Effects of exposure to Diazinon on the lung and small intestine of Guinea pig

Considerable toxicity data of chemicals have been produced in the past few years. The environmental contamination with pesticides is a problem of the regional as well as world-wide importance (EPA, 1999). The toxic chemicals have been found in a variety of environmental samples including water, air and house dust, and their presence has also been noted in the tissues of non-occupationally exposed people, particularly in the adipose tissue, blood and urine (Wagner et al. 1991).

The organophosphate insecticide Diazinon has agricultural, commercial, and household uses, but the household uses predominate. The crops using the most diazinon are almonds, berries, pecans, and nectarines (EPA, 1999). As most organophosphates, diazinon attaches to acetyl cholinesterase and prevent it from destroying acetylcholine causing over stimulation of the nerves (Ware, 2000). Also, diazinon and other organophosphates inhibitor numerous enzymes with molecular structure that are similar to AChE (Reigart and Roberts, 1999).

The prolonged exposure to the most commonly used agricultural pesticides increase the risk of the lung lesions, as well as cancer, in the farmers and commercial pesticide users. However, this increased risk was only significant for the prolonged exposure to Diazinon, dieldrin, metalochlor and pendimethalin (Dinham, 2005). Also, the histological changes in the alimentary canal of experimental animals due to pesticides exposure have been documented (Poet et al., 2003 and Manna et al., 2004 a and b).

Several studies have been made on the effect of the exposure of the experimental animals, including guinea pig, to the pesticides (Purdey, 1994; Reigart and Roberts, 1999; Dede and Chike, 2000 and Dede and Dogara, 2004). However, limited information is available on the subject of detection of the exposure effects to the minor concentrations of Diazinon insecticide on the internal organs of guinea pig, Cavia porcellus.

The aim of this work was to study the histological changes induced by the sub-lethal doses (1/20, 1/10 and 1/5 LD50) of diazinon insecticide on the lung and small intestine of guinea pig, Cavia porcellus. In addition, the changes in the total protein and carbohydrates were investigated in these tissues histochemically.

MATERIALS AND METHODS

Animals

A total of 37 male animals belonging to the common species of guinea pig Cavia porcellus were used in this study. Live and healthy specimens of these animals were collected from the markets at Cairo Province, Egypt and maintained in special cages in the animal house at room temperature for two week for acclimatization before the beginning of the experiment. The average body weight of these animals was 540±45gm. The animals were fed ad libitum daily with green meal (alfalfa) and supplied with water.

Insecticide used

The insecticide used in this study, Diazinon, was obtained from The Central Pesticides Laboratory (CPL), Ministry of Agriculture, Egypt. They were samples of that used in the fields (82% purity). The molecular formula is: C12-H21-N2-03-P-S and the structure formula is: (C2H5O)2-P(=S)-O-C4HN2-(CH3)-CH(CH3)2 (C4HN2 pyrimidine ring).

Determination of LD50

A pilot study was carried out with 25 male guinea pigs (divided into five groups) to determine the minimum dose of Diazinon that caused 100% death of the animals. The animals were observed for five days and a dead animal was removed immediately (an animal was considered dead after the stop of movements, loss of body reflexes and loss of the eye reflexes). The number of dead animals was recorded and the mortality rate was calculated as a percentages. LD50 was later calculated according to the equation of Behren and Karber (1953).

Induction of the insecticide

Acclimatized animals were diviced in four groups, three animals/ group and kept in separated cages. The first group was used as control, while the other three groups (TI, TII and TIII) received 1/20, 1/10 and 1/5 of LD50 of the insecticide, respectively, orally through gastric tube. All the cages had the same conditions mentioned at the period of the acclimatization. The animals in the treated cages as well as in control cage were observed for at least two hours after the insecticide was introduced and then were observed at time intervals. All the treated groups received eleven doses through three weeks; the dosage was day after day. After three weeks, the specimens were taken from each treated and control cages and anesthetized with ether and immediately dissected.

Histological and Histochemical Investigations

For the histological investigations, the parts of lung and small intestine from the control and the treated animals were fixed in neutral formalin. After the fixation period, the tissues were washed in the tap-water, dehydrated through a graded series of ethyl alcohol, cleared in xylene, embedded in the paraffin wax or parablast, sectioned at a thickness 5-7 microns, mounted on the glass slides and stained with haematoxylin and eosin for general morphological studies. The various slides based on the dose ranges were studied and the tissues were compared with the control, 0.9% saline injected guinea pigs.

For the histochemical investigation, polysaccharide materials were illustrated following the application of periodic acid Schiff’s (PAS) technique (Huomason, 1979). For visualization of the total proteins, the mercury bromphenol blue method of Mazia et al. (1953) was applied. The study of total protein and polysaccharide materials was done using computer image analyzing system (Leica Model). The estimation of the optical density of thirty cells in each group was made. The data obtained were statistically analyzed according to Sendecor (1987).

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