Comparison between the acute hemodynamic effects and brain penetration of atenolol and metoprolol.

Atenolol and metoprolol are beta 1-selective adrenergic receptor blockers, devoid of local anesthetic and intrinsic sympathomimetic properties. Their antihypertensive and hypotensive activities are equivalent. They differ with respect to their lipophilic character, as is apparent from their octanol/buffer (pH, 7.4 at 37 degrees C) partition coefficients: metoprolol, 1.084; atenolol, 0.012. We compared the two agents with regard to their acute hemodynamic effects and degree of penetration into the cerebrospinal fluid (CSF) and into the brain. For this purpose [3H]-metoprolol and [14C]-atenolol were injected either intravenously or into the left vertebral artery of chloralose-anesthetized cats. With both routes of administration, metoprolol, the more lipophilic of the two compounds, achieved much higher concentrations in the CSF and in the pontomedullary region than did atenolol. One hour after completion of an intravenous injection, the concentration of metoprolol in the CSF was about 6.5 times higher than that of atenolol; after administration into the vertebral artery, the difference was about ninefold. In spite of these considerable differences, the hypotensive and bradycardic activities of both drugs, administered intravenously or into the left vertebral artery, were virtually the same. These results suggest that the acute hemodynamic effects of metoprolol and atenolol are probably not due to an action within the central nervous system, but rather to a peripheral mechanism.


Sodium intake and plasma angiotensin level as modulators of adrenal and uterine angiotensin II receptors in the rat.

Angiotensin II receptors from rat adrenal cortex and myometrium were studied with the use of tritiated angiotensin under conditions where the sensitivity of the target organs for angiotensin II is modified. Sodium status was found to modulate the number of angiotensin receptors both in adrenal gland and uterus. In both target tissues low Na+ diet increases the number of receptors, while a high Na+ diet results in an increase in uterine receptors without modifying adrenal cortical receptors. However, a more markedly positive sodium balance, such as that observed in deoxycorticosterone acetate (DOCA) hypertension  and in one-kidney Goldblatt hypertension, resulted in a reduction of the adrenocortical angiotensin II binding capacity. The endogenous angiotensin II level may also regulate the number of receptor sites as demonstrated by an increased number of receptors after suppression of circulating angiotensin II. It is proposed that the number of angiotensin II receptors is determined by the combined influences of sodium status and angiotensin II concentration. Some changes in the sensitivity of the target organ can be secondary to variations in the number of angiotensin receptors. However, others cannot be so explained and stem, therefore, from events occurring beyond the hormone-receptor interaction.


Thyroid hormone regulation of alpha-adrenergic receptors: studies in rat myocardium.

The effects of alterations in thyroid state on cardiac alpha-adrenergic receptors were investigated by the binding of [3H]dihydroergocryptine (DHE), a potent alpha-adrenergic antagonist. In seven experiments, cardiac membranes from euthyroid rats bound 47 +/- 9 fmoles DHE/mg protein (mean +/- SE) at saturation and demonstrated a dissociation constant (KD) of 2.5 +/- 0.4 nM. Hyperthyroidism, produced by parenteral injection of triiodothyronine, significantly reduced the binding of DHE at all concentrations studied. Scatchard analysis showed this reduction of binding to be due largely to a decreased affinity (KD = 4.0 +/- 0.8 nM, p less than 0.05), although possibly due to a decreased number of binding sites as well (29 +/- 7 fmoles/mg protein, p less than 0.10). Hypothyroidism, produced either by oral propylthiouracil or by surgical thyroidectomy, did not produce a significant change in either the number of binding sites for DHE (56 +/- 8 fmoles/mg protein, p less than 0.40) or in binding affinity (KD = 3.1 +/- 0.5 nM, p less than 0.40). Thus, in addition to the regulation of cardiac beta-receptors by thyroid hormone that has been described previously, thyroid hormone exerts a regulatory effect on the characteristics of cardiac alpha-receptors as well. These changes provide a possible molecular mechanism for the thyroid hormone-induced alterations in cardiac responsiveness to alpha-adrenergic stimulation that have been reported previously.


Effect of cardiopulmonary bypass with heparin administration on digitoxin pharmacokinetics, serum electrolytes, free fatty acids, and renal function.

In 14 patients investigated before, during, and after extracorporeal circulation, serum digitoxin concentration fell significantly during bypass but returned to preoperative values within 24 hr. Both changes occurred in parallel with changes in hematocrit. Serum magnesium concentration fell markedly just before and during bypass and returned to preoperative values on the third and fourth postoperative days. Urine digitoxin concentration fell on the day of operation and remained low in the postoperative period, with a concomitant reduction in the renal excretion of digitoxin. Creatinine and digitoxin clearances decreased on the day of operation and returned slowly to control values during the postoperative period. In 5 other patients, serum digitoxin protein binding decreased significantly during bypass due to heparinization and was normalized by administration of protamine sulfate. Free fatty acids increased significantly immediately before bypass and returned to normal toward its end. The marked changes in digitoxin serum levels during cardiopulmonary bypass can be explained by hemodilution. Acute deterioration of renal function does not lead to accumulation of digitoxin. Heparin administration causes major changes in free fatty acids and serum digitoxin protein binding without important changes in the free digitoxin concentration. Digitoxin can thus be safely administered to patients undergoing cardiac surgery with extracorporeal circulation.


Antiarrhythmic effects of bupuranolol against canine ventricular arrhythmias induced by halothane-adrenaline or two-stage coronary ligation.

We compared the beta-adrenoceptor blocking actions of bupuranolol and propranolol using the response to isoprenaline of the strength of contraction of the canine blood-perfused ventricular muscle, of the rate and strength of contraction of the isolated guinea pig heart and of a guinea pig tracheal ring preparation. The potency ratios of bupuranolol to propranolol were 3.6, 3.0, 3.4, and 2.4, respectively. Bupuranolol decreased the maximum dV/dt of the canine ventricular action potential and had no sympathomimetic action. Bupuranolol effectively suppressed halothane-adrenaline ventricular arrhythmias in the dog, reflecting its potency as a beta-adrenoceptor antagonist. In contrast, bupuranolol did not suppress ventricular arrhythmias induced by two-stage coronary ligation in the dog. Compared to propranolol and certain other antiarrhythmic drugs, bupuranolol had weaker effects on prolonging the effective refractory period than on the maximum dV/dt of canine ventricular muscle. This suggests that lengthening of the refractory period may be important for suppressing the two-stage coronary ligation arrhythmia and that the mechanism of this arrhythmia might be reentry.


Tissue distribution and elimination of digoxin and methyldigoxin after single and multiple doses in dogs.

Glycoside concentrations in plasma and various tissues of dogs were determined after a single and repeated intravenous administration of tritium-labeled digoxin and beta-methyldigoxin. Twenty-four hours after single dosing, the highest concentrations were measured in the kidney. This was followed by the heart, adrenal gland, liver, pancreas, lung, spleen, diaphragm, and skeletal muscle. The glycoside concentration in the brain was low after a single administration of dogoxin. A higher concentration was found after a single dose of the more lipophilic methyldigoxin. After repeated daily administration, the glycoside concentration in plasma and tissues increased continuously and, except for the brain, reached a plateau level after 6 to 8 days. The accumulation factor for both digoxin and methyldigoxin was 2.6. During daily administration for 10 days, the glycoside cconcentration in the brain increased continuously. The mean accumulation factor in all brain areas amounted to 12.4 for methyldigoxin. The concentration of digoxin and methyldigoxin in the tissues decreased, with mean half-lives of 33 and 28 hr, respectively. Glycoside elimination from the brain was clearly slower. A mean half-life of 73 hr was measured for digoxin and of 154 hr for methyldigoxin. The elimination of digoxin was largely renal, while that of methyldigoxin was largely fecal.


Time courses of carbachol-induced responses in guinea pig atria under the influence of ouabain, calcium, and rate of stimulation.

The response of isolated left atria of guinea pigs to carbachol was measured in the presence and during washout of the drug. The time courses of the negative inotropic effect and of the rate of shortening in action potential duration were compared. The initial rapid decline in force of contraction was followed by a slowly developing partial recovery (fading). After removal of the drug, a transient phase of hypercontractility was observed before the control level was reached again. In contrast to the mechanical effect, the drug-induced shortening of the action potential duration proceeded monophasically. The fading and hypercontractility phenomena shown by the mechanical response to the drug were influenced by the stimulation rate and by the presence of ouabain. From the dissociation of electrical and mechanical effects it is concluded that fading and hypercontractility are not due to changes of receptor sensitivity but reflect a change in cellular calcium distribution.


Effect of adrenergic receptor blockade on the responses to isometric handgrip: studies in normal and hypertensive subjects.

Heart rate and blood pressure (BP) responses to a standardized 3 min isometric handgrip (IHG) test were measured in 32 normotensive men and compared with those found in 35 age-matched, drug-free men with established essential hypertension (BP range: 140--170/90--110). IHG testing in the normal subjects induced significant increases in heart rate (mean +/- SE: 7.6 +/- 3.3 beats/min), systolic blood pressure (19.4 +/- 5.3 mm Hg), and diastolic blood pressure (15.6 +/- 4.6 mm Hg). Although beginning from higher resting levels the hypertensive patients had similar degrees of increase in all three parameters. After chronic treatment with propranolol, the heart rate increase with IHG was suppressed in both study groups, but blood pressure responses differed, with a diminished pressor response to IHG seen in normal subjects and augmented pressor effects in the hypertensive group. Intravenous administration of phentolamine and propranolol completely abolished the pressor effects of IHG. These observations suggest that the autonomic control mechanisms mediating the responses to isometric exercise function similarly in drug-free normal and hypertensive patients and that the responses to IHG, mediated largely by endogenous catecholamine release, can be prevented by peripheral sympathetic receptor blockade.


Effects of hypertonic mannitol on contractile responses and 45Ca movements in isolated canine arteries.

The effects of hyperosmotic mannitol on vascular smooth muscle contractile responses and on 45Ca movements were examined in different isolated canine arteries. Prior exposure to 50 mM mannitol decreased contractile responses elicited with dopamine (DA) in helical strips of canine terminal mesenteric arteries (Tm) and decreased the contractile response elicited with potassium (K+) in both isolated left anterior descending (LAD) and circumflex coronary arteries. Tension responses induced by norepinephrine or DA in the Tm and K+ and prostaglandin F2 alpha in LAD were relaxed by subsequent exposure to mannitol. Mannitol increased the uptake of 45Ca in Tm arteries. Exposure of the Tm and branches of the LAD to mannitol during the washout of 45Ca resulted in a decrease in the rate of loss of 45Ca; the presence of either K+, Mg2+, or ethylenediamine tetraacetic acid during the washout did not prevent the observed mannitol-induced decrease in 45Ca efflux. These effects of mannitol on 45Ca efflux without a concomitant major change in net 45Ca uptake could be attributed to an increase in bound Ca2+ at relevant membrane sites or stores. Thus, hypertonic mannitol may alter contractile responsiveness of selected canine arteries by impeding the release of bound and/or sequestered Ca2+  and, in this manner, decreasing the Ca2+ concentration at the contractile elements.


Prazosin and renal hemodynamics: arteriolar vasodilation during therapy of essential hypertension in man.

Renal hemodynamic effects of chronic prazosin therapy were studied in 10 men and with essential hypertension. Successful antihypertensive therapy was accompanied by diminution in total renovascular resistance, probably at the level of the afferent arteriole. The resistance change was unexplained by alterations in intravascular volume, the vasodilatory renal kallikrein-kinin system, or sympathetic nervous system activity, but was associated with a diminution in plasma renin activity. Potential mechanisms for the resistance decrement are discussed, including renal perfusion autoregulation, diminished activity of the vasoconstrictive renin-angiotensin system, and direct intrarenal afferent arteriolar alpha adrenergic blockade.


Effects of atenolol, metroprolol, and pamatolol on cardiac and vascular beta-adrenoceptors in the rat.

Three beta-adrenoceptor antagonists--atenolol (A), metoprolol (M), and pamatolol (P)--were examined with respect to their effects on isoproterenol responses of isolated right atria and portal veins from rats. With all three blockers at concentrations of 10(-7), 10(-6), and 10(-5) M, the chronotropic atrial responses indicated a linear relation with slope of 1 for log (dose ratio - 1) versus log[blocker]. The following pA2 values were obtained: A = 7.14, M = 7.51, P = 7.56. Inhibition of spontaneous contractions in portal veins by isoproterenol was studied after alpha-adrenoceptor blockade by phenoxybenzamine. The beta-adrenoceptor antagonists were used at 10(-6), 10(-5), and 10(-4) M in these experiments. Only with P did the relation log (dose ratio-1) versus log[blocker] show a slope near unity, whereas A and M gave slopes significantly less than 1. Intercepts of the regression lines with the abscissa corresponded to pA2 values of A = 6.01, M = 6.51, P = 5.54. The results indicate that all three substances are "cardioselective" beta-adrenoceptor antagonists devoid of intrinsic beta-stimulatory action. A and M show a comparable degree of cardioselectivity, whereas P is more selective.


Effect of endothelial damage on prostaglandin synthesis by isolated perfused rabbit mesenteric vasculature.

Isolated perfused rabbit mesenteric blood vessels selectively metabolized arachidonic acid to prostacyclin (prostaglandin I2). However, the less lipid soluble prostaglandin endoperoxide (PGH2) administered exogenously was not metabolized by vascular prostacyclin synthetase but was partially degraded to prostaglandin E2 (PGE2). Peptide stimulation (e.g., angiotensin II, bradykinin) resulted in formation of both PGI2 and PGE2 from endogenous arachidonic acid. Denuding the blood vessels of their endothelial layer by perfusion with hypotonic fluid did not affect the metabolism of arachidonic acid or the response to peptide stimulation, suggesting that the cyclooxygenase, prostacyclin synthetase, and angiotensin II receptors are present and functional in the subendothelial smooth muscle cells of the vessel walls. In contrast, exogenous PGH2 either escaped completely unmetabolized or was converted to both PGI2 and PGE2 in the presence of vascular injury induced by hypotonic fluid.


A specific effect of lidocaine and tocainide on ventricular conduction of mid-range extrasystoles.

Lidocaine and tocainide had no effect on ventricular conduction of extrasystoles with coupling intervals longer than 500 msec in isolated blood-perfused dog hearts, but caused interval-related increases in conduction time of extrasystoles in the range of 250--400 msec, here called mid-range extrasystoles (MRE). Quinidine, procainamide, disopyramide, and methyl lidocaine increased conduction times of extrasystoles at all coupling intervals, and no additional slowing of MRE was observed. The slowing of MRE specific to lidocaine and tocainide was confirmed in the intact dog heart. During acute myocardial ischemia in the intact dog heart, conduction was slowed and additional slowing of MRE was found. Lidocaine and tocainide caused further slowing of conduction of MRE. This unique effect of lidocaine and tocainide on the conduction of MRE may be important in the suppression of reentrant arrhythmias. However, lidocaine and tocainide were also found to be arrhythmogenic when extrasystoles were introduced, after acute coronary occlusion, in those animals in which such occlusion alone did not allow demonstration of arrhythmias due to extrasystoles.


Electrophysiological actions of lorcainide in patients with cardiac disease.

Lorcainide is a new antiarrhythmic agent which effectively suppresses ventricular premature contractions. Its electrophysiological actions were studied in 15 cardiac patients with and without cardiac conduction abnormalities. In a dose of 2 mg/kg body weight, lorcainide decreased the sinus cycle length (SCL) and increased the corrected sinus node recovery time. The intra-atrial, atrioventricular (AV), and intraventricular conduction times were prolonged. A third-degree AV block developed in 2 patients with pre-existing conduction abnormalities. QRS widening showed that intramyocardial conduction was also affected. The effective refractory period of the atrioventricular node (ERP-AVN) was shortened, whereas the effect on the functional refractory period of the AVN was variable. The decrease in SCL and ERP-AVN may be due to a possible anticholinergic effect of the drug. The accessory pathway was blocked in 3 patients with a Wolff-Parkinson-White syndrome. The ERP of the ventricle was slightly prolonged. The electrophysiological profile of the drug, i.e., slowing of conduction velocity throughout the heart combined with shortening of SCL and ERP-AVN, differs from other antiarrhythmic agents.


Effects of probucol on hyperlipidemic patients with cardiac allografts.

Probucol, a well-tolerated new drug with moderate hypocholesterolemic effect in the general population, was evaluated in hypercholesterolemic (greater than 300 mg/dl) cardiac transplant recipients. Nine patients received probucol, 500 mg twice daily, in a single-blind, placebo-controlled trial lasting 20 weeks. Six patients responded with cholesterol falls of -20% (mean, -13.2%), and 1 patient showed minimal change. One patient developed cardiac rejection and was excluded, and another responded paradoxically and was returned to previous therapy. In 1 patient, probucol was readministered, and a similar favorable response occurred, which has been sustained for over 6 months. For the group completing the study, low-density lipoprotein (LDL) cholesterol fell by 15.4%, accounting for the major portion of overall change in plasma cholesterol. High-density lipoprotein (HDL) cholesterol also fell (-15.6%), and the LDL/HDL ratio was unchanged. Triglycerides and very low density lipoprotein cholesterol showed variable responses with no overall change. Probucol was universally well tolerated, without adverse clinical or laboratory effects. In conclusion, probucol may effect a moderate hypocholesterolemic response in hyperlipidemic recipients of cardiac transplantation, despite long-term maintenance therapy with corticosteroids. However, efficacy should be individually documented. The long-term effect of probucol on the natural history of coronary artery disease in these patients remains to be determined, particularly in view of its effects on HDL cholesterol.


Influence of atropine on spontaneous or catecholamine-induced arrhythmias after experimental infarction in the dog.

The effects of atropine on ventricular tachyarrhythmias induced by ligature of the anterior descending coronary artery were studied in conscious dogs. On the first day after surgery, animals received a mixture of quinidine and phenytoin orally in order to partially correct rhythm disturbances on the second day. The same animals were used on the third and fourth days after occlusion, when spontaneous regression of arrhythmias was occurring. Atropine (0.2 mg/kg i.v.) or methscopolamine (0.2 mg/kg i.v.) almost completely suppressed the ventricular arrhythmias which persisted in some animals on day 2. In animals with partially corrected tachyarrhythmias, atropine protected against aggravation of rhythm disturbances by norepinephrine and even more so against those induced by isoproterenol. The favorable effect of atropine seems to be mainly related to overdrive suppression. Atrial electrostimulation at a frequency slightly above that of the ventricular arrhythmias also normalized cardiac rhythm.


Hemodynamic consequences of administration of phentolamine or nitroprusside with dopamine in the dog.

Dopamine was administered intravenously to two groups of anesthetized dogs at rates of 2.5, 5, 10 and 20 micrograms/kg/min. After the 20 micrograms/kg/min infusion, mean arterial pressure, heart rate, renal blood flow, mean pulmonary arterial pressure, and left ventricular filling pressure were increased. The dopamine infusion was continued in both groups of animals. In Group I phentolamine was administered at rates of 2.5, 5, and 7.5 micrograms/kg/min; in Group II sodium nitroprusside at rates of 1, 2, 3, and 4 micrograms/kg/min. Phentolamine and sodium nitroprusside produced similar reductions in mean arterial blood pressure, mean pulmonary arterial pressure, left ventricular filling pressure, and similar increments in heart rate. Renal blood flow was maintained with phentolamine infusions and decreased slightly with the nitroprusside infusions. These studies demonstrated that both phentolamine and nitroprusside have the ability to reduce the vasoconstricting effects of dopamine.


Hemodynamic consequences of long-term beta-blocker therapy: a 5-year follow-up study of atenolol.

The long-term hemodynamic effects of atenolol were studied in 10 patients with previously untreated essential hypertension in WHO stage I. Oxygen consumption, heart rate, cardiac output, and intra-arterial brachial pressure were recorded at rest in supine and sitting positions and during steady-state work at 50, 100, and 150 W. The patients were treated with 100 mg atenolol daily (200 mg in 1 patient) as the sole antihypertensive drug and were restudied after 1 and 5 years. After 1 year the blood pressure was reduced approximately 18% both at rest and during exercise, and the heart rate about 25%. The cardiac output was reduced 16% at rest supine, 27% at rest sitting, and about 20% during exercise. The calculated total peripheral resistance was not decreased compared to pretreatment values. After 5 years on treatment, the hemodynamic parameters were almost identical to those seen after 1 year. There was no increase in stroke volume or cardiac output and no decrease in the total peripheral resistance. Thus prolonged beta-blocker treatment over several years does not seem to normalize hemodynamics in patients with mild to moderate essential hypertension.


Effects of prazosin and phentolamine on arterial pressure, heart rate, and renin activity: evidence in the conscious rat for the functional significance of the presynaptic alpha-receptor.

We examined the functional significance of the presynaptic alpha-adrenergic receptor, to stimulus-induced norepinephrine release. The effects of prazosin and phentolamine, alpha-adrenergic receptor antagonists with different in vitro selectivities for the presynaptic alpha-receptor, on mean arterial pressure and serum renin activity were determined in the conscious rat. Both drugs resulted in dose-related reductions in mean arterial pressure and dose-related increase in heart rate and serum renin activity. However, consistent with the greater selectivity of prazosin for the postsynaptic alpha-receptor, a given reduction in arterial pressure was associated with a lesser increment in heart rate and serum renin activity after prazosin than after the nonselective agent phentolamine. The differential effects of these agents on heart rate and serum renin activity are consistent with different degrees of blockade of a functionally significant presynaptic alpha-receptor.


Increased diastolic time: a possible important factor in the benefical effect of propranolol in patients with coronary artery disease.

Diastolic time (DT) calculated as the cycle length minus electromechanical systole (QS2) has a nonlinear relationship to heart rate (HR), increasing rapidly as rates fall below 75. The effect of propranolol on DT was studied in 150 patients with coronary artery disease. Patients were divided into three groups. Group I included patients with stable angina pectoris: propranolol (2.5 mg, i.v.) significantly increased DT from 411 +/- 18 to 527 +/- 22 msec (p less than 0.001) in 23 patients of group I; therapy with propranolol (mean daily dose 200 +/- 15 mg) increased DT from 446 +/- 29 to 766 +/- 26 msec (p less than 0.001) in 15 patients with stable angina. Group II was made up of patients with acute myocardial infarction: Propranolol (2.5 mg, i.v.) increased DT from 379 +/- 16 to 458 +/- 24 (p less than 0.001) in 18 of these patients. Group III included patients with recent coronary bypass surgery: propranolol (2.5 mg, i.v.) increased DT from 323 +/- 9 to 468 +/- 24 msec (p less than 0.001) in 14 patients 7 days after surgery. In addition, DT at 15 hr and 2 weeks after surgery was compared in 30 patients maintained on propranolol (mean daily dose, 155 +/- 11 mg preoperative and 68 +/- 9 mg postoperative) and 50 other patients who underwent coronary bypass surgery not on propranolol. DT was greater in propranolol patients (546 +/- 21 vs. 388 +/- 16 msec, p less than 0.001), preoperative and 396 +/- 15 vs. 320 +/- 12 msec, p less than 0.001, postoperative). Changes in DT after propranolol are mainly attributed to decreased HR. Changes in QS2 were much less profound and always less (p less than 0.01) than changes in DT. Thus propranolol significantly increased DT per beat in patients with coronary artery disease, which allowed more time for coronary perfusion; this effect of propranolol could well be as important as the reduction of myocardial oxygen consumption.


Interactions between clonidine and alpha-adrenoceptor blocking drugs on the tachycardic response to stimulation of the cardiac nerve in dogs.

In pentobarbital-treated dogs clonidine (10 micrograms/kg) reduced the increase in heart rate caused by electrical stimulation of the cardiac nerve (1-10 Hz). We studied the actions of six alpha-adrenoceptor blocking drugs. Yohimbine (0.3 mg/kg) and phentolamine (1 mg/kg) potentiated the effects of nerve stimulation and antagonized the inhibitory effects of clonidine. Piperoxan (1 mg/kg) increased the response to nerve stimulation but antagonized the effects of clonidine only at the lowest frequency of stimulation. Thymoxamine (1 mg/kg) and prazosin at high doses (1 mg/kg) also antagonized the effects of clonidine but failed to increase the positive chronotropic response to stimulation of the cardiac nerve. AR-C239, a new and potent alpha-adrenoceptor blocking agent, changed neither the response to nerve stimulation nor the inhibitory effect of clonidine. The effects of all these drugs were observed at doses which reduced or reversed the pressor response to adrenaline. Therefore, our results afford further evidence for a dissimilarity between postsynaptic and presynaptic alpha-adrenoceptors in the dog. In addition, they show that the failure of an alpha-adrenoceptor blocking compound to increase the response to nerve stimulation does not necessarily indicate a lack of presynaptic alpha-adrenoceptor blockade.


Correlation between pharmacokinetics and intropic and electrophysiologic responses to digitoxin in the intact dog.

We attempted to correlate inotropic and eletrophysilogic digitoxin effects with serum digitoxin concentrations during 8 hr after a single dose. Eight pentobarbital-anesthetized Labrador dogs were given 2.0 mg digitoxin intravenously. Left heart catheterization and His bundle registration were performed. acing and programed electrical stimulation were used to determine heart rate-independant changes in dP/dt, intra-atrial, atrioventricular (AV), His-Purkinje, and intraventricular conduction velocity. The effective (A-ERP) and functional (A-FRP) refractory periods of the atrium and the functional nodal refractory period (AV-FRP) were measured. Serum digitoxin concentrations were determined by radioimmunoassay. Median serum digitoxin half-time was 5.7 hr. The dP/dt increased after digitoxin, with maximum values after 2 hr. Digitoxin concentration correlated with the inotropic response after 4 hr. Heart rate fell significantly within 2 min and remained below control values for the 8 hr observation period concimitant with an increase in AV-nodal conduction time and AV-ERP increased significantly in the late elimination phase, while A-FRP increased slightly initially and remained high. No consistent correlation was found between the electrophysilogic variables and serum concentrations. We conclude that the two main effects of digitoxin, the inotropic and electrophysiologic, are dissociated in the elimination phase after a single dose.


Symbols and selectivity: a statistical analysis of native American medical ethnobotany.

Native Americans use a wide range of plants medicinally. Many of these plants have profound meaning to their users. Does this mean, as some assert, that tribal medicine is "all placebo"? Since the essential character of meaning is the arbitrariness of the sign, then insofar as this medicine is symbolic, the plants used medicinally will be a random representation of plants available in nature. Several regression analyses of plants used by native Americans on plants available to them indicate substantial selectivity in plant use. Native American medical ethnobotany is not only placebo medicine.


Disopyramide plasma and myocardial tissue concentrations as they relate to antiarrhythmic activity.

Antiarrhythmic concentrations of disopyramide in canine plasma and myocardium were determined by gas chromatography. Ventricular tachycardia was incuded in anesthetized dogs by the intravenous administration of ouabain. Disopyramide phosphate was then administered by a two-stage continuous infusion method. A rapid infusion of disopyramide (9.08 mg/kg/hr) was administered for 30 min, followed by a slow infusion (2.18 mg/kg/hr) to maintain steady-state plasma levels of 1.98-2.21, mean +/- SEM = 2.1 +/- 0.02 microgram/ml at the end of 2 hr. Myocardial tissue levels of disopyramide at steady-state plasma levels were four times those of plasma (atrial tissue, 8.91 +/- 0.10; right ventricular free wall, 8.93 +/- 0.13; left ventricular free wall, 9.11 +/- 0.16 microgram/gm wet tissue). The intravenous administration of 80 units crystalline zinc insulin produced both hypokalemia (3.78 +/- 0.22 reduced to 2.36 +/- 0.18 mEq potassium/liter plasma) and a reappearance of ventricular tachycardia despite no change in plasma and myocardial tissue concentrations of disopyramide from those which had been effective in establishing and maintaining sinus rhythm. The observations demonstrate a relationship between plasma and myocardial disopyramide concentrations such that the former can be used in assessing patient therapy. In addition, this study suggests the important of plasma potassium in determining the therapeutic effectiveness of disopyramide.


Structure-activity relationships of an aminosugar cardiac glycoside, ASI-222 HCl, in the heart-lung preparation of the dog.

ASI-222 [3-beta-O-(4-amino-4,6-dideoxy-beta-D-galactopyranosyl) digitoxigenin HCl] is a semisynthetic aminosugar cardiac glycoside that has been shown to have a greater therapeutic index than ouabain or digoxin in dogs. We have compared the effects of ASI, digitoxigenin, digitoxigenin-beta-D-galactose, and digoxin in the dog heart-lung preparation. Minute work and stroke work were calculated. Controls were obtained before and after the hearts were failed with sodium pentobarbital. ASI-222 is about three times more potent than digitoxigenin and about twice as potent as digitoxigenin-beta-D-galactose in producing similar increases in the left ventricular stroke work. ASI-222 is about three times more potent than digoxin in creasing left ventricular stroke work. Our results indicate that the addition of an aminosugar group to the genin further increased potency over that observed with the addition of a single neutral sugar and prolonged the duration of activity in failing myocardial tissue.


Cardiotonic effects of anthopleurin-A, a polypeptide from a sea anemone.

The positive inotropic effect of anthopleurin-A (AP-A) was studied in vitro on isolated cat heart papillary muscles and in vivo in anesthetized and conscious dogs. In vitro, in low Ca2+ solution (1.27 mM), AP-A increased the force of contractions of isolated cat heart papillary muscles at concentrations from 0.2 x 10(-8) M and higher; on a molar basis, AP-A was more than 200 times as potent as digoxin and on a weight basis, 33 times as potent. In vivo in anesthetized dogs, AP-A at 0.2 microgram/kg/min i.v. increased myocardial contractile force; the geometric mean dose of AP-A required to increase the contractile force by 25% was 2.6 micrograms/kg; the corresponding dose of digoxin (infused at 2.8 micrograms/kg/min) was 107.4 micrograms/kg. The geometric mean lethal dose of AP-A for 8 dogs was 19.3 and that of digoxin 263.2 micrograms/kg i.v. The therapeutic index of AP-A was significantly higher than that of digoxin. All animals that received either AP-A or digoxin died in ventricular fibrillation. The reversal of t-wave was typical for AP-A. As measured by left ventricular pressure telemetry, AP-A, 2 micrograms/kg i.v. single dose, increased LV dp/dt max in conscious dogs for longer than 2 hr.


Effects of etafenone and antiarrhythmic drugs on Na and Ca channels of guinea pig atrial muscle.

Since Ca-mediated electrophysiological phenomena seem to be important in the generation of various arrhythmias, we quantitatively compared the effects of quinidine, disopyramide, procainamide, lidocaine, propranolol, and etafenone, a new coronary vasodilator with antiarrhythmic properties, on Na and Ca channels in normal and depolarized guinea pig atria. The maximum rate of rise of the normal action potential and the amplitude of the depolarized, Ca-mediated action potential were measured to examine the Na and Ca channel activity, respectively. In both preparations, the contractile force was determined concomitantly in order to establish the role of Ca influx in the inotropic effects of the drugs. All six drugs suppressed the Na channel, the effect being strongest with etafenone, followed in order by quinidine, propranolol, disopyramide, lidocaine, and procainamide. Only quinidine, disopyramide, and propranolol suppressed the Ca channel and only in concentrations higher than those required to block the Na channel. Etafenone and procainamide decreased Ca influx, while lidocaine had no effect on it. The negative inotropic effects of propranolol and disopyramide may be due to decreases in Ca influx, but the order drugs seem to have other mechanisms for their inotropic effects.


Electrophysiological and hemodynamic disturbances in a patients overdosed with disopyramide.

Disopyramide exerts a quinidinelike effect on the heart and is a valuable drug for treating atrial and, especially, ventricular tachyarrhythmias. The therapeutic plasma concentration of disopyramide is thought to be 2.0-4.0 (5.0) microgram/ml. We here report the cardiac effects of a high dose of disopryamide in a patient with extensive coronary artery disease complicated by ventricular extrasystoles. At plasma levels above approximately 7.0 micrograms/ml, heart rate was decreased, while PQ interval, width, and QT interval were increased. At concentrations above approximately 6.0 micrograms/ml, disopyramide exerted negative intropic effects as judged by increases in mean right atrial, pulmonary arteriolar, and pulmonary capillary venous pressures and a decrease in cardiac output.


Infectivity suppressing and virus-binding activities of a membrane material isolated from tobacco leaves.

TMV binding substance (R) was isolated from a tobacco leaf membrane fraction and was purified by extraction with organic solvents and by column chromatography. Experimental results suggest that the binding of R with TMV results in inactivation of TMV. When tobacco leaves were inoculated with the R-TMV complex, it was found that the formation of polysome containing infecting viral RNA was inhibited. Model experiments showed that the mode of R-TMV adsorption to the membrane is different from that of TMV adsorption and that stripping of coat protein from TMV by SDS was inhibited by R. A possible explanation for the mechanism of this inhibition by R is that the R-TMV complex follows a pathway which does not lead to establishment of infection. Although less efficient, R was still active when it was applied after virus inoculation. Due to its affinity to coat protein, R might also interfere with a later process of viral multiplication.


Antigenicity of erythrocytes as analyzed in terms of their cross-reactivity.

The antigenicity of human erythrocytes of four different ABO blood groups and sheep erythrocytes of unknown blood type from different individual sheep were analyzed in terms of their cross-reactivity with antibody-producing cells (plaque-forming cells, PFC) and serum antibody in immunized C57BL/6 and C3H/He mice. The antigenicity of human erythrocytes of different ABO blood groups in the C57BL/6 mice, as determined by the number of specific PFC, was, in decreasing order, AB = A greater than B = O (p less than 0.005). The efficiency of immunogenicity of the human erythrocytes in terms of their cross-reactivity with PFC was, in order, AB = A = B greater than O, and the degree of reactinogenicity was, in order, AB greater than A greater than B greater than O. The order of antigenicity of sheep erythrocytes from different animals, SRBC No. 1 - No. 6, was No. 1 (= No. 2) greater than No. 3 = No. 4 = No. 5 greater than No. 6 in C57BL/6 mice and No. 1 = No. 2 = No. 3 = No. 4 = No. 6 greater than No. 5 in C3H/He mice, determined by the number of specific PFC (p less than 0.01). The cross-reactivity of SRBC No. 1 - No. 6 with PFC demonstrates that the order of immunogenicity of SRBC was No. 1 = No. 2 = No. 3 = No. 4 = No. 5 greater than No. 6 in C57BL/6 mice and No. 1 = No. 2 = No. 3 = No. 4 = No. 6 greater than No. 5 in C3H/He mice, and that of their reactinogenicity was No. 1 greater than No. 2 =No. 3 = No. 4 = No. 5 greater than No. 6 in C57BL/6 mice and No. 1 greater than No. 4 = No. 6 greater than No. 2 = No. 3 greater than No. 5 in C3H/He mice. The cross-reactivity at the antibody level was indicative of the immunologic characteristics of blood cells of low antigenicity (human group O erythrocytes and SRBC No. 5 and No. 6). SRBC No. 5 and No. 6 were somewhat opposed to each other regarding antigenicity in C57BL/6 and C3H/He mice. This signifies the presence of different immunogenic components on SRBC No. 5 and No. 6. The production of anti-SRBC No. 1 antibody reached its peak on the third day after secondary immunization. That of anti- SRBC No. 1, cross-reactive with SRBC No. 6, occurred after a longer latent period, reaching its peak on day 6. This indicates that SRBC No. 1 possesses more than one kind of immunogenic component or immunogenic determinant group on its surface.


Observations on the antigenicity and serological relationships of stocks of Trypanosoma congolense from East and West Africa.

Antigenic relationships of 4 stocks of Trypanosoma congolense from different parts of Africa were examined by immunofluorescence (IFAT) and neutralization tests. Antisera to each stock were obtained from rabbits infected with trypanosomes transmitted by Glossina morsitans. Trypanosomes for use as antigens were obtained from local skin reactions developing on rabbits infected with 2 of the stocks. Using the IFAT and antisera at end-point dilutions approximately 40% of the trypanosomes fluoresced strongly and a further 30% less intensely with homologous antisera, indicating antigenic heterogeneity among the trypanosomes developing in the skin. Using antisera at low dilutions some samples gave cross-reactions with trypanosomes of heterologous stocks, but at higher dilutions there were no cross-reactions with either of the antigens. The lack of cross-reactions at high antiserum dilutions was interpreted as indicating antigenic differences between the 4 trypanosome stocks. Using neutralization tests only homologous antisera reduced the infectivity of trypanosome suspensions. Overall, these observations indicated that there were at least 3 different strains of T. congolense among the 4 stocks examined.


Immunological differences in Eimeria maxima: effect of a mixed immunizing inoculum on heterologous challenge.

The immunological differences known to exist between laboratory strains of Eimeria maxima was confirmed. Protection against challenge with different strains or field isolates of the species could be achieved by including small numbers (25 oocysts) of each in the immunizing inoculum. Similar protection was obtained when 4 distinct populations which were allowed to interbreed were used in the immunizing inoculum. This hybrid mixture of E. maxima was used to immunize chickens against challenge with 7 new isolates of E. maxima from poultry houses in different parts of England. The results show that although immunological differences exist within E. maxima good protection against many strains of this species may be achieved by initial infection with the hybrid mixture of E. maxima.


[Convalent binding of various tetra- and tricyclic heterocycles with isothiocyanate structure to human serum albumin and bovine gamma globulin. Part 24: Immunosuppressive agent - antigen conjugates (author's transl)].

In view of their potential activity as antigen-specific immunosuppressive agents, the authors produced conjugates by treating the tri- and tetracyclic compounds 1-8 (with isothiocyanate structure) with human serum albumin (HSA) and bovine gamma globulin (BGG). In certain cases the conjugates of BGG were obtained only under conditions leading to protein degeneration. The corresponding products were devoid of antigen-specific immunosuppressive activity and must be considered to be merely repository forms of the respective low-molecular compounds. In contrast, the conjugates of HSA could be obtained under conditions under which no protein denaturation was detectable. These conjugates may be used for the planned investigation. After reaction of the isothiocyanates with the proteins, time dependent changes were observed in the ultraviolet spectra of some conjugates, the possible causes of which are discussed.


Action of substance P on neurotico-hypertensive rats.

The action of an eledoisin-hexapeptide analogue (EH) upon learning and memorising processes of 48 male Wistar laboratory rats aged between 5 and 6 months was studied and is reported in this paper. The animals suffered from neurogenic hypertension which had been experimentally induced by applying emotional stress. A comparison between the action of EH (Lys-Phe-Ile-Gly-Leu-MetNH2) and that of Substance P (Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-MetNH2) on conditioned-reflex learning in the intact rat had been reported by the authors in one of their previous papers [7]. The following results were obtained with regard to EH and its action upon rats with neurogenic hypertension. The learning process was favoured, as it had been by 2 or 3 weeks of exercise. Defective learning and memorizing process as well as impaired behavioural patterns, interpreted as neurotic phenomena, were normalized by doses of 250 microgram/kg and 500 microgram/kg. Blood pressures were reduced, depending on dosage. The action of the EH analogue used on the central nervous system was stronger than that on blood pressure. Discontinuance of peptide application was followed by the phenomenon of "state-dependent learning". The results are likely to suggest possible involvement of such peptide sequences in the regulation of processes which are relevant to the whole. That effect is of particular interest, as Substance P is under discussion as a transmitter or modulator in mammals.


[Comparison of three anti-nucleus antibody detection methods in different pathologies (author's transl)].

Anti-nucleus antibodies were detected in the blood sera of 232 patients of different pathologies by three diverse methods: a morphological method, and indirect immunofluorescent method and a radioimmunologic method. The results demonstrated the importance of the radioimmunologic method since this method displays anti-DNA antibodies which are specific in the active stage of lupus erythematosus (LE). Forthermore, the immunofluorescent method proved to be highly sensitive; and the morphological method can reveal different aspects of the LE phenomenon (such as nucleus phagocytosis) which can be indicative of an auto-aggressive process. Our survey indicates that simultaneous use of the three methods is advisable.


[Prognostic value of alpha-foetoprotein in fulminant hepatitis (author's transl)].

Serum concentrations of alpha-foetoprotein (AFP) were measured sequentially by radioimmunoassay in 32 patients with fulminant hepatitis and coma, 22 of whom died. Levels were significantly elevated in 100% of patients who survived and in 7 out of 22 patients (38.5%) who died (P = less than 0.005). In the survivors the rise of AFP levels was found early after the development of coma and subsequently in all serum samples obtained during acute phase of illness. In 2 of 7 fatal cases who had raised AFP levels and the more protracted illness, AFP levels fall after 6 days. The high levels of serum AFP observed in severe forms of hepatitis may represent active hepatocyte regeneration after extensive hepatic necrosis. Although the correlation between survival and serum concentration of AFP is not absolute, our finding indicate that the rise in serum of this protein in patients with massive hepatic necrosis may be considered a favorable prognostic sign.


[Determination of alpha-1-fetoprotein during pregnancy].

218 samples of serum of pregnant women from 144 normal and 12 risky pregnancy, are tested for AFP RIA, radial immunodiffusion and slide agglutination. The results, according to thet of other AA., as regards un normal pregnancy AFP behaviour, emphasize the necssity to have more data from the same subjects during pregnancy, in order to plot a significant curve for the interpretation of the single values. As regards the technique, the delicacy and the specificity of the research, suggest the use of very sensible methods, which leave simplicity of execution and immediacy of results, out of consideration.


Effect of LH-RH, gonadotrophins or sex hormone treatment on RNA and DNA concentration in hypothalamus, pituitary, ovary and uterus in the immature female rat.

RNA and DNA concentration was measured in hypothalamus, pituitary, ovary and uterus of immature female rats after treatment with 5 doses of LH-RH, PMSG, HCG, FSH + LH, estradiol benzoate or progesterone. All assayed hormones decreased DNA concentration and increased the [RNA]/[DNA] ratio in their target organs. These findings were interpreted as increases in cell volume and RNA synthesis in target organs after treatment. Gonadotrophins and sex hormones decreased DNA concentration and increased RNA synthesis in hypothalamus and pituitary, which revealed the stimulatory effect of both hormonal groups on the above mentioned organs.


Whipple's disease: the role of duodenal biopsy in diagnosis and follow-up.

Blind suction small intestinal biopsy is still the classical method in establishing the diagnosis of Whipple's disease. Duodenal endoscopy with directed biopsy seems however to be a better alternative after the description, in recent years, of a characteristic endoscopic picture of Whipple's disease. In clinically suspected cases, biopsy from other parts of the small intestine may be rewarding if the duodenal biopsy is negative. A case of Whipple's disease, diagnosed by duodenal biopsy, is reported and the relevant literature is discussed.


OMGE--Study on prevalence of hepatitis B surface antigen in different liver diseases.

The prevalence of hepatitis B viral infection has been evaluated by means of a questionnaire. Contributions were made by 160 institutions from 39 countries and involved more than 400 collaborators. HBsAg was identified by a variety of test kits which were available at the time of the questionnaire. Data are presented for the prevalence of HBsAg in acute viral hepatitis, chronic hepatitis, cirrhosis of the liver and primary liver cancer. Wide variations in antigenaemia were identified in different countries and between the various forms of liver disease. HBsAg is positive more often in chronic hepatitis than cirrhosis. More data using sensitive tests are needed but it appears as if at least one-fifth of the world population has had a previous hepatitis B virus infection.


Effect of subminimal inhibitory concentrations of mecillinam on the synthesis of DNA, RNA, and protein of Salmonella typhimurium: a proposed mechanism of action.

Salmonellae alter their rod-like morphology to become large ovals within 3-5 hr after being exposed to concentrations of mecillinam below the minimal inhibitory concentration. Before this overt morphologic change occurs, synthesis of DNA increases, with a concomitant decrease in total protein synthesis. Incorporation of [3H]leucine into cell wall protein is markedly inhibited by mecillinam, whereas incorporation of leucine into soluble intracellular proteins is enchanced by the drug. Electrophoretic separation of stained, soluble intracellular proteins of salmonellae exposed to mecillinam indicates that as many as 15-20 individual groups of protein occur at higher concentrations in mecillinam-exposed salmonellae than in unexposed control organisms. These results suggest that, at low concentrations, mecillinam binds selectively to cell wall components, thereby interfering with the assembly of the cell wall during growth. This interference causes a buildup of newly synthesized soluble proteins that are normally involved in cell wall assembly. The weakening of the cell wall, combined with this buildup, causes the cell to assume a three-dimensional oval structure that is the result of equal hydrostatic pressure exerted by the medium.


Effects of subminimal inhibitory concentrations of ampicillin, chloramphenicol, and nitrofurantoin on the attachment of Escherichia coli to human uroepithelial cells in vitro.

An in vitro system with epithelial cells of the human urinary tract and Escherichia coli isolated from the urine of pateints with acute pyelonephritis was used for tests of the effect of antibacterial agents on bacterial adhesion. E. coli treated with subminimal inhibitory amounts of ampicillin attached less than untreated control bacteria. Subminimal inhibitory amounts of chloramphenicol and nitrofurantoin had no effect on adhesion of the 15 strains of E. coli tested. The combination of bacterial pretreatment with ampicillin and addition of antibodies to the adhesion test tube decreased adhesion more than did either treatment separately.


Ultrastructural and functional studied on the lymphocytes of a patient with chronic lymphocytic leukaemia and nuclear extrusion.

The cells of a patient with chronic lymphocytic leukaemia (CLL) showed a peculiar phenomenon of nuclear blebbing and nuclear extrusion, observed by transmission (TEM) and scanning (SEM) electron microscopy. Protein synthesis of the lymphocytes was 3 times higher than that of cells of other 3 patients with CLL, but was lower than the synthesis of cells from a patient with acute lymphoblastic leukaemia (ALL). Similar results were observed when RNA synthesizing activity of the cells was examined. On the other hand, thymidine incorporation proceeded in a similar rate in the patient's and the ALL lymphocytes, whereas in those CLL patients it was merely detected. The morphological findings and the synthesizing activities of the cells suggest that the patient's disease represents an intermediate form between CLL and ALL.


Pernicious anaemia in association with argyrophil (Sevier-Munger) gastric carcinoid.

3 cases of pernicious anaemia in association with gastric carcinoid (in 1 case multiple) are described. All tumours were rich in argyrophil cells with the Sevier-Munger and Grimelius staining techniques, indicating that they might be of the enterochromaffin-like cell type. The possible causal relationship between pernicious anaemia and this type of argyrophil carcinoid is discussed.


Beta thromboglobulin and glycosylated haemoglobin in diabetes mellitus.

No correlation has been demonstrated between the elevated levels of plasma beta-thromboglobulin and glycosylated haemoglobin found in a group of 97 diabetic patients. The abnormality in diabetic platelets which is reflected by beta-thromboglobulin levels may not therefore be a direct result of poor metabolic control. The highest levels of beta-thromboglobulin were found in those patients with a raised blood urea. Glycosylated haemoglobin levels correlated with prevailing blood glucose and with the type of therapy employed, and were highest in the insulin dependent diabetics, intermediate in those on tablets and lowest in those on diet alone.


Hereditary alpha 2-macroglobulin deficiency.

On screening of a normal material 1 man was found (age 37) who on repeated determinations had a low alpha 2M, namely 25 % with electroimmuno assay according to Laurell. Investigation of the family revealed that the mother (age 69) and one daughter (age 5) had low values too. All other coagulation and fibrinolytic components were normal. They had no signs of increased fibrinolysis and normal levels of alpha 2-antiplasmin, alpha 1-antitrypsin and antithrombin III. Liver function tests were normal. It seems to be an inherited deficiency. The transmission is apparently autosomal dominant and the affected members heterozygotes. The defect has not caused any clinical symptoms. This family appears to be the first reported with an alpha 2M deficiency.


Macrophage-lymphocyte clusters in the immune response to soluble protein antigen in vitro. VIII. Cinephotomicrographic studies.

T cells from immune guinea-pigs produce clusters in vitro with macrophages pulsed with soluble protein antigens. The formation of clusters is antigen-specific. Time-lapse cinephotomicrography was used to study the sequence of events leading to cluster formation and the kinetics of already formed clusters. The central lymphocyte attaches through a broad base to the macrophage during the first few hours of incubation. After several hours, during which the central lymphocyte is situated alone on the surface of the macrophage, free lymphocytes begin to interact for shorter or longer periods of time through their uropods with the central lymphocyte, thus acting as peripheral lymphocytes. In spite of a frequent traffic of peripheral lymphocytes in and out, the cluster is a relatively stable structure. Occasionally the entire complex of lymphocytes leaves the macrophage, moves randomly in the culutre and attaches to a new macrophage or returns to the same macrophage.


Macrophage-lymphocyte clusters in the immune response to soluble protein antigen in vitro. IX. Antigen-pulsed macrophages as a tool for specific absorption of cluster-initiating T cells.

T-cell populations from guinea-pigs sensitized to the protein antigens purified protein derivative of Mycobacterium tuberculosis, ovalbumin, or horseradish perioxidase can be selectively depleted of cells capable of initiating antigen-specific macrophage-lymphocyte clusters in vitro. The depletion is achieved by incubating the T cells on a monolayer of antigen-pulsed macrophages in a Petri dish for some hours and then gently aspirating the cells not adhering to the bottom of the dish. When subsequently assayed, the aspirated cells were found to be depleted of cluster-initiating lymphocytes committed to horseradish peroxidase, monolayers of macrophages pulsed with that antigen must be used. The optimum time for incubation on the absorbing monolayer appears to be 4 h, and two successive incubations are more effective than one. The cell density of the absorbing monolayer and the handling of the Petri dish may be critical for effective removal of the cluster-initating lymphocytes. With optimum procedure we have achieved up to 90% depletion of specific cells with no depletion of cells committed to a control antigen.


Cell-mediated cytotoxicity during vaccinia virus revaccination in man: influence of antibodies and interferon.

Seven healthy human adults were revaccinated with vaccinia virus. Cell-mediated cytotoxicity to vaccinia virus-infected fibroblasts was investigated on the day of vaccination and on days with peak activity. Three donors were studied until day 30 after vaccination. The addition of interferon to cytotoxicity reactions resulted in an increase in killing. This increase was not seen when antibodies were added. When a mixture of lymphocytes from a revaccinated and a non-revaccinated donor was used as effector cells, the killing observed corresponded to the killing seen with lymphocytes from the revaccinated donor, when tested alone. This finding indicates that no antibodies or other soluble mediators capable of increasing cytotoxicity are released from the lymphocytes during the cytotoxicity assay.


The humoral immune response to alpha, beta and gamma crystallins of the human lens.

By radioimmunoassay the occurrence of antibodies against alpha, beta and gamma crystallins was examined in sera from groups of patients with clear lenses, cataracts or lens injuries.  In about half of the sera from each group low anti alpha crystallin activity was found. No significant difference in antibody activity was found between the groups. Antibody activity against beta or gamma crystallins was not detected in any of the sera. Four patients with lens damage were followed for 2 years; two of them developed uveitis and showed increasing anti alpha crystallin activity. It is suggested that in man immunological tolerance to lens crystallins is normally present.


The epidermal melanocyte system in individuals of Scandinavian origin, determined by DOPA-staining and TEM.

The quantitative evaluation of DOPA-positive epidermal melanocytes in 16 patients of Scandinavian origin showed both individual and regional differences in the melanocyte count. Our data is in agreement with other published studies. The distribution in the number of melanocytes varies significantly in some specimens. This is due partly to the preparation procedure and partly to normal biological variations. We believe that we have demonstrated a cyclic function of the melanocyte in the epidermis. The varying density of cells in epidermal sheets as well as their varying morphology support the theory concerning the presence of the epidermal melanin unit.


The epidermal melanocytes in patients with cutaneous malignant melanoma. A transmission electron microscopic investigation.

The epidermal melanocytes were studied in biopsies from clinically normal skin removed at various distances from cutaneous malignant melanomas. Melanocytes with some ultrastructural changes were found in a number of patients, which can be either contingent on genetic or inter/extra cellular factors. It is possible that cells with these changes are more instable and more easily transformed to malignant cells by external influence. Malignant cells were not demonstrated in either the epidermis or the dermo-epidermal junction. Activation was seen of the epidermal melanocytes lying in the immediate neighbourhood of the superficially spreading melanomas, which would suggest that this type of tumour is presumably able to influence the melanocytes. The most pronounced changes in the whole of the melanocyte system was found in patients with multiple melanomas, and in a patient with metastasizing malignant melanoma.


Antigenic relationships among thermophilic actinomycetes.

Antigens from Micropolyspora faeni, Saccharomonospora viridis, Thermoactinomyces candidus, T. sacchari and T. vulgaris were prepared by growing them on dialysate of trypticase soy broth. Sera from rabbits immunized with these antigens were used to study cross-reactivity between thermophilic actinomycetes by antigen-antibody crossed immunoelectrophoresis and by agar gel double diffusion. Mi. faeni and S. viridis showed some degree of cross-reaction, but both failed to show any cross reactivity with Thermoactinomyces species. Antigens from Thermoactinomyces cross-reacted with members within the genus, but no reactivity against S. viridis or M. faeni antisera was detected. Hence, the presence of antibodies to several thermophilic actinomycetes in the sera of patients may be attributed to the exposure of the individual to different thermophilic actinomycetes rather than to the antigenic cross-reactivity between the organisms.


Control of exocellular proteases in dermatophytes and especially Trichophyton rubrum.

The production of proteases was investigated during growth of dermatophytic fungi with special emphasis on Trichophyton rubrum. Exogenous glucose suppressed elastase production in all dermatophytes examined. The production of protease active guinea pig hair in keratin-salts broth by Microsporum gypseum. Trichophyton mentagrophytes and T. rubrum was also suppressed by glucose. Various carbohydrates added to keratin-salts broth curtailed protease production by T. rubrum as did individual amino acids but ammonium phosphate did not. Enzyme activities against guinea pig hair were compared in twenty-one diverse clinical isolates of T. rubrum cultured in keratin-salts broth. Activity also occurred towards casein, bovine serum albumin, keratin, collagen and elastin after keratin-growth. Studies concerning the properties of enzyme activities in culture filtrates of T. rubrum after keratin-growth suggested that multiple proteases occurred here. Hydrolysis of guinea pig hair and elastin were optimal at pH7 while keratinase was most active at alkaline pH. Divalent cations stimulated protease(s). Ferric ion and mercuric ion stimulated keratinase but were inhibitory to guinea pig hair hydrolysis and elastase. Chelating agents inhibited elastase and the hydrolysis of guinea pig hair more severely than keratinase and all of those effects were reversed by excess calcium. A serine-protease inhibitor, phenylmethylsulfonylfluoride (PMSF), curtailed keratinase but was less inhibitory to elastase and guinea pig hair hydrolysis. Soybean trypsin inhibitor arrested each protease.


Affinity of intramitochondrial granules for ruthenium red accompanying induced cell death in chick embryos.

To test the hypothesis that ruthenium red binding of intramitochondrial granules might reflect an altered or pathological state of membranes associated with degeneration, embryos were treated with 6-AN to induce cell death in cartilaginous skeletons of chick embryos. Cervical cartilage from normal, 6-AN-treated and nicotinamide-alleviated 6-AN embryos was examined ultrastructurally for presence of IM RR-positive granules. Mitochondria of normal cervical chondroblasts which undergo normal phenotypic expression acquire RR-positive granules, although few mature cells are observed in young embryos. Necrotic chondroblasts, chondroblasts in various stages of degeneration, and proliferating chondrogenic cells of 6-AN-treated embryos all demonstrated induced RR-positive IM granules. Foci of degenerating chondroblasts, with mitochondria demonstrating RR granules, were observed infrequently in teratogen-alleviated tissue. The cytological features induced by 6-AN confirm its lethal effect and the degenerative effect on membranes presumably "unmasks" mitochondrial Ca-affinity sites which then become RR-positive. Cytochemical observations correspond with the biochemical and structural changes induced by 6-AN and confirm the hypothesis that RR-positive sites are the result of pathological changes.


beta-thromboglobulin in antecubital vein blood--the influence of age, sex and blood group.

Samples of blood for estimation of beta-thromboglobulin (B.T.G.) in normal subjects have been obtained both from blood donors at the end of their donation and from other volunteers by standard venepuncture. The former was a satisfactory method in males only. By standard venepuncture females have on average lower values than males and in the pre-menopausal age groups the difference is statistically significant. Mean values vary little up to 59 years after which they rise slowly but steadily. There is no significant difference in the mean B.T.G. values between the ABO blood groups.


Neovasculogenesis. Triggering factors and possible mechanisms.

The possible regulating mechanism(s) of neovascularization are discussed. Experimental data supporting the view that ocular neovascularization might be triggered by a fundamental metabolic mechanism are presented. Based on his own data and scattered evidence from the literature, the author suggests a model for a possible regulating mechanism of the neovascular process in which the prostaglandins fulfill the role of neovascular mediating substances. Preliminary results demonstrating that indomethacin limits the extent of the neovascular process support this concept and may have a therapeutic implication. Further studies are in progress in order to better understand the pivotal role fulfilled by the prostaglandins in neovasculogenesis.


[Immunofluorescent diagnosis of Entamoeba histolytica trophocoites in preserved stool specimens of patients (author's transl)].

A fixative and examination technique is described for identification of Entamoeba histolytica trophocoites in faeces with the aid of the indirect immunofluorescence technique. Magnaforms in bloody-dysenteric specimens and Minutaforms in spontaneous or saline purged specimens give equally good fluorescence. The specifity of the rabbit antiserum against axenically grown E. hist. is good: There is strong positive reaction only with trophocoites of E. hist. Weak crossreactions are encountered with trophocoites of E. coli, E. hartmanni and E. polecki. Negative reactions are encountered with trophocoites of Endolimax nana, Jodamoeba bāutschlii, and Dientamoeba fragilis. Amebic cysts, flagellates, leucocytes, epithelial cells and Blastocytis give negative reactions, too.


Primary and secondary resistance of mycobacterium tuberculosis in Eastern Botswana.

Of 51 patients in Eastern Botswana who denied previous anti-tuberculosis treatment, 6 (11.8 %) were excreting tubercle bacilli resistant to one of the first-line drugs in regular use: 4 patients (7.8 %) showed resistance to isoniazid, 2 (3.9 %) to thiacetazone and none to streptomycin. Of 44 patients known to have been on previous anti-tuberculosis treatment, 31 (70.5 %) were found to show resistance to one or more of the first line drugs: 31 (70.5 %) to isoniazid, 12 (27.2 %) to streptomycin and 11 (25.9 %) to thiacetazone. No appreciable resistance was found to second line drugs. These resistance patterns, which correspond quite well with other published results from Africa, are related to the overall problem in Botswana, namely the failure of a high proportion of patients, to complete a full course of first line treatment.


Hepatocellular carcinoma with intracytoplasmic hyaline globules.

A case of hepatocellular carcinoma with intracytoplasmic hyaline globules is reported. By serological, histochemical, immunofluorescent and electron microscopic studies the authors maintain that in this case intracytoplasmic hyaline globules represent lysosomal structures. According to Von Ardenne's theory, these lysosomal globules appear related to neoplastic cells necrosis.


[Complement fixation reaction studies in rhinopneumonitis of horses].

It was established that the complement binding reaction (CBR) is a suitable and very fast method for horse rhino-pneumonitis diagnostics. Cell cultural virus produced in cell cultures of pig kidneys was used as antigen. The antigen lots tested have no anticomplementary properties. Highest complement binding activity was evident in the non-diluted antigen, which discovered specific antibodies in immune serums. The CBR specificity was tested by the aid of homologous and heterologous serums and antigens. The titers of complement binding antibodies in the serums of 255 horses recovered from the disease varied from 1:10 to 1:320, and in some cases even to 1:640. The blood samples investigated were from studs having rhinopneumonitis proven by clinical observations, by virus neutralizing reaction and by virus isolation. In almost all animals under study neutralizing and complement binding antibodies were established simultaneously.


Immunoglobulin epitopes in primates.

Non-human primate sera were investigated for the presence of various human immunoglobulin epitopes including allotypes of kappa-, gamma 1-, gamma 2-, gamma 3- and alpha 2-chains, isotypes of kappa-, gamma 1-, gamma 2-, gamma 3-, gamma 4-, alpha 1-, alpha 2- and mu-chains and iso-allotypes. The non-human primate sera comprised representatives of several species of apes, Old World Monkeys, New World monkeys and prosimians. It is concluded that non-human primate immunoglobulins have many structural similarities with those of human IgM, IgA and IgG. The extent of these similarities parallels the taxonomic classification.


[Immunological identity of neurospecific (SCP and P-2) proteins and SCP protein distribution in the nervous system].

Capacity of basic protein (SCP) to prevent the development of experimental allergic encephalomyelitis was studied in guinea pigs. This protein is shown to possess no antiencephalitogenic effect. Distribution of this protein in various areas of the central and peripheral nervous system was investigated using immune serum to the protein. The protein under study is shown to localize mainly in the peripheral nervous system: spinal roots, sciatic nerve, brachial plexus nerves, vagus and optic nerves, and in the central nervous system areas rich in nerve fibres: spinal cord, medulla oblongata, pons varolii, corpus callosum, white substance of cerebral cortex. The protein content in corpus callosum and white substance of cerebral cortex is extremely insignificant. Using the Ouchterlony reaction of double immunodiffusion the protein under study was found in the myelin membrane of the peripheral nervous system and its immunological identity to the myelin basic protein (P-2) is shown.


[Amniocentesis for antenatal diagnosis. A report on an initial series of 180 cases (author's transl)].

The results are presented of amniocentesis for diagnostic genetic purposes in 180 patients. A diagnosis was obtainable in 170 cases (94%). Genetic abnormalities were detected in 11 cases; one patient miscarried 3 days after amniocentesis. Antenatal diagnosis of certain genetic disorders can be considered a routine method nowadays and should be offered to any woman at risk. It therefore seems necessary to supply the practitioners with the relevant information on this topic.


[Effect of temperature on RNA synthesis in Escherichia coli CRT 266 (dna Bts) following infection with bacteriophage T3].

Infection of the temperature-sensitive E. coli CRT 266 (dnaBts) with T3-phages at the temperature of 30 degrees C and 35 degrees C, respectively, induced T3-specific RNA synthesis with a maximum rate at 7 min (30 degrees C) and 4.5 min (35 degrees C) after infection. At temperatures above 40 degrees C no T3-induced RNA synthesis could be observed. Infection of E. coli CR 34--45 (dnaB+) with T3 phages at 30 degrees C, 35 degrees C and at temperatures above 40 degrees C, however, produced T3-specific RNA synthesis. The maximum of T3-induced RNA synthesis could be observed between 7 min and 3 min depending on the temperature during infection. The inability to form T3-specific RNA after infection of E. coli CRT 266 at nonpermissive temperatures may be a cause for the absence of the formation of T3 phages and lysis of the host cells.


Dependence of macromolecular composition and morphology of Streptomyces hygroscopicus on specific growth rate.

The dependence of macromolecular composition and morphology of Streptomyces hygroscopicus on specific growth rate micron was investigated. The percentage of DNA on dry weight (%DNA) is constant, % protein is also nearly independent of micron whereas %RNA rises considerably with increasing micron, regarding mycelia grown in glucose-limited and ammonium-limited continuous cultures as well as in discontinuous cultures with various carbon sources. It is probable that the overall synthesis of DNA, RNA and protein is regulated in the mycelium-forming bacterium S. hygroscopicus by the same mechanisms found in unicellular bacteria like Escherichia coli because of the qualitatively similar dependence of %DNA, %RNA and %protein on micron. But differences exist in quantitative regard whereby %DNA, %RNA and %protein of S. hygroscopicus are much smaller at low micron and, with increasing micron, approach those of unicellular bacteria. The hypothesis about the increase of the hyphal regions showing high synthesis activity in S. hygroscopicus mycelia grown in glucose-limited continuous cultures with increasing micron -- derived from comparison of macromolecular composition of S. hygroscopicus and unicellular bacteria -- was confirmed autoradiographically with respect to protein synthesis. The increase of the part of mycelial regions showing high cytoplasmic activity results in an increase of mean hyphal diameter, of mean relative apical growth rate alpha and/or mean relative branching rate beta. Beta depends sigmoidally and alpha inverses sigmoidally on micron. Therefore, the morphology of the mycelium determined by alpha and beta also depends on micron. The hyphal growth unit L/N, the distance from apex to first branch Lp and the mean distance between neighbouring branches Ln decline with increasing micron and reach a minimum at micron = 0.32 (1/h). A further rise of micron is accompanied with an increase of L/N, Lp and Ln. This means that mycelia growing slowly or very quickly have a loose form whereas quickly growing mycelia are characterized by a more compact form. The complicated dependence of alpha, beta, L/N, Lp and Ln on micron indicates that the morphology is regulated by different mechanisms depending on the specific growth rate.


[Programmed stimulation for the analysis of arrhythmias].

On the basis of the examination of a left bundle-branch heart block depending on frequency, of a paroxysmal tachycardia in functional conduction dissociation in the atrioventricular node and a WPW-syndrome of type A is shown that and how by means of programmed stimulation of the right atrium in order to establish the times of conduction and the refractory periods of all conduction sections of the conduction system of the heart also under influence of medicaments the diagnostic repertoire is enriched for the analysis of disturbances of the rhythm without comprehensively or finally estimating the clinical valency of the method.


[Quantitative measurement of arterial flow by means of directional Doppler technic in hemodynamic animal experiments].

The usability of the continuous ultrasonic Doppler-method for the measurement of the blood flow in the haemodynamic animal experiment was tested. For this purpose measurements at the ascending aorta and the commune carotid artery were performed on different current conditions and compared with the electromagnetically registered blood flow. The causes for the non-conformity are discussed.


[Serum proteins and specific antibodies in a German and an alien population (author's transl)].

Serum concentrations of immunoglobulins A, M and G, of alpha 2-macroglobulin, complement component C 3 and of specific antibodies against streptolysin O, staphylolysin, influenza/parainfluenzavirus and measles virus were measured from 323 German and 167 alien pregnant women. Except from a statistically significant increase of IgG and alpha 2-macroglobulin in the alien group, no measurable differences could be seen. It is concluded that the evaluation of all measured parameters can be equal for both populations.


[Comparative examinations of high-molecular maternal serum protein bodies during pregnancy--a contribution to biochemical pregnancy control (author's transl)].

The maternal Serum concentrations of beta 1 SP 1, IgM and alpha 2-macroglobulin were determined in relation to the gestation age, using the simple radial immunodiffusion method in 102 non-pathologic pregnancies and 35 pregnancies involving risk factors. To assess the clinical relevance of these determinations, we examined to what extent the changes in concentration of beta 1 SP 1 IgM and alpha 2-macroglobulin would permit a prognostically useful conclusion on the placenta function and foetal condition. All patients with lowered beta 1 SP 1-serum concentrations were examined for their antepartua CTG-evaluation, as well as the type of termination of parturition. The normal distribution for beta 1 SP 1 showed a continuous rise in serum concentration up to the 37th pregnancy week. During the last 3 weeks, beta 1 SP 1 remained almost constant. For the cases with EPH-gestosis and placenta insufficiency, a beta 1 SP 1-concentration below the normal distribution level was found in the large majority of all cases. In diabetes mellitus during gravidity, twin gravidity and MHN, the determination of beta 1 SP 1 is not of any decisive prognostic significance. The maternal serum levels of IgM showed no significant differences when comparing normal pregnancy and risk pregnancy. The serum concentration of alpha 2-macroglobulin increased in both groups of patients with increasing gestation age. Of the three examined protein bodies, we consider beta 1 SP 1 to be a good, additional parameter for the assessment of the trophoblast function.


Serological evidence of human leptospirosis in northern Greece.

a) 1069 human blood samples were tested for the presence of C. F. antibodies to leptospirae. 98 of these sera were from patients suffering from Weil's syndrome. 13 out of 98 patients were found to be suffering from leptospirosis; one of these died. 460 sera were from patients with febrile (acute) illness of unknown origin and 511 sera from patients with respiratory disease (especially "influenza like disease"). None of these patients was found to be suffering from leptospirosis. b) 191 human sera from healthy people at risk to leptospiral infection were examined by microscopic agglutination test (MAT). A high percentage (14.1%) was found to have antibodies against one or two leptospiral serogroups. The results obtained suggest that the incidence of leptospiral infections in humans in Northern Greece is higher than previously reported and the predominant serogroups were found to be Icterohaemorrhagiae followed by Autumnalis and Canicola.


[Investigations into the biological disintegration of organic natural substances by aquatic hyphomycetes (author's transl)].

The ability of 5 aquatic Hyphomycetes to produce Amylase, Pectinase and Cellulase was investigated. All the tested strains were weak producers of Amylase and good producers of Pectinase, whereas degradation of Cellulose was only found with two strains.


Effect of 3-methylcholanthrene on the biliary excretion of bromsulphthalein and eosine in newborn rats.

The biliary excretion rates of bromsulphthalein (BSP), bromsulphthalein-glutathione conjugate (BSP-GSH) and eosine have been studied in 3-methylcholanthrene (3-MC)-pretreated (100 mg/kg i.p.) and control rats aged 10 days. Liver weight was invariably increased after 3-MC treatment, associated with enhanced biliary excretion of total BSP. The increase in the biliary excretion of total BSP was due solely to the increased excretion of BSP-GSH. Following 3-MC pretreatment, BSP-GSH and eosine appeared in the bile in the same amount as in the control rats after i.v. administration of BSP-GSH and eosine. Pretreatment with 3-MC increased the ratio of BSP-GSH to BSP in the liver and bile. Our results suggest that the increased biliary excretion of total BSP following 3-MC treatment was due to an enhanced conjugation of BSP with GSH.


Relationship between serum IgE levels and anaphylactic histamine release from isolated rat mast cells.

Inbred Hooded Lister rats were immunized with egg albumin with B. pertussis vaccine used as an adjuvant. The serum levels of total IgE and IgE antibody (egg albumin specific) were determined by radioimmunoassay techniques before and after immunization. The basic level of total IgE in serum was 560 +/- 110 ng/ml. After immunization a maximal peak at day 11 of 1940 +/- 160 ng/ml was registered. Anti-egg albumin IgE antibody showed a maximum around day 13 of 75 +/- 11 units/ml. Pleural mast cells were isolated on Ficoll between day 14 and 20 after immunization. A significant negative correlation between the basic total IgE level and histamine release by antigen (egg albumin) was found and also a significant positive correlation of specific IgE antibody (determined at day 11) and histamine release. The correlation between IgE level and histamine release was slightly improved if instead the ratio of specific IgE antibody over total IgE was used.


Effect of irradiation and chemotherapeutic agents on the capillaries of Ehrlich ascites carcinoma of mice.

The effect of irradiation and chemotherapeutic agents on the capillaries of experimental solid tumours was analysed by the resin cast method. The most evident change of the 5 vascular layers was rarefaction of the newly produced extruding capillaries on the surface of the tumour mass, which was related to the tumour growth.


Induction of interferon and radioprotective activity of polyriboguanylic-polyribocytidylic acid complex in mice.

The effect of polyriboguanylic-polyribocytidylic acid complex was investigated against acute X- and prolonged 60Co-gamma irradiation. Prophylactic administration of the polyribonucleotide complex increased endogenous spleen colony formation and the percentage of survival of irradiated BALB/c mice. The most pronounced effect was observed when the animals had been irradiated at the time of maximum interferon accumulation in blood, i.e. 24 hr after interferon induction by the polyribonucleotide complex.


Fanconi's anemia: the preanemic phase.

Anticipation of Franconi's anemia prior to the development of overt marrow aplasia may prevent some morbidity and mortality from infectious and hemorrhagic complications. A presumptive diagnosis was made in a firstborn child and radial anomalies, small size, elevated fetal hemoglobin, intermittent excessive chromosome breakage and elevated T-antigen expression in fibroblasts infected with Simian virus 40. Serial studies may help identify infants and young children at risk to develop this type of constitutional marrow hypoplasia.


Eosinophilic globules in Spitz's nevi. New findings and a diagnostic sign.

Dull pink globules were found within the epidermis in 65% of junctional, 75% of compound, and 25% of intradermal types of Spitz's nevi (the nevi of large spindle and/or epithelioid cells). These globules were PAS-positive, diastase-resistant and also were positive with the trichrome stain. Similar-appearing eosinophilic globules were noted in the epidermis in only 2% of malignant melanomas and in but 0.9% of ordinary melanocytic nevi. The globules in malignant melanomas and in ordinary melanocytic nevi were negative with PAS and trichrome stains. Therefore, the finding of PAS- and trichrome-positive eosinophilic globules within the epidermis is a helpful sign for histologic differentiation of Spitz's nevus from malignant melanoma.


Fluorescence-activated separation of cervical abnormal cells using herpesvirus antigenic markers.

Antisera to total HSV-2 (G) viral antigens (Ra-2) and to increasingly purified viral antigenic fractions ("crude" and "pure" AG-e) specifically stain HSV-2-(G)- or HSV-1-(F)-infected HEp-2 cells in indirect immunofluorescence. Anti-"crude" AG-e sera induced by HSV-1 (F) or HSV-2 (G) produce a single precipitin band of identity when reacted in gel immunodiffusion against soluble antigenic mixtures from HSV-2-infected cells (HSV-2 (G) SAM). These reactivities are lost following adsorption of the sera with HSV-1 or HSV-2-infected cells or with pelleted or dextran gradient purified virions. Ra-2 and anti-"crude" or "pure" AG-e sera stain exfoliated cervical cells from patients with herpetic cervicitis as well as atypical cells from patients with atypia, carcinoma in situ or invasive cancer. Normal squamous cells do not stain. HSV-2 viral antigens recognized by the Ra-2 and anti-AG-e sera appear to constitute specific and sensitive markers for the separation of atypical cells in a fluorescence-activated cell sorter.


Azure B-eosin staining of blood cells: the effects of variation in stain formulation and staining technique on stain performance.

A quantitative study of azure B-eosin-stained blood cells is reported. The effects of variation in stain formulation and staining technique on the binding of azure B and eosin by acidophilic, basophilic and neutrophilic substrates were measured by scanning microdensitometry. The variables considered were stain concentration (the azure B-eosin concentration ratio was constant), azure B concentration (at constant eosin concentration), eosin concentration (at constant azure B concentration), staining time, buffer pH, metal salt contamination, dye contamination, buffer concentration and fixation time. The last two were the only variables which failed to produce changes in dye uptake. Tentative explanations of the observed effects are advanced.


Evaluation of Debrisan as a treatment for leg ulcers.

Dextranomer beads, a new treatment for leg ulcerations, has been widely promoted. Comparison with simple saline dressings in an animal model demonstrated no significant difference in its ability to decrease the quantitative bacterial count of granulation tissue. In a prospective randomized clinical study it appeared that, in moist exudative venous stasis ulcers, dextranomer beads may be an effective way of decreasing bacterial contamination. In drier ulcerations caused by this ischemia, this difference was not observed. Recognized principles of treatment such as elevation, debridement, physiological protection of granulation tissue, or vascular reconstruction are as important as a particular dressing regimen and must be utilized in the treatment of leg ulcerations.


Studies on Papanicolaou staining. I. Visible-light spectra of stained cervical cells.

This paper presents visible-light absorbance spectra of the nuclei and cytoplasms of ten types of cervical cells stained by the Papanicolaou technique, together with tables summarizing the data. It is concluded that for automated image processing of Papanicolaou smears the most suitable wavelengths are 531 nm to maximize the contrast of cell against background and a wavelength in the range 560 nm to 605 nm (with a mean at 575 nm) to maximize the contrast of the nucleus against the cytoplasm.


Sample preparation variation and its effects on automated blood cell differential analysis.

Uniform and reproducible sample preparation is an indispensable ingredient in automated instruments for cell analysis. Differences in measured cell morphology resulting from nonuniform sample preparation are indistinguishable to the instrument from differences reflecting different cell types or functions. As a result, as sample preparation becomes more variable, subtle cell distinctions are first confused and then completely obscured. As part of the development of the Abbott ADC-500 differential analysis system, automatic blood film spinner and stainer modules were developed to provide uniform sample preparation. These modules were tested to quantitate the effects of variations in the spinning and staining conditions on the measured ADC-500 parameters and observed cell morphology. The previously reported increase in spin time with increasing hematocrits was confirmed for the ADC-500 spinner. Moderate variations in spin speed, acceleration, deceleration and spin chamber configuration were found to have little or not effect. Postspinning air flow over the slide had a dramatic effect on morphology distortion. Variations of the staining temperature, staining time or rinse volume were found to have very little effect on measured leukocyte parameters although staining temperature had a dramatic effect on observed erythrocyte morphology and staining. Stain:buffer ratio variations were found to alter measured leukocyte parameters, with the measured density increasing as the fraction of stain increased up to a 1:2 stain:buffer ratio. Because the operational limits for the spinning and staining parameters are much tighter than the range of variables investigated, it was concluded that the ADC-500 sample preparation modules do not introduce variations in the analyzer results.


Microfluorometric quantitation of size, deoxyribonucleic acid and viral antigenic determinants in cells productively infected with HSV-2.

The interaction of herpes simplex virus type 2 (HSV-2) with the eukaryotic human cell line (HEp-2) was investigated by flow microfluorometric analysis. The three parameters that were quantitated include cell size, deoxyribonucleic acid content and the expression of virus-specific antigens. Productively infected cells are always smaller than uninfected ones, and they resolve into two populations with respect to viral antigenic content. Consistent with viral replication, one of these two populations, displaying a higher viral antigenic content, shows morphologic features characteristic of cell degeneration. On the other hand, the second population, with a relatively lower content of viral antigens, displays morphologic features consistent with increased cell growth. Indeed, microfluorometric measurements of the HSV-2-infected cells with respect to DNA content and expression of viral antigenic determinants resolves four cell populations. Of them, one displays the lowest level of viral antigens with the highest (equivalent to 8N) DNA content, features consistent with transformation. The results demonstrate the great potential of this technique for the detection, separation and quantitation of statistically significant cell populations not otherwise identified and for the analysis of virus-host cell interactions resulting in different pathologic outcomes.


Studies on Papanicolaou staining. II. Quantitation of dye components bound to cervical cells.

Dye binding to Papanicolaou-stained cervical cells was quantitated with a spectral subtraction technique. It was shown that of the dye components of Papanicolaou stains only aluminum hematein, orange G, light green SF and eosin Y are bound to cervical cells and that their chromophores do not interact. Bismarck brown Y was not bound to cervical cells, which confirms the subjective findings of previous authors, and so it should be omitted. No evidence of substrate-related variations in dye polymerization was found. The proportions of the four dyes were found to vary considerably from substrate to substrate. Spectra are presented of (1) solutions of the component dyes of the Papanicolaou stain, and (2) cervical cells stained with these individual component dyes, for comparison with the full Papanicolaou technique.


Human tumor cell identification in short-term cultures by combined microdensitometry and autoradiography.

Several groups are using short-term cultures of human tumor specimens as models to study cancer. When cells from a biopsy are grown in this way, it is essential to confirm that the tumor cells and not the normal stromal cells have been cultured. This paper reports the use of combined microdensitometry and autoradiography to identify tumor cells in cultures prepared from different types of malignancy, especially abdominal tumors, melanomas and gliomas. Cells of known origin were analyzed to provide standards. Three main results were shown. (1) Provided that the tumor cells have DNA values which differ from normal by at least 10%, they can be identified when the cells are first put into culture, and their presence can be confirmed at later stages. (2) The proportion of normal and tumor cells can be estimated, and the presence of just 10% normal cells can be detected, so new culture methods designed to favor the growth of tumor cells can be evaluated. (3) Most types of tumor are suitable for analysis, but when biopsies contain a high proportion of stromal fibroblasts, there is a real danger that after about 20 days in culture the normal cells will overgrow the tumor cells. An attempt to distinguish tumor cells which have normal DNA values from normal cells on the basis of a different optimum hydrolysis time with hydrochloric acid was unsuccessful.


Inhibition of rat pancreatic secretion by neurotensin: mechanism of action.

The effect of neurotensin on rat pancreatic secretion was studied in the conscious animal as well as on pancreatic lobules. In vivo neurotensin induced a dose-related inhibition of both water and protein basal secretion. Protein secretion was much more depressed than fluid secretion. Neurotensin did not modify the pancreatic response to exogenous secretin or cholecystokinin-pancreozymin, or to intraduodenal infusion of HCl. On the other hand neurotensin totally inhibited the increase in volume as well as in protein output to an intraduodenal infusion of oleic acid, but did not change the delayed inhibitory effect on protein output. In vitro, neurotensin did not affect basal and cholecystokinin stimulated pancreatic secretion. These results indicate that: 1) neurotensin could interfere with the release of hormones from the gut (cholecystokinin, and possibly VIP), 2) neurotensin did not mimic the delayed protein inhibitory effect observed after administration of oleic acid.


Potentiation and inhibition of ganglionic transmission by ruthenium red.

The effect of ruthenium red, 2.5 to 5 muM, on ganglionic transmission in rat superior cervical ganglia and frog abdominal ganglia were studied in vitro. In rat ganglia, ruthenium red caused a spontaneous firing of ganglia cells, and an increase in the amplitude and duration of the compound action potential following a single stimulus volley. However, transmission following a conditioning volley or a repetitive stimulus train to the preganglionic nerve was depressed up to 60 sec. The asynchronous firing caused by bethanechol was potentiated by ruthenium red. In the frog, ruthenium red caused repetitive firing of ganglion neurons following either orthodromic or antidromic stimulation. It is suggested that the potentiation of the single potential and the spontaneous firing are due to a ruthenium red-induced increase in intracellular calcium concentration. The depression of transmission may be due to a temporary depletion of readily releasable acetylcholine. It is also suggested that ruthenium red has an effect on the postsynaptic membrane.


Interspecies antigenic determinants of structural proteins of mammalian type-C viruses as detected by a competitive sepharose bead immunofluorescence assay.

The present paper describes a competitive immunoassay using antiserum to FeLV p 27 and SSV-conjugated Sepharose beads. The assay is applied to compare the interspecies-specific antigenic determinants of the major structural proteins of type-C viruses of different mammalian species. The test proves to be highly sensitive and specific and may be used for the demonstration of viral proteins in crude cellular extracts.


Biological markers of human tumors and monitoring of cancer treatment.

The development of human tumors is accompanied very often by tumor-associated phenomena such as production of tumor-derived substances, production of certian substances in response to the tumor or immunological reactions. Up to now no of these phenomena can be used as a diagnostic cancer test but biological markers are increasingly used for monitoring progression and regression of human tumors. Basing on a number of own studies the value of the determination of CEA-serum level and urinary excretion of hydroxyprolin, spermidin and putrescin for monitoring the tumor behaviour particularly during cancer chemotherapy is demonstrated.


Biophysical properties of virions of human adenovirus of the type 6 and its DNA.

A study has been made of human adenovirus type 6 (Ad 6) which belongs to the C subgroup of the nononcogenic adenoviruses. The buoyant density of the Ad 6 virions in CsCl gradient was 1.3545 +/- 0.0010 g/cm3. The DNA content determined from the buoyant density values and direct phosphorus analysis was 11.2 +/- 0.5 and 11.5 +/- 0.5 per cent, respectively. The S20,w0 and D20,w0 values for the Ad 6 virions were 795 +/- 18S and (3.255 +/- 0.035) X 10-8 cm2/s. The hydrodynamic parameters of the viral particle and its volume as well as the specific partial volume, the content and the molecular weight of the DNA have yielded a molecular weight of the virions of (203.6 +/- 10.0) X 10(6). The sedimentation constant and the intrinsic viscosity of the DNA have been found to be 32.7 +/- 0.7S and 93.5 +/- 5.0 dl/g, respectively. The molecular weight of the DNA found from these results is (23.4 +/- 0.8) X 10(6). The buoyant density of the Ad 6 DNA measured in CsCl and Cs2SO4 density gradients was 1.7128 +/- 0.0010 and 1.427 +/- 0.001 g/cm3, respectively. The GC content found from the buoyant density and the melting temperature (90.5 +/- 0.1 degrees C in 1 SSC and 75.0 +/- 0.1 degrees C in 0.1 SSC) was 52.6 +/- 1.0 per cent, that is, it is considerably lower than the GC content of 56-60 per cent which, according to the "Green's rule", should be typical of the DNAs of all the human adenoviruses of the subgroup C (30).