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Biomedical subjects

A G Scicli

Publications and source records attributed to A G Scicli.

At least 91 records · Page 5Linked to original sources
PubMed1985

Characterization of a kininogenase from rat vascular tissue resembling tissue kallikrein.

A kininogenase resembling glandular kallikrein was partially purified from vascular tissue and characterized. Saline-perfused rat tail arteries and veins were homogenized in 0.25 M sucrose containing 10 mM Tris-HCl (pH 7.4). The homogenate was centrifuged at 105,000 g for 60 minutes, and a vascular kininogenase was purified from the supernatant by chromatofocusing, affinity chromatography on immobilized antibodies against rat urinary kallikrein, and gel filtration on Sephadex G-100. The inhibitory effects of antibodies against rat urinary kallikrein were tested with equivalent kinin-forming concentrations of rat urinary kallikrein and vascular kininogenase. Kininogenase activities of both enzymes were similarly inhibited by both polyclonal and monoclonal antibodies. Aprotinin (1,000 KIU) completely inhibited vascular kininogenase activity, while soybean trypsin inhibitor (100 micrograms) did not modify its kinin-forming activity. Vascular kininogenase and rat urinary kallikrein had the same elution volume when chromatographed on a Sephadex G-100 column, and had similar mobilities in 10% polyacrylamide gel electrophoresis. Kinins released by vascular kininogenase were identified as bradykinin by reverse-phase high performance liquid chromatography. Rat vascular kininogenase appears to be similar to glandular kallikrein. Kinins released locally by vascular kininogenase may contribute to the regulation of vascular tone.

Animals↗
PubMed1984

Lack of evidence for the participation of tonin in the pathogenesis of one-kidney, one-clip renovascular hypertension.

It has been reported that immunization against tonin normalizes blood pressure, and that sialoadenectomy, during which the tonin-rich salivary glands are removed, decreases blood pressure in one-kidney, one-clip hypertension. To investigate the role of tonin on this form of hypertension further, we actively immunized one-kidney, one-clip hypertensive rabbits with tonin and measured both the blood pressure response and the titer of antibodies raised against tonin. In addition, because sialoadenectomy may alter food intake, we assessed the effect of sialoadenectomy on the blood pressure of one-kidney, one-clip hypertensive rats fed a liquid diet to facilitate eating. After immunization, all rabbits developed antitonin-antibody titers ranging from 1:300 to 1:56,000. However, in none of the rabbits did the blood pressure decrease significantly (114 +/- 3 mm Hg before immunization; 129 +/- 6 mm Hg at 16 weeks after immunization). In one-kidney, one-clip hypertensive rats, sialoadenectomy did not lower blood pressure (179 +/- 5 mm Hg before sialoadenectomy; 202 +/- 9 mm Hg 3 weeks after sialoadenectomy). Neither blood pressure nor body weight differed between sialoadenectomy and sham-sialoadenectomy one-kidney, one-clip hypertensive rats (n = 6). In conclusion, neither active immunization against tonin in one-kidney, one-clip hypertensive rabbits nor sialoadenectomy in one-kidney, one-clip hypertensive rats significantly reduced established hypertension. These results do not support the hypothesis that tonin is involved in the pathogenesis of one-kidney, one-clip hypertension in these animal models.

Animals↗
PubMed1984

Kallikrein-kinins in the central nervous system.

We studied whether the components of the kallikrein-kinin system are present in the central nervous system. We found that human cerebrospinal fluid (CSF) contains free kinins: 53 +/- 15 pg/ml; kininogen: 10.9 +/- 2.1 ng kinin equivalent/ml, and kininogenase activity: 5.0 +/- 2.1 ng kinins/ml/minute. Kininogenase activity was 2-3 fold augmented by preincubation with trypsin. Soybean trypsin inhibitor completely inhibited untreated CSF and partially inhibited trypsin activated kininogenase. Kininogenase activity and immunoreactive glandular kallikrein were present in rat brain, and their concentrations in hypothalamus is several-fold higher than in cortex, pons-medulla, basal ganglia and cerebellum. In the hypophysis, activity in pars-intermedia was between 6- and 20-fold higher than in posterior and anterior hypophysis, respectively. High activity was also found in the pineal gland. The kallikrein-kinin system is present in the central nervous system where it may participate in modulation of nervous and neuroendocrine functions.

Animals↗
PubMed1983

Active and inactive kallikrein in rabbit connecting tubules and urine during low and normal sodium intake.

Active and total (trypsin-activated) kallikrein were measured in urine and the discrete segments of the nephron of rabbits fed low and normal sodium diets. Kallikrein was measured by its kininogenase activity, and kinins generated were measured by radioimmunoassay. The amount of inactive kallikrein was calculated as the difference between total and active kallikrein. The nephrons were microdissected and divided into eight segments: (1) glomerulus; (2) proximal convoluted tubule; (3) cortical thick ascending limb; (4) bright portion of distal convoluted tubule; (5) granular portion of distal convoluted tubule; (6) granular portion of cortical collecting tubule; (7) light portion of cortical collecting tubule; and (8) medullary collecting tubule. As we have previously described, active and inactive kallikrein were localized mainly in the granular portion of the distal convoluted and cortical collecting tubules (connecting tubule). Both active and inactive kallikrein in the granular portion of the distal convoluted and cortical collecting tubule increased markedly during low sodium intake without altering the distribution profile. Urinary excretion of active and inactive kallikrein also increased significantly in the low sodium diet. More than 50% of total kallikrein was found in the inactive form in the nephron and urine. The ratio of active to total kallikrein in the nephron and urine was not changed by sodium restriction. These results suggest that sodium restriction stimulates the biosynthesis and excretion of both active and inactive kallikrein.

Animals↗
PubMed1983

Intestinal absorption of glandular kallikrein in the rat.

We studied the possibility of intestinal absorption of glandular kallikrein in unanesthetized rats after administration of 5 mg of active or phenylmethylsulfonyl fluoride-inactivated pig pancreatic kallikrein (PPK). Immunoreactive PPK was measured in plasma and urine by radioimmunoassay using an antiserum to PPK that does not cross-react with rat glandular kallikrein. In addition, we studied the effect of intestinally administered active PPK on the intestinal blood flow distribution. Although immunoreactive PPK could not be detected (less than 10 ng/ml) in plasma after active PPK was administered, very small amounts were found in urine (20-40 ng/3 h). In the urine most of the PPK was in active form because over 75% could be bound to aprotinin-Sepharose. After inactive PPK was administered, immunoreactive PPK was detectable in the plasma of all rats, and urinary excretion was higher than after administration of active PPK. Blood flow distribution to the small intestine as a percentage of cardiac output was significantly greater 30 min after intestinal application of active PPK (5 mg) than after administration of the vehicle alone (1 ml of 0.9% saline). These results suggest that active and inactive PPK is absorbed from the gut in very small amounts. The finding of higher amounts of immunoreactive PPK in plasma and urine after inactive PPK was administered is probably due to the fact that the inactive form of the enzyme is not bound by the plasma inhibitors. These small amounts of kallikrein absorbed appear to have some effect on intestinal blood flow distribution.

Animals↗
PubMed1983

Effect of pH and amiloride on the intrarenal formation of kinins.

Changes in urinary kallikrein excretion are assumed to reflect changes in intrarenal formation of kinins. Yet factors that alter the enzymatic activity of renal kallikrein and kininases may alter the concentration of kinins in the nephron independent of amount of kallikrein excreted. In anesthetized rats, we measured excretion of urinary kallikrein (kininogenase activity) and kinin excretion during altered urinary pH and after amiloride, which reportedly inhibits urinary kallikrein. In rats fed a low sodium diet, urine was acidified by intravenous 0.28 M sodium sulfate. This decreased urinary pH from 6.1 +/- 0.09 to 5.3 +/- 0.17 and urinary kinin excretion from 28.0 +/- 9.0 to 10.5 +/- 5.0 pg/min. Urinary kallikrein excretion doubled from 43.0 +/- 5.0 to 82.5 +/- 13.5 ng/min. The optimum pH of kallikrein is congruent to 8.5, so the decreased excretion of urinary kinins is probably secondary to decreased kininogenase activity at lower urinary pH. Amiloride decreased urinary kinins from 35.5 +/- 7.3 to 18.2 +/- 2.5 pg/min and kallikrein from 18.7 +/- 4.9 to 9.3 +/- 1.8 ng/min, while urinary pH increased from 6.7 +/- 0.07 to 7.3 +/- 0.07. The depressed excretion of kallikrein and kinins with amiloride may not have been due to inhibition of kallikrein, since amiloride (1 mM) did not inhibit the kininogenase activity of rat urinary kallikrein (congruent to 1.2 nM) on dog or rat kininogen in vitro. We conclude that changes in urinary kallikrein may not reflect changes in intrarenal formation of kinins. These data also indicate that kallikrein excretion increases and kinin formation decreases when urine is acidified in the distal nephron and that there may be a link between the kallikrein-kinin system and the renal mechanisms affected by amiloride.

Amiloride↗
PubMed1983

Role of the autonomic nervous system in the release of rat submandibular gland kallikrein into the circulation.

We have previously demonstrated that the rat submandibular gland releases immunoreactive kallikrein into the circulation. To study the role of the autonomic nervous system in this release, submandibular gland blood flow and kallikrein concentration in peripheral arterial and venous blood from the gland were measured and secretion rates calculated before and after parasympathetic and sympathetic nerve stimulation (8V, 2 msec, 10 Hz) for 1 minute. Immunoreactive kallikrein in plasma was measured by radioimmunoassay, and timed collections of venous outflow were used to measure blood flow. During basal conditions, the unstimulated submandibular gland of the rat released immunoreactive kallikrein into blood at the rate of 0.92 +/- 0.07 ng/min. Parasympathetic stimulation increased blood flow 4-fold (before, 68.5 +/- 8.3 microliters/min; after, 253.5 +/- 76.2; P less than 0.05) without significantly changing immunoreactive kallikrein secretion rate. Sympathetic stimulation produced an 11-fold increase in blood flow (before, 64.9 +/- 9.3 microliters/min; after, 709.6 +/- 97.5; P less than 0.05) and a 57-fold increase in immunoreactive kallikrein secretion rate from the gland (before, 1.05 +/- 0.25 ng/min; after, 59.8 +/- 18.6; P less than 0.05). Sympathetic stimulation also produced a 4-fold increase in the concentration of immunoreactive glandular kallikrein in arterial plasma (before, 15.2 +/- 1.1 ng/ml; after, 56.2 +/- 12.9; P less than 0.05). Pretreatment with phentolamine (1 mg/kg) or prazosin (0.2 mg/kg) blocked the increase in kallikrein secretion rate produced by sympathetic stimulation. These results indicate that the sympathetic nervous system, through activation of alpha 1-adrenoreceptors, controls kallikrein secretion from the submandibular gland into the circulation. Released kallikrein may be responsible for the reactive vasodilation observed in the rat submandibular gland after sympathetic stimulation.

Adrenergic alpha-Antagonists↗
PubMed1983

Interference of converting enzyme inhibitors with the kallikrein-kinin system.

The unstimulated rat submandibular gland releases kallikrein into the circulation. This release is greatly increased by sympathetic nervous stimulation of the gland. We studied the effect of an angiotensin I converting enzyme (ACE) inhibitor (captopril) on blood flow of the unstimulated submandibular gland, and the effect of the ACE inhibitor on blood kinins and blood pressure of rats with prior sympathetic stimulation of the gland. To minimize the effects mediated by inhibition of angiotensin II formation, all rats were nephrectomized 48 hours before the studies. Administration of the ACE inhibitor caused a 2.4-fold increase in the blood flow of the unstimulated gland (p less than 0.001). Pretreatment with kinin antibodies diminished this increase by 86% (p less than 0.001). After sympathetic stimulation, blood flow to the gland increased five times, and kinin output in the venous effluent increased from 19 +/- 13 to 14,400 +/- 8,800 pg/min (p less than 0.001). Despite this conspicuous increase, arterial blood kinins did not change. Administration of the ACE inhibitor after stimulation of the gland resulted in a 10-fold increase in arterial blood kinins and in a pronounced decrease in blood pressure from 100 +/- 5 to 58 +/- 6 mmHg. This decrease was almost completely blocked by antibodies to either kallikrein or kinin. We concluded that the glandular kallikrein-kinin system may participate in the regulation of local blood flow in organs rich in glandular kallikrein such as the rat submandibular gland.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1982

Localization of active and inactive kallikrein (kininogenase activity) in the microdissected rabbit nephron.

Active and total (trypsin-activated) kallikrein were measured in discrete segments of the nephron by their kininogenase activity. The kinins generated were measured by radioimmunoassay and the amount of inactive kallikrein was calculated as the difference between total and active kallikrein. Single nephrons from collagenase-treated rabbit kidneys (N = 12) were microdissected and divided into eight segments: (1) glomerulus; (2) proximal convoluted tubule; (3) cortical thick ascending limb; (4) bright portion of distal convoluted tubule; (5) granular portion of distal convoluted tubule; (6) granular portion of cortical collecting tubule; (7) light portion of cortical collecting tubule; and (8) medullary collecting tubules. After pooled segments (approximately 50 nephron segments in each assay) were homogenized and treated with deoxycholic acid, active and total kallikrein were measured and inactive kallikrein was calculated. Active and inactive kallikrein were localized in granular portions of the distal and cortical collecting tubule, which contains more than 85% of the active and inactive kallikrein found in the total microdissected nephron. Little or no kallikrein was detected in other segments, including the bright portion of the distal segment which has the macula densa. The discrete localization of active and inactive kallikrein suggests a specific role for renal kallikrein in these nephron segments.

Animals↗
PubMed1982

Immunoreactive glandular kallikrein in rat plasma: a radioimmunoassay for its determination.

A radioimmunoassay (RIA) has been developed to measure immunoreactive glandular kallikrein in rat plasma. To prevent the binding of radioactive kallikrein to plasma inhibitors, 125I-kallikrein was inactivated with phenylmethylsulfonyl fluoride (PMSF), a procedure that maintained 125I-kallikrein immunoreactivity. Different volumes of plasma displaced 125I-PMSF-kallikrein in a parallel fashion to the kallikrein standard curve. The sensitivity of the RIA was 200 pg, and the recovery of nonradioactive active kallikrein added to plasma was 58.7%. The concentration of immunoreactive glandular kallikrein in normal rat plasma averaged 47.1 +/- 1.7 (SE) ng/ml. Bilateral nephrectomy caused a threefold increase in circulating glandular kallikrein (50 +/- 2.7 to 167 +/- 7 ng/ml; P less than 0.001). REmoval of the submandibular and sublingual glands significantly decreased its concentration from 52 +/- 2.3 to 34 +/- 1.6 ng/ml (P less than 0.001). Immunoreactive glandular kallikrein was higher in the submandibular gland vein than in arterial blood (venous: 94 +/- 10.5; arterial: 64 +/- 6.3 ng/ml; P less than 0.05) and was lower in the renal venous blood (venous: 44 +/- 2.2; arterial: 53 +/- 2.6 ng/ml; P less than 0.05). In conclusion, this study shows that the use of 125I-PMSF-kallikrein as tracer prevents the interference in the RIA caused by plasma protease inhibitors. It also indicates that the submandibular gland is an important source of the immunoreactive glandular kallikrein in rat plasma and that the kidney probably participates in its metabolism. Glandular kallikrein released by the submandibular gland into the circulation may participate in regulating local blood flow before it is inactivated by plasma inhibitors.

Animals↗
PubMed1982

Blood kinins, their concentration in normal subjects and in patients with congenital deficiency in plasma prekallikrein and kininogen.

Kinins are potent vasodilator peptides that may participate in the regulation of local blood flow and blood pressure. Here we report a new method to measure kinins in blood. For this, 6 ml of blood are collected in less than 10 sec directly into 25 ml of ethanol. Kinins are further purified by extracting lipids with ether and by removing kininogen and other interfering substances by chromatography on QAE Sephadex and BioRex 70; then they are measured by a sensitive RIA. In 22 normal subjects, after correction for recovery (50%), the kinin concentration in peripheral venous blood was 25.2 +/- 2.6 pg/ml (mean +/- S.E.M.). To determine whether the circulating kinins are formed by plasma kallikrein or other kininogenases, the concentration of blood kinins was measured in the venous blood of three patients with congenital deficiency in plasma prekallikrein (Fletcher trait) and in one patient with congenital deficiency in the substrate of plasma kallikrein, high-molecular-weight kininogen (Fitzgerald trait). In the three subjects with Fletcher trait, blood kinins were 16, 21, and 31 pg/ml, whereas in the subject with Fitzgerald trait they were 26 pg/ml. Normal subjects had concentrations in the same range (9 to 55 pg/ml), indicating that the concentration of blood kinins in normal subjects is much lower than previously reported (70 to 5000 pg/ml). These results also suggest that kininogenases other than plasma kallikrein may generate circulating kinins.

Animals↗
PubMed1979

Relation between structure and correcting activity of bovine high molecular weight kininogen upon the clotting time of Fitzgerald-trait plasma.

Bovine high molecular weight kininogen (bHMWK) partially corrects the activated plasma thromboplastin time (aPTT) of Fitzgerald trait plasma which is congenitally deficient in HMWK. The relationship between the structure and activity of HMWK was clarified by studying the effects of different fragments of bHMWK on the aPTT of Fitzgerald-trait plasma. The peptides studied were lys-bradykinin-free HMWK, bradykinin-fragment 1-2-free HMWK, heavy chain, fragment 1-2-light chain, and light chain. All fragments were tested in equimolar concentrations. Bradykinin-fragment 1-2-free HMWK, heavy chain, and light chain have little or no correcting activity upon Fitzgerald-trait plasma aPTr. Fragment 1-2 light chain has the same correcting activity as intact bHMWK, while that of lys-bradykinin-free HMWK appears to be higher. Both fragment 1-2 and fragment 2 inhibit the clotting time of normal human plasma. When compared on a molar basis, fragment 2 is a more active inhibitor than fragment 1-2. When the effects of bovine plasma kallikrein upon bHMWK and hHMWK were studied, it was found that it released kinins from both kininogens. However, while the correcting activity of bHMWK was completely destroyed after 60 min of incubation, that of hHMWK was fully retained. These data suggest that: (a) the active part of bHMWK is comprised of the fragment 1-2 light chain portion; (b) fragment 1-2 or fragment 2 is the binding site to negatively charged surfaces, while the light chain interacts with other components of the surface-mediated reactions; and (c) bovine plasma kallikrein releases kinins, but probably does not cause the release of fragment 1-2 from human HMWK.

Animals↗