Catabolism of vasoactive polypeptides by perfused rat liver.

Exsanguinated rat liver preparations perfused in situ with oxygenated saline solutions inactivated recirculating bradykinin (BK) at rates of 2.3 to 9.1 and isoleucyl5 angiotensin II (AII) at rates of 2.8 to 15.0 nmoles X min-1 X g-1 of liver, depending on the initial concentration of the peptides in the perfusion fluid (3.1 to 18.9 X 10(-6) M for BK and 8.5 to 17.0 X 10(-6) M for AII). On the other hand, at similar concentrations, recirculation of isoleucyl5 Angiotensin I (AI) for 8 min did not lead to decrease of its biological activity when assayed on the isolated rat uterus. Following a single passage through liver, picomole amounts of both BK and AII were inactivated by about 90% as revealed by assays on a superfused rat uterus. The potency ratio AI:AII, assayed on a superfused rat uterus was 1:22 and changed to 1:5 following a single passage of both peptides through liver. This finding and the separation of 4.9% of AII on carboxymethylcellulose columns following recirculation of AI through rat liver indicate a conversion of AI into AII. The dipeptides Phe-Arg, Ser-Pro and Gly-Phe were identified among the hydrolysis products of perfused BK. A peptidyldipeptide hydrolase (EC 3.4.15) may be responsible for both the BK inactivation and AI conversion. The inactivation of AII cannot be attributed to the same enzyme.

Recovery and conversion of kinins in exsanguinated rat preparations.

A rat preparation in which the perfusion route bypassed the lungs, which were substituted by an artificial oxygenator, was exsanguinated and perfused with oxygenated 4% dextran (M.W. 70,000) in Tyrode's fluid; a peristaltic pump propelled the perfusing fluid at 20-25 ml/min through the aortic arch and the perfusion medium returned to the right ventricle. Known amounts of bradykinin (BK), kallidin (lysul-bradykinin, LBK) and methionyllysylbradykinin (MLBK) were administered as single injections and samples of the perfusion fluid removed between 1.5 to 6 min following injection. Average total kinin activity remaining in the circulating fluid was calculated from assays on the isolated guinea pig ileum against respective kinin injected and found to be after 3 and 6 min respectively: 20 and 5% for BK, 54 and 21% for LBK, 60 and 30% for MLBK. When the lungs were introduced in the perfusion circuit BK recoveries decreased to 0.4% at 4 min and 0% at 6 min. In 2 experiments 1 mg MLBK and, in another two, 1 mg of LBK were recirculated for 3 to 3.5 min in the rat preparation with lung bypass; enzymic reactions were interrupted in the perfusates after removal by lowering the pH to 4.7 and placing them in a boiling water bath for 5 min. Following proper dilution, kinin activity left in the perfusates was separated on carboxymethylcellulose columns; in 3 experiments about 50% of the activity was identified as BK from its elution position and resistance to trypsin digestion. The average BK-inactivating potency of the perfusate obtained from the rat with lung bypass was 0.3 mug BK/min x ml compared to 16 mug BK/min x ml of rat plasma. The arylamidase activity on arginylnaphthylamide of the perfusate was 2 n moles NA/min x ml and it was about 25-fold lower than that of rat plasma. Rat liver was exsanguinated and perfused in situ through the portal vein and inferior cava vein using the same conditions as for the whole animal. The perfusion rate was 12 ml/min. The recovery of injected BK in this preparation was 40% after 2 min of recirculation, declining progressively in the following minutes. When MLBK was perfused in this preparation for 3 min or glycylarginyllysylbradykinin (GALBK) for 3 and 5 min, significant amounts of BK were found in the perfusates. We conclude that LBK, MLBK and GALBK may be converted at a high rate into BK by tissue aminopeptidases found in the rat preparations used. BK inactivation in the whole rat is a fast reaction, even when the pulmonary tissue is not involved in the inactivation.

Effect of sex steroids on the circulating levels of alpha 2-macroglobulin in injured rats.

The effect of sex steroids on the plasma levels of alpha 2-macroglobulin (alpha 2-M) was investigated to explain the sexual dimorphism observed in response to the injury caused by ip administration of 5 micrograms endotoxin. Mean serum alpha 2-M concentrations were lower in female (46.82 +/- 8.10 arbitrary units) than in male injured rats (82.94 +/- 9.22 arbitrary units). A reduction of plasma alpha 2-M levels was observed after orchidectomy, and the administration of 1 mg testosterone to the same animals increased the response. The same response pattern was observed in ovariectomized rats under the same treatment. The response of androgenized rats (78.93 +/- 12.81 arbitrary units) to injury was similar to the male response. These results suggest the importance of testosterone as the major determinant of this sex-dependent response.

Effect of turpentine on alpha 1-major acute phase concentration in normal and adrenalectomized rats.

Acute phase response of plasma alpha 1-major acute phase (alpha 1-MAP) was studied in normal and adrenalectomized rats. alpha 1-MAP basal levels were higher in female than in male rats. This protein, measured by radial immunodiffusion, increased significantly both in male and female rats 24 and 48 h after a turpentine stimulus, proving to be a positive acute phase protein for both sexes. In adrenalectomized male rats the increase of plasma alpha 1-MAP concentration was not different from that observed in sham-operated rats, suggesting that the acute phase response of this protein is not corticoid dependent.

Inflammatory response modulated by pinealectomy: effect of light.

In view of the known interactions between pineal and adrenal glands and the importance of glucocorticoids on the acute phase of inflammatory process, we investigated the effect of the pineal gland on the paw edema response induced by carrageenin in female Wistar rats, weighing 170-220 g, measured by the pletysmographic method. Adrenalectomy increased the rate of the inflammatory response (149.02 +/- 2.93%, N = 7 vs 113.48 +/- 2.99%, N = 7) whereas pinealectomy and sham-pinealectomy under white light had an inhibitory effect (81.73 +/- 2.16%, N = 15 and 80.21 +/- 1.85%, N = 14 vs 113.48 +/- 2.99%, N = 7). Sham-pinealectomy under red light did not modify the rate of the inflammatory response. These results indicate that the pineal gland can be affected by light during sham-pinealectomy. In view of the unaltered adrenal gland weight and corticosterone levels among the different groups, we propose an antagonism between melatonin (pineal hormone) and glucocorticoids at the peripheral level. The predominant effect of glucocorticoids over melatonin may explain the inhibition of the inflammatory response caused by pinealectomy.

Native plasma kallikrein is cleared at similar rates by mammalian and avian livers.

Rat plasma kallikrein (RPK) is a serine protease that circulates as an inactive precursor, prokallikrein, and once activated is efficiently cleared by the liver by a carbohydrate-dependent, Ca(2+)-independent mechanism. Seven hepatic lectin systems have been described for mammals but not all of these animal lectins are expressed in the avian liver. Using a liver perfusion system we compared the plasma kallikrein clearance of rats (N = 10) and pigeons (N = 4). Our results show that the lectin responsible for the hepatic clearance of plasma kallikrein is also present in pigeon liver and that this organ clears the enzyme with an efficiency (11.4 +/- 1.3 pmol/g, 20 min) similar to that of the rat liver (10.0 +/- 0.7 pmol/g, 20 min).

Inflammatory edema induced by carrageenin in monosodium glutamate-treated rats.

Rats treated with monosodium glutamate (MSG) during the neonatal period show hypothalamic lesions and multiple neuroendocrine alterations manifested as a remarkable increase in levels of circulating corticosterone and obesity. Paw edema induced by local injection of carrageenin was significantly reduced in MSG-treated rats compared to normal rats. In contrast, both adrenalectomized rats and adrenalectomized, MSG-treated rats showed an increased response to carrageenin relative to controls. These results suggest that glucocorticoids are important modulators of inflammation in this phase of the process.

Effect of endogenous pyrogen, corticosteroids and inhibitors of prostaglandins and leukotrienes on the plasma concentrations of haptoglobin and fibrinogen in rats.

Injections of endogenous pyrogen (EP) in normal rats significantly increased plasma concentration of haptoglobin and fibrinogen while in adrenalectomized animals the same treatment was ineffective. Nevertheless when cortisone was injected simultaneously with EP into adrenalectomized rats the responses of fibrinogen and haptoglobin were restored. These results suggest that biosynthesis of fibrinogen and haptoglobin is corticosteroid-dependent. Inhibition of the cyclo-oxygenase pathway with ibuprofen or the lipoxygenase pathway with BW755C had no effect on the increase of plasma fibrinogen or haptoglobin levels stimulated by either EP or endotoxin. These data indicate that neither prostaglandins nor leukotrienes are involved in the production of these acute phase reactants induced by either stimulus.