchased from Sigma-Aldrich (Darmstadt, Germany)

Cadmium:

Cadmium (Cd) was used in the form of cadmium chloride (CdCl2) of analytical grade (Merck, Darmstadt, Germany) (Product No. 655198). Cadmium was dissolved in purified water to prepare the required aqueous solution.

Experimental design:

After an appropriate acclimatization period (one-week period), rats were randomly assigned to four groups, each consisting of 20 rats. The groups were designated as Group 1, Group 2, Group 3, and Group 4. Rats in Group 1 served as the untreated control; they were not exposed to cadmium and did not receive lutein. Rats in Group 2 received daily oral doses of lutein at 250 mg/kg body weight (b.w.) in a volume of 1 mL/kg b.w. Rats in Group 3 were given an aqueous solution of cadmium chloride (CdCl2) via oral gavage at a final concentration of 5 mg/kg b.w. per day in a volume of 1 mL/kg b.w. The control rats received an equal volume of saline via the same route. Rats in Group 4 were administered cadmium orally and also received lutein at the same doses as mentioned earlier. There was a 10-hour interval between the daily administration of cadmium and lutein. The experimentation period lasted for 8 weeks, during which dry feed (commercial pellets from Nesom Distributing Envigo, USA) and drinking water were provided ad libitum. All experimental rats were observed for behavioural activity, feed consumption, water intake, and clinical signs throughout the study.

Haematological and biochemical assays:

On the last experimentation day, the rats were anaesthetized with 3% isoflurane, and blood samples were collected via cardiac puncture from all the rats in the different groups. Blood samples collected with the anticoagulant EDTA were used to estimate various hematological indices. Serum harvested from the coagulated blood samples was removed immediately and stored at -20 °C until biochemical assays were performed. The rats were then euthanized by decapitation, and their liver and kidney tissues were removed and homogenized in 150 mM NaCl. The homogenates were subsequently centrifuged at 3000 × g at 4 °C for 10 minutes. The collected supernatants were used to determine various biochemical parameters.

Blood cadmium level:

Blood cadmium levels were measured, 1 mL blood samples were digested using a mixture of HClO4 and HNO3. Blood cadmium levels were then determined using an atomic absorption spectrophotometer (CBC 906 AA).

Haematological assay:

To assess the haematological parameters, blood samples collected with the anticoagulant EDTA were used, including red blood cell (RBC) counts, total white blood cell (WBC) counts, and other erythrocytic indices such as hemoglobin (Hb) concentration and packed cell volume (PCV) percentage. Erythrocyte and total leukocyte counts were measured using a hemocytometer. The PCV percentage was determined using the micro-hematocrit method, while Hb concentration was estimated using the cyanmethemoglobin method as previously described.

Biochemical assay:

Serum samples were harvested from the coagulated blood, and used to estimate various biochemical parameters, including total proteins, albumin, globulin, creatinine, urea, blood urea nitrogen (BUN), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP). Additionally, antioxidant markers were assessed, such as total thiols, catalase, glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and total antioxidant capacity (TAC). Oxidation markers, including malondialdehyde (MDA) and hydrogen peroxide (H2O2), were also measured.

A colorimetric assay kit for total thiols (Cell Biolabs Inc., USA) (MET-5053) were used to measure the total thiols. Glutathione (GSH) levels were estimated using a reduced glutathione colorimetric assay kit (ElabScience, USA) (E-BC-K030-S). Catalase levels were determined with a catalase activity colorimetric assay kit (BioVision; Abcam, UK) (ab 83464). Superoxide dismutase (SOD) activity was assessed using a SOD activity assay kit (Sigma-Aldrich, Darmstadt, Germany). Additionally, glutathione peroxidase activity was measured using a GSH-Px activity assay kit (Elabscience, Houston, Texas, USA).

Estimation of the total antioxidant capacity (TAC) was done by employing the total antioxidant capacity assay kit (Sigma-Aldrich, Germany) (MAK 187-1 KT). This kit operates on the principle of measuring the concentration of combined protein and small-molecule antioxidants, or solely small-molecule antioxidants. In this process, Cu²⁺ ions are reduced to Cu⁺ by small molecules and proteins. However, the inclusion of a protein mask as part of the kit prevents the reduction of Cu²⁺ by proteins, allowing for the analysis of only small-molecule antioxidants. The reduced Cu⁺ ions, produced by the action of the small molecules, are then chelated with a colorimetric probe. The resulting absorbance peak is directly proportional to the antioxidant capacity.

Colourimetric assay kits were utilized to determine H₂O₂ levels (Elabscience, USA) (E-BC-K102-S) and MDA levels (Elabscience, USA) (E-BC-K028-M). ALT, AST, and ALP levels were measured using the relevant diagnostic kits (Abcam, UK) (Catalog Nos.: ALT ab105134, AST ab105135, ALP ab83369).

Blood urea nitrogen (BUN) levels were measured using a BUN colorimetric detection kit from ThermoFisher Scientific (USA) with Catalog No. EIABUN. Urea levels were determined using a colorimetric assay kit from BioVision (Biovision Incorporated, UK) with Catalog No. K375-100. Other biochemical parameters, including total proteins, creatinine, bilirubin, albumin, and globulin, were assessed using the appropriate colorimetric diagnostic kits from Interchim Diagnostics (France). The corresponding catalog numbers for these kits are as follows: total proteins FT7250, creatinine FT7040, bilirubin FT 6920, albumin FT 6760, and globulin FT 7253.

Homogenates from the liver and kidney tissues were prepared to measure the levels of total thiols, glutathione, catalase, hydrogen peroxide (H2O2), malondialdehyde (MDA), and total antioxidant capacity (TAC) in the tissues. The same assay kits were employed to assess these parameters' serum levels in the tissue homogenates.

Statistical analysis:

In the current investigation, the submitted data were expressed as means ± standard deviation (S.D.). To compare the means among multiple groups, one-way ANOVA was conducted using SPSS software (SPSS Inc., Chicago, IL, USA) for statistical analysis. The normality and homogeneity of variances were checked and the independence of observations was ensured. The normality of the data was verified using the Shapiro-Wilk test. Results with a P-value less than 0.05 (P < 0.05) were considered statistically significant.

3. Results

Rats administered with lutein (Group 2) as well as those exposed to cadmium and also treated with lutein, displayed normal behavior, activity levels, and food intake compared to the control rats that were not treated. In contrast, rats exposed to cadmium but not given lutein showed a decrease in activity and reduced food intake starting from the third week of the experiment, compared to the control group. No deaths were observed among any of the rats in the experimental groups.

Blood cadmium level in the control rats measured at 0.0019 ± 0.0001 ppm. This level significantly increased (P < 0.05) in rats exposed to cadmium, reaching 0.569 ± 0.018 ppm. Notably, blood cadmium levels were lower in rats that were exposed to cadmium but also received lutein, with a measurement of 0.224 ± 0.015 ppm, compared to those exposed solely to cadmium.

Group 2 rats that were administered with lutein had no significant differences in haematological parameters compared to the control group. However, rats exposed to cadmium without lutein (Group 3) showed notable decreases in haemoglobin (Hb) concentration and packed cell volume (PCV) %. In contrast, rats exposed to cadmium and treated with lutein (Group 4) showed improvements in their haematological parameters, approaching control levels (Table 1).

Table 1. Haematological assay of rats that received lutein, exposed to cadmium, and that were exposed to cadmium and administered with lutein, compared to the control rats. RBC count, total leucocytic count, haemoglobin concentration, and packed cell volume percentage of cadmium-exposed rats were significantly lower compared to the control rats. The measured haematological indices in lutein-administered rats exhibited no significant differences compared to the control rats. In cadmium-exposed rats and those administered with lutein, the estimated haematological parameters showed significant increments compared to cadmium-exposed rats and were closer to the control levels

Table 1. Shows the estimated haematological parameters in rats that received lutein, those exposed to cadmium, and those exposed to cadmium with lutein administration, compared to the control rats.

Regarding the biochemical profile, the rats that received lutein (Group 2) exhibited no significant changes in their biochemical parameters compared to the control group. In contrast, the rats in Group 3, which were exposed to cadmium and did not receive lutein, showed comparable decreases in the estimated levels of total proteins, albumin, and globulin. Furthermore, this group experienced a significant increase in creatinine, urea, and blood urea nitrogen (BUN) levels.

The serum levels of total thiols, glutathione, superoxide dismutase, glutathione peroxidase, and catalase significantly decreased in the group of rats exposed to cadmium that did not receive lutein (Group 3). Additionally, the total antioxidant capacity (TAC) in both serum and tissues was significantly reduced in the cadmium-exposed rats but improved in those that received lutein.

Malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) levels in the serum of cadmium-exposed rats without access to lutein were significantly higher compared to the control levels (Table 2 a, b, c).

Table 2a. Serum levels of total proteins, albumin, and globulin were significantly lower in cadmium-exposed compared to the untreated control rats. Levels of creatinine, urea, blood urea nitrogen (BUN), and bilirubin in the cadmium-exposed rats were significantly increased compared to the untreated control rats. Rats administered with lutein demonstrated no significant differences in their biochemical parameters compared to the untreated control rats. The estimated biochemical parameters in cadmium-exposed and lutein-treated rats exhibited improvements towards the control levels and were significantly different from those estimated in the cadmium-exposed rats

Table 2 (a, b, c). Shows the biochemical parameters (serum levels) in rats that were administered lutein, those exposed to cadmium, and those exposed to cadmium and given lutein, compared to the control rats.

Table 2b. Serum levels of alanine transferase (ALT), aspartate transferase (AST), and alkaline phosphatase (ALP) in the different groups. The cadmium-exposed rats had significantly increased levels of all measured biochemical parameters. No significant differences were recorded in the lutein group compared to the control group. In the group of rats exposed to cadmium and administered with lutein, the biochemical parameters displayed improvements toward the control levels and were significantly decreased compared to the cadmium-exposed rats

Table 2c. Serum levels of total thiols, glutathione (GSH), catalase, superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), total antioxidant capacity (TAC) malondialdehyde (MDA), hydrogen peroxide (H2O2), and total antioxidant capacity (TAC) in the different groups. Compared to the untreated control rats, the cadmium-exposed rats exhibited significantly decreased levels of total thiols, GSH, catalase, and TAC. The levels of malondialdehyde (MDA) and hydrogen peroxide (H2O2) were significantly increased in cadmium-exposed rats compared to the untreated control rats. Lutein-treated rats showed no significant differences in the measured biochemical parameters compared to the untreated control rats. In the rats exposed to cadmium and treated with lutein, the biochemical parameters showed improvements toward the control levels and were significantly different compared to those of the cadmium-exposed rats

In rats that were exposed to cadmium and administered lutein (Group 4), the altered biochemical parameters estimated in tissue homogenates prepared from liver and kidneys were brought closer to control levels Table 3.

Table 3. Shows the levels of total thiols, glutathione, catalase, glutathione peroxidase, superoxide dismutase, total antioxidant capacity (TAC), hydrogen peroxide (H₂O₂), and malondialdehyde (MDA) in the liver and kidney homogenates of rats in different experimental groups.

Table 3. Levels of total thiols, glutathione (GSH), catalase, superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), total antioxidant capacity (TAC), malondialdehyde (MDA), hydrogen peroxide (H2O2), and total antioxidant capacity (TAC) in the tissue homogenates of different groups. The levels of total thiols, GSH, catalase, and TAC were significantly decreased in the tissue homogenates of cadmium-exposed rats compared to untreated control rats. MDA and H2O2 levels were significantly increased in the tissue homogenates of cadmium-exposed rats compared to untreated control rats. Lutein-treated rats showed no significant differences compared to untreated control rats. The estimated biochemical parameters in tissue homogenates of rats exposed to cadmium and administered with lutein demonstrated improvements toward the control levels and were significantly different compared to those measured in the cadmium-exposed rats

4. Discussion

In the present experimental work, lutein supplement with cadmium exposure were used as an experimental model to evaluate the antioxidant properties against induced oxidative stress. The investigation aimed to test the efficacy of lutein to improve the haematological and biochemical alterations caused by cadmium-induced oxidative stress.

Some physiological metabolic processes (extra- and intra-cellular) such as cellular respiration are mediated by free radicals. However, in certain circumstances including diseases, toxicities, and environmental factors, excess amounts of free radicals are generated [28]. The body's internal antioxidant system manages excess free radicals to prevent their harmful effects. This endogenous antioxidant system encompasses enzymes such as superoxide dismutase, catalase, and glutathione peroxidase, as well as non-enzymatic molecules. Each antioxidant molecule functions to combat free radicals through specific mechanisms and mitigates their harmful effects. In this way, endogenous antioxidants help maintain the balance between oxidation reactions and antioxidant activities [29].

The hazardous effects of free radicals appear on the stage when the antioxidant system is not efficient enough to counteract them. These unstable molecules can oxidize other cellular biomolecules, resulting in oxidative stress. The oxidation reactions extend to damage cellular proteins and DNA, potentially leading to cell death. One significant outcome of this oxidative damage is lipid peroxidation, which primarily affects cell membranes.

Oxidative stress is characterized by the excessive production of reactive oxygen species (ROS), which include free radicals, such as hydroxyl and superoxide radicals, as well as non-radical compounds like hydrogen peroxide [30]. Whenever oxidative stress (oxidative overload) takes place, there is an imbalance between oxidation reactions and antioxidant activities, which is referred to as redox status. Antioxidants prevent the chain reactions initiated by free radicals mainly by stabilizing these reactive molecules. Antioxidants donate electrons to free radical molecules, which possess unpaired electrons and thus block their high reactivity [31]. Additionally, antioxidants could ameliorate the oxidative stress through multiple protective mechanisms involving scavenging of ROS [32,33,34]. However, with the progression of oxidative stress, the effectiveness of the antioxidant system is gradually diminished in managing the harmful effects of ROS. This sequence of events explain the development of oxidative stress-induced diseases and disorders.

Cadmium toxicity is a potent trigger of excess generation of ROS with eventual development of oxidative stress that leads to significant damage in the liver and kidneys [35]. Cadmium cytotoxicity contributes to oxidation of cell membrane lipids and significantly reduces the production of ATP and glutathione levels in the mitochondria. Additionally, cadmium toxicity disrupts the function of antioxidant enzymes, which exacerbates the existing oxidative stress. Ultimately, cadmium-induced toxicity triggers apoptosis through the activation of caspases [36].

To assess the antioxidative status, reliable markers including total thiols, glutathione, superoxide dismutase, glutathione peroxidase, catalase, and total antioxidant capacity (TAC) were employed in the present investigation. The levels of these marker molecules were found to be significantly decreased in rats exposed to cadmium. These decrements suggest that these antioxidants were negatively impacted by excessive oxidative stress. It is proposed that ROS generated during this process caused direct oxidation of these antioxidant molecules which start to suffer reduced capacity to combat oxidative damage [37,38]. In situations when antioxidants become oxidized and inactivated, free radicals increase, leading to heightened oxidative stress.

Antioxidant enzymes including SOD, catalase, and Gpx play a crucial role in maintaining the cellular redox balance [39,40]. Regarding the mechanism of action of endogenous antioxidants, certain endogenous antioxidants, including glutathione and catalase, could function directly to eliminate free radicals. Catalase decomposes cellular hydrogen peroxide (H₂O₂), a compound that is strongly linked to lipid peroxidation. Reduced activity of catalase allows hydrogen peroxide to exert powerful oxidative effects [40,41]. Excess production of H₂O₂ is highly cytotoxic since it triggers peroxide ions that oxidize of cell proteins, DNA, and membrane lipids [42,43]. Additionally, hydrogen peroxide contributes to the Fenton reaction, which produces the highly damaging hydroxyl radicals (OH) [30].

SOD protects from superoxide radicals by converting these radicals into H₂O₂ [44,45] which in turn decomposed by catalase [50]. Moreover, SOD was found to inhibit the apoptotic pathways [46,47].

The antioxidant efficiency of lutein is ascribed to the conjugated double bonds and hydroxyl groups in its structure [48]. The double bonds contribute greatly to the antioxidant properties of lutein by donating electrons to free radicals which become stabilized [49]. Compared to other carotenoids, lutein possesses greater antioxidant activities owing to the terminal location of its hydroxyl groups [50]. Lutein modulates the oxidative stress-induced cellular alterations via inhibition of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and prostaglandin (PG)E₂ [51,52]. Lutein has been reported to scavenge ROS such as singlet oxygen and lipid peroxy radicals, suppress ROS generated by H₂O₂, upregulate the expression of antioxidant enzymes especially SOD [53].

The significant elevation of malondialdehyde (MDA) levels in the presently cadmium-intoxicated rats could be ascribed to lipid peroxidation of cell membranes. MDA acts as a biomarker for lipid peroxidation, which results from oxidative damage [54].

ALT, AST, and ALP (serum enzymes) were significantly elevated in rats exposed to cadmium, indicating liver damage induced by cadmium. This increase may be attributed to damage to the lysosomes caused by lipid peroxidation, which releases lysosomal enzymes into the bloodstream, leading to higher enzyme levels. Furthermore, cadmium exposure also resulted in increased levels of blood urea and creatinine, suggesting kidney damage in the affected rats.

Taken together, the currently recorded biochemical alterations indicate that cadmium toxicity has substantially affected the body's natural antioxidant system and altered the blood's biochemical profile. This effect has led to a depletion of antioxidant molecules, a decrease in overall antioxidant capacity, and a disruption of the natural balance between oxidants and antioxidants in the body. When the body's natural antioxidant system is compromised, it becomes essential to introduce external sources of antioxidants to restore this balance. Antioxidants in the form of supplements are expected to protect biological systems from oxidative damage by quickly eliminating excess free radicals and enhancing the body's antioxidant system [55]. The supplemented antioxidants act synergistically with endogenous antioxidants to neutralize and eliminate free radicals. It is worth mentioning that the byproducts resulting from the reaction of antioxidant molecules with free radicals can eliminate additional radicals, thus enhancing the overall antioxidant effectiveness [56].

The significantly decreased levels of blood cadmium estimated in rats that were previously intoxicated with cadmium and then treated with lutein is ascribed to a potential chelation of cadmium. Undoubtedly, decreasing cadmium level in the blood significantly limits the initiative step of oxidative stress and contributes to the recovery of the endogenous antioxidant system.

The current haematological and biochemical assays revealed improved parameters, which were restored and brought to the control levels. These findings may support the suggested synergistic antioxidant role Lutein in reducing oxidative stress induced by cadmium toxicity. As recorded in the present experimental work, lutein exhibited the potential to significantly upregulate the expression of antioxidant enzymes, which serve as reliable biomarkers of antioxidant activity [57]. Additionally, the present results provide evidence that lutein effectively inhibits lipid peroxidation as indicated by the significant decrease in malondialdehyde (MDA) levels, the marker of lipid peroxidation [58].

The present experimental work demonstrated significant improvements in antioxidant markers (total thiols, glutathione, catalase, glutathione peroxidase, superoxide dismutase, and total antioxidant capacity) in rats exposed to cadmium and given lutein. These findings support the significant positive impact of lutein on the expression of the antioxidant molecules and highlight its contribution in enhancing the endogenous antioxidant capacity.

The results of the present study obviously express the antioxidant properties of lutein in reducing oxidative stress and damage caused by cadmium toxicity. However, further research is recommended to fully elucidate the specific molecular mechanisms by which this potent lutein exerts its antioxidant activities in the context of heavy metal toxicity.

Conclusion

Based on the presently carried out haematological and biochemical assays, it could be concluded that lutein possesses the efficient antioxidant properties to alleviate the cadmium-induced oxidative stress and restore the disrupted oxidation: antioxidation balance.

Availability of Data and Materials

Data will be made available on request

Authors' Contributions

M Z, H R, FN and AAD performed the laboratory work. MAE and SA supervised the experiment and were responsible for resources. EA and SA were responsible for the lab work, software, and statistical data analysis. M M designed the study, supervised the methodology, and wrote the manuscript.

Ethics Approval and Consent to Participate

The guidelines for the care and use of laboratory rats, as per institutional and national regulations, set by the Research Ethics Committee of Imam Mohammad Ibn Saud Islamic University (IMSIU), were diligently adhered to (LAB-rats-2024-0352). All authors were informed about their participation in the present research work.

Conflict of Interest:

The authors declare that there are no conflicts of interest.

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