Involvement of prostaglandins in paraquat intoxication

Prostaghdins 0 Longman

Leukotrienes and Essential Group UK Ltd 1990

Fatty Acids (1990) 39.213415

Involvement of Prostaglandins in Paraquat Intoxication Muslim Ali, Muhammed

Afzal and Noha Al-Sweedan

Department of Biochemistry, Faculty of Science, Kuwait University, P. 0. Box 5969, Safat Kuwait (Reprint requests to MA)

13060,

ABSTRACT. Paraquat is known to cause severe lung damage through pulmonary edema as its initial feature of toxicity. The purpose of this study was to investigate the toxicity of paraquat in rabbits intraperitoneally injected with 2 or 4 m&g/day of the herbicide for a period of 7 days. In the lung, prostagIandin levels of the intoxicated rabbits showed a significant increase in PGF 2WThis increase was dose dependent. However, a nonsignificant change in the 6-keto-PGF1, was also observed. Plasma and serum levels of thromboxanelz were also significantly elevated but the levels of 6-keto-PGF,, were affected nonsignificantly. The pathology of elevated levels of PGFza and TXBz in the lung and blood, in response to paraquat toxicity, is discussed.

INTRODUCTION

arachidonic acid or PGEz, cause elevation in both wet weight of lung and pulmonary artery. In conscious sheep an increase in plasma and lung lymph of thromboxane B2 (a stable metabolite of TXA2) and 6-keto-PGF1, (a stable metabolite of prostacyclin) as a result of PQ toxicity has been reported by Shibamota and Kobayashi (25). The vulnerability of the lung to paraquat toxicity is believed to be due to its higher concentration in lung than in other tissues (13-15). However, its mechanism of action in different species is yet to be established. For instance, rabbits are reported to be resistant while rats, guinea pigs and humans are highly sensitive to PQ toxicity (16). The object of the present investigations was to determine the effect of PQ on prostaglandin levels of rabbits in an endeavour to understand the mechanism of its resistance to this herbicide.

Paraquat (PQ) toxicity is believed to be due its redox cycle through superoxide anions and hydroxy radicals (1, 2). These radicals are not only membrane destructive but also are known to start the arachidonic acid cascade and stimulate prostaglandin synthesis. There is strong evidence that endogenous vasoactive agents from the limg act directly to mediate and/or exacerbate paraquat damage in the early phase of alveolar-capillary injury. Alveolar-capillary injury is known to be the prime target of this toxicity (3, 4) through involvement of leukocytes (5), platelets (6) and arachidonic acid metabolites (7-9). Barabas et al (5) have reported that PQ has a specific PGF2, synthesis enhancing effect which results in an increased fluid outflow to the intracellular space, leading to pulmonary edema, haemorrhage and then at later stages fibrosis (10). Loss of endothelial integrity is perhaps the primary event leading to the development of pulmonary edema. Lindenschmidt et al (11) have reported an increase in PGF2, and a 300% increase in histamine in the isolated perfused rat lung damaged by paraquat. Imamura et al (12) have demonstrated that perfusion of guinea pig lung with paraquat or PGF za, and repeated bolus injection of

MATERIALS AND METHODS Rabbits weighing 1.5-2.0 kg were maintained of fresh salad with free access to water, in a constant temperature (22-24” C) room. The animals were divided into three lots containing five animals in each group. One of this group of animals was used as controls. The animals were deprived of their diet two hours before sacrifice by anesthetizing with sodium pentothal injection. Paraquat, as gramoxane, was administered daily

Date received 15 May 1989 Date accepted 2 October 1989 213

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Prostaglandins Leukotrienes and Essential Fatty Acids

to two groups of rabbits by intraperitoneal injection of 2 or 4 mg/kg/day, for seven days. A third group of rabbits served as. controls and received normal saline intraperitoneally. After the rabbits were anesthetized, they were exsanguinated via the carotid artery. The blood was collected in tubes containing 3.8% sodium citrate. Platelet poor plasma was prepared by centrifugation of whole blood at 4” C for 15 min. at 100 Xg in a Sorval centrifuge. Blood was also collected from the carotid artery for determination of thrombin stimulated platelet TXB2 synthesis during clotting for 60 min. at 37” C (17). The lungs from experimental animals as well as controls were quickly removed and chilled on ice immediately. Thl tissue was cut into approximately one gram slices. Each slice was suspended in 0.05 M Tris-HCl buffer, pH 8.0. Prostaglandin synthesis was allowed tb proceed in the absence of added arachidonic acid for 30 min. at 37” C. All specimens were stored at -40” C until radioimmunoassay (RIA) estimation of prostanoids was carried out on these samples. Extraction, chromatography and radioimmunoassay of PGFZ,, thromboxane-Bz and 6-keto-PGF1, were carried out in the serum/plasma and lung homogenates according to the method previously described (18). The prostaglandins were measured by using radioimmunoassay kits supplied by Amersham, England. Statistical analyses were perform‘ed on the data by the two tailed student t-test.

Table 1. Synthesis of PGF;?, and&-keto-PGF,, in vitro in lung homogenates of rabbits receiving intraperitoneal injection of paraquat rip/p of

n 5 5

Treatment Control 2mg/kgPQ

5

4mg/kg PQ

rabbit lung

PGF2, 34.30+5.35 74.00rt12.2 P
6-keto-PGF,, 77.4Ok14.41 94.4Ok27.36 NS 96.50+18.60 NS

n = Number of animals in each group. NS = Not significant.

Table 2 Plasma levels of TXB, and 6-keto-PGF,, following intraperitoneal

injection of paraquat into rabbits ng/ml plasma

n Treatment 5 Control (saline) 5 2m&gPQ 5 4 mg/PQ

TXB, 0.442kO.087 1.50+0.339 P
6-keto-PGF,, 0.708fO. 129 0.904+0.107 NS 1.212kO.317 NS

n = Number of animals NS = Not significant

Table 3

Synthesis of TXB, and 6-keto-PGF,cu during blood clottingv from the animals receiving PQ intraperitoneally ng/ml serum

n Treatment 5 Control (Saline) 5 2m&gPQ 5

4m&gPQ

TXB, 163.20+50.5 1196.0+384.0 P
6-keto-PGF,, 2.32kO.281 2.40+0.316 NS 2.2420.133 NS

RESULTS Table 1 shows the levels of the PGF2, and 6-ketoPGFI, in the lungs of rabbits treated with 2 or 4 mg/kg of paraquat. There was a significant increase in PGF,, in the lung. This increase was dose dependent as shown in Table 1. However the levels of 6-keto-PGFZ, were not significantly affected. Tables 2 and 3 show the levels of TXBz and 6keto-PGfi!, in the blood of PQ treated rabbits. In plasma as well as in serum, a significant dose dependent increase in the levels of TXB;! was observed. The levels of 6-keto-PGF2, remained non significantly affected in this case also.

DISCUSSION The prime target of paraquat toxicity in the lung, where superoxide radicals initiate an arachidonic acid cascade and stimulate prostaglandin synthesis. The production of prostaglandins by the lung may be an important factor in the pathogenesis of a variety of pulmonary disease (19). Our results show a dose dependent increased level of PGF;?, in the lung. A dose dependent increase in TXBZ in blood

n = Number of animals NS = Not significant 0 = Blood was clotted at 37°C for one hour.

is also observed in our studies. The involvement of PGFZa, in an increased fluid outflow to the intracellular space, leading to pulmonary edema has been reported by Lindenschmidt et al (11, 20). An increase in plasma and lung PGFz~ in response in PQ in rats has been repotted by Giri et al (21). Paraquat mediated lung edema has also been reported by a number of other workers (10). The present study shows increased levels of PGF2, and TXB;! in PQ treated rabbits and substantiates our earlier results (22, 23) establishing the fact that rabbits, as reported by other workers, are not completely re-sistant to PQ toxicity. Contrary to the situation in rabbits, however, Shrin and Krishna (24) have reported a significantly reduced ability to synthesize PGF2, and PGE2 in paraquat treated guinea pig lungs. These workers attributed this decrease in PG to a decrease in the amount of lung microsomal protein. Our results also show a nonsignificant increase in the6-keto-PGF1. in both blood and the lungs of the

Involvement of Prostaglandins

PQ treated rabbits. However, a significant increase in plasma and lymph prostacyclin (6-keto-PGFtJ has been observed in conscious sheep treated with paraquat, indicating pulmonary endothelial damage (25). A significant increase in thromboxane A2 (as TXB2) in the blood of PQ treated rabbits may be due its release in part from the lung parenchyma or pulmonary interstitial cells, as suggested earlier (25). A modest increase in TXB2 in conscious sheep treated with PQ has been reported by Shibamoto and Kobayshi (25). In conclusion our results show that rabbits are not totally resistant to PQ toxicity; as erroneously reported earlier (16).

11.

12.

13.

14.

15. 16.

Acknowledgement This work was supported by a research grant No. SBO 21, by the Research Management Unit, University of Kuwait, for which the authors are grateful.

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