|Year : 2019 | Volume
| Issue : 1 | Page : 13-17
Lycopene restores liver function and morphology of ifosfamide-intoxicated rats
Elias Adikwu1, Bonsome Bokolo2
1 Department of Pharmacology and Toxicology, Faculty of Pharmacy, Niger Delta University, Bayelsa State, Nigeria
2 Department of Pharmacology, Faculty of Basic Medical Sciences, Niger Delta University, Bayelsa State, Nigeria
|Date of Web Publication||12-Jun-2019|
Dr. Elias Adikwu
Department of Pharmacology and Toxicology, Faculty of Pharmacy, Niger Delta University, Bayelsa State
Source of Support: None, Conflict of Interest: None
Introduction: Low incidence of liver toxicity has been anticipated with the clinical use of ifosfamide (IFO); however, there is possible hepatotoxic concern with its use. There is a paucity of effective drugs that can protect liver or regenerate hepatocytes during damage. In this light, the protective effect of lycopene (LYP) was examined against a rat model of IFO-induced liver injury. Materials and Methods: Forty adult albino rats were randomized into eight groups (A–H). Group A (control) was orally treated with water, whereas groups B–D were orally treated with 10–40 mg/kg of LYP daily for 7 days, respectively. Group E was treated with 150 mg/kg of IFO on the 7th day intraperitoneally (ip), whereas groups F–H were pretreated orally with 10, 20, and 40 mg/kg of LYP daily, respectively, before treatment with IFO on the 7th day (ip). On the 8th day, rats were sacrificed, blood was collected, and serum was separated and evaluated for biochemical parameters. Rats were dissected; liver was collected, weighed, and evaluated for biochemical parameters and histology. Results: Significant (P < 0.001) increases in aminotransferases, total bilirubin, conjugated bilirubin, gamma-glutamyl transferase, lactate dehydrogenase, and malondialdehyde levels with significant (P < 0.001) decreases in superoxide dismutase, glutathione, catalase, and glutathione peroxidase levels were obtained in IFO-treated rats when compared to control. Liver of IFO-treated rats showed periportal and pericentral necroses of hepatocytes. However, The aforementioned parameters were significantly restored in a dose-dependent manner at 10 mg/kg (P < 0.05), 20 mg/kg (P < 0.01) and 40 mg/kg (P < 0.05) of LYP-pretreated rats. Conclusion: This study showed that IFO-induced liver damage was restored in a dose-dependent manner by pretreatment with LYP.
Keywords: Ifosfamide, liver, lycopene, rats, toxicity
|How to cite this article:|
Adikwu E, Bokolo B. Lycopene restores liver function and morphology of ifosfamide-intoxicated rats. Arch Med Health Sci 2019;7:13-7
| Introduction|| |
Drug-associated liver toxicity is a common and frequent cause of acute or chronic liver injury. The manifestation of drug-associated liver injury can occur as acute hepatitis and/or cholestasis; however, any pathological pattern or type of liver injury can manifest. Over the past decade, key discovery points to drug metabolism and excretion-related events being precursors to downstream immune response-mediated liver injury. This discovery could be a function of the parent drug or its metabolites that can affect hepatocyte function and structure. Drug–protein adducts, formed by drugs or their metabolites can interact with host proteins that are presented as neoantigens by major histocompatibility complex class II, thereby triggering an immunoallergic reaction. Furthermore, a parent drug or its metabolites can cause oxidative stress, mitochondrial injury, inhibition of transporters, endoplasmic reticulum and the stimulation of inflammatory processes, leading to liver injury.
Ifosfamide (IFO) is an alkylating cytotoxic agent that has clinical application in the treatment of different types of cancers. It can be used alone or in combination with some anticancer drugs to improve the outcome and success of therapy. However, the clinical use of IFO has been associated with two primary toxicities; myelosuppression and urotoxicity. Data from manufacturers suggest a very low incidence of liver toxicity that may arise with the clinical use of IFO. However, some reports have shown possible hepatotoxic concern with its use. It has been associated with increased systemic activities of hepatic function markers and histoarchitectural damage in rats. The pathogenesis of IFO-induced liver toxicity is yet to be fully elucidated, however; findings suggested the production of chloroacetaldehyde (CAA) metabolite during its hepatic biotransformation. CAA is a toxic metabolite that can cause liver damage through the induction of oxidative, nitrosative stress and lipid peroxidation.
Lycopene (LYP) is a carotenoid present in vegetables and fruits, with tomatoes being the fruit with the most abundant carotenoid content. It has received serious attention due to its biological and physicochemical properties. In comparison to other carotenoid, it has the highest antioxidant activity and ability to terminate free radical activities in vitro and in vivo. It has a high number of conjugated dienes, which increase its ability to effectively scavenge oxidative radicals higher than α-tocopherol and β-carotene. It has the ability to directly regulate redox-sensitive signaling pathways responsible for the modulation of cell functions. The potential health benefits of LYP include prevention of cardiovascular diseases, cancer, hypertension, psychiatric disorders, diabetes, and asthma. In addition, a number of studies with animals reported potential protective effects against oxidative stress-induced diseases. LYP has shown significant anti-inflammatory, antiproliferative, antineoplastic, and neuroprotective effects and can regulate and stabilize normal cell functions. Also, studies have shown that LYP has a protective effect on nonalcoholic fatty liver disease in a rat model, however, with no study on its potential benefit against IFO-induced liver injury. Therefore, the present study examined if it can safeguard the liver of IFO-intoxicated rats.
| Materials and Methods|| |
Forty adult albino rats of 220g-240g average body weight were sourced from the Animal House of the Faculty of Pharmacy, Niger Delta University, Nigeria. The rats were randomized into five groups of five rats each in well-ventilated cages with free access to food and water ad libitum. The rats were kept in housing conditions: 12-h light/dark cycle, and a temperature of 25°C for 1 week before the study. This study was approved by the Research Ethics Committee of the Department of Pharmacology and Toxicology, Niger Delta University, Nigeria. Animals were handled according to the guidelines of the Canadian Council on Animal Care.
Dose selection and animal treatment
This study used 10, 20, and 40 mg/kg of LYP and 150 mg/kg of IFO. Rats in group A (control) were orally treated with 2 mL of water whereas rats in groups B–D were orally treated with 10, 20, and 40 mg/kg of LYP daily for 7 days, respectively. Rats in group E were treated with 150 mg/kg of IFO on the 7th day intraperitoneally (ip) whereas rats in groups F–H were pretreated orally with 10, 20, and 40 mg/kg of LYP daily, respectively, before treatment with IFO on the 7th day (ip).
On the 8th day, rats were sacrificed under diethyl ether anesthesia, blood was collected, and serum was separated and evaluated for biochemical parameters. Rats were sectioned and liver was collected and weighed and washed in ice cold 1.15% KCl solution. Liver was homogenized with 0.1 M phosphate buffer (pH 7.2), with the aid of a homogenizer. The homogenate was centrifuged at 1500 rpm for 30 min at 4°C. The resulting supernatant was decanted and used for biochemical analysis.
Limitation of the study
In this study, we quantified hepatic histological changes in treated rats using a predetermined grading or morphometry.
Evaluation of parameters
Aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), total bilirubin (TB), conjugated bilirubin (CB), gamma-glutamyl transferase (GGT), and lactate dehydrogenase (LDH) levels were determined using Randox diagnostic kits supplied by Quimica Clinica Aplicada, S. A., Spain. Liver protein was assessed as described by Gornall et al., 1949, whereas superoxide dismutase (SOD) was assayed according to Sun and Zigma, 1978. Glutathione (GSH) was analyzed as reported by Sedlak and Lindsay, 1968, whereas catalase (CAT) was evaluated as described by Aebi, 1984. Malondialdehyde (MDA) was assayed using the method of Buege and Aust, 1978, whereas glutathione peroxidase (GPX) was evaluated according to Rotruck et al., 1973.
Statistical analysis was performed with the aid of Prism 5 (GraphPad Software Inc., San Diego, California, USA). Results are represented as mean ± standard error of the mean. One-way analysis of variance was used to compare group data, followed by Tukey's multiple comparisons test. Significance was set at P < 0.05; 0.01; 0.001.
| Results|| |
Treatment with 10, 20, and 40 mg/kg of LYP did not produce significant (P > 0.05) effects on relative liver weight, and serum AST, ALT, ALP, GGT, LDH, TB, and CB levels when compared to control. In addition, effects on liver AST, ALT, ALP, GGT, and LDH levels were not significant (P > 0.05) in LYP-treated rat when compared to control. Normal (P > 0.05) liver levels of SOD, CAT, ASH, GPX, and MDA were obtained in LYP-treated rats in comparison to control. Liver weights were normal (P > 0.05) in IFO-treated rats when compared to control [Table 1], [Table 2], [Table 3], [Table 4]. In contrast, significant (P < 0.001) elevations in serum AST, ALT, ALP, GGT, LDH, CB, and TB were obtained in IFO-treated rats when compared to control. These elevations represent 266.4%, 285.0%, 296.5%, 574.7%, 319.8%, 345.5, and 360.9%, respectively [Table 2]. However, the aforementioned parameters were significantly and in a dose-dependent manner restored at 10 mg/kg (P < 0.05), 20 mg/kg (P < 0.01), and 40 mg/kg (P < 0.001) of LYP-pretreated rats when compared to IFO-treated rats [Table 2]. Furthermore, liver AST, ALT, ALP, GGT, and LDH levels were increased significantly (P < 0.001) in IFO-treated rats. The increases obtained represent 290.0%, 311.0%, 256.1%, 389.3%, and 224.5%, respectively [Table 3]. In contrast, the aforementioned parameters were significantly and in a dose-dependent manner decreased at 10 mg/kg (P < 0.05), 20 mg/kg (P < 0.01), and 40 mg/kg (P < 0.001) of LYP-pretreated rats when compared to IFO-treated rats [Table 3]. Altered liver redox status was marked by significant (P < 0.001) decreases in liver SOD, CAT, GSH, and GPX with significant (P < 0.001) increases in MDA levels in IFO-treated rats when compared to control [Table 4]. However, the aforementioned parameters were significantly restored in a dose-dependent manner at 10 mg/kg (P < 0.05), 20 mg/kg (P < 0.01), and 40 mg/kg (P < 0.001) of LYP-pretreated rats when compared to IFO-treated rats [Table 4]. Furthermore, normal liver histology was observed in the control rats [Figure 1]a. In contrast, the liver of IFO-treated rats showed periportal and pericentral necroses of hepatocytes [Figure 1]b. On the other hand, normal liver histology was observed in rats pretreated with 10, 20, and 40 mg/kg of LYP, respectively [Figure 1]c, [Figure 1]d, [Figure 1]e.
|Table 1: Effects of lycopene and ifosfamide on body and liver weights of albino rats|
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|Table 2: Effects of lycopene and ifosfamide on serum liver function biomarkers of albino rats|
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|Table 3: Effects of lycopene and ifosfamide on liver tissue biochemical parameters of albino rats|
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|Table 4: Effects of lycopene and ifosfamide on liver oxidative stress biomarkers of albino rats|
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|Figure 1: (a) Control liver showing normal hepatocytes. (b) Liver of rat treated with 150 mg/kg ifosfamide on the 7th day showing periportal and pericentral necroses of hepatocytes. (c-e) Liver of rats pretreated with 10, 20, and 40 mg/kg of lycopene daily for 7 days before treatment with ifosfamide on the 7th day showing normal hepatocytes in the pericentral zones (H and E, ×200)|
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| Discussion|| |
The involvement of hepatocytes in different metabolic functions including metabolism and excretion of xenobiotics makes the liver more vulnerable and susceptible to toxic insult. There is a paucity of effective drugs that can protect the liver or help regenerate liver cells during damage. A number of mechanisms have been proposed for liver injury including hepatic oxidative, and nitrosative stress. LYP is the most prominent carotenoid; studies suggested that it can be useful against oxidative stress-induced damage. The present study aimed at establishing a possible benefit of LYP against hepatic injury induced by IFO in albino rats. The assessment of relative liver weight, liver function, and oxidative stress markers in LYP-treated rats showed the absence of perturbations. However, in IFO-treated rats, serum and liver activities of AST, ALT, ALP, LDH, GGT, CB, and TB were increased. This observation has been earlier reported. The increases in the activities of the aforementioned parameters showed impaired functional integrity of the liver. In addition, liver damage can impair bilirubin excretion or obstruct the excretory ducts of the liver stimulating accumulation. The observed increases in TB and CB fractions and other liver function markers are symptoms of hepatic dysfunction, cholestasis, and liver injury.
However, liver function was restored in a dose-dependent manner in LYP-pretreated rats as shown by normal levels of AST, ALT, ALP, LDH, GGT, CB, and TB. This observation showed that LYP promoted parenchymal cell regeneration in the liver, protected membrane fragility, and restored liver integrity. Endogenous antioxidants such as SOD, CAT, GSH, and GPX are inhibitors of oxidative processes, even when present in small concentrations. Antioxidants have vital and diverse physiological functions in humans and animals. They are primarily the first line of defense against oxidative stress associated damage and hence are essential for sustaining maximum health and well-being. Antioxidants activities tend to decrease with increased oxidative stress activity. The current study showed that oxidative stress is a vital process in IFO-induced liver injury as supported by evident decreases in liver SOD, CAT, GSH, and GPX levels. This observation is in consonance with reported decreases in liver antioxidants in IFO-treated rats. Interestingly, antioxidant status was restored in a dose-dependent fashion in the liver of LYP-pretreated rats. MDA is the primary reactive aldehyde produced during the peroxidation of polyunsaturated fatty acids in biological membrane. Therefore, its liver level is often used as a measure of lipid peroxidation with the advent of liver damage. Liver damage observed in IFO-treated rats was characterized by lipid peroxidation as supported by elevated MDA activity. However, lipid peroxidation decreased in a dose-dependent manner in the liver of LYP-pretreated rats due to low levels of MDA. Insult to the liver cytoarchitecture can be one of the adverse consequences of the clinical use of IFO. The present study observed histological alterations in the liver of IFO-treated rats marked by periportal and pericentral necroses of hepatocytes. However, hepatic necrotic changes observed in IFO-treated rats were ameliorated in LYP-pretreated rats.
Furthermore, many suggestions have been put forward on the mechanism of IFO-induced liver injury. These suggestions include the hepatic biotransformation of IFO to CAA, a toxic metabolite leading to increased free radical formation which targets mitochondria and lysosomes causing oxidative stress in the liver. Oxidative stress can easily damage biological molecules and ultimately induce cell death either by necrosis or apoptosis., Furthermore, oxidative stress can alter the functions and structures of DNA, lipids, proteins, and carbohydrates. Also, it can inactivate transporters and enzymes and propagates chain of reactions that can oxidize polyunsaturated fatty acids. The peroxidation of membrane lipids can alter membrane fluidity which can stimulate cell lysis and death. The observed hepatoprotective effect of LYP could be attributed to its antioxidant activity which includes scavenging of free radicals which inhibits lipid peroxidation and prevents protein, lipid, and DNA damage in cells. In addition, it can stimulate cellular antioxidant defense by activating the antioxidant response element transcription system.
| Conclusion|| |
The current study showed that pretreatment with LYP attenuates IFO-induced liver injury in albino rat in a dose-dependent manner.
We appreciate the contributions of Dr. Ebinyo Clemente Nelson and Mr. Harold Adagbabina of the Faculty of Pharmacy, Niger Delta University, Nigeria.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Table 1], [Table 2], [Table 3], [Table 4]