Richard C. Talamo

α1-Antitrypsin Deficiency: A Variant with No Detectable α1-Antitrypsin

No α1-antitrypsin could be detected in the serum of a 24-year-old man with advanced pulmonary emphysema by agarose electrophoresis, immnuno-electrophoresis, double diffusion in agarose gel, or α1-antitrypsin genetic typing by a combination of starch-gel electrophoresis and crossed antigen-antibody electrophoresis. A circulating α1-antitrypsin inactivator could not be demonstrated. Evidence was obtained in family members of genetic transmission of this new α1-antitrypsin variant.

Response of the kallikrein-kinin and renin-angiotensin systems to saline infusion and upright posture.

The possibility that bradykinin, a potent vasodilator, might be a physiological antagonist of the renin-angiotensin system was investigated. 11 norman subjects, ranging in age from 21 to 33 yr were studied. Seven of the subjects were given a 10 meq sodium, 100 meq potassium, 2500 ml isocaloric diet. After metabolic balance was achieved, they were infused with either 1 liter of 5 per cent glucose over 2 h or 2 liters of 0.9 per cent saline over 4 h. During the infusions, plasma renin activity (PRA), angiotensin II (A II), prekallikrein, bradykinin, and aldosterone levels were frequently determined. Plasma prekallikrein and kallikrein inhibitor did not change during the infusion of either glucose or saline. In subjects receiving saline, plasma bradykinin fell from 3.9 plus or minus 1.5 (SEM) ng/ml at 0 min to 0.93 plus or minus 0.2 at 30 min and 0.95 plus or minus 0.3 at 120 min. These changes paralleled the decrease in PRA over the same period (7.9 plus or minus 1.3 ng/ml/h to 5.6 plus or minus 0.8 at 30 min and 3.5 plus or minus 0.7 at 120 min). Similarly, A II fell from 113 plus or minus 12 pg/ml to 62 plus or minus 10 and 48 plus or minus 5, respectively, at 30 and 120 min. In contrast, the control group infused with glucose showed no change in bradykinin, A II, or PRA. Another four subjects were given a constant 200 meq sodium/100 meq potassium isocaloric diet. After metabolic balance was achieved, they were kept supine and fasting overnight. At 9 a.m. they assumed an upright position and began walking a fixed distance (200 ft) at a normal rate (3-4 ft/s). Plasma prekallikrein and kallikrein inhibitor did not change during the posture study. The plasma bradykinin rose from a base line of 0.54 plus or minus 0.01 (SEM) ng/ml to 0.96 plus or minus 0.13 at 20 min. 0.77 plus or minus 0.18 at 60 min, and 0.96 plus or minus 0.07 at 120 min. These changes parallel the increase in PRA over the same period (1.65 plus or minus 3.3 ng/ml/h to 3.6 plus or minus 0.85 at 20 min, 5.3 plus or minus 0.9 at 60 min, and 5.35 plus or minus 0.55 at 120 min). Likewise, the A II rose from 32.5 plus or minus 1.82 pg/ml to 50.8 plus or minus 3.6 at 20 min, 54.3 plus or minus 3.2 at 60 min, and 61.3 plus or minus 5.9 at 120 min. Thus, in sodium-depleted individuals, saline infusion produces a rapid fall of plasma bradykinin at a rate similar to that observed for a II and PRA. Conversely, in sodium-loaded individuals, assumption of upright posture leads to a parallel rise in A II, TPRA, and bradykinin. These studies indicate that there is a close correlation of bradykinin levels with renin activity and angiotensin II, in both acute sodium loading and assumption of upright posture, suggesting that these two systems may be physiologically interrelated.

Antithrombin-heparin cofactor: An inhibitor of plasma kallikrein

Abstract Human antithrombin-heparin cofactor is a naturally occurring plasma inhibitor of serine proteases generated during activation of the coagulation and fibrinolytic systems. We have demonstrated that purified preparations of this inhibitor also neutralize the esterolytic activity of human kallikrein as well as the ability of the enzyme to release kinins. When an excess of inhibitor is present, the inactivation process follows pseudo-first-order kinetics. Furthermore, the addition of heparin to mixtures of kallikrein and antithrombin-heparin cofactor results in a doubling of the rate and extent of enzyme neutralization. Disc gel analysis of incubation mixtures of kallikrein and excess antithrombin-heparin cofactor, with and without heparin, revealed that the enzyme band had vanished in conjunction with the emergence of two new electrophoretic species. These two new components probably represent stoichiometric complexes of kallikrein and antithrombin-heparin cofactor since a twofold increase in the concentration of enzyme doubled the concentration of these new molecular species. In plasmas which contain adequate levels of other protease inhibitors, antithrombin-heparin cofactor does not appear to be a quantitatively important inactivator of kallikrein. This is suggested by our finding that the addition of heparin at concentrations as high as 50 units/ml did not increase the inhibitory capacity of normal plasma directed against this enzyme. However, plasma from patients with hereditary angioedema had little neutralizing activity directed against kallikrein and revealed a marked increase in this inhibitory capacity when therapeutic concentrations of heparin were added. Our observation suggests that this acidic mucopolysaccharide may prove useful in controlling acute attacks of this disorder.

Immunochemical Studies of Plasma Kallikrein

A monospecific antibody against human plasma kallikrein has been prepared in rabbits with kallikrein further purified to remove gamma globulins. The antisera produced contained antikallikrein and also anti-IgG, in spite of only 8% contamination of kallikrein preparation with IgG. The latter antibody was removed by adsorption of antisera with either Fletcher factor-deficient plasma or with purified IgG. Both kallikrein and prekallikrein (in plasma) cross-react with the antibody with no apparent difference between the precipitation arcs developed during immunoelectrophoresis and no significant difference in reactivity when quantified by radial immunodiffusion. Kallikrein antibody partially inhibits the esterolytic and fully inhibits the proteolytic activity of kallikrein. In addition, the antibody inhibits the activation of prekallikrein, as measured by esterase or kinin release. The magnitude of the inhibition is related to the molecular weight of the activator used. Thus, for the four activators tested, the greatest inhibition is observed with kaolin and factor XII(A), while large activator and the low molecular weight prekallikrein activators are less inhibited. With the kallikrein antibody, the incubation of kallikrein with either plasma or partially purified C1 esterase inactivator results in a new precipitin arc, as detected by immunoelectrophoresis. This finding provides physical evidence for the interaction of the enzyme and inhibitor. No new arc could be demonstrated between kallikrein and alpha(2)-macroglobulin, or alpha(1)-antitrypsin, although the concentration of free kallikrein antigen decreases after interaction with the former inhibitor. By radial immunodiffusion, plasma from healthy individuals contained 103+/-13 mug/ml prekallikrein antigen. Although in mild liver disease, functional and immunologic kallikrein are proportionally depressed, the levels of prekallikrein antigen in plasma samples from patients with severe liver disease remains 40% of normal, while the functional kallikrein activity was about 8%. These observations suggest that the livers of these patients have synthesized a proenzyme that cannot be converted to active kallikrein.

Kallikrein-Bradykinin System in Chronic Alcoholic Liver Disease

Abstract The kallikrein-bradykinin system was studied in patients with chronic alcoholic liver disease. Kallikreinogen levels were depressed in patients with cirrhosis when compared with levels in ...

The α1-antitrypsin in man

The recent interest in the a,-antitrypsin in man began eight years ago in Sweden, where Drs. Laurel1 and Erikssonl first noted that a deficiency of the a,-antitrypsin, the major a,-globulin in serum, could be detected on routine screening of paper electrophoretic strips. These authors noted the association between this deficiency state and chronic pulmonary disease and were first to point out that it is hereditary. In 1965, Erikssor? published a monograph which contained the clinical description of 14 families with hereditary w antitrypsin deficiency and chronic pulmonary disease. Since that time, the deficiency state has been reported in numerous families both in the United States and in several European countries.3-5 al-Antitrypsin deficiency is characteristic of some, but not all, families with an increased incidence of emphysema among siblings.

Kallikrein-Kinin System in Postgastrectomy Dumping Syndrome

Abstract The kallikrein-kinin system was studied in 10 postgastrectomy patients, 5 with the dumping syndrome and 5 patients without symptoms. After drinking 220 ml of 45% glucose, none of the asymp...

Simple Assay for Prekallikrein Activators

A simple assay has been devised for the measurement of the activity of the activators of prekallikrein. This method measures the arginine esterase activity of kallikrein, after release by the action of an activator on prekallikrein. Activator samples are incubated with the substrate (normal plasma) for exactly 60 seconds at 25°C and esterase activity of the kallikrein formed is determined by its ability to hydrolyze L-tosyl arginine methyl ester.The results obtained by this assay closely correlate with the determination of activator activity by the release of bradykinin as determined by radioimmunoassay.

PROTEASE INHIBITORS AND THE DISTAL LUNG

Interest in the role of protease inhibitors in maintaining normal tissue integrity was stimulated by the discovery by Laurel1 and Erickson in 1963 that there was a high association of diffuse chronic obstructive pulmonary disease with a severe inherited deficiency of the alpha,-antitrypsin, the major protease inhibitor in normal human serum. Much information has recently accumulated with regard to the serum protease inhibitors, and it is now generally recognized that there are at least six of them.’ In TABLE I, these inhibitors are listed, along with their in vitro inhibitory capacities. The most abundant serum protease inhibitor is the alpha,-antitrypsin, which is able to inhibit trypsin, chymotrypsin, plasmin, thrombin, collagenase. elastase and enzymes derived from neutrophiles and pulmonary alveolar macrophages. The alpha,-antitrypsin is inherited via a series of codominant alleles, which determine its concentration and physicochemical characteristics. Normal levels are in the range of 2.3 mglml, while individuals who carry two deficiency alleles have levels in the range of 10-15% of normal, and have a high risk of developing diffuse obstructive pulmonary disease at a relatively early age.’ Recently, the work of Ohlsson has demonstrated that there are important interactions between the alpha,-antitrypsin and the alpha,-macroglobulin both in vitro and in vivo; in the dog, trypsin binds mostly to the alpha,-macroglobd in , but also to the extent o€ about 18% to the alpha,-antitrypsin. Trypsin bound to the alpha,-antitrypsin is then rapidly transferred to the alpha,-macroglobulin.3 Alpha-chymotrypsin has a greater affinity for the alpha,-antitrypsin than for the alpha,-macroglobulin, and is transferred rapidly to the alpha,macr~globul in .~ It appears that complexes of either trypsin or alpha-chymotrypsin with alpha,-macroglobulin arc then rapidly removed from the circulation by the reticuloendothelial system. Much less information is available about the other serum protease inhibitors, with respect to the lung, but their further characterization with regard to enzyme specificity and interaction should be interesting in the future. It has been known for several years that a diffuse pulmonary lesion can be induced by the administration of enzymes directly into the respiratory tract. Gross made the initial observations that intratracheal administration of the enzyme papain in the rat led to emphysema-like changes. Goldring later administered the same enzyme by aerosol in the golden hamster and found more convincing emphysema-like changes with disruption of elastic tissue.‘; Lieberman has demonstrated that proteases from purulent sputum can produce emphysema in the hamster.’ Finally, Kimbel and his colleagues recently reported that homogenates of neutrophiles from the dog, man or the rabbit, as well as homogenates of pulmonary alveolar macrophages from the dog can produce emphysema when administered to dogs intratracheally.” The proteases of neutrophiles and pulmonary alveolar macrophages have