View of EFFECT OF SUB-ACUTE ORAL DOSE OF CRUDE AFLATOXINS ON WEIGHT OF LIVER AND KIDNEY OF GUINEA PIG

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Vol. 05, Issue 07,July 2020 Available Online: www.ajeee.co.in/index.php/AJEEE

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EFFECT OF SUB-ACUTE ORAL DOSE OF CRUDE AFLATOXINS ON WEIGHT OF LIVER AND KIDNEY OF GUINEA PIG

Shahda Parween¹, C.S. Varma² and Nagma Sadaf³

1,2Depatment of Botany, Veer Kunwar Singh University, Ara – 802301, India

3Department of Physiology, Nezamia Unani Medical College & Hospital, Gaya, B.R.

Ambedkar Bihar University, Muzaffarpur, India

Abstract- Objective was to evaluate the effects of ingestion of sub-acute oral dose of crude aflatoxins on liver and kidney weight of guinea pigs. Animals received about. Animals receiving sub-acute oral dose (100 ± 10 ug of crude a flatoxins daily) of crude aflatoxins exhibited dullness, hair loss, feed refusal, poor growth rate and reduced reproductive performance. A significant increase in liver and kidney weight was recorded among animals of toxin-treated group when compared to that of control group. Macroscopic pathological changes like, the appearance of pinkish lesions, enlargement in size and pale colouration kidney were observed frequently among animals of toxin-treated group. The mean weight of liver in toxin treated group was recorded to be 21.43 ± 0.08 gms. As compared to 17.75 ± 0.13 gms. in control group. Liver as percent of body was 2.88 and 4.45 in control and toxin treated group. The weight of kidney as percent of body weight also showed significant enhancement. It was 0.7 % in case of control and 1.16 % in case of toxin treated group.

Test group animals receiving amla and lemon showed improvement in respect of liver and kidney weight.

Keywords: Aflatoxin, liver, kidney, lesion, growth, hair loss.

1 INTRODUCTION

Aflatoxins are a family of extremely toxic, mutagenic, and carcinogenic compounds produced primarily by some strains of A.

flavus and by most, if not all, strains of A.

parasiticus, plus related species, A.

nomius (Peterson, et. al., 2001). In 1988, the IARC placed aflatoxin B1 on the list of human carcinogens.. Animal species respond differently in their susceptibility to the chronic and acute toxicity of aflatoxins. The toxicity can be influenced by environmental factors, exposure level, and duration of exposure, age, health, and nutritional status of diet. Aflatoxin B1 is a very potent carcinogen with liver as the primary target organ (Wan et. al.

2013). Tessari, et. al., (2006) reported an increase in the relative weight of the liver in birds fed diets containing aflatoxin. The clinical manifestation of aflatoxin in animal system involves four general areas:

1. Reduced rate of growth or productivity,

2. Liver damage,

3. Immuno-suppression and 4. Carcinogenicity.

Exposure is through gastrointestinal tract which acts as the major site for the absorption of aflatoxins. Once ingested, aflatoxins undergo metabolism i.e., biotransformation, and it is the metabolic fate of aflatoxins on which depends their

toxicological profile. Metabolism is usually catalysed by enzymes, and these are usually found most abundantly in the liver in animals. This may be the reason why most foreign compounds that enter the body via the gastrointestinal tract are directed towards the liver by the portal blood supply. The liver also receives blood draining the gastrointestinal tract, and is exposed to poisons and toxins absorbed from the gut. It is because of the central role that liver play in absorbing and metabolizing many toxic chemicals that render it liable to injury by such substances. The liver and thymus are the primary target organ of aflatoxin (Safameher et. al. 2008; Elianca et. al.

2010). Biological effects of toxins together with the response of the host to inter- current infection determine the features of the disease. Aflatoxins and their metabolites are excreted by the kidneys and have the potentiality to damage the urinary system during filtration, secretion, concentration or excretion.

Although the kidneys account for only about 0.4% of body weight, they received about onequarter of the cardiac output, resulting in heavy exposure of the kidney to potentially nephrotoxic substances.

A significant increase in the relative weight liver and kidney in animal fed with aflatoxins was reported by Smith

&Hamilton (1970) and Varma (1989). In

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21 such cases enlargement of liver and kidney and their pale colouration was recorded. In the present investigation guinea pigs were exposed to the sub-acute oral dose of crude aflatoxins in order to observe effects on their liver and kidney weight.

2 MATERIALS AND METHODS

The experimental design was totally randomised, in a 4×4 factorial arrangement with 4 treatments and 6 guinea pigs per treatment (Table 1).

Animals were given feed (assayed to be free of toxin) and water ad libitum throughout the 20 - week experimental period.

Table 1 Treatment Protocols

Oral Dose = 100 ± 10 µg/kg/day with water ad libitum Amla fruit juice 5ml/animal /day

Lemon fruit juice 5 ml/animal/day 1st group o f animals served as control and were fed with uncontaminated meal.

The second group served as treated control and were fed with same basal meal supplemented with crude a flatoxin.

For this the baker's bread was infested with the toxigenic strain (AR 105) of A.

parasiticus and then incubated at 28 ± 1°C for 10 days. The bread was therea fter dried and powdered. The concentration o f aflatoxin was determined by spectrophotometer. A known amount of this powder was thereafter fed as basal diet so that each animal among treated group was receiving about 100 ± 10 ug of crude aflatoxins daily. All the treatments were carried out simultaneously for 20 weeks. For studying the effect on body weight, a weekly mean weight for each group o f animals was recorded for 20 weeks.

3 RESULTS

A significant increase in liver and kidney weight was recorded among animals of toxin-treated group when compared to that of control group. Macroscopic pathological changes like, the appearance of pinkish lesions, enlargement in size and pale colouration were observed frequently among animals of toxin-treated

group when compared with the control group. The mean weight of liver in toxin treated group was recorded to be 21.43 ± 0.08 gms. as compared to 17.75 ± 0.31 gms. in control group (Table 2). Liver as percent of body was 2.88 and 4.45 in control and toxin treated group. The weight of kidney as percent of body weight also showed significant enhancement. It was 0.7 % in case of control and 1.16 % in case of toxin treated group (Table 2).

Test grop animals receiving amla and lemon along with aflatoxins showed improvement in respect of liver and kidney weight. Pathological signs on liver and kidney were less severe in case of test group animals.

Ingestion of aflatoxin results in a variety of clinical signs. The significant macroscopic pathological changes in liver was the appearance of non-inlammatory pinkish lesions on the dorsal surface (Fig.

1), with frequency more numerous and severity more intense among animals of treated group The macroscopic pathological signs in kidney involve enlargement in size, pale colouration (Fig.

3) and occasional appearance of bluish lesion on the surface. Fat deposition around kidney was frequent among treated group animals. Control group

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22 animals performed better in these respects

Fig. 1 Pinkish lesion on the dorsal surface of the liver of animals of aflatoxi treated group after 2 0 weeks.

Fig.2. A pair of normal kidney of the control group

Fig. 3. A pair of enlarged pale coloured kidneys in aflatoxin treated group after

2 0 - week of treatment

Fig. 4. Finger like projections from the anterior end of the gall bladder of animals of aflatoxin treated group after

20 weeks.

4 DISCUSSION

Major biochemical effects of aflatoxins involve the modification of normal metabolic and other vital processes. More severe symptoms such as diarrhea,

decreased feed consumption, decreased body weight gain, increased relative weights of liver and kidney, and liver necrosis are observed with longer lapse of time. A decrease in humoral immunity and suppression of lymphocytes were also apparent. Henry et. al. 2000). Increase in the relative weight of liver and kidney suggests cytotoxic effect of aflatoxins due to direct exposure of these organs.

Metabolism plays a major role in determining the toxicity of aflatoxin.

Studies show that aflatoxin requires metabolic activation to exert its carcinogenic effect and these effects can be modified by induction or inhibition of the mixed function oxidase system. The mode of their action appears to be based primarily on their ability to interact with macromolecules, sub-cellular organelles and organs. Many of these mycotoxin—

induced effects may be derived from and secondary to their disruption of nucleic acid or protein synthesis. Aflatoxin B1 (3 mg/kg body weight) inhibits RNA polymerase II activity only in the target tissue, liver, and not in the non-target tissues Bedard et. al., 2005).

Adverse aflatoxin reactions in liver is a reflection of its central role in the metabolism and excretion of this toxin.

Enzymes present in the liver are principally involved in the biotransformation of aflatoxins i.e., the process by which toxins are transformed into more soluble metabolites or made more suitable for conjugation with endogenous substrates such as glucuronic acid prior to excretion in the bile or in the urine. Biotransformation, thus, in general, seems directed at increasing water solubility and hence excretion. However, in case of aflatoxins the reverse occurs and a metabolite is produced which is more toxic than the parent compound. However, there are several pathways of metabolism which compete. Consequently factors which alter the balance between these pathways will alter the eventual toxicity induced by aflatoxins. Cells ability to adopt to toxin- insult may range from complete recovery to death. Between these two extremes there lie a range of cellular reactions where the cell attempts to adopt to the insult. These reactions may include hypertrophy, hyperplasia, metaplasia and the accumulation within cell of a variety of material and these may have a bearing

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23 on the organ (liver and kidney) weight.

Kidney being a principal excretary organ provides for the longer stay of the toxins in the organ enhancing, thereby, the chances of pathological damage or injury which have an effect on its weight.

Animals receiving plant extracts showed lesser degree of variation in their organ (liver and kidney) weight signifying, thereby, their protective effect in a limited way. Tessari, et. al., (2006) reported an increase in the relative weight of the liver in birds fed diets containing aflatoxin B1.

Increase in the relative weight of liver and kidney can also be due to underlying histological changes in these vital organs.

Main alterations included vacuolar degeneration and cell proliferation of bile ducts in the liver, and hydropic degeneration in renal tubules in the kidneys (Tessari, et. al, 2006).

The relative weight of the liver and kidney is increased only in guinea pigs fed diets containing aflatoxin. Other general signs of aflatoxicosis are edema of the lower extremities, abdominal pain, and vomiting. The symptoms include decrease in growth rate and immuno suppression..

Liver damage is apparent due to the yellow color that is characteristic of jaundice, and pinkish lesion on the dorsal surface of liver. The gall bladder becomes swollen with many finger like projections at the anterior end (Fig.4). Immuno- suppression is due to the reactivity of aflatoxins with T-cells, decrease in Vitamin K activities, and a decrease in phagocytic activity in macrophages. These immuno suppressive effects of aflatoxins predispose the animals to many secondary infections. Decrease in body weight gains can be attributed to poor feed utilization and a dramatic increase in alkaline phosphate activity in the gut.

Specific, disease symptoms associated with chronic exposure to aflatoxins are liver damage, feed refusal, alteration in digestion, absorption and/or metabolism of nutrients, dullness, slower rates of growth and hair loss. Aflatoxin B1 is confirmed as a potential carcinogen (IARC 1993; Manal et. al. 2012; Zeng et. al.

2013). Metabolism plays a major role in deciding the degree of toxicity (Eaton et al 1994). After ingestion, aflatoxin is metabolized by cytochrome p450 group of enzymes in the liver, where it is converted to many metabolic products like aflatoxicol, aflatoxin Q1, aflatoxin P1, and

aflatoxin M1, depending on the genetic predisposition of the species. Along with the above another metabolite called aflatoxin 8,9 epoxide is also formed. The amount of this metabolite decides the species susceptibility as this can induce mutations by intercalating in to DNA, by forming an adduct with guanine moiety in the DNA (Smela. et al 2001). This intercalation of epoxide causes a transversion at codon 249 in p53 gene in liver, which may lead to hepatic carcinoma. This was observed in most of the experimental models, and it is presumed that this is the major reason for aflatoxin carcinogenecity (Katherine et al 1997, Railey et al 1997). Moreover species susceptibility to aflatoxin mainly depends on its liver detoxification systems, genetic makeup, age and other nutritional factors (Howard et al 1990). The principal target organ for aflatoxins is the liver. After the invasion of aflatoxins into the liver, lipids infiltrate hepatocytes and leads to necrosis or liver cell death. This is mainly because aflatoxin metabolites react negatively with different cell proteins, which leads to inhibition of carbohydrate and lipid metabolism and protein synthesis. In correlation with the decrease in liver function, there is a derangement of the blood clotting mechanism, icterus (jaundice), and a decrease in essential serum proteins synthesized by the liver.

The most severe case of acute poisoning of aflatoxin was reported in north-west India in 1974 where 25% of the exposed population died after ingestion of the moulded maize with aflatoxin levels ranging from 6250 to 15600 µg/kg.

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