Protective Effects of Ginger Toward Cadmium-Induced Testes and Kidney Lipid Peroxidation and Hematological Impairment in Albino Rats

Abstract

This study was carried out to investigate the effect of oral dietary supplementation with ginger on cadmium-induced toxic effects on biochemical, hematological, and pathophysiological indices of albino rats. The effect of cadmium and cadmium/ginger treatment on lipid peroxidation was measured by malondialdehyde (MDA) levels in testes and kidney; serum activities of alkaline phosphatase (ALP), acid phosphatase (ACP), and prostatic acid phosphatase (PAP) enzyme were investigated alongside hematological indices. The results showed that cadmium induces a significant increase in both testicular and kidney MDA, whereas cadmium/ginger treatment produced a significant reversal of the effect of lipid peroxidation (P=.004). Cadmium treatment induced 75%, 78%, and 22% increases in activities of ACP, PAP, and ALP, respectively, whereas the cadmium/ginger-treated group reversed these values for enzyme activities (P=.001). Results of organ weight and hematological indices analysis in the cadmium-treated rats showed a decrease in organ weight and distortion of the hemopoietic features, whereas the cadmium/ginger-treated rats showed an improvement in organ weight and hematological indices (P=.04 and .001, respectively). The reversal of the toxic effects of cadmium in the cadmium/ginger-treated albino rats heralds the antioxidant potency of ginger toward cadmium toxicity-associated oxidative stress.

Introduction

C admium (Cd) is an extremely toxic metal commonly found in industrial workplaces, particularly where any ore is being processed or smelted. It is a highly toxic heavy metal and is distributed widely in the general environment. Clinical manifestations of chronic Cd intoxication include renal proximal tubular dysfunction, osteomalacia, and anemia. Accumulating evidence suggests that Cd toxicity may also affect various organs such as the liver, lung, testis, and the hematopoietic system.¹ In mammals, it exerts multiple toxic effects and has been classified as a human carcinogen by the International Agency for Research on Cancer. It is a well-known human carcinogen and a potent nephrotoxin. Lipid peroxidation is involved in Cd-related toxicity.² This toxic metal is a well-known nephrotoxicant, inducing kidney damage via oxidative stress coupled with gonadotoxic and spermiotoxic effects.³ Possible protective roles of onion (Allium cepa Linn.) and garlic (Allium sativum Linn.) extracts on Cd-induced testicular damage and spermiotoxicity have been suggested.⁴ Combined treatment with selenium and zinc has been shown to offer some beneficial effects over that provided by either of them alone in reversing Cd-induced oxidative stress in kidney of the rat.⁵

Zingiber officinale Roscoe, commonly called ginger, is a perennial plant with narrow, bright green, grass-like leaves and yellowish green flowers with purple markings. Ginger is cultivated in the tropics for its edible rhizome at approximately 10 months of age, with the root stocks serving a variety of purposes, including culinary and medicinal.⁶ Ginger is among the 20 top-selling herbal supplements in the United States, and today pharmacopeias of several different countries list ginger extract for various digestive diseases.⁷ The main constituents of ginger include volatile oil (β-bisabolene, cineol, phellandrene, citral, borned, citronellol, geranial, linalool, limonene, zingeberol, zingeberene, camphene, oleoresin, gingerol, and shogoal), phenol (gingerol and zingerone), proteolytic enzymes (zingibain), vitamin B₆, vitamin C, calcium, magnesium, phosphorus, potassium, and linoleic acid.⁸

The aim of this study was to investigate the potential protective effect of ginger against the Cd-induced oxidative effect in albino rats of the Wistar strain.

Materials and Methods

Animals

Thirty albino rats of the Wistar strain weighing 140–220 g were purchased from the disease-free stock of the Department of Biochemistry Animal House, University of Port Harcourt, Port Harcourt, Nigeria, and transferred to the Department of Biochemistry, University of Calabar, Calabar, Nigeria, where the study took place. They were acclimatized for 2 weeks on a commercial rat diet prior to experimentation. Permission and approval for the animal studies were obtained from the College of Medical Science Animal Ethics Committee of the University of Calabar. The rats were assigned to three study groups made up of 10 rats each on the basis of their weight: the control group was given placebo, another group was treated with Cd, and the third group was treated with Cd/ginger. The Cd/ginger-treated animals received adequate basal diet fortified with Cd (as CdCl₂) and ginger supplement at the rate of 3 mg and 0.5 g/kg of body weight, respectively. Feeding was done for 28 days. CdCl₂ was administered orally to Cd only-treated rats mixed with basal diet at the rate of 3 mg of Cd/kg of body weight/day. The rats were housed in plastic cages with wire screen tops. They were kept under adequate ventilation with room temperature and relative humidity of 29±2°C and 40–70%, respectively, with a 12-hour natural light–dark cycle. Food and water were provided ad libitum, and good hygiene was maintained by constant cleaning and removal of feces with spilled feed from cages daily.

Blood was collected by cardiac puncture, and serum was separated and further analyzed using standard colorimetric and enzyme immunoassay methods for alkaline phosphatase (ALP), acid phosphatase (ACP), and prostatic acid phosphatase (PAP) using Randox kits (Randox Clinical Diagnostic Solutions, Crumlin, United Kingdom). Malondialdehyde (MDA) levels as an index of lipid peroxidation was measured by the thiobarbituric acid assay.⁹ Hematological indices of hemoglobin (Hb), packed cell volume (PCV), white blood cell (WBC) count, and red blood cell (RBC) counts were analyzed according to methods described by Dacie and Lewis.¹⁰ Blood smears (thin film) were prepared by the push wedge technique and stained using Leishman's stain for study of RBC morphology and differential leukocyte count. Kidney and testes were surgically removed, weighed, placed in an iceberg, and later trimmed to 5 g weight and further used to ascertain the extent of lipid peroxidation.

Statistical analysis

Statistical analysis was carried out using the computer database Statistical Package for Social Sciences version 10 (SPSS Inc., Chicago, IL, USA). Data were expressed as mean±SD values. Values of P<.05 were regarded as significant for all statistical comparison. Descriptive analysis of percentages of continuous variables was reported. Comparisons were assessed using mean values and the χ² test.

Results

The MDA level was used as an index of lipid peroxidation in testes and kidney homogenates of control and experimental animals treated with Cd and ginger supplements as shown in Table 1. The mean±SD MDA levels (in nmol/g of tissue) obtained for the testes in various groups were 2.14±0.03 for control, 3.42±0.02 for Cd-treated, and 1.45±0.07 for Cd/ginger-treated rats; in the kidney tissue homogenates, the MDA levels of control and experimental groups were 2.10±0.07 for control, 2.76±0.03 for Cd-treated, and 2.53±0.06 for Cd/ginger-treated rats (P=.004). Cd treatment increased the MDA level of the testes by 60% relative to the controls that received placebo. The results further indicated a significant decrease (P=.004) in MDA levels in the Cd/ginger-treated rats as shown in Table 1. There was a 31% increase in kidney MDA levels for the groups that had Cd treatment alone. This was reversed significantly in the Cd/ginger-treated group.

Table 1.

Malondialdehyde Levels in Testes and Kidney of Control, Cadmium-Treated, and Cadmium/Ginger-Treated Albino Rats

	MDA (nmol/g of tissue)
Treatment group	Testes	Kidney	P value
Control	2.14±0.03	2.10±0.07	.004^*
Cd-treated	3.42±0.02	2.76±0.03	.04^†
Cd/ginger-treated	1.45±0.07	2.53±0.03

Data are mean±SD values.

The difference in the malondialdehyde (MDA) levels of cadmium chloride (Cd)-treated and Cd/ginger-treated rats was statistically significant: *P value for Cd versus Cd/ginger treatment; ^† P value for control versus Cd/ginger treatment.

The mean±SD values of ACP, PAP, and ALP (in U/L) for the control, Cd-treated, and Cd/ginger-treated groups were 6.05±0.26, 10.60±0.07, and 5.95±0.21, 5.10±0.04, 9.10±0.06, and 5.25±0.03, and 50.80±0.57, 62.00±0.14, and 51.3.0±0.14, respectively (P=.001). The ACP, PAP, and ALP levels of control, Cd-treated, and Cd/ginger-treated albino rats are shown in Table 2. Cd toxicity significantly increased ACP, PAP, and ALP activities in serum, whereas ginger supplementation reversed these values significantly.

Table 2.

Total Acid Phosphatase, Prostatic Acid Phosphatase, and Alkaline Phosphatase Activities in Control, Cadmium-Treated, and Cadmium/Ginger-Treated Albino Rats

Treatment group	ACP (U/L)	PAP (U/L)	ALP (U/L)	P value
Control	6.05±0.26	5.10±0.04	50.80±0.57	.001^*
Cd-treated	10.60±0.07	9.10±0.06	62.0±0.14	.05^†
Cd/ginger-treated	5.95±0.21	5.25±0.03	51.30±0.14

Data are mean±SD values.

The difference in the acid phosphatase (ACP), alkaline phosphatase (ALP), and prostatic acid phosphatase (PAP) activities of cadmium chloride (Cd)-treated and Cd/ginger-treated rats was statistically significant: *P value for Cd versus Cd/ginger treatment; ^† P value for control versus Cd/ginger treatment.

The values of organ weight (in g) for the control, Cd-treated, and Cd/ginger-treated groups were 2.12±0.14, 1.90±0.16, and 2.87±0.17, respectively, for testes. The Cd-treated rats showed a significant decrease in testes and kidney weights. In comparison, the Cd/ginger-treated group showed an increase in organ weight (P=.04). Table 3 shows the effect of Cd exposure and ginger supplementation on testes and kidney weights of albino rats.

Table 3.

Effect of Cadmium Exposure and Ginger Supplementation on Testes and Kidney Weights of Albino Rats

	Weight (g)
Treatment group	Testes	Kidney	P value
Control	2.12±0.14	1.75±0.31	.04^*
Cd-treated	1.90±0.16	1.26±0.18	.05^†
Cd/ginger-treated	2.87±0.17	1.33±0.51

Data are mean±SD values.

The difference in the testes and kidney weights of cadmium chloride (Cd)-treated and Cd/ginger-treated rats was statistically significant: *P value for Cd versus Cd/ginger treatment; ^† P value for control versus Cd/ginger treatment.

The hematological parameters assessed were Hb, PCV, WBC count, and RBC count. The mean±SD hematological values are given in Table 4. The mean values for Hb (in g/dL), PCV (in %), WBC count (in×10⁹/L), and RBC count (in×10⁹/L) in the control, Cd-treated, and Cd/ginger-treated groups were 10.68±0.41, 35±0.22, 5.8±0.41, and 7.28±0.21, 6.75±0.38, 22±0.60, 17.1±0.52, and 2.97±0.29, and 9.50±0.30, 31±0.31, 7.3±0.50, and 6.31±0.21, respectively (P=.001).

Table 4.

Effect of Cadmium Treatment and Ginger Supplementation on Hematological Parameters of Albino Rats

Treatment group	Hb (g/dL)	PCV (%)	WBCs (×10⁹/L)	RBCs (×10¹²/L)	P value
Control	10.68±0.41	35±0.22	5.8±0.41	7.28±0.21	.001^*
Cd-treated	6.75±0.38	22±0.60	17.1±0.52	2.97±0.29	.05^†
Cd/ginger-treated	9.50±0.30	31±0.31	7.3±0.50	6.31±0.21

Data are mean±SD values.

The difference in the hematological values of cadmium chloride (Cd)-treated and Cd/ginger-treated rats was statistically significant: *P value for Cd versus Cd/ginger treatment; ^† P value for control versus Cd/ginger treatment.

Hb, hemoglobin; PCV, packed cell volume; RBC, red blood cell; WBC, white blood cell.

Ginger supplementation cushioned the effect of Cd by reversing its action significantly. The total WBC count in the Cd-treated group showed a significant increase compared with the control group. The RBC profile showed marked hypochromia and microcytosis with burr, target, and crenated cells. The WBCs showed marked neutropenia and basophilia. Platelet distribution appeared significantly reduced in the blood film of Cd-treated rats (data not shown).

Discussion

This study showed that Cd treatment enhanced the extent of lipid peroxidation in the testes of rats by as much as 60% compared with that of control rats. This agrees with previous reports^2,11
–13 showing that Cd is a potent inducer of cell oxidative stress. Cd treatment with ginger supplementation produced a significant reduction in testicular MDA in comparison with Cd treatment alone. Similarly, Cd treatment alone enhanced lipid peroxidation in kidney tissues of rats by as much as 31%. Pretreatment with ginger combined with Cd treatment significantly reduced the MDA levels. These finding is consistent with results obtained in previous studies,^12,14 which reported that ginger seem to have effective antioxidant and anticancer properties against free radical scavengers. The toxic effect of Cd is indicated by significant decreases in levels of enzymatic (superoxide dismutase, catalase, glutathione peroxidase, and glutathione S-transferase) and nonenzymatic (reduced glutathione, vitamin C, and vitamin E) antioxidants.¹⁵ Cd-induced toxicity has been shown to be alleviated by other antioxidants, such as l-ascorbic acid,¹⁶ selenium,¹⁷ the natural antioxidant garlic,^18,19 and naringenin,¹³ a naturally occurring citrus flavonone. Similarly, a previous study⁴ showed that aqueous extracts of onion and garlic could offer a measure of protection against Cd-induced testicular oxidative damage and spermiotoxicity by possibly reducing lipid peroxidation and increasing the antioxidant defense mechanism in rats.

The effect of ginger on serum ACP, PAP, and ALP activities of rats given a subcutaneous dose of CdCl₂ revealed that Cd treatment caused a significant increase in serum ACP, PAP, and ALP activity in comparison with control values. Cd-treated rats showed a 75% increase in ACP, 78% in PAP, and 22% in ALP compared with the values for untreated rats. The high level and the increased activity of these clinically significant enzymes (in testes and kidney) are a result of leakage of these enzymes into the bloodstream because the integrity of the testes and kidney was substantially compromised as a result of Cd toxicity. This finding is consistent with the report by Renugadevi and Prabu,¹³ which found that liver damage induced by Cd was clearly indicated by the increased activities of serum hepatic marker enzymes (aspartate transaminase, alanine transaminase, ALP, lactate dehydrogenase, and γ-glutamyltransferase) and serum total bilirubin along with the increased level of lipid peroxidation indices (thiobarbituric acid–reactive substances, lipid hydroperoxides, and protein carbonyl contents) in the liver. Similarly, Messaodu et al.²⁰ studied the effect of Cd-induced oxidative stress on rat kidney and observed that Cd treatment decreased significantly the catalase and glutathione peroxidase activities, whereas the superoxide dismutase activity and the renal levels of lipid peroxidation (as MDA) were increased compared with values in control rats. The kidney is the critical target organ for Cd toxicity. Cd is a well-known nephrotoxicant inducing kidney damage via oxidative stress. The levels of renal lipid peroxidation and glutathione S-transferase were significantly increased in rats that received Cd alone relative to the control group.²⁰ Also, the levels of renal reduced glutathione, superoxide dismutase, catalase, and Na⁺/K⁺-ATPase were significantly decreased in rats that received Cd alone.²⁰ A reversal of the Cd-induced increase in activities of these enzymes was observed in the ginger-supplemented group. The supplementation with ginger was effective in effecting a protective role against Cd-induced increases in ACP, PAP, and ALP activities in rats. This finding of the protective antioxidant property of ginger is consistent with the previous report by Egwurugwu et al.,¹⁴ which found that ginger had a potency to alleviate Cd poisoning in rats. Ginger seems to have effective antioxidant properties against free radical scavengers.

We observed that Cd treatment in rats produced a significant reduction of testes weight and that supplementation with ginger produced a restorative and significant increase in testicular weight. Similarly, the decreased kidney organ weight observed in Cd-treated rats was effectively reversed following supplementation with ginger. This observation is in agreement with a previous report²⁰ showing that Cd exerts gonadotoxic and spermiotoxic effects producing testicular damage and spermiotoxicity. Cd intoxication significantly decreased epididymal sperm concentration and sperm progress motility and increased percentage total sperm abnormalities and live/dead cell counts.

We observed that Cd treatment resulted in a derangement of hematological indices of albino rats and that supplementation with ginger produced a restorative effect on the hemopoeitic parameters. The previous report by Asagba and Eriyamremu²¹ showed that Cd treatment significantly reduced Hb, hematocrit, and RBC counts of rats. Similarly, Mohammad²² observed in a previous study that CdCl₂ treatment in albino rats caused decreases in RBC count, hematocrit, mean cell Hb, mean cell volume, and mean cell Hb concentration and that oral administration of CdCl₂ with camel milk supplementation resulted in significant increases in RBC count, Hb, hematocrit, mean cell Hb, mean cell volume, and mean cell Hb concentration.

We conclude that organ-specific Cd-induced testicular and kidney toxicities associated with increased lipid peroxidation, raised serum activities of ALP, PAP, and ACP, and organ weight loss in Wistar rats were all reversed by ginger supplementation. It may be worthwhile by way of a randomized multicenter study to determine the protective effects of ginger in humans environmentally exposed to Cd to see if the protective role of ginger observed in this animal toward Cd-induced toxicity can be replicated in humans.

Footnotes

Author Disclosure Statement

No competing financial interests exist.

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