A perpendicular field electromagnet with a 250 mm access bore.

We present a laboratory electromagnet capable of generating magnetic fields up to ±0.48 T, specifically designed as a perpendicular flux source for thin film samples in an ambient environment. The magnet features a 250 mm diameter clear access bore above the sample plane, thus offering compatibility with a wide variety of experimental apparatus. Despite its generous size, the magnet thermally dissipates less than 1 kW at maximum field. A shaped ferromagnetic core is used to amplify and homogenize the field B, leading to an estimated uniformity of ±1.5 mT (≲0.3%) in B within a 28 mm2 zone at maximum field. The sample stage is thermally regulated and isolated from the magnet, enabling temperature control with ±5 mK precision even at elevated magnetic fields.

Erythropoiesis equivalence, pharmacokinetics and immune response following repeat hematide administration in cynomolgus monkeys.

Hematide is a synthetic PEGylated peptidic erythropoiesis stimulating agent (ESA) that is presently being developed for the correction of anemia in patients with chronic renal failure. Unlike currently marketed ESAs, Hematide does not possess any sequence homology to erythropoietin (EPO) and has not elicited moribund immune responses in animal safety studies thereby allowing the generation of a robust safety package. Animals administered marketed ESAs develop anti-EPO antibodies that null the effect of the administered ESA and neutralize endogenous EPO, resulting in severe anemia that precludes the interpretation of chronic safety studies. The primary objective of this study is to determine whether Hematide-specific antibodies are generated when male monkeys are exposed to high Hematide doses (10 mg/kg, intravenous [IV] and subcutaneous [SC]) administered at frequent dosing intervals (every two weeks) for a total of 9 doses; secondary objectives are to evaluate whether developed antibodies impact pharmacokinetics (PK) and pharmacology. In this study, no Hematide-specific antibodies were detected. Hematide exhibits a prolonged plasma half-life and slow clearance by either IV or SC administration. Hematide induced significant erythropoiesis with reticulocytosis and subsequent increases in red blood cells, hematocrit and hemoglobin (Hgb) levels. No erythropoietic differences were noted between the IV and the SC dosed groups with mean +/- SD Hgb levels of 20.9 +/- 2.5 and 20.3 +/- 2.1 g/dL, respectively, occurring on Day 48, corresponding to Hgb increases of 6.5 and 6.7 g/dL, respectively, over pre-dose levels. In conclusion, Hematide is a potent erythropoiesis stimulating agent that exhibits plasma persistence in monkeys. Similar erythropoietic responses were produced following IV and SC administration. The absence of antibody development suggests that Hematide, at the doses and regimen described, has a low immunogenic potential in cynomolgus monkeys.

Tolerability, pharmacokinetics and concentration-dependent hemodynamic effects of oral CF101, an A3 adenosine receptor agonist, in healthy young men.

OBJECTIVES: To assess safety, tolerability, pharmacokinetics and hemodynamic effects of oral CF 101, an A3 adenosine receptor (A3AR) agonist, in healthy men. METHODS: One single and 1 repeated dose, parallel-group, ascending dose, double-blind and placebo-controlled study in normal volunteers. In the single dose study, n = 15 subjects received 1, 5 or 10 mg oral CF101; in each group 1 subject received placebo, the remainder active CF101. In the repeat-dose study, n = 28 subjects received repeated 12-hourly oral doses of CF 101 (2, 3, 4 or 5 mg) for 7 days, in each group 2 subjects received placebo, the remainder active CF101. TEST MATERIALS: Single-dose study: CF101 in 30% Cremophor RH40. Multiple-dose sudy: CF101 in 0.5% methylcellulose suspension. Both studies: the corresponding vehicles were used as placebos. Galenicals were prepared remotely from the clinical study site to ensure double-blind nature of the study. RESULTS TOLERABILITY: Single doses up to 5 mg CF101 were safe and well-tolerated. However, the single dose of 10 mg CF101 was associated with flushing, tachycardia, nausea and vomiting, which were viewed as dose-limiting in normal volunteers. Single doses of CF101 (as well as the first of the multiple doses) were associated with increases in heart rate (8 - 24 beats/min after 5 mg and 18 - 55 beats/min after 10 mg). Multiple doses up to 4 mg 12-hourly for 7 days were safe and well-tolerated. However, the 5 mg multiple-dose group reported headache, drowsiness, hot flushes and dizziness on standing; this declined with dosing duration and was not dose-limiting in this study. Adverse events were commonest near t(max). RESULTS PHARMACOKINETICS: For oral CF101, the t(max) was always 1 - 2 h post-dose and t 1/2 about 9 h, in both the single- and multiple-dose studies. For a single 5 mg dose (mean +/- SD) C(max) = 81.6 +/- 23.6 ng/ml in the single dose study, and 63.6 +/- 22.0 ng/ml after the first of the multiple doses; AUC if was 904.0 +/- 221.9 ng.h/ml and 596.1 +/- 196.6 ng.h/ml for the 2 studies, respectively. After 7 days of multiple dosing there was little change, and AUC(0-24h) = 601.0 +/- 163.6 ng.h/ml. These pharmacokinetic parameters were linearly proportional to dose in the other treatment groups. RESULTS PHARMACODYNAMICS: Increases in heart rate were related to plasma concentration and evident only in the upper range of concentrations observed. There were no changes on ECG monitoring beyond sinus tachycardia, and, in particular, no evidence of PR prolongation in any subject (n = 43). In comparison with single doses, this response was almost absent after 7 days of dosing. Leucocytosis (increases up to about 1.5 x 10(9)/l after 5 and 10 mg) was similarly transient and reversible after multiple dosing. CONCLUSIONS: Single oral doses up to 5 mg CF101 and repeated doses up to 4 mg 12-hourly for 7 days were safe and well-tolerated. Multiple-dose CF101 pharmacokinetics were unchanged and predictable from single-dose estimates, and were linearly proportional to dose. Increases in heart rate and neutrophil count were reversible during multiple dosing and were not dose-limiting in the repeat dose study. CF101 warrants further study for its efficacy in treating human disease.

A fast algorithm for designing stack filters.

Stack filters are a class of nonlinear filters with excellent properties for signal restoration. Unfortunately, present algorithms for designing stack filters can only be used for small window sizes because of either their computational overhead or their serial nature. This paper presents a new adaptive algorithm for determining a stack filter that minimizes the mean absolute error criterion. The new algorithm retains the iterative nature of many current adaptive stack filtering algorithms, but significantly reduces the number of iterations required to converge to an optimal filter. This algorithm is faster than all currently available stack filter design algorithms, is simple to implement, and is shown in this paper to always converge to an optimal stack filter. Extensive comparisons between this new algorithm and all existing algorithms are provided. The comparisons are based both on the performance of the resulting filters and upon the time and space complexity of the algorithms. They demonstrate that the new algorithm has three advantages: it is faster than all other available algorithms; it can be used on standard workstations (SPARC 5 with 48 MB) to design filters with windows containing 20 or more points; and, its highly parallel structure allows very fast implementations on parallel machines. This new algorithm allows cascades of stack filters to be designed; stack filters with windows containing 72 points have been designed in a matter of minutes under this new approach.

An overview of the physiological effects of sustained high +Gz forces on human being.

In the search for the advantage in aerial combat, increasingly higher performance aircraft are being developed that impose ever greater acceleration force loads on the human pilot. The symptomatology associated with increasing +Gz acceleration forces, culminating in G-induced loss of consciousness caused by the cessation of cerebral blood circulation are described. The inability of the normal cardiovascular system mechanisms to provide adequate circulation to the brain and the eyes, and the resulting associated symptomatology and physiological changes are discussed. The normal weight, hydrostatic pressure and physiological ventilation/perfusion gradients in the lungs are exaggerated under high +Gz forces resulting in increased pulmonary arterio-venous shunting. This causes impairment of circulatory oxygenation and may also result in acceleration induced atelectasis. The effects of +Gz acceleration forces on the renal system and the limitations of the cervical musculature are also discussed. This paper serves to describe the human physiological responses to increased +Gz acceleration forces in an attempt to provide a better understanding of the body's reaction to such demanding physical stressors.

Nonpeptide endothelin receptor antagonists. VI: Pharmacological characterization of SB 217242, a potent and highly bioavailable endothelin receptor antagonist.

This study describes the pharmacological characterization of SB 217242, a highly potent orally bioavailable nonpeptide antagonist of both endothelin type A (ETA) and endothelin type B (ETB) receptors. In human cloned ETA and ETB receptors, SB 217242 produced concentration-dependent displacement of [125]I-endothelin-1 (ET-1) in both receptor subtypes with Ki values of 1.1 and 111 nM, respectively. SB 217242 produced concentration-dependent, parallel rightward shifts in the ET-1 concentration-response curves in rat isolated aorta and human isolated pulmonary artery (ETA receptor-mediated vascular contraction) with Kb values of 4.4 and 5.0 nM, respectively. SB 217242 was 4-, 62- and 125-fold more potent as an ETA receptor antagonist than the previously reported compounds BQ-123, PD 142893 and Ro 46-2005, respectively. SB 217242 (10 microM) did not produce significant effects against contraction produced by other vasoactive agents. SB 217242 produced concentration-dependent antagonism of responses produced by ETB receptor activation as demonstrated by antagonism of sarafotoxin S6c-mediated contraction in the rabbit isolated pulmonary artery with a Kb value of 352 nM. In vitro cell monolayers of Caco-2 cells had high permeability to SB 217242. In vivo pharmacokinetics in the rat confirmed that SB 217242 was rapidly absorbed from the gastrointestinal tract with a bioavailability of 66%. The SB 217242 plasma half-life in rats after intraduodenal administration was 3.3 hr, with a systemic clearance of 27.3 ml/min/kg. Orally administered SB 217242 (0.3-30 mg/kg) produced dose-dependent inhibition of the pressor response to exogenous ET-1 in conscious rats; with a dose of 30 mg/kg p.o., inhibition was observed for at least 5.5 hr. The present study demonstrates that SB 217242 is a highly potent antagonist of both ETA and ETB receptors. In addition, SB 217242 has high in vitro permeability and high oral bioavailability. SB 217242 represents a new orally active pharmacological tool that should assist in the elucidation of the chronic role of endothelin in pathophysiology.

Aspiration-induced lung injury: role of complement.

OBJECTIVES: To examine the role of complement in the development of acid aspiration-induced lung injury in the rat. It was postulated that inhibition or depletion of complement attenuates aspiration-induced lung injury. DESIGN: Controlled animal trial. SETTING: Animal Laboratory, Jefferson Medical College, Philadelphia, PA. SUBJECTS: Anesthetized rats. INTERVENTIONS: Aspiration was induced by the intratracheal administration of 0.2 mL of 0.1 N hydrochloric acid (n = 7) and lung injury was evaluated by determining water content, myeloperoxidase activity, protein concentration, and leukocyte count in bronchoalveolar lavage fluid. Muscle PO2 was directly measured using a thin-film chamber oxygen sensor and serum tumor necrosis factor-alpha was assayed by enzyme-linked immunosorbent assay. The effect of complement inhibition by recombinant human soluble complement receptor type 1 (n = 8) or complement depletion by cobra venom factor (n = 7) on lung injury was evaluated. MEASUREMENTS AND MAIN RESULTS: Acid aspiration induced pulmonary leukosequestration, edema, and a microvascular permeability defect, along with tissue hypoxia. Pretreatment with soluble complement receptor type 1 (complement inhibition) or cobra venom factor (complement depletion) significantly reduced lung edema (-61 +/- 7%; p < .05), eliminated protein accumulation in bronchoalveolar lavage fluid (p < .01), and improved (p < .05) tissue oxygenation. In contrast, there was no effect of soluble complement receptor type 1 or of cobra venom factor on leukosequestration. CONCLUSIONS: Acid aspiration induces lung injury through a complement-dependent mechanism that leads to microvascular permeability defects. Therefore, the possibility that complement inhibitors may have a salutary effect in humans with aspiration-induced lung injury should be investigated.

Characterization of the metabolites of the peptidomimetic human immunodeficiency virus type 1 protease inhibitor SK&F 107461 in rats using liquid chromatography/mass spectrometry.

The metabolic fate of SK&F 107461 [Cbz-Ala-Ala-Phe psi [CHOHCH2] Gly-Val-Val-OMe], a potent and specific inhibitor of the protease encoded by human immunodeficiency virus type 1, in male Sprague-Dawley rats is described. SK&F 107461 is a hexapeptide analog containing a hydroxyethylene linkage in place of one of the peptide bonds, and in which the amino terminus is blocked with a carbobenzyloxy group and the carboxy terminus is modified to a methyl ester. The major metabolites of SK&F 107461 found in bile and urine after intravenous administration of 3H-labeled compound were characterized by LC/MS using either thermospray or continuous flow/FAB models of ionization. Approximately 80% of the administered radioactivity was recovered in the bile of bile duct-exteriorized rats following an intravenous dose. Radiochromatographic profiling indicated that SK&F 107461 was subject to extensive biotransformation. Structures were determined for three major biliary and five major urinary metabolites. Two of the major circulating plasma metabolites observed after intravenous bolus administration had similar retention times to metabolites that were observed in both bile and urine. A pathway for the biotransformation of SK&F 107461 in the rat is proposed. The parent molecule underwent two primary modes of metabolism. Hydrolysis of the carboxy-terminal ester or hydrolysis of the Ala-Ala peptide bond near the amino terminus were the primary metabolic events. All of the other metabolites characterized can be accounted for by exopeptidase activity subsequent to one or both of these primary events. There were no major metabolites observed resulting from anything other than hydrolysis of the ester or peptide bonds in the parent molecule.

Disposition of growth hormone-releasing peptide (SK&F 110679) in rat and dog following intravenous or subcutaneous administration.

The disposition of growth hormone releasing peptide (SK&F 110679) has been studied in male Sprague-Dawley rats and in male and female beagle dogs following intravenous (iv) and subcutaneous (sc) administration. Mass balance/excretion of [3H]SK&F 110679 was assessed in bile duct-exteriorized rats from which radiolabeled biliary and urinary excreta were quantified and characterized. [3H]SK&F 110679 was excreted, predominantly in the bile, and to a large extent as intact peptide following either iv or sc administration. Although the extent of biliary excretion of radiolabel was similar following iv or sc administration (60-70% of the dose), the rate was significantly higher following iv administration. Using a specific plasma HPLC/fluorescence assay, the iv and sc pharmacokinetics of SK&F 110679 were investigated in both species. Following iv bolus administration, biphasic plasma concentration-time profiles were observed, and the initial phases were characterized by 2-4 min half-lives. Systemic plasma clearance was 27 ml/min/kg in the rat (0.4 mg/kg dose) and 17 ml/min/kg in the dog (0.5 mg/kg dose). High sc bioavailability (89-103%) was observed in both species; an apparent terminal half-life of 1 hr likely reflected slow absorption from the injection site.

Disposition of metabolically labeled recombinant soluble CD4 (sT4) in male Sprague-Dawley rats following intravenous and subcutaneous administration.

Soluble CD4 (sT4) has been metabolically labeled with [3H]leucine in Chinese hamster ovary cells and purified by S Sepharose chromatography. Over 250 microCi of high specific radioactivity [3H]sT4 (42 Ci/mmol) was prepared. The radiolabeled molecule was chemically and biologically representative of the unlabeled molecule and thus appropriate for in vivo metabolic investigations. To explore the biotransformation and disposition of a recombinant protein, this uniformly labeled [3H]sT4 was administered intravenously and subcutaneously to male Sprague-Dawley rats. Following a single dose of 0.3 mg/kg, blood samples were collected for 9 days and analyzed for total radioactivity, total plasma radioactivity, trichloroacetic acid-precipitable plasma radioactivity, sT4-related plasma radioactivity (by extraction with a Sepharose-bound polyclonal anti-sT4 antibody), and plasma sT4 concentration (by an N and C terminal-specific Leu3A/OKT4 ELISA). Excreta were analyzed for total radioactivity. The pharmacokinetic profiles of intact sT4 were as expected from the results of previous studies. sT4 was cleared rapidly from plasma with an elimination t1/2 of 7 min (intravenous), and low sT4 levels were observed following subcutaneous administration. Comparison of the kinetic profiles of total radiolabel, trichloroacetic acid-precipitable radiolabel, sT4-related radiolabel, and the isolation of plasma proteins containing tritium have led to the following conclusions. One of the major metabolic pathways for [3H]sT4 was the degradation of the polypeptide to its constituent amino acids, which were subsequently incorporated into endogenous proteins. Incorporation of tritium into blood cell proteins resulted in a prolonged radiolabel blood profile (t1/2 greater than 250 hr). Following subcutaneous administration, [3H] sT4 was significantly degraded before reaching the vascular circulation.

Role of complement in endotoxin/platelet-activating factor-induced lung injury.

C receptor-1 is a protein involved in the regulation of C3 and C5-convertases. Recombinant human soluble C receptor-1 has recently been produced and shown to reduce infarct size in a rat model of myocardial ischemia/reperfusion injury. The present study aimed to investigate whether recombinant human soluble C receptor-1 exerts any protective effect on pulmonary injury produced in a rodent model of adult respiratory distress syndrome. In this model, Escherichia coli endotoxin (LPS, 0.1 microgram/kg) combined with platelet-activating factor (1 pmol/kg/min over 60 min, n = 10) caused microvascular lung injury characterized by elevation of myeloperoxidase activity, deposition of C3 and C5b-9 on the endothelium of pulmonary vessels, and pulmonary edema. Furthermore, bronchoalveolar lavage revealed increased neutrophil count and elevated protein concentration. These pulmonary responses were associated with elevated serum TNF-alpha. Pretreatment (10 min, i.v.) with recombinant human soluble C receptor-1 at 10 mg/kg (n = 13), but not at 1 mg/kg, prevented the LPS/platelet-activating factor-induced pulmonary edema (p less than 0.01) and changes in the bronchoalveolar lavage fluid cell count (p less than 0.01) and protein concentration (p less than 0.05), and attenuated the deposition of C3 and C5b-9 to lung vessels. There was no effect on lung myeloperoxidase activity and serum TNF-alpha. Also, C depletion by cobra venom factor (500 U/kg, i.v.) eliminated the pulmonary edema and elevated leukocyte count in bronchoalveolar lavage fluid, but had no effect on lung myeloperoxidase activity and serum TNF-alpha. These data suggest that C factors may play an important role in the pathophysiology of adult respiratory distress syndrome.

Pharmacokinetic evaluation of drug interactions with anti-human immunodeficiency virus drugs. V. Effect of soluble CD4 on 2',3'-dideoxycytidine kinetics in monkeys.

This study was conducted to determine if soluble CD4 (ST4) altered the pharmacokinetics of 2',3'-dideoxycytidine (ddC) in nonhuman primates. Each of six monkeys received 5 mg/kg of ddC iv in the absence and presence of two different iv regimens of ST4. The ST4 regimens produced steady-state plasma concentrations of 10.3 micrograms/ml (N = 3) and 22.2 micrograms/ml (N = 3) for 30 min following ddC administration. Pharmacokinetic parameters for ddC and ST4 were calculated based on plasma and urine concentrations of ddC and plasma concentrations of ST4. Following combined ddC and ST4 administration, in both the low- and high-dose ST4 groups, plasma concentration-time profile of ddC were similar for each monkey, and no statistical differences were observed in the pharmacokinetic parameters compared with those obtained when ddC was given alone. Complete urinary excretion data for ddC was obtained in 3 of the 6 animals studied. At the low ST4 dose, one animal had a reduced renal clearance of ddC, whereas at the high ST4 dose two animals recorded an increased renal clearance of ddC. ST4 plasma concentrations were comparable to in vitro concentrations of antiviral activity, with pharmacokinetic parameters similar to those reported previously. The kinetic information provides a basis for rational dosage design for combination chemotherapeutic regimens of ddC and ST4 in human immunodeficiency virus infection.

Comparison of dosage schedules of rt-PA in the treatment of proximal deep vein thrombosis.

This study assessed the efficacy and safety of increasing durations of constant-dose intravenous recombinant tissue-type plasminogen activator (rt-PA) in the treatment of deep vein thrombosis. Patients with venogram-documented proximal lower limb (popliteal, iliofemoral) or upper limb (axillary, subclavian) thrombi were given an initial 2-hour rt-PA infusion at 4 micrograms/kg/min, followed by a maintenance infusion of 1 microgram/kg/min for an additional 4, 22, or 33 hours (mean total rt-PA dosages of 54, 127, and 185 mg). A new quantitative venogram scoring system was applied to the study, based on measurements of thrombus volume before and after completion of treatment. Whereas none of the seven patients given treatment for 6 hours and only one of four given treatment for 24 hours showed significant lysis, four of seven who received a prolonged infusion for 35 hours showed lysis of more than 40% of the original thrombus. Overall, the prolonged 35-hour infusion induced 51% lysis of original thrombus, representing a thrombus volume of 16.7 ml dissolved. Hemorrhagic complications were common in all three groups, with four of 18 patients having significant bleeding, including one massive gastrointestinal hemorrhage, two patients with a decrease in hematocrit of more than 10%, and one patient with an intracranial hemorrhage who recovered completely. Pharmacokinetics of the rt-PA showed a steady state antigen concentration of 240 ng/ml and activity of 200 IU/ml during the initial 2-hour infusion and a postinfusion half-life of 5 minutes. Plasma fibrinogen concentrations decreased to approximately 40% to 50% of initial values with all three treatment regimens, but the nadir fibrinogen concentrations did not correlate with either therapeutic efficacy or bleeding complications. One patient with systemic lupus erythematosus had an unusual allergic reaction that manifested primarily as angioedema. This study suggests that rt-PA infusion of 35 hours induces greater thrombolysis of deep vein thrombosis than does a shorter course of 6 or 24 hours, without an increase in hemorrhagic complications.

Myeloma-like cast nephropathy caused by human recombinant soluble CD4 (sCD4) in monkeys.

CD4 is the receptor for human immunodeficiency virus (HIV) on lymphocytes and macrophages. Soluble CD4 (sCD4), a recombinant truncated form of CD4, has been shown to inhibit HIV-1 in vitro and is being tested as a therapy for AIDS. Preclinical studies in cynomolgus monkeys revealed a protein cast nephropathy after four daily intravenous doses of 100 mg/kg/day. Renal lesions were not found in monkeys that received 10 mg/kg/day. The renal lesions consisted of proteinaceous tubular casts associated with multinucleate giant cells and neutrophils located in the tubules of the distal nephron. The affected tubules were surrounded by an interstitial mixed inflammatory cell infiltrate. By electron microscopy, the casts were composed of moderately electron dense, paracrystalline material. Immunostaining demonstrated that the casts contained sCD4-derived material and Tamm-Horsfall protein. Moreover, biochemical analysis of urine showed that a portion of sCD4 was excreted as intact protein. Because infection with HIV-1 can be associated with clinically significant nephropathy, these data suggest that renal function should be closely monitored in patients receiving soluble forms of CD4.

Internalization of recombinant tissue-type plasminogen activator by isolated rat hepatocytes is via coated pits.

The uptake and internalization of tissue-type plasminogen activator (t-PA) by freshly isolated rat hepatocytes was investigated. Electron microscopic examination of the uptake of t-PA-colloidal gold conjugates (t-PA-gold) by isolated rat hepatocytes showed that t-PA-gold was internalized via coated pits. This was inhibited with excess t-PA. Uptake of 125I-t-PA by isolated rat hepatocytes was a rapid, saturable, and specific process. The initial rate of specific uptake was 0.1 fmol/10(6) cells per min. The specific uptake plateaued at 1.4 fmol/10(6) cells by 30 min and declined to 0.8 fmol/10(6) cells at 2 h. Depletion of cellular ATP by 85-90% did not affect the initial rate of specific uptake. However, specific uptake by ATP-depleted hepatocytes at 30 min was reduced by 37%. By 2 h specific uptake by ATP-depleted hepatocytes was only 5% lower than by untreated hepatocytes, suggesting that processing of t-PA and/or its receptor is ATP-dependent. Uptake of 125I-t-PA was temperature dependent. Specific uptake was reduced by approximately 20% at 22 degrees C and by 70% at temperatures below 16 degrees C. Finally, inhibition of coated pit formation by K(+)-depletion with nigericin decreased the uptake of 125I-t-PA. This inhibition was shown to be K(+)-specific since treatment with nigericin in the presence of K+ did not inhibit coated pit formation or 125I-t-PA uptake. A threshold K(+)-depletion level for inhibition of coated pit formation was also demonstrated since treatment under conditions that reduced cellular K+ by only 54% had no effect on coated pit formation or 125I-t-PA uptake. These data support our hypothesis that internalization of t-PA by isolated rat hepatocytes is via coated pits and suggest that uptake of t-PA is a receptor-mediated process.

Uptake of human recombinant tissue-type plasminogen activator by rat hepatocytes in vivo: an electron microscope autoradiographic study.

The hepatic uptake of recombinant human tissue-type plasminogen activator (tPA) has been studied by electron microscope autoradiography (EMARG) of serial hepatic biopsies taken from anaesthetized, laparotomized rats following intravenous injection of 125I labeled tPA. Serial blood samples showed both radiolabel and biologic activity to be eliminated from circulation with an initial half-life of approximately two minutes. Grain half-distance distribution profiles and grain density analysis showed that the para-sinusoidal region of the hepatic parenchymal cell is the only site in the liver to concentrate radiolabeled tPA after intravenous injection. These data support the hypothesis that the parenchymal cell is the principal cell responsible for hepatic clearance of tPA from circulation and suggest that receptor mediated endocytosis may be the mechanism of cellular uptake.

Evaluation of the acute hemodynamic effects and pharmacokinetics of coronary thrombolysis produced by intravenous tissue-type plasminogen activator in the anesthetized dog.

The thrombolytic dose-response effectiveness and pharmacokinetics of tissue-type plasminogen activator (rt-PA) was evaluated in anesthetized, open-chest dogs instrumented for the measurement of systemic hemodynamics. Intracoronary thrombi were formed by injecting thrombin (100 U) and CaCl2 (50 microM) into a cannulated, isolated segment of the left anterior descending coronary artery (LAD). Coronary blood flow was measured by placing an electromagnetic flow probe proximal to the LAD thrombus. Thirty minutes after formation of a stable LAD thrombus, intravenous infusion of rt-PA was given at rates of 0.5, 1, 2, 4, or 8 micrograms/kg/min (n = 8/dose) for 60-90 min, and the animals were followed for an additional 30 min. In vehicle-treated animals, residual thrombus wet weight, determined at the end of the experiment, was 30 +/- 4 mg (mean +/- SEM, n = 8) and spontaneous reperfusion did not occur. The rt-PA produced a dose-related increase in the number of animals reperfusing, a decrease in the time to reperfusion, and a decrease in residual thrombus weight, but had no effect on systemic hemodynamics. The increase in infusion rate from 0.5 to 4 micrograms/kg/min resulted in a linear increase in both the steady-state rt-PA plasma concentration and the area under the rt-PA plasma concentration versus time curve (n = 3-5 animals/dose); between the infusion rates of 4 and 8 microgram/kg/min there was a disproportionate increase in both these parameters that was due to a decrease in the total systemic clearance of rt-PA. The postdosing elimination half-life (t1/2 alpha) did not differ significantly at any dose of rt-PA, and the pooled half-life (t1/2 alpha) for all doses of rt-PA was 2.36 +/- 0.12 min (n = 19).(ABSTRACT TRUNCATED AT 250 WORDS)

Dose-dependent pharmacokinetics of recombinant tissue-type plasminogen activator in anesthetized dogs following intravenous infusion.

The pharmacokinetics of SK&F recombinant two-chain tissue-type plasminogen activator (tPA) following intravenous (iv) infusion were characterized in anesthetized, open chested mongrel dogs in which artificial intracoronary thrombi were formed. SK&F tPA was infused at rates of 0.5, 1, 2, 4, and 8 micrograms/kg/min (N = 3 to 5 per dose) for 90 min, and arterial blood samples were withdrawn during and after infusion for determination of functionally active tPA concentrations using a modified and validated S-2251 chromogenic assay. At all doses studied, steady state active tPA plasma concentrations were achieved 10-20 min after the onset of infusion. Upon cessation of infusion, active tPA plasma concentrations declined rapidly with a t1/2 of 2-3 min. The active tPA plasma concentration at steady state (Css) and the area under the tPA plasma concentration-time curve (AUC) increased linearly with the dose in the range of 0.5-4 micrograms/kg/min. However, as the dose was increased 2-fold from 4 to 8 micrograms/kg/min, the AUC and the Css increased 2.5-fold. The systemic clearance ranged from 15-16 ml/min/kg at doses of 0.5-4 micrograms/kg/min, but decreased to 11.7 ml/min/kg at the 8 micrograms/kg/min dose. With exceptions in three dogs, the volume of distribution at steady state approached or slightly exceeded the blood volume. Plasma tPA antigen concentrations were also determined in the dogs receiving the 2 micrograms/kg/min dose. At steady state, active tPA accounted for 40-60% of the total tPA antigen. The postinfusion t1/2 of the tPA antigen was considerably longer (13.46 +/- 5.94 min) than that of active tPA.(ABSTRACT TRUNCATED AT 250 WORDS)